Artykuły w czasopismach na temat „12/14/12-helix”

The Orsiro device is a hybrid drug-eluting stent that represents a new strategy in the treatment of coronary artery stenosis. Orsiro features a hybrid coating of passive and active components: the PROBIO passive coating seals the metal surface of the stent and prevents interaction with the surrounding blood and tissue, while the BIOlute active coating contains a highly biocompatible polymer that delivers a -limus drug over 12–14 weeks and degrades gently over one to two years, thereby avoiding increased inflammation. The stent backbone is the PRO-Kinetic Energy platform, which has a double-helix stent design and thin struts, bringing flexibility and ease of deliverability.

We have isolated and characterized four collagen-related c-DNA clones (N-COL 1, N-COL 2, N-COL 3, N-COL 4) that are highly expressed in developing nematocytes in hydra. All four c-DNAs as well as their corresponding transcripts are small in size (600-1,000 bp). The deduced amino acid sequences show that they contain a central region consisting of 14 to 16 Gly-X-Y triplets. This region is flanked amino-terminal by a stretch of 14-23 proline residues and carboxy-terminal by a stretch of 6-9 prolines. At the NH2- and COOH-termini are repeated patterns of cysteine residues that are highly conserved between the molecules. A model is proposed which consists of a central stable collagen triple helix of 12-14 nm length from which three 9-22 nm long polyproline II type helices emerge at both ends. Disulfide linkage between cysteine-rich segments in these helices could lead to the formation of oligomeric network structures. Electrophoretic characterization of nematocyst extracts allows resolution of small proline-rich polypeptides that correspond in size to the cloned sequences.

Annexin A11 is one of the 12 vertebrate subfamilies in the annexin superfamily of calcium/phospholipid-binding proteins, distinguishable by long, non-homologous N-termini rich in proline, glycine and tyrosine residues. As there is negligible structural information concerning this annexin subfamily apart from primary sequence data, we have cloned, expressed and purified recombinant mouse annexin A11 to investigate its structural and functional properties. CD spectroscopy reveals two main secondary-structure contributions, α-helix and random coil (approx. 30% each), corresponding mainly to the annexin C-terminal tetrad and the N-terminus respectively. On calcium binding, an increase in α-helix and a decrease in random coil are detected. Fluorescence spectroscopy reveals that its only tryptophan residue, located at the N-terminus, is completely exposed to the solvent; calcium binding promotes a change in tertiary structure, which does not affect this tryptophan residue but involves the movement of approximately four tyrosine residues to a more hydrophobic environment. These calcium-induced structural changes produce a significant thermal stabilization, with an increase of approx. 14 °C in the melting temperature. Annexin A11 binds to acidic phospholipids and to phosphatidylethanolamine in the presence of calcium; weaker calcium-independent binding to phosphatidylserine, phosphatidic acid and phosphatidylethanolamine was also observed. The calcium-dependent binding to phosphatidylserine is accompanied by an increase in α-helix and a decrease in random-coil contents, with translocation of the tryptophan residue towards a more hydrophobic environment. This protein induces vesicle aggregation but requires non-physiological calcium concentrations in vitro. A three-dimensional model, consistent with these data, was generated to conceptualize annexin A11 structure–function relationships.

Inhibitor of differentiation-2 (Id2) is a basic helix-loop-helix protein that acts as a negative regulator of the myogenic regulatory transcription factor family, but Id2 has also been implicated in apoptosis in several cell lines. In this study, we tested the hypothesis that Id2 has a role in both apoptosis-associated muscle atrophy and muscle hypertrophy. A weight corresponding to 12% of the body weight was attached to one wing of Japanese quail to induce hypertrophy in the patagialis (PAT) muscle. Birds in group 1 were killed after 5 ( n = 8), 7 ( n = 10), or 14 days ( n = 10) of loading. The left wing was loaded for 14 days in group 2 birds, and then the weight was removed and the PAT was examined after 7 ( n = 10), 14 ( n = 10), or 21 ( n = 5) days of unloading. A time-released bromodeoxyuridine (BrdU) pellet was implanted subcutaneously with wing weighting to identify activated satellite cells during loading. The left wing was loaded for 14 days, unloaded for 14 days, and then the weight was reattached for a subsequent 7 ( n = 10) or 14 days ( n = 10) in group 3 birds. BrdU was implanted on the second loading phase in this group. Id2 mRNA as measured by kinetic PCR increased by 3.9-, 2.7-, and 1.6-fold, relative to control levels after 7, 14, and 21 days of unloading ( group 2). Id2 protein as estimated by Western blots increased by 1.5-, 1.4-, and 0.75-fold after 7, 14, and 21 days of unloading ( group 2). Muscle unloading induced apoptosis, because poly(ADP-ribose) polymerase-(PARP)-positive nuclei increased and caspase 8 levels increased by 2.6- and 1.7-fold after 7 or 14 days of unloading, respectively ( group 2). Although BrdU-positive nuclei increased during loading ( groups 1 and 3), 50% failed to survive during unloading ( group 2). Id2 mRNA increased by 2.2- and 1.8-fold after 5 and 7 days of loading, respectively, but decreased to control levels by 14 days of loading in group 1. Id2 protein levels increased 2.1-fold after 5 days of loading ( group 1). In contrast, Id2 did not increase in reloaded muscles of group 3 birds. These data suggest that Id2 may have a role in apoptosis-associated atrophy of skeletal muscles, but its role in muscle hypertrophy is less clear.

To assess changes of metabolite content and regulation mechanism of the phenolic acid biosynthesis pathway at different developmental stages of leaves, this study performed a combined metabolome and transcriptome analysis of Cyclocarya paliurus leaves at different developmental stages. Metabolite and transcript profiling were conducted by ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometer and high-throughput RNA sequencing, respectively. Transcriptome identification showed that 58 genes were involved in the biosynthesis of phenolic acid. Among them, 10 differentially expressed genes were detected between every two developmental stages. Identification and quantification of metabolites indicated that 14 metabolites were located in the phenolic acid biosynthetic pathway. Among them, eight differentially accumulated metabolites were detected between every two developmental stages. Association analysis between metabolome and transcriptome showed that six differentially expressed structural genes were significantly positively correlated with metabolite accumulation and showed similar expression trends. A total of 128 transcription factors were identified that may be involved in the regulation of phenolic acid biosynthesis; these include 12 MYBs and 10 basic helix–loop–helix (bHLH) transcription factors. A regulatory network of the phenolic acid biosynthesis was established to visualize differentially expressed candidate genes that are involved in the accumulation of metabolites with significant differences. The results of this study contribute to the further understanding of phenolic acid biosynthesis during the development of leaves of C. paliurus.

We report the synthesis, characterization, X-ray crystal structure and Hirshfeld surface analysis of Ni(II) perchlorate complex (1, Ni2L3·4ClO4·2CH3CN) of 1,2-bis(pyridin-2-ylmethylene)hydrazine (L) ligand. The X-ray crystallographic study of complex 1 reveals that in the presence of Ni(II) ions,the ligand L forms a dimeric triple helix with a Ni(II)-Ni(II) distance of 3.794 Å. Crystal data for C40H36Cl4N14Ni2O16: Monoclinic, space group P21/c (no. 14), a = 20.7558(19) Å, b = 13.1937(12) Å, c = 20.0181(18) Å, β = 96.9510(10)°, V = 5441.6(9) Å3, Z = 4, T = 293.15 K, μ(MoKα) = 0.965 mm-1, Dcalc = 1.498 g/cm3, 38075 reflections measured (1.976° ≤ 2Θ ≤ 43.728°), 6557 unique (Rint = 0.0695, Rsigma = 0.0466) which were used in all calculations. The final R1 was 0.0518 (I > 2σ(I)) and wR2 was 0.1270 (all data). The Hirshfeld surface analysis of complex 1 shows that C···H, H···H, N···H and O···H interactions of 10.9, 26.4, 6.7, and 33.4%; respectively, which exposed that the main intermolecular interactions were H···H intermolecular interactions.

In nerve and muscle cells, the voltage-gated opening and closing of cation-selective ion channels is accompanied by the translocation of 12–14 elementary charges across the membrane's electric field. Although most of these charges are carried by residues in the S4 helix of the gating module of these channels, the precise nature of their physical movement is currently the topic of spirited debate. Broadly speaking, two classes of models have emerged: those that suggest that small-scale motions can account for the extensive charge displacement, and those that invoke a much larger physical movement. In the most recent incarnation of the latter type of model, which is based on structural and functional data from the archaebacterial K+ channel KvAP, a “voltage-sensor paddle” comprising a helix-turn-helix of S3–S4 translocates ∼20 Å through the bilayer during the gating cycle (Jiang, Y., A. Lee, J. Chen, V. Ruta, M. Cadene, B.T. Chait, and R. MacKinnon. 2003. Nature. 423:33–41; Jiang, Y., V. Ruta, J. Chen, A. Lee, and R. MacKinnon. 2003. Nature. 423:42–48.; Ruta, V., J. Chen, and R. MacKinnon. 2005. Cell. 123:463–475). We used two methods to test for analogous motions in the Shaker K+ channel, each examining the aqueous exposure of residues near S3. In the first, we employed a pore-blocking maleimide reagent (Blaustein, R.O., P.A. Cole, C. Williams, and C. Miller. 2000. Nat. Struct. Biol. 7:309–311) to probe for state-dependent changes in the chemical reactivity of substituted cysteines; in the second, we tested the state-dependent accessibility of a tethered biotin to external streptavidin (Qiu, X.Q., K.S. Jakes, A. Finkelstein, and S.L. Slatin. 1994. J. Biol. Chem. 269:7483–7488; Slatin, S.L., X.Q. Qiu, K.S. Jakes, and A. Finkelstein. 1994. Nature. 371:158–161). In both types of experiments, residues predicted to lie near the top of S3 did not exhibit any change in aqueous exposure during the gating cycle. This lack of state dependence argues against large-scale movements, either axially or radially, of Shaker's S3–S4 voltage-sensor paddle.

Abstract The t(12;13)(p11-13:q11-14) is a recurring chromosomal abnormality found in different types of hematological malignancies. Three genes on 13q12-14, CDX2, FLT3, and TTL have been identified as fusion partners of ETV6(TEL) on 12p13. These genes create different forms of fusion transcripts with ETV6. In the present study, we identified novel types of ETV6-TTL fusion transcripts in a case of a 9 year-old female who developed myeloid/NK cell precursor acute leukemia with t(12;13)(p13;q14). Cytogenetic analyses of the leukemic cells of the patient using a regular G-banding revealed 47, XX, del(2)(q?), -9, der(12)t(12;13)(p13;q14), add(13)(q1?2), +mar1, +mar2 [13/20]. Fluorescence in situ hybridization using YAC clone revealed that ETV6 gene was split in metaphase chromosomes of patient’s leukemic cells. We performed reverse-transcriptase-polymerase chain reaction (RT-PCR) analysis to detect the fusion transcripts of ETV6-CDX2, ETV6-FLT3 or TTL-ETV6, however, no fusion transcripts of previously described types were detected. We next performed RT-PCR analysis using various sets of primers to detect unknown types of fusion transcripts involving these genes, and detected novel types of fusion transcripts of ETV6-TTL. These fusion transcripts consisted of exons 1 to 5 of ETV6 and exons 5 to 8 of TTL, and exons 1 to 6 of ETV6 and exon 9 or exons 8 to 9 of TTL. No reciprocal fusion transcripts were detected. Predicted fusion proteins consisted of N-terminal ETV6 lacking whole or part of ETS-binding domain and C-terminal TTL. Previous report showed major type of TTL-ETV6 fusion transcript consisted of exons 1 to 5 of TTL and exons 2 to 8 of ETV6 which contained both helix-loop-helix and ETS-binding domains. These results suggested that novel types of ETV6-TTL act as different fusion proteins from previously reported TTL-ETV6 in leukemogenesis. At the cytogenetic level, it may be difficult to distinguish ETV6-CDX2, ETV6-FLT3, and ETV6-TTL. Further accumulation of the patients with t(12;13) and further analysis of these novel types of fusion transcripts may clarify the pathogenesis of t(12;13)-leukemia.

Oligomers of -amino acids ( -peptides), which are readily available by standard meth­ ods either in solution or on solid support, adopt a large variety of different secondary structures in solution and in the solid state. -Peptides 4, 5 and 10 fold into a helix with 3 residues per turn and 14-membered H-bonded rings (314 helix) that is left-handed for 5 and 10 and right-handed for 2 (due to the reversal of the chirality of the building blocks), as was clearly demonstrated by two-dimensional NMR-spectroscopy. This helix thermally is very stable in methanol solution upon heating. As shown by NMR- and CO-spectroscopy, it is partially populated even at 1oo•c (Figure 3). Another helix was discovered for ,mixed' -peptide 8 in methanol solution: it is characterized by 12- and 10- membered turns (Figure 4, left) and its central 10-membered turn has been found in the solid state of a geminally disubtituted -peptide (Figure 4, right). This central 10-membered turn was used as a scaffold to attach -amino acid residues that prefer a linear (non-helical) conformation ( -peptide 21) : a hairpin (pleated sheet-turn-pleated sheet) structure was determined in solution by NMR-spectroscopy (Figure 5). In contrast to this antiparallel pleated-sheet, a parallel pleated sheet was found for a -tripeptide in the solid state. For the first time it was possible to observe reversible peptide folding in MD simulations by studying -peptides (Figure 6) and to determine folding pathways and intermediates. -Peptides are a new class of promising peptidomimetics. They are resistant against the. degradation by proteolytic enzymes such as pepsin, elastase, carboxypeptidase A, pronase or proteasom 208. A variety of -amino acids (27-34) was shown to be non- mutagenic by Ames' tests and -peptides 47 and 48 reveal large elimination half-lives of 3 h (for 47) and 10 h (for 48) in the serum of rodents (Figure 7). Conjugates of a- and - peptides are efficient ligands for the HLA*B27 MHC Class I protein, a five fold increase of binding (2.0 J.!M for 55) compared to a natural peptidic ligand 51 was observed. Furthermore, -peptides are able to mimic natural a.­ peptidic hormones such as somatostatin. The cyclo- -tetrapeptide 57 binds to the five human somatostatin receptors in the micromolar range. In addition, several other non-natural oligomers such as -peptide nucleic acids (built from 58 and 59), -peptoids (60), oligomers of anthranilic acids and -sulfonamide peptides are discussed.

