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Статті в журналах з теми "Human skin explant"

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Автор: Grafiati

Опубліковано: 14 березня 2023

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1

Garg, H. G., E. W. Lippay, E. A. Carter, M. B. Donelan, and J. P. Remensnyder. "Proteoglycan synthesis in human skin and burn scar explant cultures." Burns 17, no. 6 (December 1991): 452–57. http://dx.doi.org/10.1016/0305-4179(91)90070-w.

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2

Lemarquand, Alice, Vincent Gauthier, Nicolas Wilkie-Chancellier, and Stéphane Serfaty. "Mesoscopic Monitoring of Human Skin Explants Viscoelastic Properties." Cosmetics 10, no. 1 (January 10, 2023): 13. http://dx.doi.org/10.3390/cosmetics10010013.

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The investigation of the mechanical properties of skin is of great interest for monitoring physiological and pathological changes in the cutaneous barrier function for dermatological and cosmetic issues. Skin constitutes a complex tissue because of its multi-layered organisation. From a rheological point of view, it can be considered to be a soft tissue with viscoelastic properties. In order to characterise ex vivo mechanical properties of skin on the mesoscopic scale, a biosensor including a thickness shear mode transducer (TSM) in contact with a skin explant was used. A specific experimental set-up was developed to monitor continuously and in real-time human skin explants, including the dermis and the epidermis. These were kept alive for up to 8 days. Skin viscoelastic evolutions can be quantified with a multi-frequency impedance measurement (from 5 MHz to 45 MHz) combined with a dedicated fractional calculus model. Two relevant parameters for the non-destructive mesoscopic characterisation of skin explants were extracted: the structural parameter αapp and the apparent viscosity ηapp. In this study, the validity of the biosensor, including repeatability and viability, was controlled. A typical signature of the viscoelastic evolutions of the different cutaneous layers was identified. Finally, monitoring was carried out on stripped explants mimicking a weakened barrier function.

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3

Wolf Horrell, Erin, and John D'Orazio. "UV-independent induction of beta defensin 3 in neonatal human skin explants." F1000Research 3 (November 21, 2014): 288. http://dx.doi.org/10.12688/f1000research.5794.1.

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In order to determine the effect of UV radiation on β-defensin 3 (BD3) expression in human skin, freshly-isolated UV-naïve skin was obtained from newborn male infants undergoing planned circumcision. Skin explants sustained ex vivo dermis side down on RPMI media were exposed to 0.5 kJ/m2 UVB, and biopsies were taken from the explant through 72 hours after radiation. mRNA expression was measured by qRTPCR and normalized to TATA-binding protein. BD3 expression at each time point was compared with an untreated control taken at time 0 within each skin sample. Extensive variability in both the timing and magnitude of BD3 induction across individuals was noted and was not predicted by skin pigment phenotype, suggesting that BD3 induction was not influenced by epidermal melanization. However, a mock-irradiated time course demonstrated UV-independent BD3 mRNA increases across multiple donors which was not further augmented by treatment with UV radiation, suggesting that factors other than UV damage promoted increased BD3 expression in the skin explants. We conclude that BD3 expression is induced in a UV-independent manner in human skin explants processed and maintained in standard culture conditions, and that neonatal skin explants are an inappropriate model with which to study the effects of UV on BD3 induction in whole human skin.

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4

Wolf Horrell, Erin, and John D'Orazio. "UV-independent induction of beta defensin 3 in neonatal human skin explants." F1000Research 3 (February 19, 2015): 288. http://dx.doi.org/10.12688/f1000research.5794.2.

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Анотація:

In order to determine the effect of UV radiation on β-defensin 3 (BD3) expression in human skin, freshly-isolated UV-naïve skin was obtained from newborn male infants undergoing planned circumcision. Skin explants sustained ex vivo dermis side down on RPMI media were exposed to 0.5 kJ/m2 UVB, and biopsies were taken from the explant through 72 hours after radiation. mRNA expression was measured by qRTPCR and normalized to TATA-binding protein. BD3 expression at each time point was compared with an untreated control taken at time 0 within each skin sample. Extensive variability in both the timing and magnitude of BD3 induction across individuals was noted and was not predicted by skin pigment phenotype, suggesting that BD3 induction was not influenced by epidermal melanization. However, a mock-irradiated time course demonstrated UV-independent BD3 mRNA increases across multiple donors which was not further augmented by treatment with UV radiation, suggesting that factors other than UV damage promoted increased BD3 expression in the skin explants. We conclude that BD3 expression is induced in a UV-independent manner in human skin explants processed and maintained in standard culture conditions, and that neonatal skin explants are an inappropriate model with which to study the effects of UV on BD3 induction in whole human skin.

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5

Frade, Marco Andrey Cipriani, Thiago Antônio Moretti de Andrade, Andréia Fernanda Carvalho Leone Aguiar, Flávia Araújo Guedes, Marcel Nani Leite, Williane Rodrigues Passos, Eduardo Barbosa Coelho, and Pranab Kummar Das. "Prolonged viability of human organotypic skin explant in culture method (hOSEC)." Anais Brasileiros de Dermatologia 90, no. 3 (June 2015): 347–50. http://dx.doi.org/10.1590/abd1806-4841.20153645.

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6

Stoll, Stefan W., Sanjay Kansra, and James T. Elder. "Keratinocyte outgrowth from human skin explant cultures is dependent upon p38 signaling." Wound Repair and Regeneration 11, no. 5 (September 2003): 346–53. http://dx.doi.org/10.1046/j.1524-475x.2003.11506.x.

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7

Lindsay, CD, and P. Rice. "Assessment of the biochemical effects of percutaneous exposure of sulphur mustard in an in vitro human skin system." Human & Experimental Toxicology 15, no. 3 (March 1996): 237–44. http://dx.doi.org/10.1177/096032719601500309.

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1 Sulphur mustard (HD) is a potent chemical warfare agent which causes incapacitating blisters on human skin. There is no specific pretreatment nor therapy against this agent and the mechanism of dermo-epidermal cleavage is unclear. The aim of this study was to use a human skin explant system to determine the consequences of percuta neous exposure to HD. 2 Increased activities of serine proteases associated with blistering disorders in humans were detected from human skin explants after exposure to HD. The most consistent response and the highest protease activities measured were found for trypsin. This class of enzyme is therefore implicated in the dermo-epidermal separation which is associated with blistering in humans following exposure to HD. 3 An inflammatory response was observed in the skin explants exposed to HD. At low doses of HD it was characterised by the presence of neutrophils in the papillary dermis, culminating in the infiltration of the epidermis by these inflammatory cells at higher concen trations of HD. A variety of other histopathological changes in the explants was found such as focal dermo- epidermal separation, nuclear pyknosis and perinuclear vacuolation. 4 The study indicates that full thickness human skin explants can be used to investigate various aspects of the possible pathogenesis of HD-induced skin damage, in cluding the associated inflammatory response.

