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Articles de revues sur le sujet "Ex Vivo Vein Culture System"
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Cialdai, Francesca, Stefano Bacci, Virginia Zizi, Aleandro Norfini, Michele Balsamo, Valerio Ciccone, Lucia Morbidelli et al. « Optimization of an Ex-Vivo Human Skin/Vein Model for Long-Term Wound Healing Studies. Ground Preparatory Activities for the ‘Suture in Space’ Experiment Onboard the International Space Station ». International Journal of Molecular Sciences 23, no 22 (16 novembre 2022) : 14123. http://dx.doi.org/10.3390/ijms232214123.
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This study is preliminary to an experiment to be performed onboard the International Space Station (ISS) and on Earth to investigate how low gravity influences the healing of sutured human skin and vein wounds. Its objective was to ascertain whether these tissue explants could be maintained to be viable ex vivo for long periods of time, mimicking the experimental conditions onboard the ISS. We developed an automated tissue culture chamber, reproducing and monitoring the physiological tensile forces over time, and a culture medium enriched with serelaxin (60 ng/mL) and (Zn(PipNONO)Cl) (28 ng/mL), known to extend viability of explanted organs for transplantation. The results show that the human skin and vein specimens remained viable for more than 4 weeks, with no substantial signs of damage in their tissues and cells. As a further clue about cell viability, some typical events associated with wound repair were observed in the tissue areas close to the wound, namely remodeling of collagen fibers in the papillary dermis and of elastic fibers in the vein wall, proliferation of keratinocyte stem cells, and expression of the endothelial functional markers eNOS and FGF-2. These findings validate the suitability of this new ex vivo organ culture system for wound healing studies, not only for the scheduled space experiment but also for applications on Earth, such as drug discovery purposes.
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Prandi, Francesca, Marco Piola, Monica Soncini, Claudia Colussi, Yuri D’Alessandra, Eleonora Penza, Marco Agrifoglio et al. « Adventitial Vessel Growth and Progenitor Cells Activation in an Ex Vivo Culture System Mimicking Human Saphenous Vein Wall Strain after Coronary Artery Bypass Grafting ». PLOS ONE 10, no 2 (17 février 2015) : e0117409. http://dx.doi.org/10.1371/journal.pone.0117409.
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Maffei, S., G. Galeati, G. Pennarossa, T. A. L. Brevini et G. Gandolfi. « 188 DEVELOPMENT OF AN EFFECTIVE WHOLE-OVARY PERFUSION SYSTEM ». Reproduction, Fertility and Development 27, no 1 (2015) : 185. http://dx.doi.org/10.1071/rdv27n1ab188.
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The different structures of a mammalian ovary require complex 3-dimensional interactions to function properly. It is difficult to access the ovary in vivo and to study its physiology in vitro, it is necessary to dissect its different parts and culture them individually. Although informative, this approach prevents the understanding of the role played by their interactions. Perfusion systems are available for ovaries of laboratory animals while organs of larger species have been maintained in culture only for a few hours. This has prompted us to develop a system that can preserve the function of a whole sheep ovary for a few days ex vivo so that it is available for analysis in controlled conditions. Twenty-four sheep ovaries were collected at the local abattoir; 18 were assigned randomly to 3 experimental groups (media A, B, and C) and 6 were immediately fixed in 10% formaldehyde and used as fresh controls. Whole ovaries were cultured for up to 4 days using a semi-open perfusion system. Organs were perfused through the ovarian artery, at a flow rate of 1.5 mL min–1 with basal medium (M199, 25 mM HEPES, 2 mM l-glutamine and 100 µg mL–1 antibiotic-antimycotic solution) supplemented with 0.4% fatty acid free BSA (medium A); or 0.4% BSA heat shock fraction (medium B); or 10% FBS, 50 ng mL–1 IGF-1, and 50 mg bovine insulin (medium C). Ovaries were stimulated with FSH (Folltropin®-V, Bioniche Animal Health Inc., Belleville, Ontario, Canada) changing medium in a pulsatile manner (1 mg mL–1 for 2 h; 0.5 mg mL–1 for 2 h; 0 mg mL–1 for 20 h), with the same cycle repeated each day of culture. At every change, aliquots were collected for oestradiol (E2) and progesterone (P4) quantification. After culture, ovaries were examined for follicular morphology, cell proliferation, and apoptotic rate. Statistical analysis was performed using one-way ANOVA (SPSS 20, IBM, Armonk, NY, USA). In media A and B, all morphological parameters showed a small but significant decrease compared to fresh control, only after 3 days of culture. The different BSA in medium B did not affect follicle morphology but significantly increased cell proliferation (medium A, 28.59 ± 3.26%; medium B, 32.04 ± 2.67%) and decreased apoptosis (medium A, 32.51 ± 5.92%; medium B, 24.55 ± 2.55%). In both media, steroid concentration increased after FSH pulses (E2 range 1.95–10.50 pg mL–1; P4 range 0.34–3.08 ng mL–1), reaching levels similar to those measurable in peripheral plasma. The presence of FBS, IGF-1, and insulin in medium C allowed extension of the culture period to 4 days with a percentage of intact follicles comparable to that observed after 3 days in media A and B. Moreover, proliferation rates were comparable to fresh controls. Steroid pattern changed with P4 values dropping close to zero (range 0.03–1.18 ng mL–1) and E2 level (range 23.59–94.98 pg mL–1) increasing 10-fold, achieving a concentration similar to that measured in the ovarian vein around oestrous. Our data indicate that it is possible to support viability of large animal whole ovaries for up to 4 days, providing a physiologically relevant model for studying ovarian functions in vitro. Research was supported by AIRC IG 10376 and by the Carraresi Foundation.
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Chan, Tim, Cheryl Bolinger, Sean Scott, Mengyan Du, Carol Poortman, Byron Koenitzer, Taranjit Athwal et al. « Abstract 2821 : Incorporation of intrinsic checkpoint blockade enhances functionality of multigenic autologous UltraCAR-T® cells manufactured using non-viral gene delivery and rapid manufacturing process ». Cancer Research 82, no 12_Supplement (15 juin 2022) : 2821. http://dx.doi.org/10.1158/1538-7445.am2022-2821.
