| 1. |
- Asif, Sana, et al.
(författare)
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Oxygen-charged HTK-F6H8 emulsion reduces ischemia : reperfusion injury in kidneys from brain-dead pigs
- 2012
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Ingår i: Journal of Surgical Research. - : Elsevier BV. - 0022-4804 .- 1095-8673. ; 178:2, s. 959-967
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Tidskriftsartikel (refereegranskat)abstract
- Background:Prolonged cold ischemia is frequently associated with a greater risk of delayed graft function and enhanced graft failure. We hypothesized that media, combining a high oxygen-dissolving capacity with specific qualities of organ preservation solutions, would be more efficient in reducing immediate ischemia-reperfusion injury from organs stored long term compared with standard preservation media.Methods:Kidneys retrieved from brain-dead pigs were flushed using either cold histidine-tryptophan-ketoglutarate (HTK) or oxygen-precharged emulsion composed of 75% HTK and 25% perfluorohexyloctane. After 18 h of cold ischemia the kidneys were transplanted into allogeneic recipients and assessed for adenosine triphosphate content, morphology, and expression of genes related to hypoxia, environmental stress, inflammation, and apoptosis.Results:Compared with HTK-flushed kidneys, organs preserved using oxygen-precharged HTK-perfluorohexyloctane emulsion had increased elevated adenosine triphosphate content and a significantly lower gene expression of hypoxia inducible factor-1 alpha, vascular endothelial growth factor, interleukin-1 alpha, tumor necrosis factor-alpha, interferon-alpha, JNK-1, p38, cytochrome-c, Bax, caspase-8, and caspase-3 at all time points assessed. In contrast, the mRNA expression of Bcl-2 was significantly increased.Conclusions:The present study has demonstrated that in brain-dead pigs the perfusion of kidneys with oxygen-precharged HTK-perfluorohexyloctane emulsion results in significantly reduced inflammation, hypoxic injury, and apoptosis and cellular integrity and energy content are well maintained. Histologic examination revealed less tubular, vascular, and glomerular changes in the emulsion-perfused tissue compared with the HTK-perfused counterparts. The concept of perfusing organs with oxygen-precharged emulsion based on organ preservation media represents an efficient alternative for improved organ preservation.
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| 2. |
- Babiker, Adil A., et al.
(författare)
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Mapping pro- and antiangiogenic factors on the surface of prostasomes of normal and malignant cell origin
- 2010
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Ingår i: The Prostate. - : Wiley. - 0270-4137 .- 1097-0045. ; 70:8, s. 834-847
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Angiogenesis is the formation of new blood vessels by capillary sprouting from pre-existing vessels. Tumor growth is angiogenesis-dependent and the formation of new blood vessels is associated with the increased expression of angiogenic factors. Prostasomes are secretory granules produced, stored and released by the glandular epithelial cells of the prostate. We investigated the expression of selected angiogenic and anti-angiogenic factors on the surface of prostasomes of different origins as well as the direct effect of prostasomes on angiogenesis. METHODS: VEGF, endothelin-1, endostatin, and thrombospondin-1 were determined on prostasomes from seminal fluid and human prostate cancer cell lines (DU145,PC-3,LNCaP) using different immunochemical techniques. Human dermal microvascular endothelial cells were incubated with seminal and DU145 cell-prostasomes and with radioactive thymidine. The effect of prostasomes on angiogenesis was judged by measuring the uptake of labeled thymidine. The presence of any deleterious effects of prostasomes on the endothelial cells was investigated using thymidine assay and confocal laser microscopy. RESULTS: VEGF and endothelin-1 were determined on malignant cell-prostasomes (no difference between cell lines) but not determined on seminal prostasomes. The same applies for the expression of endostatin but with much higher expression on malignant cell-prostasomes with obvious differences between them. Seminal and DU145 cell-prostasomes were found to have anti-angiogenic effect which was more expressed by DU145 cell-prostasomes. No deleterious effect of prostasomes on endothelial function was detected using either thymidine assay or microscopy. CONCLUSIONS: Prostasomes contain pro- and anti-angiogenic factors that function to counteract each other unless the impact from one side exceeds the other to bring about dysequilibrium.
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| 3. |
- Bekkhus, Tove, et al.