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8

Jacobs, J. J. L., C. Lehé, K. D. A. Cammans, P. K. Das, and G. R. Elliott. "An in vitro model for detecting skin irritants: methyl green-pyronine staining of human skin explant cultures." Toxicology in Vitro 16, no. 5 (October 2002): 581–88. http://dx.doi.org/10.1016/s0887-2333(02)00039-5.

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9

Roychowdhury, Sanjoy, Albert E. Cram, Al Aly, and Craig K. Svensson. "Detection of Haptenated Proteins in Organotypic Human Skin Explant Cultures Exposed to Dapsone." Drug Metabolism and Disposition 35, no. 9 (June 6, 2007): 1463–65. http://dx.doi.org/10.1124/dmd.107.015560.

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10

Danso, Mogbekeloluwa O., Vincent van Drongelen, Aat Mulder, Gert Gooris, Jeroen van Smeden, Abdoelwaheb El Ghalbzouri, and Joke A. Bouwstra. "Exploring the potentials of nurture: 2nd and 3rd generation explant human skin equivalents." Journal of Dermatological Science 77, no. 2 (February 2015): 102–9. http://dx.doi.org/10.1016/j.jdermsci.2014.12.002.

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11

Gonnet, J., H. Perrin, A. J. Hutton, D. Boccara, O. Bonduelle, M. Mimoun, M. Atlan, A. Soria, and B. Combadière. "Interleukin-32 promotes detachment and activation of human Langerhans cells in a human skin explant model." British Journal of Dermatology 179, no. 1 (May 28, 2018): 145–53. http://dx.doi.org/10.1111/bjd.16721.

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12

Nikolić, Nenad, Philip Kienzl, Pooja Tajpara, Martin Vierhapper, Johannes Matiasek, and Adelheid Elbe-Bürger. "The Antiseptic Octenidine Inhibits Langerhans Cell Activation and Modulates Cytokine Expression upon Superficial Wounding with Tape Stripping." Journal of Immunology Research 2019 (March 3, 2019): 1–11. http://dx.doi.org/10.1155/2019/5143635.

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Анотація:

Ideal agents for the topical treatment of skin wounds should have antimicrobial efficacy without negative influence on wound healing. Octenidine (OCT) has become a widely used antiseptic in professional wound care, but its influence on several components of the wound healing process remains unclear. In the present study, we have used a superficial wound model using tape stripping on human full-thickness skin ex vivo to investigate the influence of OCT on epidermal Langerhans cells (LCs) and cytokine secretion pattern of skin cells during wound healing in a model without disruption of the normal skin structure. Histological and immunofluorescence studies showed that OCT neither altered human skin architecture nor the viability of skin cells upon 48 hours of culture in unwounded or wounded skin. The epidermis of explants and LCs remained morphologically intact throughout the whole culture period upon OCT treatment. OCT inhibited the upregulation of the maturation marker CD83 on LCs and prevented their emigration in wounded skin. Furthermore, OCT reduced both pro- and anti-inflammatory mediators (IL-8, IL-33, and IL-10), while angiogenesis and growth factor mediators (VEGF and TGF-β1) remained unchanged in skin explant cultures. Our data provide novel insights into the host response to OCT in the biologically relevant environment of viable human (wounded) skin.

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13

Oliver, E. "P192 DEVELOPMENT OF A FUNCTIONAL HUMAN SKIN EXPLANT MODEL TO STUDY MAST CELL ACTIVATION." Annals of Allergy, Asthma & Immunology 127, no. 5 (November 2021): S54. http://dx.doi.org/10.1016/j.anai.2021.08.163.

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14

Forsberg, Sofi, and Ola Rollman. "Re-epithelialization from human skin explant cultures is promoted by ligand-activated HER3 receptor." Journal of Dermatological Science 59, no. 1 (July 2010): 7–15. http://dx.doi.org/10.1016/j.jdermsci.2010.03.017.

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15

Reisch, Nicole, Angela E. Taylor, Edson F. Nogueira, Daniel J. Asby, Vivek Dhir, Andrew Berry, Nils Krone, et al. "Alternative pathway androgen biosynthesis and human fetal female virilization." Proceedings of the National Academy of Sciences 116, no. 44 (October 14, 2019): 22294–99. http://dx.doi.org/10.1073/pnas.1906623116.

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Androgen biosynthesis in the human fetus proceeds through the adrenal sex steroid precursor dehydroepiandrosterone, which is converted to testosterone in the gonads, followed by further activation to 5α-dihydrotestosterone in genital skin, thereby facilitating male external genital differentiation. Congenital adrenal hyperplasia due to P450 oxidoreductase deficiency results in disrupted dehydroepiandrosterone biosynthesis, explaining undervirilization in affected boys. However, many affected girls are born virilized, despite low circulating androgens. We hypothesized that this is due to a prenatally active, alternative androgen biosynthesis pathway from 17α-hydroxyprogesterone to 5α-dihydrotestosterone, which bypasses dehydroepiandrosterone and testosterone, with increased activity in congenital adrenal hyperplasia variants associated with 17α-hydroxyprogesterone accumulation. Here we employ explant cultures of human fetal organs (adrenals, gonads, genital skin) from the major period of sexual differentiation and show that alternative pathway androgen biosynthesis is active in the fetus, as assessed by liquid chromatography–tandem mass spectrometry. We found androgen receptor expression in male and female genital skin using immunohistochemistry and demonstrated that both 5α-dihydrotestosterone and adrenal explant culture supernatant induce nuclear translocation of the androgen receptor in female genital skin primary cultures. Analyzing urinary steroid excretion by gas chromatography–mass spectrometry, we show that neonates with P450 oxidoreductase deficiency produce androgens through the alternative androgen pathway during the first weeks of life. We provide quantitative in vitro evidence that the corresponding P450 oxidoreductase mutations predominantly support alternative pathway androgen biosynthesis. These results indicate a key role of alternative pathway androgen biosynthesis in the prenatal virilization of girls affected by congenital adrenal hyperplasia due to P450 oxidoreductase deficiency.

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16

Boccara, David, Béhazine Combadière, and Angèle Soria. "Maintenance of human skin tissue and cellularity after engraftment in immunodeficient mouse model." Journal of Experimental and Applied Animal Sciences 2, no. 2 (June 6, 2017): 165. http://dx.doi.org/10.20454/jeaas.2017.1280.

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A lack of an appropriate animal model to study human skin in vivo remains a major problem. Human skin explant can be conserved only few days in culture medium. For these reasons, it is necessary to developed humanized mice model with human skin graft for studying of skin innate and adaptive responses. Humanized mice model with human skin engraftment have been Non-obese diabetic-SCID gamma chain null mice (NSG, NOD.Cg-PrkdcscidIL2rg tmlWjl/Sz) were engrafted with fresh human skin obtained from healthy volunteers undergoing plastic surgery of breast, abdomen or face lifting. Eighty-two mice were engrafted with 16 different human skin with standard operating procedure. A failure of human skin engraftment was found in 21 (25.6%) of the engrafted mice, due to partial or total human skin necrosis. We observed very good or good engrafted skin behavior in 61 humanized mice (78.6%), with preservation of the structures of the human skin compared to non-engrafted human skin and conservation of the major skin resident immune Antigen Presenting Cells 1 to 3-week post-graft. Here, we described the standard operating procedure, and performance of human skin engraftment in NOD-Scid IL2rgamma(null) mouse model with the maintenance of skin epidermal tissue and cells

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17

Helbig, Doris, Anne Moebius, Jan C. Simon, and Uwe Paasch. "Nonablative skin rejuvenation devices and the role of heat shock protein 70: results of a human skin explant model." Journal of Biomedical Optics 15, no. 3 (2010): 038002. http://dx.doi.org/10.1117/1.3449736.