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Abstract Current chimeric antigen receptor (CAR)-T manufacturing utilizes viral vectors and extensive ex vivo expansion at large central facilities leading to an exhausted CAR-T phenotype, high costs and long vein-to-vein times. While allogeneic CAR-T can reduce delays in patient treatment, they require extensive manipulation of donor cells, severe lymphodepletion and demonstrate short persistence limiting their therapeutic window. The UltraCAR-T® platform is designed to overcome these limitations by utilizing non-viral multigene delivery and a rapid, decentralized manufacturing process without ex vivo activation or expansion of T cells. Patient’s own T cells are collected and manufactured at the medical center and re-infused one day after gene transfer. Here we describe the next generation UltraCAR-T platform that addresses the inhibitory tumor microenvironment by incorporating a novel mechanism for intrinsic downregulation of one or more checkpoint inhibitor (CPI) genes. Our design achieves intrinsic CPI blockade without gene editing and is aimed at avoiding systemic toxicity and the high cost of combining CPI antibodies. Next generation UltraCAR-T cells simultaneously express CAR, membrane bound IL-15 (mbIL15), kill switch, and incorporate intrinsic CPI blockade. To illustrate the ability of this platform, we designed exemplary non-viral transposons to generate UltraCAR-T cells against multiple tumor targets incorporating intrinsic blockade of either one (PD-1) or two (PD-1 and TIGIT) CPI genes. Healthy donor T cells were transfected using the UltraPorator™ electroporation system to manufacture UltraCAR-T cells without ex vivo activation or expansion. The co-expression of CAR, mbIL15 and kill switch was confirmed by flow cytometry and western blot. Activated UltraCAR-T showed significant reduction in CPI gene expression compared to control CAR-T cells lacking the CPI blockade and did not show unintended off-target activity. Downregulation of CPI gene(s) on UltraCAR-T enhanced cytotoxicity and inflammatory cytokine production, especially at low effector to target (E:T) cell ratios, when co-cultured with PD-L1+/CD155+ tumor cells. Single-cell cytokine proteomics showed significant increase in polyfunctionality of UltraCAR-T with intrinsic downregulation of CPI gene(s). In vivo, a single infusion of receptor tyrosine kinase-like orphan receptor 1 (ROR1)-specific UltraCAR-T with intrinsic PD-1 blockade resulted in rapid expansion, an increase in preferred T cell memory (TSCM/TCM) populations, and significantly improved overall survival of ROR1+ PD-L1+ tumor bearing mice. These preclinical data highlight the improved efficacy of incorporating intrinsic CPI blockade in UltraCAR-T cells using non-viral gene delivery and an established rapid, decentralized manufacturing process. Citation Format: Tim Chan, Cheryl Bolinger, Sean Scott, Mengyan Du, Carol Poortman, Byron Koenitzer, Taranjit Athwal, Lindsey Shepard, R. Daniel Slone, Shourik Dutta, Steven Zilko, James M. Dunleavey, Giorgio Zenere, Jacques Plummer, Bernward Klocke, Christian Zinser, Shamim Ahmad, Douglas E. Brough, Rutul R. Shah, Helen Sabzevari. Incorporation of intrinsic checkpoint blockade enhances functionality of multigenic autologous UltraCAR-T® cells manufactured using non-viral gene delivery and rapid manufacturing process [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2821.
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Jackman, Rachael P., Marcus O. Muench, John W. Heitman, Susanne Marschner, Raymond P. Goodrich et Philip J. Norris. « Prevention of Alloimmunization and Induction of Partial Tolerance Following Transfusion with Pathogen Reduced Platelets in Mice ». Blood 118, no 21 (18 novembre 2011) : 718. http://dx.doi.org/10.1182/blood.v118.21.718.718.
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Abstract Abstract 718 Introduction: The presence of donor white blood cells (WBC) in transfused blood products can induce alloimmunization, and reducing or eliminating this response may prove to be of clinical benefit. The use of a pathogen reduction method based on UV light illumination in the presence of riboflavin has been shown to induce changes in WBCs that result in a failure to bind to, or induce proliferation of allogeneic PBMCs in vitro. In addition, a study in rats has shown a reduction in alloimmunization in vivo using this treatment. Transfusion of cells illuminated with UV light at other doses without riboflavin has been shown to induce some degree of tolerance with a reduced antibody response to subsequent allogeneic transfusions. We sought to assess both the degree of alloimmunization in mice given pathogen reduced versus untreated allogeneic platelets, as well as determine if cells from mice given pathogen reduced platelets exhibited signs of tolerance ex vivo. Methods: Peripheral blood was collected from C57Bl/6 and Balb/cJ mice into CPDA-1, and platelet rich plasma (PRP) was prepared by gentle centrifugation. WBCs were isolated from the remainder of the blood and were added back to a portion of the PRP to generate either WBC-enriched or WBC-poor PRP. These products were either left untreated or pathogen reduced using the Mirasol pathogen reduction technology system, which uses a combination of riboflavin and UV illumination. These products were transfused via tail vein injection into Balb/cJ mice. Two weeks after transfusion the treated mice were sacrificed, and peripheral blood and spleens were collected. Serum levels of circulating alloantibodies were measured by flow cytometry. Splenocytes were cultured for 48 hours in the presence or absence of C57Bl/6 splenocytes, and levels of secreted cytokines were measured in culture supernatants using multiplexing techniques. Groups were compared using one-way ANOVA with Tukey's multiple comparison post-test, α=0.05. Results: Mice given allogeneic PRP transfusions had significantly elevated levels of alloantibodies compared with non-transfused control mice, whereas mice given syngeneic PRP or pathogen reduced PRP did not. Mice given either the WBC-enriched PRP or WBC-poor PRP generated alloantibodies, though higher levels of antibodies were observed with WBC-enriched PRP. Levels of IFN-γ, TNF-α, IL-10 and GM-CSF were significantly higher following secondary allogeneic challenge of cells from mice given untreated allogeneic PRP compared with those given no transfusion or syngeneic PRP, but not with those given pathogen reduced PRP. Levels of IL-1β, IL-4, IL-5, IL-6, IL-12(p70), and IL-13 were significantly reduced following secondary allogeneic challenge of cells from mice given pathogen reduced allogeneic PRP compared with those given no transfusion or syngeneic PRP. Conclusions: Treatment of allogeneic PRP with riboflavin and UV light prior to transfusion blocks alloimmunization in mice. Furthermore, secondary cytokine responses to allogeneic cells ex vivo are reduced, in some cases bellow the levels observed in cells from mice without prior exposure, suggesting induction of tolerance. Disclosures: Marschner: CaridianBCT Biotechnologies: Employment. Goodrich:CaridianBCT Biotechnologies: Employment. Norris:CaridianBCT Biotechnologies: Consultancy, Research Funding.