The aim of this article is to present a clinical case report where a female patient, 62 years old, presented with element 14 poorly positioned and mobility. The initial planning was immediate implantation with immediate provisionalization. The case described was carried out with a minimally invasive extraction, implant placement of the 3.75 x 11.5 mm (Helix, GrandMorse, Acqua, Neodent®, Brazil), immediate loading with a universal healing abutment (3.3 x 2.5 mm, Neodent®, Brazil) and a provisional cylinder of the universal healing abutment click (3.3 x 6.0 mm, Neodent®, Brasil) to approach of capture from a denture teeth and thus, the cementation of provisional implant crown. For modification of the gingival area, a subepithelial connective tissue graft was performed through the tunneling technique to improve peri-implant tissue conditioning. A 6-month postoperative follow-up was performed, showing satisfactory clinical and radiographic conditions during the most important phase of healing, and finally, a final follow-up of 12 months with the definitive cemented prosthesis. According to the literature and clinical experience, rehabilitation with immediate implant placement is one of the most challenging procedures when associated with immediate loading, where many criteria must be evaluated before the surgical phase, it means, and excellent initial planning.

The bacterioferritin (BFR) of Escherichia coli is an iron-storage protein containing 24 identical subunits and between three and 11 protohaem IX groups per molecule. Titration with additional haem gave a maximum loading of 12-14 haems per molecule. The e.p.r. spectra and magnetic c.d. spectra of the protein-bound haem show it to be low-spin Fe(III), and coordinated by two methionine residues as previously reported for BFRs isolated from Pseudomonas aeruginosa and Azotobacter vinelandii [Cheesman, Thomson, Greenwood, Moore and Kadir, Nature (London) (1990) 346, 771-773]. A recent sequence alignment indicated that BFR may be structurally related to ferritin. The molecular model proposed for E. coli BFR has a four-alpha-helix-bundle subunit conformation and a quaternary structure similar to those of mammalian ferritins. In this model there are two types of hydrophobic pocket within which two methionine residues are correctly disposed to bind haem. The e.p.r. spectra also reveal a monomeric non-haem Fe(III) species with spin, S = 5/2. On the basis of sequence comparisons, a ferroxidase centre has recently been proposed to be present in BFR [Andrews, Smith, Yewdall, Guest and Harrison (1991) FEBS Lett. 293, 164-168] and the possibility that this Fe(III) ion may reside at or near the ferroxidase centre is discussed.

OBJECTIVE To determine the frequency and types of craniofacial abnormalities observed in patients with trisomy 18 or Edwards syndrome (ES). METHODS This descriptive and retrospective study of a case series included all patients diagnosed with ES in a Clinical Genetics Service of a reference hospital in Southern Brazil from 1975 to 2008. The results of the karyotypic analysis, along with clinical data, were collected from medical records. RESULTS: The sample consisted of 50 patients, of which 66% were female. The median age at first evaluation was 14 days. Regarding the karyotypes, full trisomy of chromosome 18 was the main alteration (90%). Mosaicism was observed in 10%. The main craniofacial abnormalities were: microretrognathia (76%), abnormalities of the ear helix/dysplastic ears (70%), prominent occiput (52%), posteriorly rotated (46%) and low set ears (44%), and short palpebral fissures/blepharophimosis (46%). Other uncommon - but relevant - abnormalities included: microtia (18%), orofacial clefts (12%), preauricular tags (10%), facial palsy (4%), encephalocele (4%), absence of external auditory canal (2%) and asymmetric face (2%). One patient had an initial suspicion of oculo-auriculo-vertebral spectrum (OAVS) or Goldenhar syndrome. CONCLUSIONS: Despite the literature description of a characteristic clinical presentation for ES, craniofacial alterations may be variable among these patients. The OAVS findings in this sample are noteworthy. The association of ES with OAVS has been reported once in the literature.

The seed metering process of a fluted force-feed seeder was simulated using the Discrete Element Method and its parameters optimized using the Box–Behnken Design of Experiments and the Response Surface Method. The rotational speed of the feed roller, the lead (helix) angle of the flutes, and the number of flutes were the independent variables, while the response value was the seeding uniformity index. Two regression models were investigated, and the following conclusions drawn. For the flute lead angle between 0 and 10 degrees, and the number of flutes between 10 and 14, it was found that the number of flutes and the lead angle influenced the seeding performance the most, with the order of importance being the (i) number of flutes, (ii) lead angle and (iii) roller speed. For the flute lead angle between 5 and 15 degrees, and the number of flutes between 12 and 16, it was found that the roller speed and the number of flutes influenced the seeding performance the most, with the order of importance being the (i) roller speed, (ii) number of flutes and (iii) flute lead angle. The two regression models were then minimized for the seeding uniformity index and the corresponding optima verified experimentally on a conveyor belt test stand fitted with an image recognition system.

Objectives: This study aims at the identification of the distribution of basal cell carcinomas (BCCs) in the auricle in correlation with the currently most credited sites of the embryonic fusion planes of the auricle. Methods: An overall number of 69 patients with 72 BCCs of the auricle were enrolled in the study over a period of 14 years, from June 2003 to October 2017. All the cases underwent medical preoperative digital photography and the specific location of each BCC was coded on an original full-size anatomical diagram of the auricle derived from the reports by Streeter, Wood-Jones, Park, Porter, and Minoux showing the currently most credited sites of the embryonic fusion planes arbitrarily featured as two 5-mm-wide ribbon-like areas: (1) the hyoid-mandibular fusion plane (HM-FP) running from the upper margin of the tragus toward the concha and then deflecting toward the lower margin of the tragus and (2) the free ear fold-hyoid fusion plane (FEFH-FP) running from the cranial-most portion of the helix to the mid-portion of the ascending helix. The latter fusion planes were comprehensively termed embryological fusion planes (EFP) while all of the remaining surface of the auricle was comprehensively termed non-fusion area (NFA). The surfaces of all of the latter areas were calculated using the ImageJ software. Results: According to our data, the greatest number of BCCs was observed within the currently most credited sites of the embryonic fusion planes of the auricle. The latter sites displayed a 12-fold increased tumor incidence in comparison with the remaining surface of the ear. Conclusions: A correspondence between the sites of onset of BCCs and the sites of merging and/or fusion of embryonal processes was demonstrated in the auricle. Therefore, the latter sites might be considered as high-risk areas for the development of a BCC. Such an evidence provides further support to the hypothesis of an embryological pathogenesis of BCC.

Abstract Non-Hodgkin’s lymphoma (NHL) is a heterogeneous clinicopathologic entity characterized by distinct cells of origin, cytogenetic and molecular aberrations. In vivo studies recently showed that mice deficient in Period (PER) 2 developed lymphomas. Furthermore, we and others have recently suggested that the core circadian clock genes Period (Per)1 and Per2 have been linked to DNA damage response pathways. We showed that both genes are involved in tumor suppression by regulating cell cycle- and apoptosis-related genes. In mammals and other vertebrates, heterodimers of the basic helix-loop-helix-PAS transcription factors, CLOCK and BMAL1, activate transcription of the Period and Cryptochrome (Cry1 and Cry2) genes via binding to E-box sequences in their promoters. Subsequently, the PER and CRY repressor proteins accumulate in the nucleus and inhibit CLOCK:BMAL1 complexes, thereby inhibiting their own gene expression. This forms the major negative circadian feedback loop. In the present study, we focused on Per1 and Per2 and their role in different types of NHL. Real-time reverse transcriptase-polymerase chain reaction (real time RT-PCR) was performed to determine the mRNA expression levels of Per1 and Per2 in patients with diffuse large B-cell lymphoma (DLBCL; n = 37), mantle cell lymphoma (MCL; n = 12), follicular lymphoma (FL; n = 12), and Burkitt’s Lymphoma (n = 6) compared to human normal tonsil samples (n = 10). We further tested their expression in 14 different human lymphoma cell lines. The relative expression of Per2 was markedly down-regulated in all DLBCL samples compared to normal tonsils (mean fold change: − 20.5 ± 28.7; p < 0.001). On the other hand, levels of expression of Per2 in samples of either MCL (2.5 ± 2.6; p = 0.1), FL (1.1 ± 10; p = 0.9), or Burkitt’s Lymphoma (− 1.1 ± 2; p = 0.07) showed no significant change compared to the normal tonsils. Moreover, we found no significant regulation of the mRNA levels of Per1 in any of the four NHL subtypes. We confirmed our results by showing concordant results in human lymphoma cell lines. Again, Per2 expression levels were persistently down-regulated in the 7 DLBCL cell lines (OCI-Ly1, -Ly4, -Ly7, -LY10, and SUDHL-4, -6, -16; mean fold change: − 8.6 ± 5.7; p < 0.001) compared to normal tonsils, but not in MCL (SP-49, Jeko-1, NCEB-1), FL (FLK-1), Burkitt’s lymphoma (Daudi), and B-precursor acute lymphoblastic leukemia (Nalm-6, BALL-1) cell lines (p > 0.05). Western blot protein analyses showed easily detectable levels of PER2 in tonsil samples, but the protein was either not expressed or significantly reduced in expression in human DLBCL cell lines. In summary, our results strongly suggest for the first time that the disruption of the key circadian clock gene Per2 may play a role in the development of DLBCL.

Abstract Dysregulation of receptor tyrosine kinase FGFR3 has been implicated to play a pathogenic role in a number of human hematopoietic malignancies and solid tumors. These include t(4;14) multiple myeloma associated with ectopic expression of FGFR3 and t(4;12)(p16;p13) acute myeloid leukemia associated with expression of a constitutively activated fusion tyrosine kinase TEL-FGFR3. We recently reported that FGFR3 directly tyrosine phosphorylates p90 Ribosomal S6 Kinase2(RSK2) at Y529, which consequently regulates RSK2 activation [Kang et al, Cancer Cell 2007 Sep;12(3):201–14]. Here we identified Y707 as an additional tyrosine site of RSK2 that is phosphorylated by FGFR3. Phosphorylation at Y707 contributes to RSK2 activation, through a putative disruption of the autoinhibitory αL-helix on the C-terminus of RSK2, unlike Y529 phosphorylation that facilitates ERK binding. To elucidate the role of tyrosine phosphorylation at Y707 induced by FGFR3 in RSK2 activation, we characterized the RSK2 mutants with single Y→A and Y→F substitutions at Y707. RSK2 Y707F demonstrated decreased kinase activity, suggesting substitution of Y707 attenuates activation of RSK2 induced by FGFR3. Tyrosine phosphorylation at Y529 by FGFR3 regulates RSK2 activation by facilitating inactive ERK binding, whereas substitution of Y707 in RSK2 does not similarly attenuate inactive ERK binding to RSK2. Phosphorylation at Y707 may regulate RSK2 activation by affecting the structure of the autoinhibitory C-terminal domain of RSK2 since the Y707 is localized at the C-terminal tail region which represents a conserved putative auto-inhibitory alpha helix. Since other tyrosine kinases including FGFR1 and Src also phosphorylate RSK2 at Y529 and Y707, tyrosine phosphorylation may be a general requirement for RSK2 activation through the ERK/MAPK pathway. Together, our current and previous findings represent a paradigm for tyrosine phosphorylation-dependent regulation of serine-threonine kinases. Moreover, we found that FGFR3 interacts with RSK2 through residue W332 in the linker region of RSK2, and that this association is required for FGFR3-dependent phosphorylation of RSK2 at Y529 and Y707, and subsequent RSK2 activation. Furthermore, in a murine bone marrow transplant assay, genetic deficiency in RSK2 resulted in a significantly delayed and attenuated myeloproliferative syndrome induced by TEL-FGFR3 as compared with wild type cells, suggesting a critical role of RSK2 in FGFR3-induced hematopoietic transformation.

Background and Objectives Keloids are benign dermal fibrous growth and excessive collagen deposition that occur usually after trauma or surgery. In the treatment of keloids, the recurrence rate is relatively high after surgical excision. Fillet flap is known to be a good surgical method for keloid lesions. The purpose of this study is to find out manifestation and compare the results of ear keloids after the surgery by fillet flap.Subjects and Method We retrospectively evaluated 22 patients with ear keloids (n=31) who underwent core excision with fillet flap at the Department of Dermatology, Chosun University Hospital from May 2010 to June 2018.Results With the 22 of treated patients and 31 ear keloid lesions, the average size of keloid lesions was 0.75×1.05 cm2 . The frequencies of occurrence with respect to the location of keloids according to the anatomical structure of the ear were 12 lobule (38.7%), 17 helix (54.8%), 1 antihelical fold (3.2%), and 1 postauricle (3.2%), respectively. There were 14 lobular types (45.2%), 9 dumbbell types (29.0%), 5 button types (16.1%), and 3 wrap-around type (9.7%). Recurrence was found in 8 keloid lesions (25.8%) and 5 patients (22.7%) after the surgery. Earlobe lesions and dumbbell shaped recurred with the highest recurrence rate. Among the 5 patients who relapsed, 4 had family history of keloids.Conclusion The earlobe and dumbbell shaped types showed the highest recurrence rate and family history was also an important risk factor for recurrence. Also, surgical excision with fillet flap can be very effective and is a good way to treat ear keloids.

Abstract Thyroid hormones (THs) are transported across cell membranes by different transmembrane transporter proteins. In previous studies, we showed marked 3,3′-diiodothyronine (3,3′-T2) but moderate T3 uptake by the L-type amino acid transporter 2 (Lat2). We have now studied the structure-function relationships of this transporter and TH-like molecules. Our Lat2 homology model is based on 2 crystal structures of the homologous 12-transmembrane helix transporters arginine/agmatine antiporter and amino acid/polyamine/organocation transporter. Model-driven mutagenesis of residues lining an extracellular recognition site and a TH-traversing channel identified 9 sensitive residues. Using Xenopus laevis oocytes as expression system, we found that side chain shortening (N51S, N133S, N248S, and Y130A) expanded the channel and increased 3,3′-T2 transport. Side chain enlargements (T140F, Y130R, and I137M) decreased 3,3′-T2 uptake, indicating channel obstructions. The opposite results with mutations maintaining (F242W) or impairing (F242V) uptake suggest that F242 may have a gating function. Competitive inhibition studies of 14 TH-like compounds revealed that recognition by Lat2 requires amino and carboxylic acid groups. The size of the adjacent hydrophobic group is restricted. Bulky substituents in positions 3 and 5 of the tyrosine ring are allowed. The phenolic ring may be enlarged, provided that the whole molecule is flexible enough to fit into the distinctly shaped TH-traversing channel of Lat2. Taken together, the next Lat2 features were identified 1) TH recognition site; 2) TH-traversing channel in the center of Lat2; and 3) switch site that potentially facilitates intracellular substrate release. Together with identified substrate features, these data help to elucidate the molecular mechanisms and role of Lat2 in T2 transport.