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18

Marshall, Scott R., Xiao N. Wang, Chris M. McArthy, Graham Jackson, and Anne M. Dickinson. "Downregulation of Interleukin-13 by PUVA Treatment in a Human Model of Graft-Versus-Host Disease May Highlight the Importance of This Cytokine in the Mechanism of Extracorporeal Phototherapy Treatment." Blood 106, no. 11 (November 16, 2005): 5234. http://dx.doi.org/10.1182/blood.v106.11.5234.5234.

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Abstract Graft-versus-Host disease (GvHD) remains the most serious complication following haemopoietic stem cell transplantation, with an incidence of 40–60% and can be fatal in up to 50% of cases. Extracorporeal phototherapy (ECP) is a novel treatment of both acute and chronic GvHD involving psoralen and UVA (PUVA) treatment of peripheral blood cells with high reported success even in those resistant to conventional immunosuppressive treatments. ECP appears to induce selective immune suppression without increased rates of infection or disease relapse. It is well tolerated by patients with minimal side effects and therefore is emerging as a highly desirable therapy. However, its mechanism of action remains poorly understood. An in vitro human skin explant model for GvHD has been used to study the role of cytokine changes induced by ECP. The model involves sensitising donor lymphocytes with recipient lymphocytes in vitro in a primary mixed lymphocyte reaction and then evaluating the secondary response on recipient skin biopsies by grading the graft-versus-host reactivity (grades I-IV) histopathologically using the Lerner grading system for GvHD. The assay has been proved to be superior to other assays used to predict clinical GvHD and has been used to investigate the pathophysiology of GvHD. Skin explant supernatants were collected and stored at −80 C and measured in batches using a multiplex Th1/Th2 cytokine assay. In 20 experiments where PUVA treatment of responder cells caused down regulation of GvHR in the skin explant model, statistically significant downregulation of IL-13, IL-5 and IFNgamma (p=<0.05) was demonstrated. No changes of IL-6, IL-1b, IL-10, IL-2, IL-4 or TNFalpha levels were demonstrated. Further evidence for the importance of IL-13 was highlighted in 3 skin explant assays where no downregulation of GvHR was seen with PUVA treatment and IL-13 levels remained high. Jordon et Al (Blood Jan2004) have documented the association between IL-13 production and the incidence of acute GvHD. Our data adds further evidence to challenge the notion that acute GvHD is purely a Th1-type cytokine response and that there may be a significant link between the Th2-type cytokine IL-13 with acute GvHD. Furthermore, downregulation of IL-13 and by PUVA may be important in the control of GvHD. More skin explant assays will be performed to consolidate these provisional findings.

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19

Anderson, Michele J., Maren L. David, Matt Scholz, Sally J. Bull, Dan Morse, Michelle Hulse-Stevens, and Marnie L. Peterson. "Efficacy of Skin and Nasal Povidone-Iodine Preparation against Mupirocin-Resistant Methicillin-Resistant Staphylococcus aureus and S. aureus within the Anterior Nares." Antimicrobial Agents and Chemotherapy 59, no. 5 (March 2, 2015): 2765–73. http://dx.doi.org/10.1128/aac.04624-14.

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ABSTRACTMupirocin decolonization of nasalStaphylococcus aureusprior to surgery decreases surgical-site infections; however, treatment requires 5 days, compliance is low, and resistance occurs. In 2010, 3M Company introduced povidone-iodine (PVP-I)-based skin and nasal antiseptic (Skin and Nasal Prep [SNP]). SNP has rapid, broad-spectrum antimicrobial activity. We tested SNP's efficacy using full-thickness tissue (porcine mucosal [PM] and human skin) explant models and human subjects. Prior to or following infection with methicillin-resistantStaphylococcus aureus(MRSA) (mupirocin sensitive and resistant), explants were treated with Betadine ophthalmic preparation (Bet), SNP, or mupirocin (Bactroban nasal ointment [BN]) or left untreated. One hour posttreatment, explants were washed with phosphate-buffered saline (PBS) plus 2% mucin. One, 6, or 12 h later, bacteria were recovered and enumerated. Alternatively, following baseline sampling, human subjects applied two consecutive applications of SNP or saline to their anterior nares. One, 6, and 12 h after application of the preparation (postprep), nasal swabs were obtained, andS. aureuswas enumerated. We observed that treatment of infected PM or human skin explants with SNP resulted in >2.0 log10CFU reduction in MRSA, regardless of mupirocin sensitivity, which was significantly different from the values for BN- and Bet-treated explants and untreated controls 1 h, 6 h, and 12 h after being washed with PBS plus mucin. Swabbing the anterior nares of human subjects with SNP significantly reduced residentS. aureuscompared to saline 1, 6, and 12 h postprep. Finally, pretreatment of PM explants with SNP, followed by a mucin rinse prior to infection, completely prevented MRSA infection. We conclude that SNP may be an attractive alternative for reducing the bioburden of anterior nares prior to surgery.

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20

Sviland, L., and A. M. Dickinson. "A human skin explant model for predicting graft-versus-host disease following bone marrow transplantation." Journal of Clinical Pathology 52, no. 12 (December 1, 1999): 910–13. http://dx.doi.org/10.1136/jcp.52.12.910.

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21

Kim, Hayeon, Hyunju Woo, Seoungwoo Shin, Deokhoon Park, and Eunsun Jung. "The Potential Application of Ecklonia cava Extract in Scalp Protection." Cosmetics 7, no. 1 (January 20, 2020): 9. http://dx.doi.org/10.3390/cosmetics7010009.

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The scalp is exposed to environmental hazards including airborne pollutants, which exert adverse effects on skin health. Therefore, compounds for defending skin from pollutants have attracted interest in the cosmeceutical community. We investigated whether Ecklonia cava exhibited prophylactic effects against urban pollutants by measuring cell viability and cell cycle distribution in human follicle dermal papilla cells (HFDPC). The effect of E. cava on pollutant-induced damage to skin barrier was determined by measuring filaggrin and MMP-1 expression in both keratinocytes and in a skin explant model. In a clinical trial, the effect of E. cava on scalp skin of patients with scalp scale was observed by evaluating hydration and redness after 4 weeks of daily treatment with a shampoo containing E. cava extract. E. cava extract recovered the loss of cell viability and abnormal cell cycle distribution induced by urban pollutants in HFDPCs. It also attenuated pollutant-induced damage to skin barrier by decreasing MMP-1 and increasing filaggrin expression in keratinocytes and the epidermis of skin explants. Moreover, E. cava showed soothing effects on human scalp by increasing hydration and decreasing redness in a clinical trial. Collectively, E. cava extract may be a good candidate for therapeutic applications designed to repair or protect hair scalp.