(författare)
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Stromal transdifferentiation drives lipomatosis and induces extensive vascular remodeling in the aging human lymph node
- 2023
-
Ingår i: Journal of Pathology. - : John Wiley & Sons. - 0022-3417 .- 1096-9896. ; 259:3, s. 236-253
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Tidskriftsartikel (refereegranskat)abstract
- Lymph node (LN) lipomatosis is a common, but rarely discussed phenomenon, associated with aging, involving a gradual exchange of the LN parenchyma into adipose tissue. The mechanisms behind these changes and the effects on the LN are unknown. We show that LN lipomatosis starts in the medullary regions of the human LN and link the initiation of lipomatosis to transdifferentiation of LN fibroblasts into adipocytes. The latter is associated with a downregulation of lymphotoxin beta expression. We also show that, isolated medullary and CD34+ fibroblast, in contrast to the reticular cells of the T-cell zone, display an inherent higher sensitivity for adipogenesis. Progression of lipomatosis leads to a gradual loss of the medullary lymphatic network, but at later stages, collecting-like lymphatic vessels, are found inside the adipose tissue. The stromal dysregulation includes a dramatic remodeling and dilation of the high endothelial venules associated with reduced density of naïve T-cells. Abnormal clustering of plasma cells is also observed. Thus, LN lipomatosis causes widespread stromal dysfunction with consequences for the immune contexture of the human LN. Our data warrant an increased awareness of LN lipomatosis as a factor contributing to decreased immune functions in the elderly and in disease.
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| 4. |
- Cabric, Sanja, et al.
(författare)
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Anchoring of vascular endothelial growth factor to surface-immobilized heparin on pancreatic islets : implications for stimulating islet angiogenesis
- 2010
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Ingår i: Tissue engineering. Part A. - : Mary Ann Liebert Inc. - 1937-3341 .- 1937-335X. ; 16:3, s. 961-970
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Tidskriftsartikel (refereegranskat)abstract
- In pancreatic islet transplantation, early revascularization is necessary for long-term graft function. We have shown in in vitro and in vivo models that modification with surface-attached heparin protects the islets from acute attack by the innate immune system of the blood following intraportal islet transplantation. In this study, we have investigated the ability of an immobilized conjugate composed of heparin to bind the angiogenic growth factor vascular endothelial growth factor-A (VEGF-A) as a means of attracting endothelial cells (ECs) to induce angiogenesis and revascularization. We analyzed the capacity of VEGF-A to bind to immobilized heparin and how this affected the proliferation and adherence of ECs to both artificial glass surfaces and islets. Quartz crystal microbalance with dissipation monitoring and slot-blot demonstrated the binding of VEGF-A to heparin-coated surfaces upon which ECs showed protein-dependent proliferation. Also, ECs cultured on heparin-coated glass surfaces exhibited effects upon focal contacts. Heparinized islets combined with VEGF-A demonstrated unaffected insulin release. Further, covering islets with heparin also increased the adhesion of ECs to the islet surface. Immobilized heparin on the islet surface may be a useful anchor molecule for achieving complete coverage of islets with angiogenic growth factors, ultimately improving islet revascularization and engraftment in pancreatic islet transplantation.
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| 5. |
- Duprez, Ida Rasmusson, et al.
(författare)
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Preparatory studies of composite mesenchymal stem cell islets for application in intraportal islet transplantation
- 2011
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Ingår i: Upsala Journal of Medical Sciences. - : Uppsala Medical Society. - 0300-9734 .- 2000-1967. ; 116:1, s. 8-17
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Tidskriftsartikel (refereegranskat)abstract
- Abstract Background. Low engraftment and adverse immune reactions hamper the success rate of clinical islet transplantation. In this study, we investigated the capacity of human mesenchymal stem cells (MSCs) to adhere to human islets of Langerhans and their effects in immune modulation and during blood interactions in vitro. Methods. Composite MSC-islets were formed by suspension co-culture, and the phenotype was evaluated by confocal microscopy. Islet function was assessed by dynamic insulin release in response to glucose in vitro. Mixed lymphocyte-islet reactions (MLIR) and the tubing blood loop model were utilized as in vitro tools to analyse the effect of MSCs on the innate and adaptive immune reactions triggered by the islets. Results. MSCs rapidly adhered to islets and spread out to cover the islet surface. Insulin expression and secretion were sustained with the MSC coating. MSC-coated islets showed unaffected reactions with blood in vitro in comparison to control islets. Furthermore, MSCs suppressed lymphocyte proliferation induced by islet cells in MLIR. Conclusion. We conclude that it is possible to create composite MSC-islets to enable delivery of the MSCs by utilizing the adhesive capacity of the MSCs. This could have beneficial immunosuppressive effects in optimizing pancreatic islet transplantation.
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| 6. |
- Edin, Fredrik, et al.