In this study, a comparative evaluation was performed in batch esterification reactions under conventional heating (CH) and assisted by microwave irradiation (MW) using bioprinted lipases. Microwave-irradiation-assisted reactions generally provide higher productivities and improve synthesis performance in terms of increased rate and reduced reaction times, resulting in higher interest yields in less time. Productivity was calculated with the enzymes: Burkholderia cepacia lipase (BCL), Candida rugosa lipase (CRL), and porcine pancreas lipase (PPL) using different fatty acids (lauric acid (12:0), myristic acid (14:0), palmitic acid (16:0), stearic acid (18:0), and oleic acid (18:1)) and alcohols at a molar ratio of 1:8. The microwave reactor was operated at a temperature of 45 °C, and power varied between 50 W and 200 W. Bioprinted BCL (bBCL) showed the highest productivity among the tested lipases. In the reaction with the best result, bBCL with lauric acid under MW, the reaction time decreased from 24 h (CH) to 25 min (MW) and the productivity increased 33 times compared with the reactions under CH. The increase in productivity demonstrates its activation that occurred as a result of conformational changes of the enzyme in the bioprinting process, confirmed by Fourier transform infrared (FTIR) spectrometric analysis, which reduces the content of bBCL α-helix with lauric acid. The biocatalyst showed high operational stability over eight cycles, while losing only 19% of its initial activity with half-life times of 12.8 batches. The storage time was five weeks, maintaining ≈80% activity. The results demonstrate the prospect of a new enzymatic route to obtain hyperactive catalysts, with the use of bioprinted lipases in esterification reactions under microwave irradiation, for the synthesis of esters with a view to large-scale industrial application.

Abstract Abstract 3396 The tertiary structure of cytokine-receptor complex provides information about molecular interactions trigger the receptor activation, the primary event of physiological response. Despite the low homology in primary structures, cytokines share a left-handed antiparallel four-helix bundle fold. Cytokine receptors are composed of a common domains and motifs and some cytokine receptors share functional and conservational subunits. We can discuss about the structural similarities and differences, however, the structural features caused a specific molecular recognition of individual cytokines are difficult to clarify. To further our understanding of structural basis of cytokines, cross-species comparison of a cytokine structure is an important clue. When we identified erythropoietin (EPO) in African clawed frog, Xenopus laevis, namely xlEPO, it was our surprise that xlEPO shares only 38% of the amino acid sequence with human EPO (hEPO) and lacks N-glycosylations. Dispite the low molecular similarity, xlEPO could stimulate proliferation and differentiation of erythrocytic cells transcending species. Meanwhile the activity of hEPO, sharing 45% homology with the functional N-terminal domain of human thrombopoietin (hTPO), is distinct from that of hTPO. These findings lead us to explore a particular topology specific to EPO-EPOR binding by solving the tertiary structure of xlEPO that enables us to compare that of EPO. Since xl EPO has no carbohydrate structures, we purified biologically active recombinant xlEPO expressed in Escherichia coli to homogeneity. To examine the in vivo effects, frogs were administrated xlEPO (0.15–0.25 mg/kg/day B.W.) for consecutive 8 days. As a result, immature erythrocytes were appeared in the circulation on day 7, and gradually increased to account for 20% in total peripheral blood cells on day 11. Structural stability of xlEPO was measured by circular dichroism spectrometry, resulting that non-glycosylated xlEPO is much more stable against thermal denaturation than heavily-glycosylated hEPO. Furthermore, we acquired the structural information about xlEPO by X-ray crystal structure analysis. Unbound form of xlEPO was crystallized and diffracted to 2.9 Å. The structure was determined by molecular replacement method with hEPO complexed to its receptor (PDB ID: 1EER) as search model. This is the first crystal structure of unbound form of EPO. Overall tertiary structure of xlEPO shows a left-handed antiparallel four helical bundle fold. xlEPO is composed of four long α-helices (H1, P5-S28; H2, Q58-F80; H3, L86-L107 and H4, F128-R152) arranged in an “up-up-down-down” topology, connected by two long cross-over loops (H1-2 and H3-4) and one short loop (2-3). H1 and H4 are bridged by disulfide bond through C7 and C151. In addition to the four long α-helices, two much shorter helices are observed in the long connecting loops. One is located at between H1 and H2 (V48-K53), and one at between H3 and H4 (Q110-Q114). One short stretch of antiparallel β-sheet is formed within long cross-over loops: strand 1, I39-P42 lies on between H1 and H2; strand 2, T122-V125 between helices 3 and 4. The α-carbon RMSD (root mean square distance) between xlEPO and hEPO are 2.35 Å and 3.06 Å, with 1EER and the NMR structure (PDB ID: 1BUY), respectively. Residues involved in receptor binding were highly conserved with 0.78 Å for 12 residues of site 1, 0.65 Å for 12 residues of site 2, calculating with 1EER. These conformational similarities in receptor binding sites are thought to be responsible for cross-reactivity between human and Xenopus EPO. One distinct structural feature is that xlEPO has a shorter loop length than hEPO. Especially, long loop between H3 and H4 of xlEPO is consisted of 7 residues (T118-K124), much shorter than 14 residues (I119-T132) of hEPO. Eventually the secondary structure content of xlEPO showed 71.5%, higher than 63.3%, that of hEPO. In addition, the number of glycine in α-helix, which is known to destabilize the helix, is 6 residues in hEPO, larger than in xlEPO, 3 residues. These structural features would account for the large difference in structural stability between xlEPO and hEPO. Together with the crystal structure of unbound form of xlEPO, the tertiary structure of receptor-binding form of xlEPO would enable us to understand the conformational change of xlEPO, and to design EPO mimetics based on newer concepts. Disclosures: No relevant conflicts of interest to declare.

Abstract The most frequent oncogenic activation events characterized in childhood T acute lymphoblastic leukemia (T-ALL) result in the transcriptional activation of genes coding for transcription factors. The main genes are TAL1/SCL, a member of the basic region helix-loop-helix gene family, HOX11L2 a member of the homeobox-containing protein family. Conflicting results have been reported concerning molecular epidemiology and prognostic values of these markers (Cavé Blood2003:103442–445, Ferrando Cancer Cell 2002, Dicciani Blood 2003 abs 67) we therefore analysed retrospectively 200 pts treated in the French protocol FRALLE 93 for T-ALL between 11/93 and 12/99. Pts were stratified according to prednisone response at D8 ( good or poor : GPR or PPR) and bone marrow at D21. Pts with D8 PPR or M3 received an intensified treatment with genoidentical or autologous transplant in CR1. Molecular analysis was possible for 79/200 T-ALL samples. Clinical caracteristics were not significantly different between population with or without molecular analysis male (n=121) (69% vs 70%), median age 8.4y (range1.1–19.5) vs 9.2y, median leucocytosis 140.109 (0.6–736) vs 171.915 109 (<50 n=23, 50–99 n=8, >100 n=48), mediastinal involvement 72% vs 71% ,CNS+ 4% vs 5%, CD 10 neg 54% vs 52%. Steroid response PPR n=37/73, GPR n=36/73 and D21 bone marrow status (M3 n=10, M2 n=11) were similar. CR was obtained in 72/79 pts (91%) after first induction therapy and 2 deaths occurs during induction treatment. With a median follow-up of 63 months (2–123), 5 y OS , EFS and DFS is 62 %± 9 and 54% ±10. SIL-TAL1/SCL fusion was detected in 20/79 (25%) pts; expression of HOX11L2 was observed in 14/79 (17%) pts. These activations are mutually exclusive and they allow the subclassification of 42 % of the patients. Median leucocytosis was significantly higher in SIL-TAL1/SCL pts (p=0.01) but other significant features ( ie median age, D8 response, D21 status) were not significantly different between each group. OS and EFS for TAL1/SCL , HOX11L2+ and none of these were respectively 81%±10, 43%±13, 62%± 7 and 80%±10 , 42%± 13, 55%± 7. OS and EFS D8 PPR/GPR were 47%± 9 vs 78% ±7 (p=0.009) and 41%±8 vs 71%±7 (p= 0.008); OS and EFS M2M3 vs M1 D21 bone marrow status were 45%± 15 vs 70% ±6 (p=0.05) and 36%±10 vs 66%± 6 (p= 0.018). In multivariate Cox model analysis, D8 steroid response and HOX11L2 expression were significantly associated with adverse event (failure or relapse) Risk Ratio :3 and 2.6 - p=0.008 and 0.03 and a higher risk of death Risk Ratio : 4.1 and 3.4 -p=0.061 and 0.05. Finally HOX11L2 expression and D8 PPR are independently associated with poor outcome in FRALLE 93 protocol analysis which confirms our previous report. Discrepancies among series may be explained by confounding factors such as differences in median leucocytes value(140.109 in our study) and treatment.

Abstract Patients with type 4 hemochromatosis exhibit iron accumulation in the reticuloendothelial system and a tendency to develop anemia following venesection. These patients are heterozygous for mutations in the intestinal and macrophage iron exporter, ferroportin1 (fpn1), however it is not known whether these mutations cause a gain or loss of function. Of current animal models, only the Tp85c allele of the zebrafish anemia mutant weissherbst (weh) encodes a missense mutation (L167F) in the conserved region of fpn1, between transmembrane helix 3 and 4, where several missense mutations have been identified in individuals with type 4 hemochromatosis. Tp85c homozygote zebrafish die within 10–14 days post-fertilization, however administering iron dextran injections allows them to survive. To explore the effects of fpn1 deficiency on blood development and iron homeostasis in adult zebrafish, we raised Tp85c homozygotes along with control cohorts of injected and uninjected heterozygotes and wild types. Iron dextran injections were discontinued at 4 months of age in all cohorts. Although iron accumulation was observed in all iron-injected fish, only homozygotes exhibited iron accumulation in the intestinal epithelium on Perl’s staining, consistent with a block in enterocyte export. At 6 months of age, none of the cohorts were anemic, however at 12 months of age, only the homozygotes developed hypochromic anemia, characterized by reduced levels of hemoglobin per erythrocyte and impaired late erythroid maturation. Histologic evaluation of the anemic homozygotes revealed increased numbers of iron-loaded macrophages in the liver and in the kidney, the zebrafish bone marrow equivalent. Quantitative realtime rt-PCR, performed on liver and intestinal tissues at 1 year of age, revealed that homozygotes had increased intestinal expression of fpn1 compared to the other cohorts. Hepatic transcript levels of the secreted iron-regulator hepcidin were profoundly decreased in homozygotes, consistent with response to anemia. In contrast, hepatic transcript levels for transferrin, fpn1, and DMT1 were not significantly different among the cohorts. In summary, we provide evidence that Tp85c is a loss of function mutation affecting enterocytes and macrophages. After iron dextran injections were discontinued, Tp85c homozygotes developed anemia, despite increased macrophage iron stores. These studies support the hypothesis that fpn1-deficiency impairs iron cycling and suggest this as a possible mechanism in type 4 hemochromatosis.

Abstract Abstract 2414 Introduction: Neurofibromatosis 1 (NF1) is an autosomal dominant disease, caused by mutation of neurofibromin (NF1) gene, located in the 17q11.2 chromosomal region. Children affected by NF1 have an increased risk to develop tumors, including leukemia. It has been suggested that NF1 aberrancy is a tumor predisposing factor, and that secondary events in somatic cells gives rise to the formation of neoplasm. However, the clonal evolution from NF1 mutation to tumor development has never been elucidated. Monozygotic twins constitute an ideal tool to understand the clonal evolution of leukemia in the context of a common genetic background. Patients: We performed the genomic characterization of a pair of monozygotic twins with diagnosis of NF1, who developed concordant B-cell precursor ALL at the age of 6 (Twin 1, T1) and 6.5 years (T2). The diagnostic and relapse samples showed common and unique lesions, allowing us to hypothesize a model for leukemia clonal evolution. At diagnosis, T1 had common ALL with 45,XX,-7,del(9)(p12),del(10)(q23)[8]/46,XX[12] whereas T2 had common ALL with 45,XX,-7,del(10)(q22)[14]. Both were treated according to AIEOP-BFM ALL 2000 protocol; T1 was classified as MRD high risk (HR), and T2 as MRD Intermediate risk (IR). T1 underwent BMT 8 months after diagnosis, and achieved clinical remission, still persisting, whilst T2 relapsed 13 months after diagnosis with 47,XX,del(9)(p21),+21[11]/46,XX[5]; she entered AIEOP REC 2003 protocol and, after achieving second complete remission and negativity for MRD, underwent allogeneic HLA-compatible BMT; she is in clinical remission 3.5 years after BMT. Results: Genome-wide copy number alteration (CNA) analysis by Cytogenetics Whole Genome 2.7M Arrays (Affymetrix) indicated that T1 and T2 shared a copy number neutral Loss Of Heterozygosity (LOH) of 17q arm, were the NF1 gene is located. At diagnosis, both had chromosome 7 monosomy but showed completely different Ig/TCR rearrangements. Three additional clonal rearrangements were found in T2 relapse sample. By backtracking the rearrangements, the relapse clone was detected in about 1% of T2 diagnostic cells, but not in T1 diagnosis sample. Several twin-specific abnormalities were detected: both had a deletion involving the q arm of chromosome 10, with different extent: del(10)(q23.33) in T1 and del(10)(q23.1) in T2. Moreover, T1 carried hemizygous deletions on chromosome 9 and a focal loss of 6q15 locus, containing the B-cell specific transcriptional regulator BACH2. Genomic analysis of T2 revealed a gain in the telomeric region of chromosome 2 and heterozygous losses in 14q32.13 and 15q21.3 regions, involving BX247990 (human full-length cDNA clone of B cells) and TCF12 (transcription factor 12) genes, respectively. The T2 relapse showed LOH 17q and del(15)(q21.3), as found in her diagnostic sample, and several additional CNAs, including the heterozygous deletions of IKZF1 (7p12.2), ETV6 (12p13.2) and C20orf94 (20p12.2) genes, trisomy of chromosome 21 and LOH of chromosome 20. Discussion: This is the first study that describes the natural history of events in NF1 patients (with 17q LOH) who developed ALL. The unique feature common to both twins in diagnosis and relapse samples was 17q LOH, that probably arose prenatally in a common hematopoietic progenitor (before somatic recombination) of one twin and spread to the other twin through intraplacental circulation. After birth, an independent clonal evolution is sustained by different Ig/TCR rearrangements as well as specific and different oncogenetic lesions. This model support a preleukemic state followed by at least-two-step mechanism for progression to leukemia. The absence of monosomy 7 at relapse indicates an independent and postnatal acquisition of this lesion; and that relapse in T2 occurs from a pre-leukemic clone before monosomy 7, which further evolved through acquisition of additional abnormailities, commom in BCP-ALL (i.e. ETV6 and Ikaros deletions). The role of BACH2 and TCF12 gene deletions in NF1 and ALL must be further explored. BACH2 is a human B-cell specific transcriptional repressor already described as tumor suppressor gene, involved in IgH@ translocations. TCF12 (also named HEB) is a basic helix-loop-helix transcription factor involved in B and T cell commitment. The altered expression of these genes could contribute to the differentiation arrest and the uncontrolled proliferation of leukemic blasts. Disclosures: No relevant conflicts of interest to declare.