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22

Hsieh, Ming-Fa. "ID3024 Polymeric nanoparticles display bactericidal effect and selective fermentation for the treatment of acne vulgaris." Biomedical Research and Therapy 4, S (September 5, 2017): 34. http://dx.doi.org/10.15419/bmrat.v4is.244.

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The use of antibiotics in the treatment of acne in specific group (pregnant women) of patients can lead to serious complications. We have previously demonstrated that the nanoparticles made of block copolymers of poly (ethylene glycol) and poly(e-caprolactone) can inhibit the growth of Propionibacterium acnes (P. acnes), a bacterium highly associated with the progress of acne vulgaris in the human skin [Polymers 2016; 8, 321]. To reduce the amount of antibiotics used in the treatment of skin acne, we have further demonstrated that a bacterium in the human skin microbiome can utilize PEG-based polymers to produce various short-chain fatty acids (SCFAs) which suppressed the growth of P. acnes. PEG-based polymers were chosen as selective fermentation initiators which specifically induced the fermentation of the skin commensal bacterium but not P. acnes. Interestingly, PEG-based polymers can efficiently suppress the growth of P. acnes. An acne ex vivo explant was established by using acne biopsies collected from patients with acne vulgaris at the early and middle stages. The levels of pro-inflammatory interleukin (IL)-8 cytokine in early- and middle-staged acnes were significantly higher than those in healthy skins. Incubation of acne ex vivo explants with sucrose remarkably reduced the level of IL-8 and the number of P. acnes. Results from mouse studies revealed that PEG-based polymer functions as antibiotic adjuvants which can considerably reduce the effective doses of clindamycin, a clinically-used acne antibiotic

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23

Chato-Astrain, Jesús, David Sánchez-Porras, Óscar Darío García-García, Claudia Vairo, María Villar-Vidal, Silvia Villullas, Indalecio Sánchez-Montesinos, Fernando Campos, Ingrid Garzón, and Miguel Alaminos. "Improvement of Cell Culture Methods for the Successful Generation of Human Keratinocyte Primary Cell Cultures Using EGF-Loaded Nanostructured Lipid Carriers." Biomedicines 9, no. 11 (November 6, 2021): 1634. http://dx.doi.org/10.3390/biomedicines9111634.

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Human skin keratinocyte primary cultures can be established from skin biopsies with culture media containing epithelial growth factor (EGF). Although current methods are efficient, optimization is required to accelerate the procedure and obtain these cultures in less time. In the present study, we evaluated the effect of novel formulations based on EGF-loaded nanostructured lipid carriers (NLC). First, biosafety of NLC containing recombinant human EGF (NLC-rhEGF) was verified in immortalized skin keratinocytes and cornea epithelial cells, and in two epithelial cancer cell lines, by quantifying free DNA released to the culture medium. Then we established primary cell cultures of human skin keratinocytes with basal culture media (BM) and BM supplemented with NLC-rhEGF, liquid EGF (L-rhEGF), or NLC alone (NLC-blank). The results showed that cells isolated by enzymatic digestion and cultured with or without a feeder layer had a similar growth rate regardless of the medium used. However, the explant technique showed higher efficiency when NLC-rhEGF culture medium was used, compared to BM, L-rhEGF, or NLC-blank. Gene expression analysis showed that NLC-rhEGF was able to increase EGFR gene expression, along with that of other genes related to cytokeratins, cell–cell junctions, and keratinocyte maturation and differentiation. In summary, these results support the use of NLC-rhEGF to improve the efficiency of explant-based methods in the efficient generation of human keratinocyte primary cell cultures for tissue engineering use.

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24

Wang, Xiao N., Lisbet Sviland, Adebo J. Ademokun, Janice Dunn, Gary Cavanagh, Stephen J. Proctor, and Anne M. Dickinson. "CELLULAR ALLOREACTIVITY OF HUMAN CORD BLOOD CELLS DETECTED BY T-CELL FREQUENCY ANALYSIS AND A HUMAN SKIN EXPLANT MODEL1." Transplantation 66, no. 7 (October 1998): 903–9. http://dx.doi.org/10.1097/00007890-199810150-00015.

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25

Lehé, Cynthia L., John J. L. Jacobs, Graham R. Elliott, and Pranab K. Das. "A Two-centre Evaluation of the Human Organotypic Skin Explant Culture Model for Screening Contact Allergens." Alternatives to Laboratory Animals 31, no. 6 (November 2003): 553–61. http://dx.doi.org/10.1177/026119290303100604.

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26

Sørensen, Isabella S., Christian Janfelt, Mette Marie B. Nielsen, Rasmus W. Mortensen, Nina Ø. Knudsen, André H. Eriksson, Anders J. Pedersen, and Kim T. Nielsen. "Combination of MALDI-MSI and cassette dosing for evaluation of drug distribution in human skin explant." Analytical and Bioanalytical Chemistry 409, no. 21 (July 7, 2017): 4993–5005. http://dx.doi.org/10.1007/s00216-017-0443-2.

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27

Wang, X. N., M. Collin, L. Sviland, S. R. Marshall, G. H. Jackson, U. Schulz, E. Holler, et al. "Skin explant model of human GVHD: Prediction of clinical outcome and correlation with biological risk factors." Biology of Blood and Marrow Transplantation 12, no. 2 (February 2006): 60–61. http://dx.doi.org/10.1016/j.bbmt.2005.11.188.

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28

Lemaire, Géraldine, Malvina Olivero, Virginie Rouquet, Alain Moga, Aurélie Pagnon, Valérie Cenizo, and Pascal Portes. "Neryl acetate, the major component of Corsican Helichrysum italicum essential oil, mediates its biological activities on skin barrier." PLOS ONE 18, no. 3 (March 3, 2023): e0268384. http://dx.doi.org/10.1371/journal.pone.0268384.

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Corsican Helichrysum italicum essential oil (HIEO) is characterized by high concentrations of neryl acetate, and we previously demonstrated that Corsican HIEO increases the expression of genes that are part of the differentiation complex (involucrin, small proline rich proteins, late cornified envelope, S100 protein family). The biological activities of HIEO and neryl acetate (NA) were compared to identify how NA contributes to HIEO activity on human skin. NA, as a part component of HIEO, was tested on skin explant models for 24 hours and 5 days in comparison with HIEO. We analyzed the biological regulations in the skin explant by transcriptomic analysis, skin barrier protein immunofluorescence, lipid staining and ceramide analysis by liquid chromatography-mass spectrometry. Transcriptomic analysis revealed that 41.5% of HIEO-modulated genes were also regulated by NA and a selected panel of genes were confirmed by qquantitative reverse transcription PCR analysis. Those genes are involved in epidermal differentiation, skin barrier formation and ceramide synthesis. Involucrin (IVL), involved in formation of the cornified envelope (CE), was upregulated at both gene and protein levels after 24 hours and 5 days respectively. After 5 days of treatment, total lipids and ceramides were also increased. Our results demonstrate that NA mediates a large part of Corsican HIEO activity on skin barrier formation.