(författare)
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3-D gel culture and time-lapse video microscopy of the human vestibular nerve
- 2014
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Ingår i: Acta Oto-Laryngologica. - : Informa UK Limited. - 0001-6489 .- 1651-2251. ; 134:12, s. 1211-1218
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Tidskriftsartikel (refereegranskat)abstract
- UNLABELLED: Abstract Conclusions: Human inner ear neurons have an innate regenerative capacity and can be cultured in vitro in a 3-D gel. The culture technique is valuable for experimental investigations of human inner ear neuron signaling and regeneration.OBJECTIVES: To establish a new in vitro model to study human inner ear nerve signaling and regeneration.METHODS: Human superior vestibular ganglion (SVG) was harvested during translabyrinthine surgery for removal of vestibular schwannoma. After dissection tissue explants were embedded and cultured in a laminin-based 3-D matrix (Matrigel™). 3-D growth cone (GC) expansion was analyzed using time-lapse video microscopy (TLVM). Neural marker expression was appraised using immunocytochemistry with fluorescence and laser confocal microscopy.RESULTS: Tissue explants from adult human SVG could be cultured in 3-D in a gel, indicating an innate potential for regeneration. Cultured GCs were found to expand dynamically in the gel. Growth cone expansion and axonal Schwann cell alignment were documented using TLVM. Neurons were identified morphologically and through immunohistochemical staining.
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| 7. |
- Edin, Fredrik, et al.
(författare)
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Differentiation of human neural progenitor cell-derived spiral ganglion-like neurons : a time-lapse video study
- 2014
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Ingår i: Acta Oto-Laryngologica. - : Informa UK Limited. - 0001-6489 .- 1651-2251. ; 134:5, s. 441-447
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Tidskriftsartikel (refereegranskat)abstract
- Conclusions: Human neural progenitor cells can differentiate into spiral ganglion-like cells when exposed to inner ear-associated growth factors. The phenotype bears resemblance to human sphere-derived neurons. Objective: To establish an in vitro model for the human auditory nerve to replace and complement in vivo animal experiments and ultimately human in vivo transplantation. Methods: Human neural progenitors were differentiated under conditions developed for in vitro survival of human primary spiral ganglion culture with media containing growth factors associated with inner ear development. Differentiation was documented using time-lapse video microscopy. Time-dependent marker expression was evaluated using immunocytochemistry with fluorescence and laser confocal microscopy. Results: Within 14 days of differentiation, neural progenitors adopted neural phenotype and expressed spiral ganglion-associated markers.
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| 8. |
- Fransson, Moa, et al.
(författare)
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CAR/FoxP3-engineered T regulatory cells target the CNS and suppress EAE upon intranasal delivery
- 2012
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Ingår i: Journal of Neuroinflammation. - : Springer Science and Business Media LLC. - 1742-2094. ; 9, s. 112-
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:Multiple sclerosis (MS) is an autoimmune disease of the central nervous system (CNS). In the murine experimental autoimmune encephalomyelitis (EAE) model of MS, T regulatory (Treg) cell therapy has proved to be beneficial, but generation of stable CNS-targeting Tregs needs further development. Here, we propose gene engineering to achieve CNS-targeting Tregs from naive CD4 cells and demonstrate their efficacy in the EAE model.METHODSCD4+T cells were modified utilizing a lentiviral vector system to express a chimeric antigen receptor (CAR) targeting myelin oligodendrocyte glycoprotein (MOG) in trans with the murine FoxP3 gene that drives Treg differentiation. The cells were evaluated in vitro for suppressive capacity and in C57BL/6 mice to treat EAE. Cells were administered by intranasal (i.n.) cell delivery.RESULTSThe engineered Tregs demonstrated suppressive capacity in vitro and could efficiently access various regions in the brain via i.n cell delivery. Clinical score 3 EAE mice were treated and the engineered Tregs suppressed ongoing encephalomyelitis as demonstrated by reduced disease symptoms as well as decreased IL-12 and IFNgamma mRNAs in brain tissue. Immunohistochemical markers for myelination (MBP) and reactive astrogliosis (GFAP) confirmed recovery in mice treated with engineered Tregs compared to controls. Symptomfree mice were echallenged with a second EAE-inducing inoculum but remained healthy, demonstrating the sustained effect of engineered Tregs.CONCLUSIONCNS-targeting Tregs delivered i.n. localized to the CNS and efficiently suppressed ongoing inflammation leading to diminished disease symptoms.
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| 9. |
- Fransson, Moa, et al.