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Gardner, Rebecca, Corinne Summers, Dan Weber, Michael C. Jensen et Colleen Delaney. « Lentiviral Transduction Of Notch Ligand Expanded Cord Blood HSC’s To Express a CD19-Specific CAR Generates NK and Myeloid Progeny With Antitumor Activity ». Blood 122, no 21 (15 novembre 2013) : 900. http://dx.doi.org/10.1182/blood.v122.21.900.900.
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Abstract Over the past decade, significant progress has been made using chimeric antigen receptors (CAR) to redirect the specificity of T cells against an array of cell surface tumor antigens. Using this same technology, we hypothesized that CD34+ hematopoietic stem and progenitor cells (HSPC) derived from cord blood (CB) could be transduced with lentivirus to efficiently express a CAR specific for CD19 that would enable targeted lysis through its continued expression in differentiated progeny effector cells. As previously reported, the Notch-mediated ex vivo expansion of CB HSPC is a clinically validated cell therapy product that is well tolerated, can be given off the shelf without HLA matching, and provides transient myeloid engraftment in both the HCT and intensive chemotherapy setting. CB-derived CD34+ cells are selected using the Miltenyi CliniMACS or AutoMACS platform and plated in StemSpan serum free expansion media supplemented with IL-6, IL-3, TPO, Flt3L, and SCF and in the presence of immobilized Notch ligand and retronectin. The cells are cultured for 14-16 days resulting in >150 fold expansion of CD34+ cells on average and a final heterogeneous cell product consisting of 14.5% (range 6.2-26) CD34+, 20.5% (6-36) CD15+, 11.3 (1.8-23) CD14+, 5.4%(2.2-13.6) CD56+CD16+, and 0.1(0-0.2)CD20+ which is cryopreserved for future clinical use. Importantly, no T cells are derived from this culture system. Off the shelf expanded units have been infused into 45 patients and no serious adverse events have been noted except for one allergic reaction attributed to DMSO. Additionally, there has been no persistent engraftment beyond day 180 in the HCT setting and 14 days post infusion in the chemotherapy setting. To engineer anti-CD19 activity into this product, cells are transduced at an MOI of 3 on day 3 of culture using a self inactivating lentivirus that encodes for a second generation CD19 CAR with 41BB costimulation as well as an EGFR tag which lacks an intracellular signaling domain and serves as a marker of transduction and a selection marker. Transduction efficiency ranges from 20-40% and equally transduces the CD34+ and CD34- populations. Copy number analysis demonstrates between 1-4 copies/cell. Expression of the transgene does not affect the final culture phenotype at 14 days and transgene expression is seen in all cell subsets and is stable over the culture period. Due to concerns of expression of the CAR on HSC with potential signaling capacity, irradiated LCL was added on day 7 of culture at a 1:1 ratio to provide antigen stimulation and did not negatively affect the growth of the cultures, nor the expression of the transgene. Functional NK cells were derived in vitro from the end culture population by an additional week of culture with RPMI supplemented with L-glutamine, human serum, IL-2 and IL-15. The CD19-CAR expressing NK cells had enhanced cytotoxic activity in a CRA against LCL compared with non-transduced NK cells (50 v 30%) whereas both killed K562 targets equally (75 v 80%). Ongoing experiments are being conducted to further evaluate the expression, signaling capacity and function of the CAR on each of the cell compartments. The NOG mouse model was used for in vivo functional assays to look at effects of CD19 CAR expression on human repopulating ability and to assess anti-CD19 activity. NOG mice underwent sub-lethal irradiation followed by tail vein injection of the expanded progeny of 20,000 selected CD34+ CB cells. There was no effect on early engraftment of transduced cells, however there was no detectable CD19 engraftment in the mice transplanted with CD19 CAR expressing cells compared with untransduced cells, in which the CD19 population was >20% of the engrafted cells, indicating anti-CD19 activity. NK cell populations were increased using NS0-IL15 secreting cells, irradiated and injected subcutaneously three times per week starting at week 3 to provide enhanced effector function. Current studies are ongoing to evaluate the anti-CD19 activity using the Raji CD19+ tumor model in the NOG mice. These initial results are promising as a way to engineer a graft versus leukemia effect into cord blood transplant. Furthermore, transduction of a CD19 CAR into our existing universal donor expanded CB cell therapy product allows for infusion of an anti-CD19 cell product to be given immediately following identification of a patient with clinical need for therapy, for example one in relapse or with persistent MRD. Disclosures: Jensen: ZetaRx: Consultancy, Equity Ownership, Membership on an entity’s Board of Directors or advisory committees, Patents & Royalties, Research Funding.
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Nishimoto, Mitsutaka, Ke Zeng, Meixian Huang, Mi-Ae Lyu, Nina D. Shah, Swaminathan P. Iyer, Robert Z. Orlowski et Simrit Parmar. « Cord Blood Regulatory T Cells Prevent Mutiple Myeloma Progression By Suppressing Inflammation ». Blood 134, Supplement_1 (13 novembre 2019) : 3095. http://dx.doi.org/10.1182/blood-2019-128418.