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Klicznik, Maria M., Peter A. Morawski, Barbara Höllbacher, Suraj R. Varkhande, Samantha J. Motley, Leticia Kuri-Cervantes, Eileen Goodwin, et al. "Human CD4+CD103+ cutaneous resident memory T cells are found in the circulation of healthy individuals." Science Immunology 4, no. 37 (July 5, 2019): eaav8995. http://dx.doi.org/10.1126/sciimmunol.aav8995.

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Tissue-resident memory T cells (TRM) persist locally in nonlymphoid tissues where they provide frontline defense against recurring insults. TRM at barrier surfaces express the markers CD103 and/or CD69, which function to retain them in epithelial tissues. In humans, neither the long-term migratory behavior of TRM nor their ability to reenter the circulation and potentially migrate to distant tissue sites has been investigated. Using tissue explant cultures, we found that CD4+CD69+CD103+ TRM in human skin can down-regulate CD69 and exit the tissue. In addition, we identified a skin-tropic CD4+CD69−CD103+ population in human lymph and blood that is transcriptionally, functionally, and clonally related to the CD4+CD69+CD103+ TRM population in the skin. Using a skin xenograft model, we confirmed that a fraction of the human cutaneous CD4+CD103+ TRM population can reenter circulation and migrate to secondary human skin sites where they reassume a TRM phenotype. Thus, our data challenge current concepts regarding the strict tissue compartmentalization of CD4+ T cell memory in humans.

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Broady, Raewyn, Sarah Q. Crome, Jessie Yu, Jan P. Dutz, and Megan K. Levings. "Development of a Modified Skin Explant Assay to Study Treg Suppression of Th17 Cell Mediated GvHD in the Skin." Blood 112, no. 11 (November 16, 2008): 5434. http://dx.doi.org/10.1182/blood.v112.11.5434.5434.

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Abstract Acute graft versus host disease (aGVHD) following haematopoietic stem cell transplantation (HCT) occurs when donor T cells infused with the graft recognise and react to histo-incompatible recipient antigens causing tissue damage. Historically, the inflammatory response in aGVHD was attributed to alloreactive CD4+ T helper and CD8+ cytotoxic T cells and alterations in cytokine production. Recently, a new CD4+ T cell subset, characterised by IL-17 production has been identified. TH17 cells produce high levels of proinflammatory cytokines, including IL-17A, IL-17F, and IL-22, and have been implicated in solid organ rejection and more recently a number of murine studies suggest that Th17 cells play a role in the development of aGVHD. It is well known that FOXP3+ regulatory T cells (Tregs) are critical for the maintenance of self-tolerance, and control the immune response to alloantigens. Murine studies have shown that adoptive transfer of these cells can prevent acute GVHD whereas selective depletion leads to an increased severity. In humans, Tregs also appear to control acute GVHD as they occur at a lower frequency in the peripheral blood patients with aGVHD compared to patients without GVHD. These findings have led to active interest into the use of these cells to prevent or decrease GVHD following allogeneic HCT. It has been reported that in vitro, Th17 cells are resistant to Treg cell mediated suppression of proliferation and IL-17 production, suggesting that the effector functions of Th17 cells might not be susceptible to Treg-cell-mediated inhibition. If true, this would suggest that Treg-based therapies might not be effective at limiting Th17-cell-mediated tissue damage. However, there is currently no evidence regarding whether Treg cells affect the phenotype or function of Th17 cells in tissues. Understanding the interactions between suppressive Tregs and pro-inflammatory T effectors in tissues that are targets of aGVHD, such as the skin, is critical to better define the potential of Tregs as adoptive therapy for the prevention or treatment of aGVHD. In order to address this question, we developed two methods to generate human Th17 cells, one based on over-expression of RORC2 and the other on sorting CCR4+CCR6+CD4+ T cells. We found that ectopic expression of RORC induces a cytokine and chemokine receptor profile analogous to in vivo differentiated Th17 cells. Although expression of RORC2 made CD4+ T cells resistant to Treg-cell mediated suppression of proliferation and IL-17 production, production of IFN-g, TNF-a and IL-6 could be suppressed in these Th17-like cells. In order to further delineate the functional consequence of the interaction between Treg and Th17 cells in tissues we developed a modified the human skin explant model that involves culture of 4 mm punch biopsies of skin with ex vivo Th17 cells (CCR4+CCR6+CD4+ T cells), RORC2 transduced CD4+ T cells, or controls, in the presence or absence of Treg and grading the graft-versus-host reactivity (grades I–IV) histopathologically. Preliminary data suggest that Th17 cells cause significant tissue destruction in this skin explant model, and experiments are ongoing to determine whether Treg cells can counteract these effects.

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Jarvis, M., U. Schulz, A. M. Dickinson, L. Sviland, G. Jackson, A. Konur, X. N. Wang, et al. "The detection of apoptosis in a human in vitro skin explant assay for graft versus host reactions." Journal of Clinical Pathology 55, no. 2 (February 1, 2002): 127–32. http://dx.doi.org/10.1136/jcp.55.2.127.

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Kim, Kang-Hoon, Ji Hoon Jung, Won-Seok Chung, Chang-Hun Lee та Hyeung-Jin Jang. "Ferulic Acid Induces Keratin 6α via Inhibition of Nuclear β-Catenin Accumulation and Activation of Nrf2 in Wound-Induced Inflammation". Biomedicines 9, № 5 (22 квітня 2021): 459. http://dx.doi.org/10.3390/biomedicines9050459.

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Injured tissue triggers complex interactions through biological process associated with keratins. Rapid recovery is most important for protection against secondary infection and inflammatory pain. For rapid wound healing with minimal pain and side effects, shilajit has been used as an ayurvedic medicine. However, the mechanisms of rapid wound closure are unknown. Here, we found that shilajit induced wound closure in an acute wound model and induced migration in skin explant cultures through evaluation of transcriptomics via microarray testing. In addition, ferulic acid (FA), as a bioactive compound, induced migration via modulation of keratin 6α (K6α) and inhibition of β-catenin in primary keratinocytes of skin explant culture and injured full-thickness skin, because accumulation of β-catenin into the nucleus acts as a negative regulator and disturbs migration in human epidermal keratinocytes. Furthermore, FA alleviated wound-induced inflammation via activation of nuclear factor erythroid-2-related factor 2 (Nrf2) at the wound edge. These findings show that FA is a novel therapeutic agent for wound healing that acts via inhibition of β-catenin in keratinocytes and by activation of Nrf2 in wound-induced inflammation.

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Zhang, Bin, Ruenn Chai Lai, Wei Kian Sim, Andre Boon Hwa Choo, Ellen Birgit Lane, and Sai Kiang Lim. "Topical Application of Mesenchymal Stem Cell Exosomes Alleviates the Imiquimod Induced Psoriasis-Like Inflammation." International Journal of Molecular Sciences 22, no. 2 (January 13, 2021): 720. http://dx.doi.org/10.3390/ijms22020720.