(författare)
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Intranasal Delivery of CNS-Retargeted Human Mesenchymal Stromal Cells Prolongs Treatment Efficacy of Experimental Autoimmune Encephalomyelitis
- 2014
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Ingår i: Immunology. - : Wiley. - 0019-2805 .- 1365-2567. ; 142:3, s. 431-441
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Tidskriftsartikel (refereegranskat)abstract
- Treatment with mesenchymal stromal cells (MSC) is currently of interest for a number of diseases including multiple sclerosis (MS). MSCs is well known to target inflamed tissues however, in a therapeutic scenery, systemic administration will lead to few cells reaching the brain. We hypothesized that MSCs may target the brain upon intranasal (i.n) administration and persist in CNS tissue if expressing a CNS-targeting receptor. To demonstrate proof of concept, MSCs were genetically engineered to express a myelin oligodendrocyte glycoprotein (MOG)-specific receptor. Engineered MSCs retained their immunosuppressive capacity, infiltrated into the brain upon i.n. cell administration, and were able to significantly reduce disease symptoms of experimental autoimmune encephalomyelitis (EAE). The mice treated with CNS-targeting MSCs were resistant to further EAE induction whereas non-targeted MSC did not give such persistent effects. Histological analysis revealed increased brain restoration in engineered MSC-treated mice. In conclusion, MSCs can be genetically engineered to target the brain and prolong therapeutic efficacy in an EAE model.
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| 10. |
- Globisch, Maria A., et al.
(författare)
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Immunothrombosis and vascular heterogeneity in cerebral cavernous malformation
- 2022
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Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 140:20, s. 2154-2169
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Tidskriftsartikel (refereegranskat)abstract
- Cerebral cavernous malformation (CCM) is a neurovascular disease that results in various neurological symptoms. Thrombi have been reported in surgically resected CCM patient biopsies, but the molecular signatures of these thrombi remain elusive. Here, we investigated the kinetics of thrombi formation in CCM and how thrombi affect the vasculature and contribute to cerebral hypoxia. We used RNA sequencing to investigate the transcriptome of mouse brain endothelial cells with an inducible endothelial-specific Ccm3 knock-out (Ccm3-iECKO). We found that Ccm3-deficient brain endothelial cells had a higher expression of genes related to the coagulation cascade and hypoxia when compared with wild-type brain endothelial cells. Immunofluorescent assays identified key molecular signatures of thrombi such as fibrin, von Willebrand factor, and activated platelets in Ccm3-iECKO mice and human CCM biopsies. Notably, we identified polyhedrocytes in Ccm3-iECKO mice and human CCM biopsies and report it for the first time. We also found that the parenchyma surrounding CCM lesions is hypoxic and that more thrombi correlate with higher levels of hypoxia. We created an in vitro model to study CCM pathology and found that human brain endothelial cells deficient for CCM3 expressed elevated levels of plasminogen activator inhibitor-1 and had a redistribution of von Willebrand factor. With transcriptomics, comprehensive imaging, and an in vitro CCM preclinical model, this study provides experimental evidence that genes and proteins related to the coagulation cascade affect the brain vasculature and promote neurological side effects such as hypoxia in CCMs. This study supports the concept that antithrombotic therapy may be beneficial for patients with CCM.
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| 11. |
- Huang, Hua, 1986-, et al.
(författare)
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ELTD1-deletion reduces vascular abnormality and improves T-cell recruitment after PD-1 blockade in glioma.
- 2021
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Ingår i: Neuro-Oncology. - : Oxford University Press. - 1522-8517 .- 1523-5866. ; 24:3, s. 398-411
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Tumor vessels in glioma are molecularly and functionally abnormal, contributing to treatment resistance. Proteins differentially expressed in glioma vessels can change vessel phenotype and be targeted for therapy. ELTD1 (Adgrl4) is an orphan member of the adhesion G-protein-coupled receptor family upregulated in glioma vessels, and has been suggested as a potential therapeutic target. However, the role of ELTD1 in regulating vessel function in glioblastoma is poorly understood.METHODS: ELTD1 expression in human gliomas and its association with patient survival was determined using tissue microarrays and public databases. The role of ELTD1 in regulating tumor vessel phenotype was analyzed using orthotopic glioma models and ELTD1 -/- mice. Endothelial cells isolated from murine gliomas were transcriptionally profiled to determine differentially expressed genes and pathways. The consequence of ELTD1-deletion on glioma immunity was determined by treating tumor bearing mice with PD-1-blocking antibodies.RESULTS: ELTD1 levels were upregulated in human glioma vessels, increased with tumor malignancy, and were associated with poor patient survival. Progression of orthotopic gliomas was not affected by ELTD1-deletion, however, tumor vascular function was improved in ELTD1 -/- mice. Bioinformatic analysis of differentially expressed genes indicated increased inflammatory response and decreased proliferation in tumor endothelium in ELTD1 -/- mice. Consistent with an enhanced inflammatory response, ELTD1-deletion improved T-cell infiltration in GL261-bearing mice after PD-1 checkpoint blockade.CONCLUSION: Our data demonstrate that ELTD1 participates in inducing vascular dysfunction in glioma, and suggests that targeting of ELTD1 may normalize the vessels and improve the response to immunotherapy.