Abstract Background: I-mfa has been identified as an inhibitor of MyoD and other related myogenic basic helix-loop-helix proteins. I-mfa contains a cysteine-rich C-terminal domain, and has been reported to function as transcriptional regulator of different pathways including Wnt signaling, c-jun N-terminal kinase signaling, and the regulatory properties of I-mfa depend on the C-terminal domain. Furthermore, recent studies have found that the I-mfa domain may have a close correlation with the development of myeloid neoplasms, however the role of I-mfa in adult patients with de novo acute myeloid leukemia still remain unclear. Aims: The aim of this study was to determine I-mfa expression in adult patients with de novo acute myeloid leukemia and its clinical significance. Methods: BM samples form 110 adult patients with de novo AML were analyzed. Of the 110 AML patients, 66 were males and 44 were females, with a mean age of 32 years( range from 12 to 77 years). Among them, 1 out of 110 patients was M1, 49 were M2, 14 were M4, 28 were M5, 1was M6 and 17 were acute unclassified leukemia. All patients received 1 to 2 cycles of induction of standard-dose cytarabine continuous infusion×7 days with idarubicin or daunorubicin×3days, fellowed by consolidation therapy with HiDAC and then stem cell transplantation according to patient’s condition. Real-time reverse transcription-polymerase chain reaction(RT-PCR) was used to detect the expression of I-mfa gene in 110 de novo adult AML patients, and the patients were divided into high and low I-mfa expression groups accordint to the median expression of I-mfa mRNA. Comparisons were performed using Mann-Whitney U test, Chi-square test and Kaplan-Meier method. Results:Distribution of I-mfa gene expression in different FAB subtypes was with no significant differences (P=0.169). The median age of AML pateints in low and high I-mfa gene epxression groups were 35 and 40 years old(P=0.162), and the median expression of I-mfa in 44 female patients and 66 male patients was 0.018 and 0.013 separately(P=0.728). What’s more, there was no significant difference of WBC, Hb level, PLT, bone marrow blast counts between the two groups (P＞0.05), and the I-mfa expression level was also not correlated with chromosome risk stratification and the expression of CD34 (P＞0.05). High I-mfa expression group had a lower complete remission rate than that in the low expression group (81.8% vs 63.6%, P=0.032), However, the overall survival rate was with no significant difference in the low and hign I-mfa gene expression groups(76.4% vs 76.4%, P=0.471). Conclusions: Our results showed high I-mfa expression correlates with a poor treatment response, the OS rate was with no significant difference in the two groups. There is somewhat correlation between the expression level of I-mfa gene and prognosis and the expression of I-mfa may be a prognostic factor for adult patients with de novo acute myeloid leukemia. Disclosures No relevant conflicts of interest to declare.

Abstract Background: Nuclear factor erythroid-2 related factor-2(Nrf2), a helix-loop-helix basic leucine zipper transcription factor, is a key regulator in the cellular defense system against oxidative stress. Nrf2 is binding with Keap1 in a physiological manner, activation of Nrf2 by ROS results in the induction of a series of anti-oxidative stress/detoxifying enzymes and proteins, such as HO-1, NQO-1, UGT and GST. Previous studies suggest that, Nrf2 is aberrant activated in many cancer cells, which is playing an important role in the drug-resistance of the patients. What’s more, it was reported that there’s also an aberrant activation of Nrf2 in the AML cells, which leading to the chemotherapy resistance of AML patients. But the value of Nrf2 in prognosis of adult acute myeloid leukemia(AML) still remain unknown. Aims: The aim of this study was to determine Nrf2 expression in adult patients with de novo acute myeloid leukemia and its clinical significance. Methods: BM samples form 110 adult patients with de novo AML were analyzed. Of the 110 AML patients, 66 were males and 44 were females, with a mean age of 32 years (range from 12 to 77 years). Among them, 1 out of 110 patients was M1, 49 were M2, 14 were M4, 28 were M5, 1was M6 and 17 were acute unclassified leukemia. All patients received 1 to 2 cycles of induction of standard-dose cytarabine continuous infusion×7 days with idarubicin or daunorubicin×3days, fellowed by consolidation therapy with HiDAC and then stem cell transplantation according to patient’s condition. Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of Nrf2 gene in 110 de novo adult AML patients, and the patients were divided into high and low Nrf2 expression groups accordint to the median expression of Nrf2 mRNA. Comparisons were performed using Mann-Whitney U test, Chi-square test and Kaplan-Meier method. Results: Distribution of Nrf2 gene expression in different FAB subtypes was with no significant differences (P=0.0.385). The median age of AML pateints in low and high Nrf2 gene epxression groups were 35 and 39 years old (P=0.385), and the median expression of Nrf2 in 44 female patients and 66 male patients was 0.034 and 0.030 separately (P=0.696). What’s more, there was no significant difference of WBC, Hb level, PLT, bone marrow blast counts between the two groups (P＞0.05), and the Nrf2 expression level was also not correlated with chromosome risk stratification and the expression of CD34 (P＞0.05). The patients in high Nrf2 expression group got the same complete remission (72.7%) with the low Nrf2 epxression group. The survival curve gave the clues that the median survival period of hig and low Nrf2 expression groups were 218d and 361d, the overall survival rate was also with no significant difference in the two groups(72.7% vs 80.0%，P=0.667). Conclusions: Our results showed that no matter the expression level of Nrf2 gene, there are no significant difference of CR rate and OS rate in the patients with de novo acute myeloid leukemia. Disclosures No relevant conflicts of interest to declare.

Abstract Background The transcription factor Hairy enhancer of split1 (Hes1) is well characterized as a downstream target of Notch signaling. Hes1 is a basic helix-loop-helix-type protein, and represses target gene expression. Notch signaling has been proposed to play both pro- and anti-tumorigenic roles; it promotes development of T-cell acute lymphoblastic leukemia (T-ALL), while serves as a tumor suppressor for acute myeloid leukemia (AML). Hes1 has been proven as an essential mediator of Notch signaling in T-ALL development. In contrast, we reported, in the last annual meeting, that Hes1 functions as a tumor suppressor against AML development, using a mouse model of AML induced by the MLL-AF9 fusion protein. We further explored the mechanism of Hes1-mediated suppression of AML development. Methods Common myeloid progenitors (CMPs) purified from RBP-Jf/f mouse bone marrow (BM) were serially transduced with MLL-AF9 and Cre recombinase (iCre) using retroviral vectors, and transplanted into lethally irradiated syngenic mice. CMPs from Hes1-/- mouse fetal liver were also retrovirally transduced with MLL-AF9 and transplanted after multiple rounds of replating, and then, expression levels of downstream targets were evaluated by cDNA array. Next Hes1 was retrovirally re-expressed in MLL-AF9/Hes1-/- cells and these cells were transplanted. MLL-AF9-transduced cells were treated with a hamster anti-mouse Notch agonistic antibody (Notch Ab). Results Mice transplanted with MLL-AF9/RBP-J-/- cells developed leukemia at shorter latencies than those with MLL-AF9/RBPJ+/+ cells. MLL-AF9-transduced Hes1-/- cells formed the higher number of colonies at third replating compared with MLL-AF9-transduced Hes1+/+ cells. When infused into irradiated syngenic mice, MLL-AF9/Hes1-/- cells developed leukemia at shorter latencies than MLL-AF9/ Hes1+/+ cells (MLL-AF9/Hes1-/-, 7-10 weeks, n=18 vs MLL-AF9/Hes1+/+, 10-14 weeks, n=18; p<0.001). When Hes1 was retrovirally re-expressed in MLL-AF9/Hes1-/- cells, these cells developed leukemia in recipient mice at longer latencies than mock-transduced MLL-AF9/Hes1-/- cells (Hes1/MLL-AF9, 12 weeks, n=8 vs mock/MLL-AF9, 5-7 weeks, n=7 p<0.001). When treated with an anti-Notch2 Ab, MLL-AF9/Hes1+/+ cells underwent apoptosis, whereas MLL-AF9/Hes1-/- cells did not. These results indicate that Hes1 is a definitive downstream mediator for Notch signaling-mediated suppression of AML. Among the genes with different expression levels between MLL-AF9/Hes1-/- and MLL-AF9/Hes1+/+ leukemia cells, FMS-like tyrosine kinase 3 (FLT3) was expressed at significantly higher levels in MLL-AF9/Hes1-/- leukemia cells as well as RBP-J-null Background. It was also demonstrated that FLT3 and ERK were phosphorylated with FLT3 ligand stimulation in the MLL-AF9-immortalized cells specifically with the Hes1-/- Background. An FLT3 inhibitor efficiently abrogated the proliferation of MLL-AF9/Hes1-/- leukemia cells. We accessed the independent database containing mRNA expression profiles and found that the expression level of Flt3 mRNA was negatively correlated with those of Hes1 in AML samples. Conclusion Canonical Notch signaling serves as a tumor suppressor in MLL-AF9-induced AML through upregulation of Hes1. Hes1 is an essential Notch signaling mediator for AML suppression. At least a part of Hes1 function might be explained by repression of FLT3. Disclosures: No relevant conflicts of interest to declare.

ABSTRACTTherapeutic options are limited forNeisseria gonorrhoeaeinfection, especially for oral drugs. The purpose of this study was to investigate the susceptibility ofN. gonorrhoeaeto oral azithromycin (AZM) and the correlation between AZM resistance-related gene mutations and MIC. We examined the AZM MICs of clinical strains ofN. gonorrhoeae, sequenced the peptidyltransferase loop in domain V of 23S rRNA, and investigated the statistical correlation between AZM MIC and the presence and number of the mutations. Among 59N. gonorrhoeaestrains, our statistical data showed that a deletion mutation was seen significantly more often in the higher-MIC group (0.5 μg/ml or higher) (35/37; 94.6%) than in the lower-MIC group (0.25 μg/ml or less) (4/22; 18.2%) (P< 0.0001). However, a mutation of codon 40 (Ala→Asp) in themtrRgene (helix-turn-helix) was seen significantly more often in the lower-MIC group (12/22; 54.5%) (P< 0.0001). InN. gonorrhoeaemultiantigen sequence typing (NG-MAST) analyses, ST4777 was representative of the lower-MIC group and ST1407, ST6798, and ST6800 were representative of the higher-MIC group. NG-MAST type 1407 was detected as the most prevalent type in AZM-resistant or -intermediate strains, as previously described. In conclusion, a deletion mutation in themtrRpromoter region may be a significant indicator for higher MIC (0.5 μg/ml or higher). ST4777 was often seen in the lower-MIC group, and ST1407, ST6798, and ST6800 were characteristic of the higher-MIC group. Further research with a greater number of strains would help elucidate the mechanism of AZM resistance inN. gonorrhoeaeinfection.

ABSTRACTFlavivirus RNA synthesis is mediated by a multiprotein complex associated with the endoplasmic reticulum membrane, named the replication complex (RC). Within the flavivirus RC, NS4B, an integral membrane protein with a role in virulence and regulation of the innate immune response, binds to the NS3 protease-helicase. NS4B modulates the RNA helicase activity of NS3, but the molecular details of their interaction remain elusive. Here, we used dengue virus (DENV) to map the determinants for the NS3-NS4B interaction. Coimmunoprecipitation and anin situproximity ligation assay confirmed that NS3 colocalizes with NS4B in both DENV-infected cells and cells coexpressing both proteins. Surface plasmon resonance demonstrated that subdomains 2 and 3 of the NS3 helicase region and the cytoplasmic loop of NS4B are required for binding. Using nuclear magnetic resonance (NMR), we found that the isolated cytoplasmic loop of NS4B is flexible, with a tendency to form a three-turn α-helix and two short β-strands. Upon binding to the NS3 helicase, 12 amino acids within the cytoplasmic loop of NS4B exhibited line broadening, suggesting a participation in the interaction. Sequence alignment showed that 4 of these 12 residues are strictly conserved across different flaviviruses. Mutagenesis analysis showed that three (Q134, G140, and N144) of the four evolutionarily conserved NS4B residues are essential for DENV replication. The mapping of the NS3/NS4B-interacting regions described here can assist the design of inhibitors that disrupt their interface for antiviral therapy.IMPORTANCENS3 and NS4B are essential components of the flavivirus RC. Using DENV as a model, we mapped the interaction between the viral NS3 and NS4B proteins. The subdomains 2 and 3 of NS3 helicase as well as the cytoplasmic loop of NS4B are critical for the interaction. Functional analysis delineated residues within the NS4B cytoplasmic loop that are crucial for DENV replication. Our findings reveal molecular details of how flavivirus NS3 protein cooperates with NS4B within the RC. In addition, this study has established the rationale and assays to search for inhibitors disrupting the NS3-NS4B interaction for antiviral drug discovery.