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Introduction: Regulatory T cells (Tregs) are potent immunosuppressors and are being developed as adoptive immunotherapy for inflammatory disorders and autoimmune diseases. Specifically, cord blood (CB) Tregs are emerging as a promising treatment for graft vs. host disease (GVHD) as well as immune mediated bone marrow failure disorders including aplastic anemia, hypoplastic myelodysplasia and primary myelofibrosis. We and others have shown that when compared to peripheral blood (PB), CB Tregs have a higher expression of Helios, a marker of thymic Tregs; exert superior suppression on proliferating conventional T cells and do not secrete inflammatory IL-17 or express RORүt under stressful conditions. Such a stable suppressor profile coupled with their ready availability as an off-the-shelf product positions CB Tregs to be a very attractive therapeutic agent. Since multiple myeloma, a plasma cell clonal malignancy, exists in an inflammatory microenvironment of hematopoietic and non-hematopoietic cells which is likely related to disease progression, we hypothesized that adoptive therapy with CB Tregs may block myeloma progression by exerting their anti-inflammatory function. Methods: We used the NOD-SCID IL2rγnull (NSG) mice to first establish the myeloma tumor model, where mice were injected intravenously via tail vein with 3 x 106 Firefly luciferase labeled MM1S cells. For generating CB Tregs for adoptive therapy, Treg cell enrichment was performed from the whole CB unit using CD25+ magnetic beads and LS column (MIltenyl) followed by continuous culture in the presence of CD3/28 T-cell activator beads, and human recombinant interleukin-2 for 14 days. Expanded CB Treg phenotype was confirmed using flow cytometry analysis and the in-vitro suppressive capacity was assessed by CellTrace Violet Cell Proliferation assay. The mice were injected intravenously via tail vein with Firefly luciferase labeled MM1S cells on day 0 with or without 10 x 106 ex-vivo expanded CB Treg cells on day -1. We monitored the progression of MM1S cells by in vivo bioluminescence imaging (BLI) using the IVIS imaging system (Xenogen) 10 minutes after intraperitoneal injection of D-luciferin. Imaging data were analyzed and quantified with Living Image Software (Xenogen). We also assessed the weight and survival as well as the plasma cytokines levels. At the time of euthanasia, organs were harvested and analyzed for myeloma burden by using immunohistochemistry as well as flow analysis. Results: The expanded CB Tregs showed a consistent phenotype of CD4+25+127-FoxP3high and more than 80% suppression of the proliferating conventional T cells (Tcons). The progression of MM1S cells in the mice injected with CB Treg (n=9) was significantly delayed as shown in figure 1A, where minimal evidence of myeloma cells is visualized on day 31 compared to widespread tumor in the control arm (n=9). CB Treg recipients preserved their weight for a longer time (Figure 1B) and had improved overall survival (p=0.039) (Figure 1C). The prevention of development of myeloma by CB Tregs correlated with the lower levels of the inflammatory cytokine, interleukin-6 (IL-6) (murine) especially on day 35 post myeloma inoculation (Figure 1D). The circulating myeloma cells in the CB Treg recipients were significantly lower as compared to control mice. Upon euthanasia on day 25 post inoculation, myeloma cells were barely detectable in bone marrow and spleen in the CB Treg recipients as compared to large tumor burden in the control arm (Figure 1E). Conclusions: Our results demonstrate that ex-vivo expanded CB Treg cells prevent the growth of myeloma cells in a xenogenic model via suppression of inflammatory cytokines such as IL-6. These data provide additional insight to the underpinnings of myeloma progression and may contribute to the development of the novel immunotherapies in the future. Figure 1 Disclosures Shah: University of California, San Francisco: Employment; Genentech, Seattle Genetics, Oncopeptides, Karoypharm, Surface Oncology, Precision biosciences GSK, Nektar, Amgen, Indapta Therapeutics, Sanofi: Membership on an entity's Board of Directors or advisory committees; Indapta Therapeutics: Equity Ownership; Celgene, Janssen, Bluebird Bio, Sutro Biopharma: Research Funding; Poseida: Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Nkarta: Consultancy, Membership on an entity's Board of Directors or advisory committees; Kite: Consultancy, Membership on an entity's Board of Directors or advisory committees; Teneobio: Consultancy, Membership on an entity's Board of Directors or advisory committees. Orlowski:Poseida Therapeutics, Inc.: Research Funding. Parmar:Cellenkos Inc.: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Research Funding.
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Choi, Yoonsu, Zheng Li, Xukui Wang, Hui Li, Feng Li, Daniel Y. Lee, Parijat Bhatnagar et al. « Combination of Gene Therapy and Nanoparticle Imaging for Improving T-Cell Therapy. » Blood 116, no 21 (19 novembre 2010) : 1479. http://dx.doi.org/10.1182/blood.v116.21.1479.1479.
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Abstract Abstract 1479 Despite a functioning immune system, malignant tumors develop strategies to avoid immune recognition and elimination by T cells. Gene therapy can be used to render T cells capable of targeting tumors in an antigen-specific manner. The ability to genetically manipulate a T cell ex vivo provides new opportunities to enhance their biological activity in vivo. To explore new treatment approaches we propose a novel paradigm in which T cells are developed as biological delivery vehicles for desired genes and nanoparticles. We have combined gene therapy with nanotechnology to generate T cells that express a chimeric antigen receptor (CAR) selectively expressed on T cells operating within the tumor microenvironment and to introduce radiolabeled gold nanoparticles for tracking T cells in vivo. T cells can home to tumor microenvironments to exert their immunoreceptor-dependent cytolytic activity. However, despite the ability of T-cell receptors (TCRs) and CAR to recognize tumor cells, the anti-tumor effect can be incomplete. The ability to transfer a wide range of materials into T cells via electroporation can be exploited to adapt T cells as biological delivery agents capable of targeting specific tissues. This method may also circumvent the limited biodistribution of “raw” nanomaterials due to rapid clearance by the liver and other reticuloendothelial system (RES) organs. Gold is an attractive biocompatible nanomaterial which can be synthesized in compliance with current good manufacturing practice (cGMP) guidelines required for clinical-grade production and can be chemically functionalized for specific imaging or therapeutic functions. We first tested whether commercially available nanoparticles can be electro-transferred into cultured and primary human T cells. 43 nm diameter latex nanoparticles and 7 nm gold nanoparticles (GNPs) were electro-transferred into T cells and visualized by TEM and confocal imaging (Figure 1). We have modified GNPs on the surface with the chelator, diethylenetriaminepentaacetic acid (DTPA), for stable coordination with 111In followed by GNP PEGylation. We have shown that these radiolabeled GNPs can be readily electro-transferred into T cells suitable for combined single-photon emission computed tomography (SPECT) and computed tomography (CT) (Figure 2). We used a clinical SPECT/CT scanner to detect 111In-GNP in T cells (∼2.1 × 104 nanoparticles/cell) at a sensitivity of ∼760 cells/mL. After developing electroporation protocols of nanoparticles and in vivo imaging of 111In-GNPs, additional sensitivity was achieved by modifying the chelating chemistry using the macrocyclic chelator, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), to bind 64Cu to GNPs, increasing the number of gold particles/cell, and using 64Cu-labeled GNPs for imaging by positron emission tomography (PET). Before tail vein injection to a mouse, 11.4 mCi was detected from 10 million T cells (suspended in 300 μL PBS) electroporated using a BTX ECM830 device with the following settings: 1 kV/cm, 4 ms duration, single square pulse (Figure 3). The estimated concentration of nanoparticles transferred into T cells was ∼2.3 × 105 nanoparticles/cell as determined by a gamma counter (2470 Wizard, PerkinElmer) and nanoparticle titration. While 20 nm GNPs were used for 111In labeling, 7 nm GNPs were chosen for 64Cu labeling because of the improved (10-fold) electroporation efficiency. We are currently investigating whether multi-functional GNPs encapsulating chemotherapy drugs can add both in vivo T-cell imaging capability and enhanced cytotoxicity to CAR-redirected T cells. In aggregate, this may improve the potency of clinical-grade genetically modified T cells as a vehicle with the imaging capability for targeted delivery of drug-loaded GNPs to the tumor microenvironment. Disclosures: No relevant conflicts of interest to declare.