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Severe psoriasis, a chronic inflammatory skin disease is increasingly being effectively managed by targeted immunotherapy but long-term immunotherapy poses health risk and loss of response. Therefore, there is a need for alternative therapy strategies. Mesenchymal stem/stromal cell (MSC) exosomes are widely known for their potent immunomodulatory properties. Here we investigated if topically applied MSC exosomes could alleviate psoriasis-associated inflammation. Topically applied fluorescent exosomes on human skin explants were confined primarily to the stratum corneum with <1% input fluorescence exiting the explant over a 24-h period. Nevertheless, topically applied MSC exosomes in a mouse model of imiquimod (IMQ) psoriasis significantly reduced IL-17 and terminal complement activation complex C5b-9 in the mouse skin. MSC exosomes were previously shown to inhibit complement activation, specifically C5b-9 complex formation through CD59. Infiltration of neutrophils into the stratum corneum is characteristic of psoriasis and neutrophils are a major cellular source of IL-17 in psoriasis through the release of neutrophil extracellular traps (NETs). We propose that topically applied MSC exosomes inhibit complement activation in the stratum corneum and this alleviates IL-17 release by NETS from neutrophils that accumulate in and beneath the stratum corneum.

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Wawersik, Matthew J., Stacy Mazzalupo, Diem Nguyen, and Pierre A. Coulombe. "Increased Levels of Keratin 16 Alter Epithelialization Potential of Mouse Skin Keratinocytes In Vivo and Ex Vivo." Molecular Biology of the Cell 12, no. 11 (November 2001): 3439–50. http://dx.doi.org/10.1091/mbc.12.11.3439.

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The process of wound repair in adult skin is complex, involving dermal contraction and epithelial migration to repair the lesion and restore the skin's barrier properties. At the wound edge, keratinocytes undergo many changes that engender an epithelialization behavior. The type II keratin 6 and type I keratins 16 and 17 are induced well before cell migration begins, but the role of these proteins is not understood. Forced expression of human K16 in skin epithelia of transgenic mice has been shown to cause dose-dependent skin lesions concomitant with alterations in keratin filament organization and in cell adhesion. Here we show, with the use of a quantitative assay, that these transgenic mice show a delay in the closure of full-thickness skin wounds in situ compared with wild-type and low-expressing K16 transgenic mice. We adapted and validated an ex vivo skin explant culture system to better assess epithelialization in a wound-like environment. Transgenic K16 explants exhibit a significant reduction of keratinocyte outgrowth in this setting. This delay is transgene dose-dependent, and is more severe when K16 is expressed in mitotic compared with post-mitotic keratinocytes. Various lines of evidence suggest that the mechanism(s) involved is complex and not strictly cell autonomous. These findings have important implications for the function of K16 in vivo.

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Swamy, Vinutha Eshwara, Nikhil Shetty, Jayaprakasha Shetty, Veena Shetty, Tonita Noronha, and Mohana Kumar Basavarajappa. "Growth kinetics and phenotypic markers expression in human dermal stem cells." Research Journal of Biotechnology 16, no. 12 (November 25, 2021): 24–29. http://dx.doi.org/10.25303/1612rjbt2429.

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Human dermal stem cells (DSCs) have generated significant interest in the field of regenerative medicine due to their prospects of autologous transplantation. The present study evaluated the growth kinetics and phenotypic markers expression in human DSCs. The primary cultures of DSCs (n=3) were established by explant culture and characterization of the cells was carried out by assessing morphology, viability, proliferation rate, population doubling time (PDT), cell cycle status and the expression of cell surface markers such as CD29, CD73, CD90 and CD166. The cells released from tissue explants showed spindleshaped fibroblast morphology with the mean percentage viability varying between 93.43% and 100% from passages 1 to 4. DSCs displayed a strong and steady proliferative potential with an average PDT of 42.55 hrs. Cell cycle profile of DSCs demonstrated the majority of cells (59.80% to 76.29%) at G0/G1 phase. Further, the phenotypic profile of markers confirmed the stromal origin of DSCs by exhibiting positivity for CD29, CD73, CD90 and CD166. In conclusion, the growth kinetics and expression of phenotypic markers are consistent with the notion that skin dermis contains a population of stem cells and can serve as a potential autologous source for therapeutic applications.

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36

Bibby, L., A. Ribeiro, S. Ahmed, and A. M. Dickinson. "A novel in-vitro human skin explant test to predict adverse immune reactions to biologics and aggregated monoclonal antibodies." Toxicology Letters 295 (October 2018): S66—S67. http://dx.doi.org/10.1016/j.toxlet.2018.06.072.

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37

Helbig, Doris, Marc Bodendorf, Ulf Anderegg, Jan C. Simon, and Uwe Paasch. "A human skin explant model to study molecular changes in response to fractional photothermolysis: Spatio-temporal expression of HSP70." Medical Laser Application 25, no. 3 (August 2010): 173–80. http://dx.doi.org/10.1016/j.mla.2009.12.002.

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38

Wang, Xiao-nong, Matthew Collin, Lisbet Sviland, Scott Marshall, Graham Jackson, Ute Schulz, Ernst Holler, et al. "Skin Explant Model of Human Graft-versus-Host Disease: Prediction of Clinical Outcome and Correlation with Biological Risk Factors." Biology of Blood and Marrow Transplantation 12, no. 2 (February 2006): 152–59. http://dx.doi.org/10.1016/j.bbmt.2005.09.018.

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39

Patrick, B., G. Camille, J. Carine, D. Leslie, R. Nicolas, B. Pascale, D. Hélène, and B. Sandrine. "646 Anti-pollution and anti-oxidant effects of two formulations containing Taraxacum officinalis root extract on human skin explant." Journal of Investigative Dermatology 138, no. 5 (May 2018): S110. http://dx.doi.org/10.1016/j.jid.2018.03.655.

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40

Donaldson, D. J., J. T. Mahan, and G. N. Smith. "Newt epidermal cell migration in vitro and in vivo appears to involve Arg-Gly-Asp-Ser receptors." Journal of Cell Science 87, no. 4 (May 1, 1987): 525–34. http://dx.doi.org/10.1242/jcs.87.4.525.