Venetoclax (Ven) is an effective element of treatments for chronic lymphocytic leukemia (CLL) with high response rates observed in the upfront and relapsed/refractory (R/R) settings. In addition to inducing apoptosis in CLL cells, Ven also induces apoptosis within normal and malignant myeloid lineage populations (accounting for its efficacy in the treatment of acute myeloid leukemia). We investigated the effects of Ven outside the target tumor compartment in patients (pts) with CLL receiving long-term continuous Ven and make the novel observation of the development of BAX-mutated clonal hematopoiesis in this heavily pre-treated patient group. 92 pts with CLL receiving continuous non time-limited Ven have been treated at our institutions on clinical trials. Of these, 41 had sufficient (&gt;6 mo) follow up (median 70; range 14-95 mo) and suitable samples available for further analysis. 38/41 (93%) pts had received previous treatment with alkylators and/or fludarabine. In order to assess the non-CLL compartment in these 41 pts we identified those with peripheral blood or bone marrow aspirate samples taken during deep response to Ven demonstrating either minimal (&lt;5%) or no CLL involvement by flow cytometry (sensitivity 10-4). We initially performed unique molecular index (UMI)-based targeted next generation sequencing of apoptosis pathway genes as well a panel of 60 genes recurrently mutated in lymphoid and myeloid malignancy. From these 41 pts we identified mutations in the apoptosis effector BAX in samples from 12 (29%). 20 different BAX mutations were observed across these 12 pts at variant allele frequencies (VAF) consistent with their occurrence in the non-CLL compartment. Mutations included frameshift, nonsense, canonical splice site and missense mutations occurring in key structural elements of BAX consistent with a loss-of-function mechanism (Fig 1A). Interestingly, an enrichment of missense and truncating mutations predicted to escape nonsense mediated decay were observed at the C-terminus of the BAX protein affecting the critical α9 helix. Mutations in this region have previously been shown in cell lines to cause aberrant intracellular BAX localization and abrogation of normal BAX function in apoptosis (Fresquet Blood 2014; Kuwana J Biol Chem 2020). For comparison, NGS targeted sequencing for BAX mutations was performed on samples from cohorts of pts with (i) myeloid or lymphoid malignancy (n=80) or (ii) R/R CLL treated with BTK inhibitors (n=15) after a similar extent of preceding chemotherapy. Neither of these cohorts had previous exposure to Ven. BAX mutations were not detected in any samples from these pts. Longitudinal sampling from pts on Ven harboring BAX mutations in the non-CLL compartment was performed to further understand compartment dynamics over time (in 9 pts over 21-93 months of follow up). Multiple pts demonstrated a progressive increase in VAF of single BAX mutations over time to become clonally dominant within the non-CLL compartment and with observed VAFs consistent with their presence in the myeloid compartment. Mutations in other genes implicated in clonal hematopoiesis and myeloid malignancy including ASXL1, DNMT3A, TET2, U2AF1 and ZRSR2 were also detected in these pts samples. Targeted amplicon single cell sequencing (Mission Bio) demonstrated the co-occurrence of clonally progressive BAX mutations within the same clones as mutations in DNMT3A and ASXL1 as well as the existence of further BAX mutations at low VAF outside these dominant clones which remained non-progressive over time (Fig 1B). In addition, fluctuations in the presence and VAF of myeloid-disease associated mutations was noted with Ven exposure. In aggregate these data are consistent with the existence of a selective pressure within the myeloid compartment of these pts and an interplay of BAX with other mutations in determining survival and enrichment of these clones over time with ongoing Ven therapy. In summary, we have observed the development of BAX-mutated clonal hematopoiesis specifically in pts with CLL treated with long-term Ven. These data are consistent with a multi-lineage pharmacological effect of Ven leading to a survival advantage for clones harboring BAX mutations within the myeloid compartment during chronic Ven exposure. Finally, our data support the further investigation of BAX mutations as a potential resistance mechanism in myeloid malignancies treated with Ven. Disclosures Blombery: Invivoscribe: Honoraria; Amgen: Consultancy; Janssen: Honoraria; Novartis: Consultancy. Anderson:Walter and Eliza Hall Institute: Patents & Royalties: milestone and royalty payments related to venetoclax.. Seymour:Celgene: Consultancy, Honoraria, Research Funding; F. Hoffmann-La Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Janssen: Consultancy, Honoraria, Research Funding; AstraZeneca: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy; Mei Pharma: Consultancy, Honoraria; Morphosys: Consultancy, Honoraria; Nurix: Honoraria; AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Tam:Janssen: Honoraria, Research Funding; AbbVie: Honoraria, Research Funding; BeiGene: Honoraria. Huang:Servier: Research Funding; Walter and Eliza Hall Institute: Patents & Royalties: milestone and royalty payments related to venetoclax.; Genentech: Research Funding. Wei:Janssen: Honoraria, Other; Walter and Eliza Hall Institute: Patents & Royalties; AMGEN: Honoraria, Other: Advisory committee, Research Funding; Novartis: Honoraria, Research Funding, Speakers Bureau; Astellas: Honoraria, Other: Advisory committee; Pfizer: Honoraria, Other: Advisory committee; Macrogenics: Honoraria, Other: Advisory committee; Abbvie: Honoraria, Other: Advisory committee, Research Funding, Speakers Bureau; Genentech: Honoraria, Other: Advisory committee; Servier: Consultancy, Honoraria, Other: Advisory committee; Celgene: Honoraria, Other: Advisory committee, Speakers Bureau; Astra-Zeneca: Honoraria, Other: Advisory committee, Research Funding. Roberts:Janssen: Research Funding; Servier: Research Funding; AbbVie: Research Funding; Genentech: Patents & Royalties: for venetoclax to one of my employers (Walter & Eliza Hall Institute); I receive a share of these royalties.

AbstractLarge-area helix microstructures intended for metamaterials were fabricated using a negative photoresist, SU-8 using a two photon absorption direct laser writing (TPA-DLW). Two types of helix structures were fabricated. One type is those with no neighboring distance. In this case, compact helix structures with radius of 2.5 and 1.0 μm were fabricated. Another type is those with enough neighboring distance. The helix structures with shorter neighboring distance below 6.0 μm were collapsed, whereas those with longer neighboring distance more than 6.5 μm, free-standing helix structures could successfully be built. To stabilize the fabricated free-standing helix microstructures with a 1 μm radius, circular foundations with a radius of 1.3 μm and elevation angle of 10, 12, or 14° were built in advance. The foundation is useful to avoid collapsing the helix microstructures. Due to the useful foundation, over 18,000 helical structures were fabricated in a large area. The fabricated helical structures were coated with silver using an electroless plating method to produce 3D metallic helix structures. Silver coating was measured using a EDX measurement. The obtained helical structures have the potential for metamaterials to control the handedness of a circularly polarized infrared beam.

Abstract Chimeric plants composed of green and albino tissues have great ornamental value. To unveil the functional genes responsible for albino phenotypes in chimeric plants, we inspected the complete plastid genomes (plastomes) in green and albino leaf tissues from 23 ornamental chimeric plants belonging to 20 species, including monocots, dicots, and gymnosperms. In nine chimeric plants, plastomes were identical between green and albino tissues. Meanwhile, another 14 chimeric plants were heteroplasmic, showing a mutation between green and albino tissues. We identified 14 different point mutations in eight functional plastid genes related to plastid-encoded RNA polymerase (rpo) or photosystems which caused albinism in the chimeric plants. Among them, 12 were deleterious mutations in the target genes, in which early termination appeared due to small deletion-mediated frameshift or single nucleotide substitution. Another was single nucleotide substitution in an intron of the ycf3 and the other was a missense mutation in coding region of the rpoC2 gene. We inspected chlorophyll structure, protein functional model of the rpoC2, and expression levels of the related genes in green and albino tissues of Reynoutria japonica. A single amino acid change, histidine-to-proline substitution, in the rpoC2 protein may destabilize the peripheral helix of plastid-encoded RNA polymerase, impairing the biosynthesis of the photosynthesis system in the albino tissue of R. japonica chimera plant.

Abstract Background ACYL-LIPID THIOESTERASES (ALTs) are a subclass of plastid-localized, fatty acyl-acyl carrier protein (ACP) thioesterase enzymes from plants. They belong to the single hot dog-fold protein family. ALT enzymes generate medium-chain (C6-C14) and C16 fatty acids, methylketone precursors (β-keto fatty acids), and 3-hydroxy fatty acids when expressed heterologously in E. coli. The diverse substrate chain-length and oxidation state preferences of ALTs set them apart from other plant acyl-ACP thioesterases, and ALTs show promise as metabolic engineering tools to produce high-value medium-chain fatty acids and methylketones in bacterial or plant systems. Here, we used a targeted motif-swapping approach to explore connections between ALT protein sequence and substrate specificity. Guided by comparative motif searches and computational modelling, we exchanged regions of amino acid sequence between ALT-type thioesterases from Arabidopsis thaliana, Medicago truncatula, and Zea mays to create chimeric ALT proteins. Results Comparing the activity profiles of chimeric ALTs in E. coli to their wild-type counterparts led to the identification of interacting regions within the thioesterase domain that shape substrate specificity and enzyme activity. Notably, the presence of a 31-CQH[G/C]RH-36 motif on the central α-helix was shown to shift chain-length specificity towards 12–14 carbon chains, and to be a core determinant of substrate specificity in ALT-type thioesterases with preference for 12–14 carbon 3-hydroxyacyl- and β-ketoacyl-ACP substrates. For an ALT containing this motif to be functional, an additional 108-KXXA-111 motif and compatible sequence spanning aa77–93 of the surrounding β-sheet must also be present, demonstrating that interactions between residues in these regions of the catalytic domain are critical to thioesterase activity. The behaviour of chimeric enzymes in E. coli also indicated that aa77–93 play a significant role in dictating whether an ALT will prefer ≤10-carbon or ≥ 12-carbon acyl chain-lengths, and aa91–96 influence selectivity for substrates of fully or partially reduced oxidation states. Additionally, aa64–67 on the hot dog-fold β-sheet were shown to be important for enabling an ALT to act on 3-hydroxy fatty acyl-ACP substrates. Conclusions By revealing connections between thioesterase sequence and substrate specificity, this study is an advancement towards engineering recombinant ALTs with product profiles suited for specific applications.

It is believed that the interactions of high density lipoproteins (HDL) with different plasma enzymes are partly determined by the site specific solvent exposure of apolipoproteins in HDL. Ultra-centrifugally isolated human HDL was separated into subpopulations containing both major apolipoproteins apoA-I and apoA-II (termed LpA-I/A-II) and only apoA-I (termed LpA-I) using thiopropyl sepharose chromatography. LpA-I/A-II and LpA-I were further separated into three sub-fractions based on the particle diameter utilizing fast performance liquid chromatography. Individual sub-fractions were subjected to partial acetylation; the extent of acetylation that represents the solvent exposure was quantified by mass spectrometry (MS). LpA-I of ∼85 Å, demonstrated solvent exposure variations for Lys residues in apoA-I in the range of 9-55%. Lys 133 (helix 5) and Lys 226 (helix 10) showed the highest solvent exposure of 54-55%. Surprisingly Lys residues 238 and 239 showed the lowest exposure of 9-10%. The rest of the Lys residues located in N-term through helix 10 ranged from 12-40% demonstrating distinctive differences among specific sites of exposure. The presence of apoA-II in comparable diameter LpA-I/A-II showed similar solvent exposure for apoA-I, implying the non-perturbing nature of apoA-II on apoA-I conformation. Limited proteolysis experiments resulted in similar fragment patterns for apoA-I in LpA-I and LpA-I/A-II, agreeing with mass spectrometry observations. Furthermore, proteolysis demonstrated that apoA-II in HDL is less susceptible to proteolysis compared to apoA-I in HDL. Confirming this observation further, the extent of acetylation of individual Lys residues of apoA-II in LpA-I/A-II demonstrated overall low solvent exposure (9-14%) by MS. Subtle solvent exposure differences were seen among particles of different diameter (75 Å vs 110 [[Unable to Display Character: &#506;]]). These novel findings bring out conformational details of major apolipoprotein in different HDL particles. Assessment of site specific solvent exposure of minor HDL apolipoproteins including apoE, apoD, apoCs, apoM and determination of their specific preference for LpA-I vs LpA-I/A-II are currently underway implementing high throughput mass spectrometry techniques.