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Hartman, Robert A., Kevin M. Bell, Richard E. Debski, James D. Kang et Gwendolyn A. Sowa. « Novel ex-vivo mechanobiological intervertebral disc culture system ». Journal of Biomechanics 45, no 2 (janvier 2012) : 382–85. http://dx.doi.org/10.1016/j.jbiomech.2011.10.036.
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Davies, CM, DB Jones, MJ Stoddart, K. Koller, E. Smith, CW Archer et RG Richards. « Mechanically loaded ex vivo bone culture system 'Zetos' : Systems and culture preparation ». European Cells and Materials 11 (12 avril 2006) : 57–75. http://dx.doi.org/10.22203/ecm.v011a07.
Texte intégralThèses sur le sujet "Ex Vivo Vein Culture System"
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PRANDI, FRANCESCA. « Identification of early pathophysiological events underlying venous coronary bypass stenosis by a mechano-biology approach ». Doctoral thesis, Università degli Studi di Milano-Bicocca, 2014. http://hdl.handle.net/10281/50493.
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Saphenous vein graft disease represents an unresolved problem in coronary artery bypass grafting. After surgery, progressive modification in the vein wall occurs, leading to occlusion of the graft lumen. This process, called intima hyperplasia, involves the participation of vein-resident cells as well as the recruitment of vein-extrinsic cells. Arterial wall strain has recently emerged as one of the factor that can contribute to the pathogenesis of the vein graft disease. Therefore, in collaboration with Department of Bioengineering, Politecnico di Milano we developed a culture system for the ex vivo pressure stimulation of vein segments. This new ex vivo vein culture system is able to reproduce the wall strain typical of the arterial circulation. The ex vivo vein culture system (ECVS) adopted in this project has been validated and proved as a valuable, reliable, easy handling and versatile tool for studying arterial pressure events triggered in VGD.
The biological data achieved confirm an important contribution of the arterial-like wall strain in SV structural and biochemical changes, activation of vessel resident cells and in the expression of molecular signals involved in the pathogenesis of IH.
Using this system, we found that either venous- or arterial-specific pressure regimens induced vein pro-pathologic commitment involving upregulation of Matrix Metallo-Proteases 2/9, and induction of microRNAs-21/146a/221. By contrast, arterial-like pressure caused a significant morphological rearrangement of the vein, a suppression of Tissue Inhibitor of Metallo Protease-1, an enhanced expression of TGF-β1 and BMP-2 mRNAs and, finally, the upregulation of microRNAs-138/200b/200c. In coronary-pressure stimulated vessels, the density of the adventitial vasa vasorum was significantly increased. This was accompanied by an increased presence of cells co-expressing NG2, CD44 and SM22α markers in the adventitia, identifying them as multipotent mesenchymal cells/smooth muscle cells progenitors with a pericyte origin. An increase in Histone H3 Lysine 4 methylation and histone H4 Lysines 9/16 acetylation levels was finally found in adventitial cells and vasa vasorum. The present findings suggest a mechanistic role of the arterial-like pulsatile pressure in reinforcement of SV-resident cells pro-pathologic commitment in vein bypass failure, by activation of mechanical-dependent transcriptional circuitries and of pericyte-derived cells located in the vessel adventitia.
The ultimate goal of this project is to find a treatment that can prevent, avoid or reduce the incidence of the vein graft disease in patients subjected to bypass surgery with saphenous vein. This treatment could include one or more targets identified in this work focusing on the early stage of the pathological adaptation of the SV to the new hemodynamic environment.
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Toutain, Hervé. « Développement et caractérisation de modèles expérimentaux pour l'étude ex-vivo et in-vitro de la cellule tubulaire proximale de rein de lapin ». Rouen, 1989. http://www.theses.fr/1989ROUES036.
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Une suspension de cellules tubulaires proximales de rein de lapin est isolée par une nouvelle méthode qui ne fait pas intervenir d'enzymes protéolytiques. Ces cellules, après une caractérisation biochimique, morphologique, et métabolique, sont séparées en deux populations hautement purifiées par une technique d'électrophorèse ou flux libre en veine liquide. Présentation d'un modèle de culture primaire de cellules tubulaires proximales de rein de lapin
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Morrison, Kristen M. « Ex Vivo Salivary Gland Culture as a Novel System to Evaluate HCMV Infection ». University of Cincinnati / OhioLINK, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=ucin1470044093.
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Hammarbäck, Madelene. « Development of a dynamic ex vivo culture system for human islets of langerhans ». Thesis, Uppsala universitet, Institutionen för kvinnors och barns hälsa, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-353398.
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Type 1 diabetes(T1D)is a disease that only gets more common. The etiology of the disease is not known but there are many existing theories about what the cause is. These different theories have been tested in vivoin rodents or invitro. The resultsfrom experiments done in those waysarenotall realistic because rodents differnotablyfrom humans,and when studies areperformed in vitrowith human isletsof Langerhans different hormones will accumulate. The aim of this studywas to establisha dynamic ex vivosystem in which stimulation of human islets of Langerhans can be performed in a more lifelike environment. To study islets in this system couldin the future lead to increased knowledge in the etiology of T1D.The perifusion system PERI-4.2 from Biorep Technologies together with an incubator with 37°Cand5% CO2were used to arrangethe ex vivosystem. An Insulin ELISA from Mercodia was performedto analyze the insulin secretion from the islets. Fourdifferent set ups for the system were tested and the last one showed the best results.In conclusion this study has shown that it is possible to preserve human islets of Langerhans in a dynamic ex vivosystem with a constant medium exchange if it is done under conditionswhere the islets are protected from shear forces from the supplying medium,together with a medium exchange rate which replaces the whole medium in at least one hour.
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Tashiro, Takahiro. « In vivo and ex vivo cetuximab sensitivity assay using three-dimensional primary culture system to stratify KRAS mutant colorectal cancer ». Kyoto University, 2018. http://hdl.handle.net/2433/232473.