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The effect of a synthetic peptide consisting of Arg-Gly-Asp-Ser (RGDS), the amino acid sequence representing the fibroblast attachment site in fibronectin (FN), was tested on migrating newt epidermal cells. In one approach, skin explants were placed on the bottom of plastic dishes coated with human FN, human fibrinogen (FGN), human serum spreading factor (SF), or bovine type I collagen. The explants were then incubated overnight in serum-free medium with or without RGDS. In these experiments exposure to 50 micrograms ml-1 of RGDS reduced migration over FN, FGN and SF to 2–7% of control levels. Two peptides structurally dissimilar to RGDS (Val-Gly-Ser-Glu and Thr-Pro-Arg-Lys), and two that are structurally similar (Lys-Gly-Asp-Ser and Arg-Gly-Glu-Ser), had no effect on explant migration even when used at concentrations higher than 50 micrograms ml-1. Upon removal of the RGDS peptide, inhibited explants quickly recovered. In collagen-coated dishes 50 micrograms ml-1 of RGDS was much less effective than in dishes coated with the other substrates. Raising the RGDS concentration in collagen-coated dishes tenfold did not greatly increase the RGDS effect. When added to the medium bathing wounded limbs, 50 micrograms ml-1 of RGDS only moderately inhibited wound closure. This concentration of peptide, however, severely inhibited migration from skin explants in newt-plasma-coated-dishes and migration over pieces of newt-plasma-coated plastic placed under one edge of a skin wound. Increasing the RGDS concentration to 500 micrograms ml-1 resulted in almost total suppression of wound closure. Wounds exposed to this same concentration of Lys-Gly-Asp-Ser closed normally. These results indicate that newt epidermal cells possess RGDS receptors and that these receptors are involved in epidermal wound closure in vivo and in migration from skin explants onto plastic coated with FN, FGN, SF and collagen. The relative RGDS-insensitivity of wound closure in vivo and in migration from explants onto collagen may reflect in these instances the presence of a relatively high density of RGDS receptor binding sites on the substrate; the presence of RGDS receptor binding sites of relatively high affinity; or the participation of receptors other than those involved in migration over plastic coated with FN, FGN or SF.

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41

Peramo, Antonio, Cynthia L. Marcelo, Steven A. Goldstein, and David C. Martin. "Improved Preservation of the Tissue Surrounding Percutaneous Devices by Hyaluronic Acid and Dermatan Sulfate in a Human Skin Explant Model." Annals of Biomedical Engineering 38, no. 3 (December 18, 2009): 1098–110. http://dx.doi.org/10.1007/s10439-009-9872-1.

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42

Gonzalez-Aspajo, German, Haouaria Belkhelfa, Laïla Haddioui-Hbabi, Geneviève Bourdy, and Eric Deharo. "Sacha Inchi Oil ( Plukenetia volubilis L.), effect on adherence of Staphylococus aureus to human skin explant and keratinocytes in vitro." Journal of Ethnopharmacology 171 (August 2015): 330–34. http://dx.doi.org/10.1016/j.jep.2015.06.009.

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43

Caserta, A. V., J. E. Neil, J. Volmer, L. Ring, J. Lenn, and M. Brown. "932 Development of an early wound model in human ex vivo skin explant as a preclinical model of wound healing." Journal of Investigative Dermatology 139, no. 5 (May 2019): S161. http://dx.doi.org/10.1016/j.jid.2019.03.1008.

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44

Kalyuzhnaya, L. I., M. O. Sokolova, V. E. Chernov, D. A. Zemlyanoy, S. V. Chebotarev, N. I. Chalisova, A. A. Kondratenko, Yu S. Grechanaya, N. V. Edomenko, and E. I. Alexander-Sinclair. "The effect of the cell-free matrix of the human umbilical cord on the growth dynamics and viability of cultured human and animal cells ex vivo." Genes & Cells 16, no. 3 (September 15, 2021): 72–79. http://dx.doi.org/10.23868/202110010.

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Due to its extraembryonic origin, the umbilical cord retains regenerative molecules of the fetal phenotype The cell-free matrix of the human umbilical cord as the basis of various tissue-engineering structures (TES) for regenerative medicine can contribute to the repopulation of transplanted matrices by patient-specific cells. The aim of the work was to study in vitro the cytotoxic properties of a tissue-engineered lyophilized cell-free matrix made by the detergent method from the human umbilical cord Warton jelly with respect to human skin fibroblasts, as well as the effect of this matrix on the viability and growth of cells of the cerebral cortex, liver, spleen, articular cartilage, heart and skin of various laboratory animals during organotypic cultivation. Using the MTT test, it was revealed that the tissue-engineered acellular lyophilized matrix of the human umbilical cord does not show a toxic effect on human dermal fibroblast. When using the method of organ cultures, differences were revealed in the influence of the tissue-engineered matrix on the migration of cells from explants of tissues of different organs of various laboratory animals, their growth dynamics and viability, which may be due to species specificity, which must be taken into account when choosing test systems. The most noticeable positive effect of the matrix on the growth dynamics and viability of articular cartilage explant cells was found. Adhesion to the matrix of cells of the cerebral cortex of the Vietnamese lop-bellied pig as well as cells of the spleen of a guinea pig was revealed. The cell-free human umbilical cord matrix can be used in the creation of acellular matrices for the treatment of deep dermal and articular cartilage damages.

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Singh, Mahipal, and Arunachala Shiv Kumar Reddy Kutagulla. "PSVI-7 Postmortem in Vitro Culture of Cells from Visceral Organs Stored at 4°C in Small Ruminants." Journal of Animal Science 100, Supplement_3 (September 21, 2022): 403. http://dx.doi.org/10.1093/jas/skac247.738.

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Abstract Exploration of cell viability and their regenerative capacity in postmortem tissues has a huge potential for tissue/organ transplantation and cell therapies in both human and animals. Proliferative stem cells have been recovered from refrigerated postmortem skin tissue in bovine, goat and sheep, and from ligaments in horses. However, it is not clearly known whether live and proliferative cells can be recovered from visceral organs that stay comparatively warmer than other tissues after death, and whether the recovery time can be extended after refrigeration of individual organs. To achieve this goal, six different organ tissues i.e. lung, heart, oviduct, liver, kidney and skin from goats were procured from slaughter house and stored in the laboratory at 4°C. Ten small sized (2-3mm2) explants from each of the five organs were then cultured in two 35mm dishes for each time point in DMEM media supplemented with 10% fetal bovine serum. The tissues were cultured on day 0, 3, 6, 9 and 12. The dishes were observed under inverted microscope and any explant exhibiting outgrowth of cells on day 10-12 of culture was considered positive. Our results exhibited outgrowth of cells around 3 (lung, oviduct, skin) of the 6 organs tested. Kidney, liver and heart tissues did not show any outgrowth. Interestingly, oviduct and skin, tissues exhibited outgrowth even after 9 days of postmortem storage in refrigerator, whereas lung tissues show growth only up to 6 days of storage. Experiments to test the cell survival limits on more animals of different age groups and characterization of recovered cells are underway.

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Dickinson, A. M., J. Cavet, H. Cullup, X. N. Wang, L. Sviland, and P. G. Middleton. "GvHD risk assessment in hematopoietic stem cell transplantation: role of cytokine gene polymorphisms and an in vitro human skin explant model." Human Immunology 62, no. 11 (November 2001): 1266–76. http://dx.doi.org/10.1016/s0198-8859(01)00324-x.

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47

Tajpara, P., C. Schuster, P. Kienzl, M. Mildner, and A. Elbe-Buerger. "366 Epidermal delivery of polyinosinic-polycytidylic acid in a human skin explant model activates the MDA5/MAVS pathway in Langerhans cells." Journal of Investigative Dermatology 137, no. 10 (October 2017): S255. http://dx.doi.org/10.1016/j.jid.2017.07.561.