Abstract Introduction: Aryl hydrocarbon nuclear translocator 2 (ARNT2) is a basic helix-loop-helix (bHLH)-PAS (Per/Arnt/Sim) transcription factor shown to be critical to the development of paraventricular nucleus of the hypothalamus (PVN), key region for energy homeostasis and feeding response. In vivo and in vitro studies have shown that ARNT2 is an obligate heterodimer for SIM1, known cause of monogenic obesity. Null mutations in Arnt2 in animals are not viable, but hypomorphic mutation results in hyperphagic obesity and its associated consequences (1). Due to the critical role of ARNT2 in the development of PVN, we hypothesize that hypomorphic mutations may result in early onset obesity in humans. Methods: The Genetics of Early Childhood Obesity (GECO) study recruits children with severe obesity (BMI &gt; 120% of 95th percentile) of early onset (&lt; 6 years). Whole exome sequencing (WES) was performed in a subset of proband-parent trios. The functional validation of the mutation(s) in ARNT2 is ongoing with co-transfection of tagged Arnt2 and Sim1 in HEK293 cells, with the induction of a luciferase reporter gene under the control of 6 repeats of bHLH-PAS core binding element by the Arnt2-Sim1 complex. Results: Two adolescents from unrelated families were found to have genetic variants in ARNT2. Subject 1 has a novel de novo heterozygous coding variant in ARNT2, c.388 C&gt;G (p.P130A, CADD 25), predicted to be deleterious by 8/12 in silico algorithms. She is a 14-year old Caucasian girl with severe early onset obesity, BMI 28.1 kg/m2 (BMIz +4.72) at 2.5 years of age that has increased to 53.54 kg/m2 (BMIz + 3.25) at 14-years, and height &gt; 95th %tile. She is non-dysmorphic, has developmental delay, absence seizures, behavior abnormalities & glucose intolerance/dyslipidemia secondary to obesity. Using genematcher, we identified another proband with the phenotype of obesity: an African American girl (BMIz +1.9) with biallelic inherited heterozygous variants in ARNT2, c.1228T&gt;A (p.W410R, CADD 29) and c.916G&gt;A (p.G306S, CADD 22). An only child conceived by IVF, she is non-dysmorphic and on treatment for bilateral focal epilepsy. All 3 variants are rare, with mean allele frequency &lt; 0.005 in population-based databases such as gNOMAD. Both the patients have early onset obesity and a significant neurological phenotype. ARNT2 is a highly constrained gene of 717 amino acids with a significant depletion of missense variants in the N-terminus (1-244 aa) and overall fewer loss of function variants in ~282,644 alleles sequenced in gNOMAD. Conclusions: We propose that hypomorphic mutations in ARNT2 could be a potential novel cause of monogenic obesity in humans. Future studies will investigate the molecular mechanisms causing weight dysregulation in patient specific disease relevant hypothalamic neurons. Reference: (1) Turer et al., Dis Model Mech. 2018; 11(12)

Introduction: Pilomatrixomas are benign neoplasms derived from hair follicle matrix cells. They are among the most common soft tissue head and neck tumors of childhood. Pilomatrixomas are typically isolated, slow-growing, firm, nontender masses that are adherent to the epidermis but mobile in the subcutaneous plane. This clinical presentation is so characteristic that many experienced surgeons will excise suspected pilomatrixomas without prior imaging. We reviewed the results of this approach to determine whether physical examination alone differentiates pilomatrixomas from other similar soft tissue lesions of the pediatric head and neck. Methods: Computerized review of all pilomatrixomas over a 20-year period in a single academic pediatric otolaryngology practice. Results: 18 patients presented to our pediatric otolaryngology practice between 2001 and 2021 with historical and physical findings consistent with pilomatrixoma. Of the 18 patients, 7 were male and 11 were female. Ages ranged from 1.5 to 14 years, with a mean of 7.5 years. Most of the lesions (12) were located in the head and face, while the rest (6) were found in the neck. All patients were treated with complete surgical excision. Pathology confirmed pilomatrixoma in 15 patients. The remaining 3 children were found to have an epidermal inclusion cyst, a ruptured trichilemmal cyst, and a giant molluscum contagiosum lesion, respectively. One additional patient presented with a small lesion of the auricular helix that was thought to be a dermoid cyst, but proved to be a pilomatrixoma on histologic examination. Discussion: As pilomatrixomas are common and have a very characteristic presentation, surgical excision without prior diagnostic imaging will lead to correct treatment in the majority of cases. High resolution ultrasonography can help to confirm the diagnosis preoperatively, but is not definitive in large case series. Most of the cystic lesions that imitate pilomatrixoma will ultimately require surgical excision.

This study develops a high performance liquid chromatography with ultraviolet detection (HPLC-UV) for simultaneous quantification of hederacoside C and α-hederin in Hedera nepalensis K. Koch. The method proposed in this study was validated in terms of the analytical parameters such as high repeatability, high accuracy and good sensitivity. The method was used to determine the content of hederacoside C and α-hederin in Hedera nepalensis K. Koch, which had been collected in Ha Giang, Lao Cai and Lai Chau. The study results show that the content of hederacoside C and the content of α-hederin ranged from 0.40 to 4.01% and 0.21 – 0.54% based on absolute dry mass, respectively. Keywords Hedera nepalensis K. Koch, hederacoside C, α-hederin, HPLC-UV. References [1] L. Jafri, et al, In vitro assessment of antioxidant potential and determination of polyphenolic compounds of Hedera nepalensis K. Koch, Arabian Journal of Chemistry. 10 (2017) 3699-3706. https://doi.org/10.1016/j.arabjc.2014.05.002.[2] S. Saleem, et al, Plants Fagonia cretica L, and Hedera nepalensis K. Koch contain natural compounds with potent dipeptidyl peptidase-4 (DPP-4) inhibitory activity, Journal of ethnopharmacology. 156 (2014) 26-32. https://doi.org/10.1016/j.jep.2014.08.017[3] D.H. Bich, Medicinal plants and animals for medicine in Vietnam, Vol 1, Science and Technics Publishing House, Hanoi, 2006 (in Vietnamese).[4] National Institute Of Medicinal Materials, List of medicinal plants in Vietnam, Science and Technics Publishing House, Hanoi, 2016 (in Vietnamese).[5] L. Jafri, et al, Hedera nepalensis K. Koch: A Novel Source of Natural Cancer Chemopreventive and Anticancerous Compounds, Phytotherapy research. 30(3) (2016) 447-453. https://doi.org/10.1002/ptr.5546. [6] S. Kanwal, et al, Antioxidant, antitumor activities and phytochemical investigation of Hedera nepalensis K. Koch, an important medicinal plant from Pakistan, Pakistan Journal of Botany. 43 (2011) 85-89. [7] G. Uddin, et al, Biological screening of ethyl acetate extract of Hedera nepalensis stem, African Journal of Pharmacy and Pharmacology. 6(42) (2012) 2934-2937. https://doi.org/10.5897/AJPP12.828 [8] H. Kizu, et al, Studies on Nepalese Crude Drugs, III, On the Saponins of Hedera nepalensis K. Koch, Chemical and Pharmaceutical Bulletin. 33(8) (1985) 3324-3329. https://doi.org/0.1248/cpb.33.3324[9] X. Tong, et al, Extraction and GC-MS Analysis of Volatile Oil from Hedera nepalensis var sinensis, Fine Chemicals. 24(6) (2007) 559-561. [10] EDQM, European Pharmacopoeia, fifth ed., Council of Europe, France, 2015. [11] N.T.H. Mai, et al, Simultaneous Quantification of Hederacoside C and α-Hederin from the Leaves of Hedera helix L. by HPLC, Journal of Medicinal Material. 21(6) (2016). (in Vietnamese).[12] L. Havlíková, et al, Rapid Determination of α-Hederin and Hederacoside C in Extracts of Hedera helix Leaves Available in the Czech Republic and Poland, Natural product communications. 10(9) (2015). https://doi.org/10.1177/1934578X1501000910[13] M. Yu, et al, Determination of Saponins and Flavonoids in Ivy Leaf Extracts Using HPLC-DAD, Journal of Chromatographic Science. 53(4) (2014) 478-483. https://doi.org/10.1093/chromsci/bmu068.[14] EMEA, Validation of analytical procedures: text and methodology Q2 (R1), in International conference on harmonization, Geneva, Switzerland, 2005. [15] W. Horwitz, Official methods of analysis, 12 ed., Vol 1, Association of Official Analytical Chemists, Washington DC, 1975.

Abstract Background and Aims Alport syndrome (AS) is an inherited nephropathy caused by pathogenic variants in COL4A3 (autosomal dominant -AD- and autosomal recessive -AR- inheritance), COL4A4 (AR) and COL4A5 (X-linked dominant -XLD). It is characterized by glomerular nephropathy with hematuria progressing to end-stage renal disease, frequently associated with sensorineural deafness and ocular anomalies. In thin basement membrane disease (TBMD) most patients are asymptomatic and are incidentally noted to have microhematuria, mild proteinuria, and occasionally gross hematuria, with normal renal function. This condition is caused by pathogenic variants in COL4A3 and COL4A4 (AD). We have systematically applied genetic testing to pediatric and adult patients with clinical features suggestive of AS (Table 1). Method Clinical exome sequencing (CES) was performed on a cohort of 95 patients referred to our center from 2019 to 2022. Analyses were performed on an in-silico designed panel including 523 renal genes. Whenever possible, identified variants were segregated by Sanger sequencing. Results We identified causative (C4/C5) variants in 43 (45.3%) patients. Among them, genetic diagnosis of AS was obtained in 25 patients (58.1%): one (4.0%) with biallelic variants in COL4A4, 13 (52.0%) with COL4A5 variants (6 females and 7 males), 5 (20.0%) with COL4A3 variants (4 heterozygous and 1 homozygous) and 6 (24.0%) cases with coexisting variants in one or more other collagen genes. In 16 (37.2%) patients, we found causative monoallelic variants compatible with TBMD: 12 (75.0%) patients with variants in COL4A4 and 4 (25.0%) in COL4A3. Lastly, in 2 cases, we found variants in COL4A1 (#120130) and MYH9 (#160775), both genes associated with other renal diseases with a clinical presentation partially overlapping with AS. In additional 16 cases (16.8%), we identified C3 variants in collagen genes. In the remaining patients, genetic testing was negative (14 patients, 14.7%) or inconclusive (22, 23.2%). Of note, we identified 6 families with digenic AS. In 4 of them - 3 with COL4A5/COL4A4 variants and one with COL4A3/COL4A4 variants – the co-existence of two variants was associated to earlier renal failure, compared to family members bearing a single variant. In contrast, in the remaining 2 families (33.3%), members with COL4A5/COL4A3 missense variants had a milder phenotype compared to male family members with the single variant COL4A5 variant. In families with digenic AS, at least one variant involved the substitution of a glycine with another amino acid in the triple helix domain, frameshift indel variants in the triple helix domain or inframe indel in the collagen IV domain non-collagenous. To better understand the impact of multiple variants on collagen structure, computational studies are currently ongoing. Conclusion CES is a powerful tool to clearly define the diagnosis when AS or TBMD are suspected, as witnessed by a detection rate of 43.2%. Genetic diagnosis allows to identify the causative gene offering the possibility of extending diagnosis to other family members, also in the prenatal setting. In addition, it offers the possibility to identify multiple variants in collagen genes associated with digenic AS, as well as to find variants in other genes implicated in kidney disease. Notably, the presence of digenic variants is not necessarily predictive of a worse disease outcome. For all these reasons, molecular diagnosis can be useful to improve clinical management, to calculate recurrence risk and to better define the prognosis of the patient.

The purpose of the paper is to research some solutions of cooperation between universities and enterprises in training vocational skills for students. Through the survey on assessment of vocational skills of students and the need for cooperation between Enterprises and Electric Power University, the research proposes some solutions between two elements of the mechanism of cooperation between University and Enterprises in training vocational skills for students to ensure graduates can meet the requirements of recruitment agencies as well as the requirements of socio-economic development and employment. Keywords Volcational skill, cooperation, universities, enterprises, Electric Power University References [1] Mạnh Xuân, Gắn kết trường đại học và doanh nghiệp trong đào tạo nhân lực, Nhân dân điện tử, 2015. http://www.nhandan.com.vn/giaoduc/tin-tuc/item/25807602-gan-ket-truong-dai-hoc-va-doanh-nghiep-trong-dao-tao-nhan-luc.html [2] Nguyễn Đình Luận, Sự gắn kết giữa nhà trường và doanh nghiệp trong đào tạo nguồn nhân lực phục vụ phát triển kinh tế xã hội ở Việt Nam: Thực trạng và Khuyến nghị, Tạp chí Phát triển và hội nhập, số 22 (32), Tháng 5-6/2015.[3] Nguyễn Thị Thanh Dần, Động lực hợp tác giữa nhà trường và doanh nghiệp trong việc nâng cao kỹ năng nghề cho sinh viên, Tạp chí giáo dục, Số đặc biệt 11/2016[4] Nguyễn Tiến Long, Phạm Hồng Hạnh. Xây dựng kho dữ liệu kĩ thuật, ứng dụng cho nghiên cứu khoa học và dạy học tại trường sư phạm kỹ thuật – đạo tạo nghề. 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In: Research Policy, 29, pag. 109-123 [14] Harris, R., Guthrie, H., Hobart B., & Lundberg, D. (1995). Competency based education and training: Between a Rock and a Whirlpool. South Melbourne: Macmillan Education Australia.[15] Henry Etzkowitz (2008). The triple helix: university-industry-government innovation. Routledge 270 Madison Ave, New York, NY 10016, ISBN 0-203-92960-8 Master e-book ISBN [16] Jones, L., & Moore, R. (1995). Appropriating competence. British Journal of Education and Work, 8(2) 78-92[17] Kathleen Santopietro Weddel (2006), Competency Based Education and Content Standards, Northern Colorado Literacy Resource Center, USA.[18] Leydesdorff, L., Etzkowitz, H. (1996): Emergence of a Triple Helix of University-Industry-Government Relations, Science and Public Policy, 23, pp.279-286. [19] Mihaela & Cornelia Dan (2013). Why Should University and Business Cooperate? 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Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): British Heart Foundation Background The microstructural changes following myocardial infarction (MI) can be characterised in-vivo with cardiac diffusion tensor imaging (cDTI) imaging, using mean diffusivity (MD), fractional anisotropy (FA), secondary eigenvector angle (E2A) and helix angle (HA) maps. In this study, we use cDTI to explore the microstructural differences between subendocardial and transmural chronic infarct segments. Method Twenty STEMI patients (15 men, 5 women, mean age 59) underwent 3T CMR scan at 3 months following presentation (mean interval 107 ± 18 days). Scan protocol included: second order motion compensated (M012) free-breathing spin echo DTI (3 slices, 18 diffusion directions at b-values 100s/mm2[3], 200s/mm2[3] and 500s/mm2[12], acquired resolution was 2.20x2.27x8mm3; cine gradient echo and LGE imaging. Average MD, FA, E2A and HA parameters were calculated on a 16-AHA-segmental level. HA maps were described by dividing values into left-handed HA (LHM, -90° &lt; HA &lt; -30°), circumferential HA (CM, -30° &lt; HA &lt; 30°), and right-handed HA (RHM, 30° &lt; HA &lt; 90°) and reported as relative proportions. Infarct segments were identified using LGE; patients were categorised according to the maximal transmurality of their infarct segments, into subendocardial (&lt;50% LGE) or transmural (&gt;50% LGE) MI. Results DTI acquisition was successful in all patients (acquisition time 13 ± 5mins). Ten patients had transmural MI. The results are shown in table 1. Transmurally infarcted segments had significantly lower FA (FA subendocardial MI = 0.27 ± 0.04, FA transmural MI = 0.23 ± 0.02, p &lt; 0.01), lower E2A (E2A subendocardial MI = 47 ± 7°, E2A transmural MI = 38 ± 6°, p &lt; 0.01) and lower proportions of right-handed cardiomyocytes (RHM subendocardial MI = 21 ± 5%, RHM transmural MI = 14 ± 5%, p &lt; 0.01) than subendocardial infarct segments. Conclusion Compared to subendocardial MI segments, the diffusion of water molecules is more isotropic in transmurally infarcted myocardium as evidenced by lower FA values, signifying increased structural disarray. The significantly lower E2A values suggest that laminar sheetlets of transmural infarct segments remain fixed at shallower angles during systole and are unable to reach their usual contractile configuration. The lower proportions of RHM on HA maps highlight the significantly greater loss of subendocardial cardiomyocytes in transmural infarct segments. Further studies are required to assess if these segmental changes can be predictive of long-term LV remodelling.