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Roberts, Jessica L. « Development of an ex-vivo co-culture system to model pulpal infection by Streptococcus anginosus group bacteria ». Thesis, Cardiff University, 2010. http://orca.cf.ac.uk/55492/.
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A reproducible model of infection of dental tissues by SAG bacteria has been produced which shows attachment patterns of bacteria and the effect on host tissues. This model can be used to further investigate processes involved in endodontic infections, including expression of virulence factors by bacteria and host response from the dental tissues. It may also be used in the future for testing novel antimicrobials for use in treating pulpal disease.
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Karekla, Ellie. « Improving therapeutic approaches for the treatment of non-small cell lung cancer using an ex-vivo explant culture system ». Thesis, University of Leicester, 2015. http://hdl.handle.net/2381/39930.
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Lung Cancer is the leading cause of cancer death worldwide in both males and females. Non-small cell lung cancer (NSCLC) accounts for ~80-85% of lung cancers. NSCLC is a very heterogeneous disease, both genetically and histologically, and there is an increasing list of mutations and copy number alterations in cancer associated genes. Several drugs that could potentially improve lung cancer outcomes are in development and some have entered clinical trials. However, the current established preclinical models, particularly animal xenografts, are not always predictive of patient outcome and there has been a large attrition of clinical candidate drugs at the Phase III stage. The aim of this project was to establish a primary NSCLC explant culture system with the view to developing a better platform to test the efficacy of existing drugs as well as novel drug combinations. The tissue architecture and tumour heterogeneity of individual NSCLC patients can be examined in an ex-vivo NSCLC explant culture system which maintains viability and proliferation in a short period of 24 hours + recovery (16-20 hours). Even though there is a moderate effect of cultivation, the ex-vivo NSCLC explant culture system can be used for assessing in situ drug responses over short periods. Responses of explants were assessed after treatment with cisplatin, MEK and PI3K inhibitors singly and in combination and TRAIL and ABT-737 singly and in combination in the presence or absence of cisplatin. This model points towards being more predictive of patient outcome in clinical studies than in vitro studies or animal models. The data show that the explant system has the potential to improve on current preclinical models for lung cancer or other solid cancers and help the drug development process achieve greater successes in the clinic. The model could provide a platform for personalising treatment to each patient and for identifying effective biomarkers for drug responses.
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Konduri, Suchitra. « The Influence of normal physiological forces on porcine aortic heart valves in a sterile ex-vivo pulsatile organ culture system ». Thesis, Available online, Georgia Institute of Technology, 2005, 2005. http://etd.gatech.edu/theses/available/etd-03042005-135623/unrestricted/konduri%5Fsuchitra%5F200505%5Fmast.pdf.
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Thesis (M. S.)--Chemical and Biomolecular Engineering, Georgia Institute of Technology, 2005.Dr. Athanassios Sambanis, Committee Member ; Dr. Timothy M. Wick, Committee Member ; Dr. Ajit P.Yoganathan, Committee Chair. Includes bibliographical references.
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Fernandes, Darcy. « Estudo ex vivo e in vitro da modulação da imunidade inata e adaptativa por queratinócitos displásicos em leucoplasia oral e leucoplasia verrucosa proliferativa / ». Araraquara, 2020. http://hdl.handle.net/11449/192432.
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Orientador: Andreia BufalinoResumo: O carcinoma espinocelular (CEC) representa mais de 95% de todas as neoplasias malignas que acometem a cavidade oral e muitas vezes estes tumores são precedidos por alterações clínicas que apresentam um evidente potencial de transformação maligna, as quais são chamadas de desordens potencialmente malignas orais (DPMOs). Dentre estas, a leucoplasia oral (LO) possui taxa de transformação maligna que varia de 0,2% até 17,5%; contudo, uma outra DPMO conhecida como leucoplasia verrucosa proliferativa (LVP) apresenta um comportamento persistente e progressivo para malignidade, com taxa de transformação maligna maior que 70%. Diferente da LO, fator de risco como tabaco, álcool e noz de areca não parecem estar associados com o desenvolvimento da LVP. Adicionalmente, a LVP frequentemente apresenta resposta inadequada a todas as modalidades de tratamento, sofre rápida disseminação pelos sítios orais e muitas vezes demonstra recorrência. Estudos recentes sugerem que o infiltrado inflamatório associado às lesões leucoplásicas de paciente com LVP pode estar relacionado a etiologia e/ou comportamento clínico agressivo desta DPMO. Assim, foi realizada uma análise comparativa entre amostras de LO e LVP que consistiu em: (1) avaliar a porcentagem e identificar os subtipos de linfócitos T auxiliares e estado de ativação de linfócitos T citotóxicos, (2) avaliar a densidade e estado de ativação das células dendríticas, e (3) determinar o efeito de produtos solúveis de células displásicas na modul... (Resumo completo, clicar acesso eletrônico abaixo)
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Tobin, Desmond J. « Ex vivo organ culture of human hair follicles : a model epithelial-neuroectodermal-mesenchymal interaction system ». 2010. http://hdl.handle.net/10454/7452.
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noThe development of hair follicle organ culture techniques is a significant milestone in cutaneous biology research. The hair follicle, or more accurately the "pilo-sebaceous unit", encapsulates all the important physiologic processes found in the human body; controlled cell growth/death, interactions between cells of different histologic type, cell differentiation and migration, and hormone responsitivity to name a few. Thus, the value of the hair follicle as a model for biological scientific research goes way beyond its scope for cutaneous biology or dermatology alone. Indeed, the recent and dramatic upturn in interest in hair follicle biology has focused principally on the pursuit of two of biology's holy grails; post-embryonic morphogenesis and control of cyclical tissue activity. The hair follicle organ culture model, pioneered by Philpott and colleagues, ushered in an exceptionally accessible way to assess how cells of epithelial (e.g., keratinocytes), mesenchymal (e.g., fibroblasts), and neuroectodermal (e.g., melanocytes) origin interact in a three-dimensional manner. Moreover, this assay system allows us to assess how various natural and pharmacologic agents affect complex tissues for growth modulation. In this article, I focus on the culture of the human hair follicle mini-organ, discussing both the practical issues involved and some possible research applications of this assay.