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48

Caldwell, C., A. Krug, N. Goldstein, M. Koster, B. Ward, D. R. Roop, D. Norris, and S. Birlea. "1250 Development of an ex vivo human skin explant model to examine candidate gene functions in the hair follicle and epidermis." Journal of Investigative Dermatology 138, no. 5 (May 2018): S212. http://dx.doi.org/10.1016/j.jid.2018.03.1265.

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Turner, Brie, Shaheda Ahmed, Sarah Pagan, Jean Norden, Matthew Collin, Anne M. Dickinson, and Xiao Nong Wang. "Regulatory T Cells Prevent Effector T Cell Infiltration Into GvHD Target Tissue by Affecting Chemokine Signaling." Blood 114, no. 22 (November 20, 2009): 1341. http://dx.doi.org/10.1182/blood.v114.22.1341.1341.

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Abstract Abstract 1341 Poster Board I-363 Introduction Graft versus host disease (GVHD) following haematopoietic stem cell transplantation is often first observed in the skin; a primary target organ of GVHD. GVH related tissue damage in the skin is mainly driven by infiltrating alloreactive cytotoxic effector T cells, facilitated by a cascade of cytokines and chemokines. Our recently published observations showed that addition of regulatory T cells (Treg) suppressed skin GVH tissue damage mediated by alloreactive CD8+ T cells in an in vitro human GVHD skin explant model [1]. The current study investigated the role of Treg in modulating effector T cell infiltration into skin, it's consequence on the severity of skin GVH histopathology and the possible changes of effector T cell production and expression of chemokines and chemokine receptors. Methods CD8+ T cells, monocyte derived dendritic cells (mDC) and natural Treg (CD4+CD2highFOXP3+) were generated as previously described [1]. In an in vitro human GVHD skin explant model, CD8+ T cells and ex vivo expanded Treg obtained from buffy coats were used as “donor” cells. mDC and skin biopsies obtained from HLA unmatched unrelated normal volunteers acted as “recipient” tissues. “Donor” CD8+ T cells primed with “recipient” mDC in the presence or absence of Treg were co-cultured with “recipient” skin. The severity of histopathological GVH skin damage was scored as grade 0 to grade IV using a clinically validated scoring system. The number of infiltrating CD8+ T cells in skin was evaluated using immunohistochemistry then correlated to the severity of skin GVH histopathology. The gene expression of selected chemokines and chemokine receptors in alloreactive CD8+ effector T cells was analysed using quantitative RT-PCR. The effector T cell expression of chemokine receptors was assessed using flow cytometry. The secretion of selected chemokines into the culture supernatants was quantified using BD cytometric bead array kit. Results The percentage of infiltrating effector T cells in skin was significantly associated with the severity of skin GVH histopathology (2.6±0.8, 12.6±3.1 and 27.2±2.7 for skin sections with GVH histopathology grade I, II and III-IV; p=0.017 and 0.021 respectively). The percentage of skin infiltrating CD8+ T cells was significantly reduced by the presence of Treg (24.8±3.7 vs 11.58±1.8, p=0.011, n=13) which correlated with Treg mediated suppression of skin GVH histopathology (p<0.0001, n=13). The presence of Treg also down-regulated effector T cell expression of chemokine/chemokine receptor genes (CCL3, CCL5, CCR4, CCR5, CXCL10 and CXCL11) that are involved in the recruitment of effector T cells to GVH target tissues. Further analysis indicated a trend toward reduced effector T cell surface expression of CCR4 and CCR5 (31.9±4.3 vs 12.5±2.7; 13.7±1.8 vs 8.8±4.1 respectively, n=3) in the presence of Treg. There was also a reduction in CX3CR1 and cutaneous lymphocyte associated antigen (CLA) (11.1±1.9 vs 2.2±0.6; 34.6%±7.0 vs 11.6±2.2 respectively, n=3). The effector T cell surface expression of CXCR3, CCR2 and CCR10 was very low regardless of the presence or absence of Treg. Release of CCL3, CCL5, CXCL9 and CXCL10 into the culture supernatant was strongly suppressed by the presence of Treg (813±122 vs 77±30; 816±248 vs 405±148; 5517±967 vs 974±540 and 858±209 vs 195±57, p=0.001, 0.065, 0.002 and 0.015 respectively, n=8). However, CXCL11 and CX3CL1 levels were below detectable limits regardless of the presence or absence of Treg. Conclusions Ex vivo expanded natural Treg can inhibit CD8+ effector T cell infiltration into skin which correlated with Treg suppression of cytotoxic T cell mediated skin GVH histopathology in an in vitro human GVHD model. This effect may be attributed to a decrease in chemokine and chemokine receptor interactions mediated by Treg. These observations indicate a potential mechanism for Treg mediated GVHD suppression. [1] X N Wang et al., Transplantation 2009 Disclosures No relevant conflicts of interest to declare.

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Kawamura, Tatsuyoshi, Shannon E. Bruce, Awet Abraha, Makoto Sugaya, Oliver Hartley, Robin E. Offord, Eric J. Arts, Peter A. Zimmerman, and Andrew Blauvelt. "PSC-RANTES Blocks R5 Human Immunodeficiency Virus Infection of Langerhans Cells Isolated from Individuals with a Variety of CCR5 Diplotypes." Journal of Virology 78, no. 14 (July 15, 2004): 7602–9. http://dx.doi.org/10.1128/jvi.78.14.7602-7609.2004.

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ABSTRACT Topical microbicides that effectively block interactions between CCR5+ immature Langerhans cells (LC) residing within genital epithelia and R5 human immunodeficiency virus (HIV) may decrease sexual transmission of HIV. Here, we investigated the ability of synthetic RANTES analogues (AOP-, NNY-, and PSC-RANTES) to block R5 HIV infection of human immature LC by using a skin explant model. In initial experiments using activated peripheral blood mononuclear cells, each analogue compound demonstrated marked antiviral activity against two R5 HIV isolates. Next, we found that 20-min preincubation of skin explants with each RANTES analogue blocked R5 HIV infection of LC in a dose-dependent manner (1 to 100 nM) and that PSC-RANTES was the most potent of these compounds. Similarly, preincubation of LC with each analogue was able to block LC-mediated infection of cocultured CD4+ T cells. Competition experiments between primary R5 and X4 HIV isolates showed blocking of R5 HIV by PSC-RANTES and no evidence of increased propagation of X4 HIV, data that are consistent with the specificity of PSC-RANTES for CCR5 and the CCR5+ CXCR4− phenotype of immature LC. Finally, when CCR5 genetic polymorphism data were integrated with results from the in vitro LC infection studies, PSC-RANTES was found to be equally effective in inhibiting R5 HIV in LC isolated from individuals with CCR5 diplotypes known to be associated with low, intermediate, and high cell surface levels of CCR5. In summary, PSC-RANTES is a potent inhibitor of R5 HIV infection in immature LC, suggesting that it may be useful as a topical microbicide to block sexual transmission of HIV.

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