Today's universities are transforming into the entrepreneurial university model. Along with that is a strong innovation in governance towards autonomy and associated with entrepreneurship, innovation and creativity. The article presents research results on the model of the entrepreneurial university and the advanced university governance in terms of structure and management methods to adapt to this model in the world. Through the review of studies on the current situation, the article contributes a number of policy proposals to meet the requirements of university governance innovation for Vietnamese public universities in the context of transition to a model of entrepreneurial university. Keywords University, Entrepreneurial university, University governance, Vietnam public universities. References [1] D.V. Toan, 2020, Factors Affecting Third Mission Implementation and The Challenges for Vietnam’s Universities in The Transitioning Period. 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Urbano, The development of an entrepreneurial university, The Journal of Technology Transfer 37(1) (2010) 43-74. DOI: 10.1007/s10961-010-9171-x.[24] L.K. Sooreh, Salamzadeh, A., Safarzadeh, H. Salamzadeh, Y., Defining and Measuring Entrepreneurial Universities: A Study in Iranian Context Using Importance-Performance Analysis and TOPSIS Technique, Global Business and Management Research: An International Journal, 3(2) (2011) 182-199. [25] J.Y. Farsi, N. Imanipour and A. Salamzadeh, Entrepreneurial university conceptualization: case of developing countries, Global Business and Management Research, 4(2) (2012) 193-204. [26] Y.C. Chang, P.Y. Yang, B.R. Martin, H.R. Chi, T.F. Tsai-Lin, Entrepreneurial universities and research ambidexterity: A multilevel analysis, Technovation 54 (2016) 7-21. http://dx.doi.org/10.1016/j.technovation.2016.02.006[27] G. Dalmarco, W. Hulsink, G.V. Blois, Creating entrepreneurial universities in an emerging economy: Evidencefrom Brazil, Technological Forecasting & Social Change 135 (2018) 99-111. doi:10.1016/j.techfore.2018.04.015.[28] S. Boffo, A. Cocorullo, University Fourth Mission: Spin-offs and Academic Entrenreneurship: Connecting Public Policies with new missions and management issues of universities, Higher Education Forum 16 (2019) 125-142.[29] D.V. Toan, Entrepreneurial Universities and the Development Model for Public Universities in Vietnam, International Journal of Entrepreneurship, 24(1) 2020 1-16. [30] J. Röpke, The Entrepreneurial University, Innovation, academic knowledge creation and regional development in a globalized economy, Working Paper Department of Economics, Philipps- Universität Marburg, Germany: 15, 1998[31] D.V. Toan, H.V. Hai, N.P. Mai, The Role of Entrepreneurship Development in Universities to Promote Knowledge Sharing: The Case of Vietnam National University Hanoi, Proceedings of Asia Pacific Conference on Information Management “Common Platform to A Sustainable Society In The Dynamic Asia Pacific”, VNU Press, Hanoi, October, 2016. [32] D.V. Toan, Development of enterprises in universities and policy implications for university governance reform in Vietnam VNU Journal of Science: Economics and Business, 35(1) (2019) 83-96 (in Vietnamese).[33] P. Zgaga, Higher Education in Transition - Reconsiderations on Higher Education in Europe at the Turn of Millennium, Monographs on Journal of Research in Teacher Education, Ed. Gun-Marie Frånberg, Publisher: Umeå University, 2007. ISBN: 978-91-7264-505-9.[34] J. Fielden, Global Trends in University Governance. Education Working Paper Series, number 9, World Bank, Washington, 2008.[35] A.H. Dooley, The role of academic boards in university governance, Policy paper formulated at the National Conference of Chairs of Academic Boards and Senates, The University of New South Wales, October 2005.[36] A. Lizzio, Student participation in university governance: the role conceptions and sense of efficacy of student representatives on departmental committees, Studies in Higher Education Journal, Taylor & Francis 34(1) (2009) 69-84. https://doi.org/10.1080/03075070802602000.[37] D.V. Toan, Development of Enterprises in Universities: From International Experience to Practices in Vietnam, Vietnam National University Press, Hanoi, 2019, 49-64 (in Vietnamese),.[38] D.V. Toan, H.T.C. Thuong, International experience in university governance and lessons for Vietnam, Economy and Forecast Review 20 (2020) 41-45. [39] D.V. Toan, Business development in universities: International experience and policy recomendation for Vietnam Economy and Forecast Review 35 (2018) 58-60 (in Vietnamese). [40] D.V. Toan, Entrepreneurship in public universities in Vietnam in the context of transition to autonomy (in Vietnamese), Economy and Forecast Review 30 (2019) 111-116.[41] D.V. Toan, University - Enterprise Cooperation in International Context and Implications for Vietnam (in Vietnamese), VNU Journal of Science: Economics and Business 32 (4) (2016) 32-44.

This study develops procedures for cloning ITS and matK genes on six specimens in order to exploit and conserve the genetic resources of H. nepalensis and evaluate its genetic diversity based on molecular markers. The study methods include DNA extraction from dried leaf samples, amplification of ITS and matK regions using PCR, sequencing and comparing with the sequences on Genbank. The study results include a successfully-established process of cloning ITS and matK genes; successful amplification and sequencing of the ITS and matK regions. The results also show that four samples (N1-N4) were 100% homologous to H. nepalensis and H1and H2 samples were 100% homologous to H. helix. The results provide data and tools for further studies of exploitation and development of the H. nepalensis K. Koch genetic resources in Vietnam. Keywords ITS, matK, Hedera nepalensis K. Koch, PCR References [1] V.V. Chi. Dictionary of Vietnamese Medicinal Plants, Publ. House Medicine, Ho Chi Minh City, 2012 (in Vietnamese).[2] D.H. Bich, D.Q. Cuong, B.X. Chuong, N. Thuong, D. T. Dam. The medicinal plants and animals in Vietnam, Hanoi Sci. Technol. Publ. House Hanoi, 2006 (in Vietnamese).[3] A. Sadat, M. Alam, A. Rauf, W. Ullah, Biological screening of ethyl acetate extract of Hedera nepalensis stem, Afr J Pharm Pharmacol, 6 (2012) 2934-2937. https://doi.org/10.5897/AJPP12.828.[4] T. Li, H. Pan, Y. Feng, H. Li, Y. Zhao, Bioactivity-guided isolation of anticancer constituents from Hedera nepalensis K. Koch, S Afr J Bot, 100 (2015) 87-93. https://doi.org/10.1016/j.sajb.2015.05.011.[5] L. Jafri, S. Saleem, N. Ullah, B. Mirza, In vitro assessment of antioxidant potential and determination of polyphenolic compounds of Hedera nepalensis K. Koch, Arab J Chem, 10 (2017) 3699-3706. https://doi.org/10.1016/j.arabjc.2014.05.002. [6] S. Saleem, L. Jafri, I. ul Haq, L.C. Chang, D. Calderwood, B.D. Green, B. Mirza, Plants Fagonia cretica L. and Hedera nepalensis K. Koch contain natural compounds with potent dipeptidyl peptidase-4 (DPP-4) inhibitory activity, J Ethnopharmacol, 156 (2014) 26-32. https://doi.org/10.1016/j.jep.2014.08.017.[7] W.J. Hashmi, H. Ismail, F. Mehmood, B. Mirza, Neuroprotective, antidiabetic and antioxidant effect of Hedera nepalensis and lupeol against STZ+ AlCl 3 induced rats model, DARU, 26 (2018) 179-190. https://doi.org/10.1007/s40199-018-0223-3.[8] H. Ismail, A. Rasheed, I.-u. Haq, L. Jafri, N. Ullah, E. Dilshad, M. Sajid, B. Mirza, Five indigenous plants of Pakistan with Antinociceptive, anti-inflammatory, antidepressant, and anticoagulant properties in Sprague Dawley rats, Evid Based Complement Alternat Med, 2017 (2017). https://doi.org/10.1155/2017/7849501[9] N.D. Thanh. DNA marker techniques in study and selection of plant. Journal of Biology. 36 (2014) 265-294 (in Vietnamese). https://doi.org/10.15625/0866-7160/v36n3.5974.[10] P.Z. Goldstein, R. DeSalle, Review and interpretation of trends in DNA barcoding, Front Ecol Evol, 7 (2019) 302. https://doi.org/10.3389/fevo.2019.00302.[11] S. Abugalieva, L. Volkova, Y. Genievskaya, A. Ivaschenko, Y. Kotukhov, G. Sakauova, Y. Turuspekov, Taxonomic assessment of Allium species from Kazakhstan based on ITS and matK markers, BMC plant biol, 17 (2017) 258. https://doi.org/10.1186/s12870-017-1194-0.[12] R.M. Bhagwat, B.B. Dholakia, N.Y. Kadoo, M. Balasundaran, V.S. Gupta, Two new potential barcodes to discriminate Dalbergia species, PloS one, 10 (2015) e0142965. https://doi.org/10.1371/journal.pone.0142965[13] D. Grivet, R. Petit, Phylogeography of the common ivy (Hedera sp.) in Europe: genetic differentiation through space and time, Mol Ecol, 11 (2002) 1351-1362. https://doi.org/10.1046/j.1365294x.2002.01522.x.[14] R. Li, J. Wen, Phylogeny and biogeography of Dendropanax (Araliaceae), an amphi-Pacific disjunct genus between tropical/subtropical Asia and the Neotropics, Syst Bot, 38 (2013) 536-551. https://doi.org/10.1600/036364413X666606.[15] Y. Sun, D. Skinner, G. Liang, S. Hulbert, Phylogenetic analysis of Sorghum and related taxa using internal transcribed spacers of nuclear ribosomal DNA, ‎Theor Appl Genet, 89 (1994) 26-32. https://doi.org/10.1007/BF00226978[16] P. Cuénoud, V. Savolainen, L.W. Chatrou, M. Powell, R.J. Grayer, M.W. Chase, Molecular phylogenetics of Caryophyllales based on nuclear 18S rDNA and plastid rbcL, atpB, and matK DNA sequences, Am J Bot, 89 (2002) 132-144. https://doi.org/10.3732/ajb.89.1.132.[17] D. Bošeľová, J. Žiarovská, L. Hlavačková, K. Ražná, M. Bežo, Comparative analysis of different methods of Hedera helix DNA extraction and molecular evidence of the functionality in PCR Acta fytotechn zootechn, 19 (2016) 144-149. https://doi.org/10.15414/afz.2016.19.04.144-149.[18] D.D. Long, Comparative analysis of different DNA extraction methods and preliminary analysis of genetic diversity of Hedera nepalensis K. Koch. in Vietnam based on GBSSI marker, VNU Journal of Science: Medical and Pharmaceutical Sciences, 35 (2019) 88-95 (in Vietnamese). https://doi.org/10.25073/2588-1132/vnumps.4165 [19] J.H. Cota-Sánchez, K. Remarchuk, K. Ubayasena, Ready-to-use DNA extracted with a CTAB method adapted for herbarium specimens and mucilaginous plant tissue, Plant Mol Biol Rep, 24 (2006)161. https://doi.org/10.1007/BF02914055.[20] S. Xu, D. Li, J. Li, X. Xiang, W. Jin, W. Huang, X. Jin, L. Huang, Evaluation of the DNA barcodes in Dendrobium (Orchidaceae) from mainland Asia, PloS one, 10 (2015) e0115168. https://doi.org/10.1371/journal.pone.0115168.[21] P. Vargas, H.A. McAllister, C. Morton, S.L. Jury, M.J. Wilkinson, Polyploid speciation in Hedera (Araliaceae): Phylogenetic and biogeographic insights based on chromosome counts and ITS sequences, Pl Syst Evol, 219 (1999) 165-179. https://doi.org/10.1007/BF00985577[22] X. Lei, Y.W. Wang, S.Y. Guan, L.J. Xie, L. Xin, C.Y. Sun, Prospects and problems for identification of poisonous plants in China using DNA barcodes, Biomed Environ Sci, 27 (2014) 794-806. https://doi.org/10.3967/bes2014.115.[23] H. Sun, W. McLewin, M.F. Fay, Molecular phylogeny of Helleborus (Ranunculaceae), with an emphasis on the East Asian‐Mediterranean disjunction, Taxon, 50 (2001) 1001-1018. https://doi.org/10.2307/1224717.