Chapitres de livres sur le sujet "Ex Vivo Vein Culture System"
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Tobin, Desmond J. « Ex Vivo Organ Culture of Human Hair Follicles : A Model Epithelial–Neuroectodermal–Mesenchymal Interaction System ». Dans Methods in Molecular Biology, 213–27. Totowa, NJ : Humana Press, 2010. http://dx.doi.org/10.1007/978-1-60761-984-0_14.
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Heine, Claudia, et Heike Franke. « Organotypic Slice Co-culture Systems to Study Axon Regeneration in the Dopaminergic System Ex Vivo ». Dans Methods in Molecular Biology, 97–111. New York, NY : Springer New York, 2014. http://dx.doi.org/10.1007/978-1-4939-0777-9_8.
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Chua, Jie Shi, Kuberan Balagurunathan et Yukio Saijoh. « Manipulation of Glycosaminoglycans Using Synthetic to Study Their Roles in in Ex Vivo Lung Culture System ». Dans Methods in Molecular Biology, 645–53. New York, NY : Springer US, 2021. http://dx.doi.org/10.1007/978-1-0716-1398-6_49.
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Thayakumar Basanthakumar, Allen. « Application of Ex-Vivo/3D Organoid Models in COVID-19 Research ». Dans Biotechnology to Combat COVID-19 [Working Title]. IntechOpen, 2021. http://dx.doi.org/10.5772/intechopen.99100.
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COVID-19 treatment methods based on 3D organoids and ex-vivo platforms are analyzed in this chapter. Initially, the platforms available for cell culture and its working characteristics are explained. Subsequently discusses the organoids with their definition and included their uses in various applications. Further, the chapter extends to describe the uses of different organoids with their use in different stages. Most of these methods utilized the 3D ex-vivo cell culture method to develop organoids and test them over infected tissues. Based on the study in this chapter, it is found that the demonstration of active replication of the human organoids culture system of lungs is found to be more helpful for COVID-19 treatment.
Actes de conférences sur le sujet "Ex Vivo Vein Culture System"
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Zhang, W., W. Van Weerden, S. Erkens-Schulze, R. Kanaar, D. Van Gent et J. Nonnekens. « PO-442 Ex vivo prostate cancer culture system recapitulates in vivo response to androgen antagonist ». Dans Abstracts of the 25th Biennial Congress of the European Association for Cancer Research, Amsterdam, The Netherlands, 30 June – 3 July 2018. BMJ Publishing Group Ltd, 2018. http://dx.doi.org/10.1136/esmoopen-2018-eacr25.953.
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Arreola, Alexandra, et W. Kimryn Rathmell. « Abstract 4276 : Characterization of an ex vivo primary multicellular renal cell culture as a potential model system for renal cell carcinoma tumorigenesis ». Dans Proceedings : AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011 ; Orlando, FL. American Association for Cancer Research, 2011. http://dx.doi.org/10.1158/1538-7445.am2011-4276.
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Han, Hai-Chao, Raymond P. Vito, Kristin Michael et David N. Ku. « Axial Stretch Increases Cell Proliferation in Arteries in Organ Culture ». Dans ASME 2000 International Mechanical Engineering Congress and Exposition. American Society of Mechanical Engineers, 2000. http://dx.doi.org/10.1115/imece2000-2516.
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Abstract To study the effect of axial stretch on vascular function and wall remodeling, porcine carotid arteries were cultured under conditions of physiological flow and elevated axial stretch in an ex vivo organ culture system. Smooth muscle cell proliferation was measured by bromodeoxyuridine index. Results showed that cell proliferation was significantly increased in the highly stretched arteries when compared to the normally stretched arteries. This may indicate the feasibility of stimulating new arterial growth by stretching natural arteries.
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Nakano, Michitaka, Hioshi Ariyama, Shingo Tamura, Taichi Isobe, Kohta Miyawaki, Yuta Okumura, Hitoshi Kusaba, Takashi Ueki, Eishi Baba et Koichi Akashi. « Abstract 1520 : Plasticity of CD44+ colorectal cancer stem cells depends on TGF-beta-induced epithelial mesenchymal transition (EMT) : evidences from ex vivo culture system ». Dans Proceedings : AACR 106th Annual Meeting 2015 ; April 18-22, 2015 ; Philadelphia, PA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1538-7445.am2015-1520.
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Banda, Malathi, Karen L. McKim et Barbara Parsons. « Abstract A15 : Establishing an ex-vivo, tumor spheroid culture system to assess molecular-targeted therapies for personalized treatment of non-small cell lung cancer ». Dans Abstracts : Patient-Derived Cancer Models : Present and Future Applications from Basic Science to the Clinic ; February 11-14, 2016 ; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1557-3265.pdx16-a15.
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Quee, Shawn Chin, Hai-Chao Han et David N. Ku. « Bench-Top Validation Tests for Tissue-Engineered Arteries ». Dans ASME 2000 International Mechanical Engineering Congress and Exposition. American Society of Mechanical Engineers, 2000. http://dx.doi.org/10.1115/imece2000-2514.
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Abstract Standard tests are needed for evaluating and comparing the mechanical and biological functions of tissue engineered arteries and other vascular grafts. We propose an ex vivo organ culture system as a living system for testing tissue-engineered vascular grafts. This bench-top organ culture system mimics the physiological environment of arteries including the flow, pressure, and the axial stretch. Arterial mechanical properties and physiologic functions including compliance, burst pressure, and contractile functions can be assessed before an expensive long-term animal test is initiated. Test results of natural arteries indicate that organ culture is a valid model for comprehensive evaluation of tissue-engineered vascular grafts.
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Inauen, W., T. Bombeli, H. R. Baumgartner, A. Haeberli et P. W. Straub. « EFFECTS OF HEPARIN AND COUMARIN ON DEPOSITION OF FIBRIN, PLATELETS AND PLATELET THROMBI ON RABBIT AORTA SUBENDOTHELIUM EXPOSED TO FLOWING HUMAN BLOOD ». Dans XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643876.