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| 12. |
- Johansson, Ulrika, 1974-, et al.
(författare)
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Formation of composite endothelial cell-mesenchymal stem cell islets : a novel approach to promote islet revascularization
- 2008
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 57:9, s. 2393-2401
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE: Mesenchymal stem cells (MSCs) contribute to endothelial cell (EC) migration by producing proteases, thereby paving the way into the tissues for ECs. MSCs were added to our previously described composite EC islets as a potential means to improve their capacity for islet angiogenesis. RESEARCH DESIGN AND METHODS: Human islets were coated with primary human bone marrow-derived MSCs and dermal microvascular ECs. The capacity of ECs, with or without MSCs, to adhere to and grow into human islets was analyzed. The survival and functionality of these composite islets were evaluated in a dynamic perifusion assay, and their capacity for angiogenesis in vitro was assessed in a three-dimensional fibrin gel assay. RESULTS: ECs proliferated after culture in MSC-conditioned medium, and MSCs improved the EC coverage threefold compared with EC islets alone. Islet survival in vitro and the functionality of the composite islets after culture were equal to those of control islets. The EC-MSC islets showed a twofold increase in total sprout formation compared with EC islets, and vascular sprouts emanating from the EC-MSC-islet surface showed migration of ECs into the islets and also into the surrounding matrix, either alone or in concert with MSCs. CONCLUSIONS: EC proliferation, sprout formation, and ingrowth of ECs into the islets were enhanced by MSCs. The use of composite EC-MSC islets may have beneficial effects on revascularization and immune regulation. The technique presented allows for pretreatment of donor islets with recipient-derived ECs and MSCs as a means of improving islet engraftment.
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| 13. |
- Kourtzelis, Iannis, et al.
(författare)
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Regulation of Instant Blood Mediated Infl ammatoryReaction (IBMIR) in Pancreatic Islet Xeno-Transplantation : Points for Therapeutic Interventions
- 2015
-
Ingår i: Immune Responses to Biosurfaces. - Cham : Springer. - 9783319186023 - 9783319186030 ; , s. 171-188
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Bokkapitel (refereegranskat)abstract
- Xeno-transplantation of pancreatic islets represents a promising therapeuticalternative for the treatment of type 1 diabetes mellitus. However, potentinnate immune responses induced shortly after the transplantation of donor islets tothe recipient, comprising the Instant Blood Mediated Immune Reaction (IBMIR),exert detrimental actions on islet graft function. The coagulation and complementcascades together with the leukocyte and platelet populations are the major playersin IBMIR. This innate immune attack affects dramatically islet integrity and leadsto signifi cant loss of function of the xenograft. In the present review, we focus on themechanisms contributing to IBMIR components and address therapeutic interventionapproaches to limit IBMIR by administering inhibitors in circulation, by coatingthe islet surface with inhibitors or by generating transgenic donor animals; theseapproaches could result in improved xenograft survival.
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| 14. |
- Kourtzelis, Ioannis, et al.
(författare)
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Regulation of Instant Blood Mediated Inflammatory Reaction (IBMIR) in Pancreatic Islet Xeno-Transplantation : Points for Therapeutic Interventions
- 2015
-
Ingår i: IMMUNE RESPONSES TO BIOSURFACES. - Cham : Springer International Publishing. - 9783319186030 - 9783319186023 ; , s. 171-188
-
Konferensbidrag (refereegranskat)abstract
- Xeno-transplantation of pancreatic islets represents a promising therapeutic alternative for the treatment of type 1 diabetes mellitus. However, potent innate immune responses induced shortly after the transplantation of donor islets to the recipient, comprising the Instant Blood Mediated Immune Reaction (IBMIR), exert detrimental actions on islet graft function. The coagulation and complement cascades together with the leukocyte and platelet populations are the major players in IBMIR. This innate immune attack affects dramatically islet integrity and leads to significant loss of function of the xenograft. In the present review, we focus on the mechanisms contributing to IBMIR components and address therapeutic intervention approaches to limit IBMIR by administering inhibitors in circulation, by coating the islet surface with inhibitors or by generating transgenic donor animals; these approaches could result in improved xenograft survival.
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| 15. |
- Magnusson, Peetra U, et al.