Critique of Ability In July 2008, we could be on the eve of an enormously important shift in disability in Australia. One sign of change is the entry into force on 3 May 2008 of the United Nations convention on the Rights of Persons with Disabilities, which will now be adopted by the Rudd Labor government. Through this, and other proposed measures, the Rudd government has indicated its desire for a seachange in the area of disability. Bill Shorten MP, the new Parliamentary Secretary for Disabilities and Children’s Services has been at pains to underline his commitment to a rights-based approach to disability. In this inaugural speech to Parliament, Senator Shorten declared: I believe the challenge for government is not to fit people with disabilities around programs but for programs to fit the lives, needs and ambitions of people with disabilities. The challenge for all of us is to abolish once and for all the second-class status that too often accompanies Australians living with disabilities. (Shorten, “Address in reply”; see also Shorten, ”Speaking up”) Yet if we listen to the voices of people with disability, we face fundamental issues of justice, democracy, equality and how we understand the deepest aspects of ourselves and our community. This is a situation that remains dire and palpably unjust, as many people with disabilities have attested. Elsewhere I have argued (Goggin and Newell) that disability constitutes a systemic form of exclusion and othering tantamount to a “social apartheid” . While there have been improvements and small gains since then, the system that reigns in Australia is still fundamentally oppressive. Nonetheless, I would suggest that through the rise of the many stranded movements of disability, the demographic, economic and social changes concerning impairment, we are seeing significant changes in how we understand impairment and ability (Barnes, Oliver and Barton; Goggin and Newell, Disability in Australia; Snyder, Brueggemann, and Garland-Thomson; Shakespeare; Stiker). There is now considerable, if still incomplete, recognition of disability as a category that is constituted through social, cultural, and political logics, as well as through complex facets of impairment, bodies (Corker and Shakespeare), experiences, discourses (Fulcher), and modes of materiality and subjectivity (Butler), identity and government (Tremain). Also there is growing awareness of the imbrication of disability and other categories such as sex and gender (Fine and Asch; Thomas), race, age, culture, class and distribution of wealth (Carrier; Cole; Davis, Bending over Backwards, and Enforcing Normalcy; Oliver; Rosenblum and Travis), ecology and war (Bourke; Gerber; Muir). There are rich and wide-ranging debates that offer fundamental challenges to the suffocating grip of the dominant biomedical model of disability (that conceives disability as individual deficit — for early critiques see: Borsay; Walker), as well as the still influential and important (if at times limiting) social model of disability (Oliver; Barnes and Mercer; Shakespeare). All in all,there have been many efforts to transform the social and political relations of disability. If disability has been subject to considerable examination, there has not yet been an extended, concomitant critique of ability. Nor have we witnessed a thoroughgoing recognition of unmarked, yet powerful operations of ability in our lives and thought, and the potential implications of challenging these. Certainly there have been important attempts to reframe the relationship between “ability” and “disability” (for example, see Jones and Mark). And we are all familiar with the mocking response to some neologisms that seek to capture this, such as the awkward yet pointed “differently-abled.” Despite such efforts we lack still a profound critique of ability, an exploration of “able”, the topic that this special issue invites us to consider. If we think of the impact and significance of “whiteness”, as a way to open up space for how to critically think about and change concepts of race; or of “masculinity” as a project for thinking about gender and sexuality — we can see that this interrogation of the unmarked category of “able” and “ability” is much needed (for one such attempt, see White). In this paper I would like to make a small contribution to such a critique of ability, by considering what the concept of innovation and its contemporary rhetorics have to offer for reframing disability. Innovation is an important discourse in contemporary life. It offers interesting possibilities for rethinking ability — and indeed disability. And it is this relatively unexplored prospect that this paper seeks to explore. Beyond Access, Equity & Diversity In this scene of disability, there is attention being given to making long over-due reforms. Yet the framing of many of these reforms, such as the strengthening of national and international legal frameworks, for instance, also carry with them considerable problems. Disability is too often still seen as something in need of remediation, or special treatment. Access, equity, and anti-discrimination frameworks offer important resources for challenging this “special” treatment, so too do the diversity approaches which have supplemented or supplanted them (Goggin and Newell, “Diversity as if Disability Mattered”). In what new ways can we approach disability and policies relevant to it? In a surprisingly wide range of areas, innovation has featured as a new, cross-sectoral approach. Innovation has been a long-standing topic in science, technology and economics. However, its emergence as master-theme comes from its ability to straddle and yoke together previously diverse fields. Current discussions of innovation bring together and extend work on the information society, the knowledge economy, and the relationships between science and technology. We are now familiar for instance with arguments about how digital networked information and communications technologies and their consumption are creating new forms of innovation (Benkler; McPherson; Passiante, Elia, and Massari). Innovation discourse has extended to many other unfamiliar realms too, notably the area of social and community development, where a new concept of social innovation is now proposed (Mulgan), often aligned with new ideas of social entrepreneurship that go beyond earlier accounts of corporate social responsibility. We can see the importance of innovation in the ‘creative industries’ discourses and initiatives which have emerged since the 1990s. Here previously distinct endeavours of arts and culture have become reframed in a way that puts their central achievement of creativity to the fore, and recognises its importance across all sorts of service and manufacturing industries, in particular. More recently, theorists of creative industries, such as Cunningham, have begun to talk about “social network markets,” as a way to understand the new hybrid of creativity, innovation, digital technology, and new economic logics now being constituted (Cunningham and Potts). Innovation is being regarded as a cardinal priority for societies and their governments. Accordingly, the Australian government has commissioned a Review of The National Innovation System, led by Dr Terry Cutler, due to report in the second half of 2008. The Cutler review is especially focussed upon gaps and weaknesses in the Australian innovation system. Disability has the potential to figure very strongly in this innovation talk, however there has been little discussion of disability in the innovation discourse to date. The significance of disability in relation to innovation was touched upon some years ago, in a report on Disablism from the UK Demos Foundation (Miller, Parker and Gillinson). In a chapter entitled “The engine of difference: disability, innovation and creativity,” the authors discuss the area of inclusive design, and make the argument for the “involvement of disabled people to create a stronger model of user design”:Disabled people represented a market of 8.6 million customers at the last count and their experiences aren’t yet feeding through into processes of innovation. But the role of disabled people as innovators can and should be more active; we should include disabled people in the design process because they are good at it. (57) There are two reasons given for this expertise of disabled people in design. Firstly, “disabled people are often outstanding problem solvers because they have to be … life for disabled people at the moment is a series of challenges to be overcome” (57). Secondly, “innovative ideas are more likely to come from those who have a new or different angle on old problems” (57). The paradox in this argument is that as life becomes more equitable for people with disabilities, then these ‘advantages’ should disappear” (58). Accordingly, Miller et al. make a qualified argument, namely that “greater participation of disabled people in innovation in the short term may just be the necessary trigger for creating an altogether different, and better, system of innovation for everyone in the future” (58). The Demos Disablism report was written at a time when rhetorics of innovation were just beginning to become more generalized and mainstream. This was also at a time in the UK, when there was hope that new critical approaches to disability would see it become embraced as a part of the diverse society that Blair’s New Labor Britain had been indicating. The argument Disablism offers about disability and innovation is in some ways a more formalized version of vernacular theory (McLaughlin, 1996). In the disability movement we often hear, with good reason, that people with disability, by dint of their experience and knowledge are well positioned to develop and offer particular kinds of expertise. However, Miller et al. also gesture towards a more generalized account of disability and innovation, one that would intersect with the emerging frameworks around innovation. It is this possibility that I wish to take up and briefly explore here. I want to consider the prospects for a fully-fledged encounter between disability and innovation. I would like to have a better sense of whether this is worth pursuing, and what it would add to our understanding of both disability and innovation? Would the disability perspective be integrated as a long-term part of our systems of innovation rather than, as Miller et al. imply, deployed temporarily to develop better innovation systems? What pitfalls might be bound up with, or indeed be the conditions of, such a union between disability and innovation? The All-Too-Able User A leading area where disability figures profoundly in innovation is in the field of technology — especially digital technology. There is now a considerable literature and body of practice on disability and digital technology (Annable, Goggin, and Stienstra; Goggin and Newell, Digital Disability; National Council on Disability), however for my purposes here I would like to focus upon the user, the abilities ascribed to various kinds of users, and the user with disability in particular. Digital technologies are replete with challenges and opportunities; they are multi-layered, multi-media, and global in their manifestation and function. In Australia, Britain, Canada, the US, and Europe, there have been some significant digital technology initiatives which have resulted in improved accessibility for many users and populations (Annable, Goggin, and Stienstra; National Council on Disability) . There are a range of examples of ways in which users with disability are intervening and making a difference in design. There is also a substantial body of literature that clarifies why we need to include the perspective of the disabled if we are to be truly innovative in our design practices (Annable, Goggin and Stienstra; Goggin and Newell, “Disability, Identity and Interdependence”). I want to propose, however, that there is merit in going beyond recognition of the role of people with disability in technology design (vital and overlooked as it remains), to consider how disability can enrich contemporary discourses on innovation. There is a very desirable cross-over to be promoted between the emphasis on the user-as-expert in the sphere of disability and technology, and on the integral role of disability groups in the design process, on the one hand, and the rise of the user in digital culture generally, on the other. Surprisingly, such connections are nowhere near as widespread and systematic as they should be. It may be that contemporary debates about the user, and about the user as co-creator, or producer, of technology (Haddon et al.; von Hippel) actually reinstate particular notions of ability, and the able user, understood with reference to notions of disability. The current emphasis on the productive user, based as it is on changing understandings of ability and disability, provides rich material for critical revision of the field and those assumptions surrounding ability. It opens up possibilities for engaging more fully with disability and incorporating disability into the new forms and relations of digital technology that celebrate the user (Goggin and Newell, Digital Disability). While a more detailed consideration of these possibilities require more time than this essay allows, let us consider for a moment the idea of a genuine encounter between the activated user springing from the disability movement, and the much feted user in contemporary digital culture and theories of innovation. People with disability are using these technologies in innovative ways, so have much to contribute to wider discussions of digital technology (Annable, Goggin and Stienstra). The Innovation Turn Innovation policy, the argument goes, is important because it stands to increase productivity, which in turn leads to greater international competitiveness and economic benefit. Especially with the emergence of capitalism (Gleeson), productivity has strong links to particular notions of which types of production and produce are valued. Productivity is also strongly conditioned by how we understand ability and, last in a long chain of strong associations, how we as a society understand and value those kinds of people and bodies believed to contain and exercise the ordained and rewarded types of ability, produce, and productivity. Disability is often seen as antithetical to productivity (a revealing text on the contradictions of disability and productivity is the 2004 Productivity Commission Review of the Disability Discrimination Act). When we think about the history of disability, we quickly realize that productivity, and by extension, innovation, are strongly ideological. Ideological, that is, in the sense that these fields of human endeavour and our understanding of them are shaped by power relations, and are built upon implicit ‘ableist’ assumptions about productivity. In this case, the power relations of disability go right to the heart of the matter, highlighting who and what are perceived to be of value, contributing economically and in other ways to society, and who and what are considered as liabilities, as less valued and uneconomical. A stark recent example of this is the Howard government workplace and welfare reforms, which further disenfranchised, controlled, and impoverished people with disability. If we need to rethink our ideas of productivity and ability in the light of new notions of disability, then so too do we need to rethink our ideas about innovation and disability. Here the new discourses of innovation may actually be useful, but also contain limited formulations and assumptions about ability and disability that need to be challenged. The existing problems of a fresh approach to disability and innovation can be clearly observed in the touchstones of national science and technology “success.” Beyond One-Sided Innovation Disability does actually feature quite prominently in the annals of innovation. Take, for instance, the celebrated case of the so-called “bionic ear” (or cochlear implant) hailed as one of Australia’s great scientific inventions of the past few decades. This is something we can find on display in the Powerhouse Museum of Technology and Design, in Sydney. Yet the politics of the cochlear implant are highly controversial, not least as it is seen by many (for instance, large parts of the Deaf community) as not involving people with disabilities, nor being informed by their desires (Campbell, also see “Social and Ethical Aspects of Cochlear Implants”). A key problem with the cochlear implant and many other technologies is that they are premised on the abolition or overcoming of disability — rather than being shaped as technology that acknowledges and is informed by disabled users in their diverse guises. The failure to learn the lessons of the cochlear implant for disability and innovation can be seen in the fact that we are being urged now to band together to support the design of a “bionic eye” by the year 2020, as a mark of distinction of achieving a great nation (2020 Summit Initial Report). Again, there is no doubting the innovation and achievement in these artefacts and their technological systems. But their development has been marked by a distinct lack of consultation and engagement with people with disabilities; or rather the involvement has been limited to a framework that positions them as passive users of technology, rather than as “producer/users”. Further, what notions of disability and ability are inscribed in these technological systems, and what do they represent and symbolize in the wider political and social field? Unfortunately, such technologies have the effect of reproducing an ableist framework, “enforcing normalcy” (Davis), rather than building in, creating and contributing to new modes of living, which embrace difference and diversity. I would argue that this represents a one-sided logic of innovation. A two-sided logic of innovation, indeed what we might call a double helix (at least) of innovation would be the sustained, genuine interaction between different users, different notions of ability, disability and impairment, and the processes of design. If such a two-sided (or indeed many-sided logic) is to emerge there is good reason to think it could more easily do so in the field of digital cultures and technologies, than say, biotechnology. The reason for this is the emphasis in digital communication technologies on decentralized, participatory, user-determined governance and design, coming from many sources. Certainly this productive, democratic, participatory conception of the user is prevalent in Internet cultures. Innovation here is being reshaped to harness the contribution and knowledge of users, and could easily be extended to embrace pioneering efforts in disability. Innovating with Disability In this paper I have tried to indicate why it is productive for discourses of innovation to consider disability; the relationship between disability and innovation is rich and complex, deserving careful elaboration and interrogation. In suggesting this, I am aware that there are also fundamental problems that innovation raises in its new policy forms. There are the issues of what is at stake when the state is redefining its traditional obligations towards citizens through innovation frameworks and discourses. And there is the troubling question of whether particular forms of activity are normatively judged to be innovative — whereas other less valued forms are not seen as innovative. By way of conclusion, however, I would note that there are now quite basic, and increasingly accepted ways, to embed innovation in design frameworks, and while they certainly have been adopted in the disability and technology area, there is much greater scope for this. However, a few things do need to change before this potential for disability to enrich innovation is adequately realized. Firstly, we need further research and theorization to clarify the contribution of disability to innovation, work that should be undertaken and directed by people with disability themselves. Secondly, there is a lack of resources for supporting disability and technology organisations, and the development of training and expertise in this area (especially to provide viable career paths for experts with disability to enter the field and sustain their work). If this is addressed, the economic benefits stand to be considerable, not to mention the implications for innovation and productivity. Thirdly, we need to think about how we can intensify existing systems of participatory design, or, better still, introduce new user-driven approaches into strategically important places in the design processes of ICTs (and indeed in the national innovation system). Finally, there is an opportunity for new approaches to governance in ICTs at a general level, informed by disability. New modes of organising, networking, and governance associated with digital technology have attracted much attention, also featuring recently in the Australia 2020 Summit. Less well recognised are new ideas about governance that come from the disability community, such as the work of Queensland Advocacy Incorporated, Rhonda Galbally’s Our Community, disability theorists such as Christopher Newell (Newell), or the Canadian DIS-IT alliance (see, for instance, Stienstra). The combination of new ideas in governance from digital culture, new ideas from the disability movement and disability studies, and new approaches to innovation could be a very powerful cocktail indeed.Dedication This paper is dedicated to my beloved friend and collaborator, Professor Christopher Newell AM (1964-2008), whose extraordinary legacy will inspire us all to continue exploring and questioning the idea of able. 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