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The effect of heparin and phenprocoumon on thrombogenesis induced by rabbit aorta subendothelium (SE) was investigated in 20 volunteers using an ex vivo perfusion system. Blood was drawn directly from an antecubital vein through an annular chamber with exposed SE at lOml/min flow rate (650sec−1 shear rate) for 5min. Following buffer perfusion for 15sec, the middle portion of SE was removed for plasmin digestion and adjacent segments were fixed and embedded for morphometric analysis. Perfusions were performed 20 min after i.v. injection of heparin 1000, 2500 and 5000 IU, respectively; and during the decline and steady-state of prothrombin activity during a 2 weeks treatment with phenprocoumon to target INR of 5.0.The amount of fibrin attached to SE, as measured by fragment E RIA in plasmin digests, correlated negatively with the dose of heparin (r=−0.83, P<0.001, n=48) and with INR during coumarin intake (r=−0.58, P<0.01, n=40). After high doses of either heparin or coumarin fibrin deposition on SE was virtually abolished (table). Platelet adhesion was increased. Platelet thrombus volumes and heights were reduced by heparin and coumarin.We conclude that both heparin and coumarin dose-dependently inhibit fibrin formation induced by SE. In addition, both drugs impair platelet thrombus growth and/or stability indicating that these processes may also depend on the coagulation mechanism.
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Li, Lei, Xuetao Shi, Xiaoqing Lv et Jing Liu. « A Biomimetic Microfluidic Device for the Study of the Response of Endothelial Cells Under Mechanical Forces ». Dans ASME 2014 International Mechanical Engineering Congress and Exposition. American Society of Mechanical Engineers, 2014. http://dx.doi.org/10.1115/imece2014-36430.
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Vascular science is an active area of medicine and biological research. In recent years, intensive research has been focused on the reaction of endothelial cells (ECs) to relevant biological, chemical, or physical cues in vitro. The primary thing of these studies is to make a biomimic environment of ECs which is closer to the in vivo conditions. Here we developed a microfluidic system and fabricated a grooved micropattern thin film to simulate inner blood vessel wall. The micropattern structure was generated by using the elastic biocompatible material poly(dimethylsiloxane) (PDMS). Human umbilical vein endothelial cells (HUVECs) were cultured on the grooved micropattern film. After the cells reached confluence, the thin PDMS film with cells was inserted into the biological grade plastic tube. Then cell culture medium was perfused into the tube and the cellular responses under shear stress and pressure were investigated. The F-actin cytoskeleton and the nuclei of the cells were stained for examination. This microfluidic system provides a convenient and cost-effective platform for the studies of cellular response to mechanical forces. Moreover, this system could also be used for studying cellular responses to drugs under mechanical forces.
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Fujimori, T., T. Saeki, K. Harada, M. Sato et N. Ohshima. « ANTI-THROMBOTIC EFFECTS OF E-5510 IN EXPERIMENTAL THROMBOSIS MODELS ». Dans XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643429.
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A new agent developed in our laboratory, 4-cyano-5,5-bis(4-methoxyphenyl)-4-pentenoic acid (E-5510), suppressed various human platelet functions in vitro. The compound also showed quite potent ex vivo anti-platelet effects in many experimentalanimals. It is well known that anti-platelet effects are not always parallel to anti-thrombotic effects. Thus, in order to predict the efficacy of E-5510 in thrombotic disorders, the anti-thrombotic effects were examined in 3 different animal models of thrombosis.(1) Anti-thrombotic effect in an extracorporeal shunt model Two hrs after oral administration of the drug to guinea pigs,an extracorporeal shunt between the right carotid artery and the left jugular vein was performed. The thrombus formation on a silk thread inserted in the shunt tube was quantitated by measuring the time from the onset of circulation to the stenosis of blood flow. E-5510 dose-dependently inhibited thrombus formation, the minimum effective dose being 0.03 mg/kg.(2) Effect on microthrombus formation in mesenteric arteriole In order to evaluate the effect on intravascular platelet thrombus formation, thrombosis was induced in vivo in mesenteric arteriole in guinea pigs with filtered light from a mercury lamp and an intravenous fluorescent dye in an intravital microscope system (M. Sato and N. Ohshima, Thromb. Res.,35, 319, 1984). The thrombus formation was quantitated by measuring the time taken for circulating platelets to begin to adhere to vessel wall and the time taken for blood flow to stop completely due to fully developed thrombus. Both the time required for platelet adhesion to vessel wall and for platelet thrombus formation were significantly prolonged after oral administration of E-5510.(3) Effect on pulmonary thromboembolism Acute pulmonary thromboembolism was induced in mice by a bolus intravenous injection of arachidonic acid, and mortality was determined 3 min later. E-5510 dose-dependently reduced pulmonary thromboembolic mortality, and its ED50 was 0.11 mg/kg. The results described above indicate thatE-5510 may have beneficial effects in clinical treatments of thrombotic disease.
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Boffa, M. C., B. Burke et C. C. Haudenschild. « THROMBOMODULIN ON EXTRAVASCULAR MEMBRANES ». Dans XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643965.
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The distribution of thrombomodulin (TM) antigen (Ag) was studied in the rabbit using an affinity-purified antibody raised against rabbit TM in a goat. Tissues were obtained from 8 New Zealand rabbits. Paraffin-embedded sections were prepared from various organs. Staining was performed using the avidin-biotin peroxi dase method. TM was found to be best and most consistently preserved after perfusion of the vasculature with fixative (buffered formalin) i.e. perfusion of the systemic vasculature via the ascending aorta and individual perfusion of the pulmonary, portal, coronary vasculature via the appropriate artery or vein. A positive reaction was observed on the entire endothelial surface of the vascular system: arteries, veins ahd capillaries. In contrast, parenchyma, secretory epithelia, connective tissue, cartilage, bone and nerve tissue were not stained.Interestingly, TM antigen was also found on the serosae: peritoneum, pericardium and epicardium and in the intralobular folds of the pleura, on the synovial membrane of the knee joint and on the entire surface of the spinal and cerebral arachnoid. The reaction was maximal on the intima of the large arteries, on the arachnoid and on the synovial membrane. The intensity of the reac tion did not depend on the organ examined but on the perfusion quality, except for arachnoidal and synovial membranes which were stained even without initial perfusion. The presence of TM on the membranes of body cavities was confirmed by the recovery of TM activity (as cofactor of protein C activation by thrombin in a chromogenic assay) in intraperitoneal lavage in vivo and in fluid used to rinse the brain arachnoidal surface ex vivo.These findings suggest the presence of potent anticoagulant mechanisms in the systems of slow circulating fluids such as the cerebrospinal and synovial fluids and in the virtual spaces of the serosae. While no thrombin is present there in physiological conditions, anticoagulant activity in these locations may be very important in case of even mild permeability changes, such as in inflammation. As anticoagulant systems assure the fluidity of blood in the vasculature, the TM(-PC) system on body cavities linings may assure free mobility and absence of adhesion of the membranes.