(författare)
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FGFR-1 regulates angiogenesis through cytokines interleukin-4 and pleiotrophin
- 2007
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Ingår i: Blood. - : American Society of Hematology. - 0006-4971 .- 1528-0020. ; 110:13, s. 4214-4222
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Tidskriftsartikel (refereegranskat)abstract
- The role of fibroblast growth factors (FGFs) in blood vessel formation has remained unclear. We used differentiating stem-cell cultures (embryoid bodies) and teratomas to show that FGIF receptor-1 (FGFR-1) exerts a negative regulatory effect on endothelial cell function in these models. Embryoid bodies lacking expression of FGFR-1 as a result of gene targeting (Fgfr-1(-/-)) displayed increased vascularization and a distinct, elongated vessel morphology. Teratomas derived from FGFR-1-deficient stem cells were characterized by an increased growth rate and abundant, morphologically distinct vessels. Transmission electron microscopy of the Fgfr-1(-/-) teratomas showed a compact and voluminous but functional endothelium, which anastomosed with the host circulation. The increased vascularization and altered endothelial cell morphology was dependent on secreted factor(s), based on the transfer of the Fgfr-1(+/-) vascular phenotype by conditioned medium to Fgfr-1(+/-) embryoid bodies. Antibody and transcript arrays showed down-regulation of interleukin-4 (IL-4) and up-regulation of pleiotrophin in Fgfr-1(-/-) embryoid bodies, compared with the heterozygous cultures. We used neutralizing antibodies to show that IL-4 and pleiotrophin act as negative and positive angiogenic regulators, respectively. We conclude that FGFR-1 negatively regulates endothelial cell function by altering the balance of modulatory cytokines.
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| 16. |
- Magnusson, Peetra U., et al.
(författare)
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Platelet-derived growth factor receptor-beta constitutive activity promotes angiogenesis in vivo and in vitro
- 2007
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Ingår i: Arteriosclerosis, Thrombosis and Vascular Biology. - 1079-5642 .- 1524-4636. ; 27:10, s. 2142-2149
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE - Knockout studies have demonstrated crucial roles for the platelet-derived growth factor-B and its cognate receptor, platelet-derived growth factor receptor-β (PDGFR-β), in blood vessel maturation, that is, the coverage of newly formed vessels with mural cells/pericytes. This study describes the consequences of a constitutively activating mutation of the PDGFR-β (Pdgfrb) introduced into embryonic stem cells with respect to vasculogenesis/angiogenesis in vitro and in vivo. METHODS AND RESULTS - Embryonic stem cells were induced to either form teratomas in vivo or embryoid bodies, an in vitro model for mouse embryogenesis. Western blotting studies on embryoid bodies showed that expression of a single allele of the mutant Pdgfrb led to increased levels of PDGFR-β tyrosine phosphorylation and augmented downstream signal transduction. This was accompanied by enhanced vascular development, followed by exaggerated angiogenic sprouting with abundant pericyte coating as shown by immunohistochemistry/immunofluorescence. Pdgfrb embryoid bodies were characterized by increased expression of vascular endothelial growth factor (VEGF)-A and VEGF receptor-2; neutralizing antibodies against VEGF-A/VEGF receptor-2 blocked vasculogenesis and angiogenesis in mutant embryoid bodies. Moreover, Pdgfrb embryonic stem cell-derived teratomas in nude mice were more densely vascularized than wild-type teratomas. CONCLUSION - Increased PDGFR-β kinase activity is associated with elevated expression of VEGF-A and VEGF receptor-2, acting directly on endothelial cells and resulting in increased vessel formation.
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| 17. |
- Malinverno, Matteo, et al.
(författare)
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Endothelial cell clonal expansion in the development of cerebral cavernous malformations
- 2019
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Ingår i: Nature Communications. - : NATURE PUBLISHING GROUP. - 2041-1723. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- Cerebral cavernous malformation (CCM) is a neurovascular familial or sporadic disease that is characterised by capillary-venous cavernomas, and is due to loss-of-function mutations to any one of three CCM genes. Familial CCM follows a two-hit mechanism similar to that of tumour suppressor genes, while in sporadic cavernomas only a small fraction of endothelial cells shows mutated CCM genes. We reported that in mouse models and in human patients, endothelial cells lining the lesions have different features from the surrounding endothelium, as they express mesenchymal/stem-cell markers. Here we show that cavernomas originate from clonal expansion of few Ccm3-null endothelial cells that express mesenchymal/stem-cell markers. These cells then attract surrounding wild-type endothelial cells, inducing them to express mesenchymal/stem-cell markers and to contribute to cavernoma growth. These characteristics of Ccm3-null cells are reminiscent of the tumour-initiating cells that are responsible for tumour growth. Our data support the concept that CCM has benign tumour characteristics.
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| 18. |
- Moll, Guido, et al.
(författare)
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Are Therapeutic Human Mesenchymal Stromal Cells Compatible with Human Blood?
- 2012
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Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1066-5099 .- 1549-4918. ; 30:7, s. 1565-1574
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Tidskriftsartikel (refereegranskat)abstract
- Multipotent mesenchymal stromal cells (MSCs) are tested in numerous clinical trials. Questions have been raised concerning fate and function of these therapeutic cells after systemic infusion. We therefore asked whether culture-expanded human MSCs elicit an innate immune attack, termed instant blood-mediated inflammatory reaction (IBMIR), which has previously been shown to compromise the survival and function of systemically infused islet cells and hepatocytes. We found that MSCs expressed hemostatic regulators similar to those produced by endothelial cells but displayed higher amounts of prothrombotic tissue/stromal factors on their surface, which triggered the IBMIR after blood exposure, as characterized by formation of blood activation markers. This process was dependent on the cell dose, the choice of MSC donor, and particularly the cell-passage number. Short-term expanded MSCs triggered only weak blood responses in vitro, whereas extended culture and coculture with activated lymphocytes increased their prothrombotic properties. After systemic infusion to patients, we found increased formation of blood activation markers, but no formation of hyperfibrinolysis marker D-dimer or acute-phase reactants with the currently applied dose of 1.0-3.0 x 10(6) cells per kilogram. Culture-expanded MSCs trigger the IBMIR in vitro and in vivo. Induction of IBMIR is dose-dependent and increases after prolonged ex vivo expansion. Currently applied doses of low-passage clinical-grade MSCs elicit only minor systemic effects, but higher cell doses and particularly higher passage cells should be handled with care. This deleterious reaction can compromise the survival, engraftment, and function of these therapeutic cells.
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| 19. |
- Nilsson, Per H., et al.
(författare)
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Autoregulation of thromboinflammation on biomaterial surfaces by a multicomponent therapeutic coating
- 2013
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Ingår i: Biomaterials. - : Elsevier BV. - 0142-9612 .- 1878-5905. ; 34:4, s. 985-994
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Tidskriftsartikel (refereegranskat)abstract
- Activation of the thrombotic and complement systems is the main recognition and effector mechanisms in the multiple adverse biological responses triggered when biomaterials or therapeutic cells come into blood contact. We have created a surface which is auto-protective to human innate immunity by combining three fundamentally different strategies, all developed by us previously, which have been shown to induce substantial, but incomplete hemocompatibility when used separately. In summary, we have conjugated a factor H-binding peptide; and an ADP-degrading enzyme; using a PEG linker on both material and cellular surfaces. When exposed to human whole blood, factor H was specifically recruited to the modified surfaces and inhibited complement attack. In addition, activation of platelets and coagulation was efficiently attenuated, by degrading ADP. Thus, by inhibiting thromboinflammation using a multicomponent approach, we have created a hybrid surface with the potential to greatly reduce incompatibility reactions involving biomaterials and transplantation.
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| 20. |
- Nordling, Sofia, 1985-, et al.
(författare)
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Enhanced protection of the renal vascular endothelium improves early outcome in kidney transplantation : Preclinical investigations in pig and mouse
- 2018
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Ingår i: Scientific Reports. - : Nature Publishing Group. - 2045-2322. ; 8
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Tidskriftsartikel (refereegranskat)abstract
- Ischemia reperfusion injury is one of the major complications responsible for delayed graft function in kidney transplantation. Applications to reduce reperfusion injury are essential due to the widespread use of kidneys from deceased organ donors where the risk for delayed graft function is especially prominent. We have recently shown that coating of inflamed or damaged endothelial cells with a unique heparin conjugate reduces thrombosis and leukocyte recruitment. In this study we evaluated the binding capacity of the heparin conjugate to cultured human endothelial cells, to kidneys from brain-dead porcine donors, and to murine kidneys during static cold storage. The heparin conjugate was able to stably bind cultured endothelial cells with high avidity, and to the renal vasculature of explanted kidneys from pigs and mice. Treatment of murine kidneys prior to transplantation reduced platelet deposition and leukocyte infiltration 24 hours post-transplantation, and significantly improved graft function. The present study thus shows the benefits of enhanced protection of the renal vasculature during cold storage, whereby increasing the antithrombotic and anti-adhesive properties of the vascular endothelium yields improved renal function early after transplantation.
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| 21. |
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| 22. |
- Nordling, Sofia, et al.
(författare)
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Vascular repair utilising immobilised heparin conjugate for protection against early activation of inflammation and coagulation
- 2015
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Ingår i: Thrombosis and Haemostasis. - 0340-6245 .- 2567-689X. ; 113:6, s. 1312-1322
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Tidskriftsartikel (refereegranskat)abstract
- Ischaemia-reperfusion injury (IRI) poses a major challenge in many thrombotic conditions and in whole organ transplantation. Activation of the endothelial cells and shedding of the protective vascular glycocalyx during IRI increase the risk of innate immune activation, cell infiltration and severe thrombus formation, promoting damage to the tissue. Here, we present a novel one-step strategy to protect the vas, culature by immobilisation of a unique multi-arm heparin conjugate to the endothelium. Applying a new in vitro blood endothelial cell chamber model, the heparin conjugate was found to bind not only to primary human endothelial cells but also directly to the collagen to which the cells adhered. Incubation of hypoxic endothelial cells with freshly drawn human blood in the blood chambers elicited coagulation activation reflected by thrombin anti-thrombin formation and binding of platelets and neutrophils. Immobilisation of the heparin conjugate to the hypoxic endothelial cells created a protective coating, leading to a Significant reduction of the recruitment of blood cells and coagulation activation compared to untreated hypoxic endothelial cells. This novel approach of immobilising multi-arm heparin conjugates on the endothelial cells and collagen of the basement membrane ensures to protect the endothelium against IRI in thrombotic disorders and in transplantation.
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| 23. |
- Rezai Jahromi, Behnam, et al.
(författare)
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Slow-Closing Clip for the Treatment of Nonsaccular Vertebrobasilar Aneurysms : A Retrospective Case Series
- 2022
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Ingår i: World Neurosurgery. - : Elsevier. - 1878-8750 .- 1878-8769. ; 168, s. e645-e665
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Tidskriftsartikel (refereegranskat)abstract
- ObjectiveVertebrobasilar artery nonsaccular aneurysms (VBANSAs) are associated with a 13% annual mortality. Revascularization and flow diversion are life-saving options in select cases; technical failures and rapid hemodynamic changes may contribute to unwanted outcomes. We describe a technique and report clinical outcomes of patients treated with an experimental slow-closing clip (SCC).MethodsAn experimental SCC was created to gradually close the parent artery of aneurysms. Clinical, radiographic, and outcome data from patients with VBANSAs who underwent experimental treatment with the SCC were retrospectively analyzed.ResultsAmong 10 patients (7 men; mean age, 49.5 years; range, 18–73 years), 6 presented with mass effect symptoms, 1 with ischemic stroke, 2 with subarachnoid hemorrhage, and 1 with hydrocephalus. Five patients underwent revascularization plus SCC application, and 5 were treated with SCC alone. The mean follow-up was 6.7 years. The expected mortality among patients with unruptured VBANSAs with previous treatment options in this period was 52.7%, whereas the observed rate was 20%. Four patients died within 12 months after treatment. Causes of death were brainstem ischemic stroke, poor-grade subarachnoid hemorrhage, poor clinical presentation, and unknown. Six patients were alive at last follow-up, with unchanged or improved modified Rankin Scale scores. Mortality was associated with posterior-projecting aneurysms and late-stage treatment.ConclusionsIn this small case series, use of SCC overcame the natural history of VBANSAs when treatment timing and aneurysm anatomy were suitable. The SCC potentially favors aneurysm thrombosis and collateral reactivation. More studies are necessary to better develop the SCC.
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| 24. |
- Selvarajan, Ilakya, et al.
(författare)
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Coronary Artery Disease Risk Variant Dampens the Expression of CALCRL by Reducing HSF Binding to Shear Stress Responsive Enhancer in Endothelial Cells In Vitro
- 2024
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Ingår i: Arteriosclerosis, Thrombosis and Vascular Biology. - : Lippincott Williams & Wilkins. - 1079-5642 .- 1524-4636. ; 44:6, s. 1330-1345
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND:CALCRL (calcitonin receptor-like) protein is an important mediator of the endothelial fluid shear stress response, which is associated with the genetic risk of coronary artery disease. In this study, we functionally characterized the noncoding regulatory elements carrying coronary artery disease that risks single-nucleotide polymorphisms and studied their role in the regulation of CALCRL expression in endothelial cells.METHODS:To functionally characterize the coronary artery disease single-nucleotide polymorphisms harbored around the gene CALCRL, we applied an integrative approach encompassing statistical, transcriptional (RNA-seq), and epigenetic (ATAC-seq [transposase-accessible chromatin with sequencing], chromatin immunoprecipitation assay-quantitative polymerase chain reaction, and electromobility shift assay) analyses, alongside luciferase reporter assays, and targeted gene and enhancer perturbations (siRNA and clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeat-associated 9) in human aortic endothelial cells.RESULTS:We demonstrate that the regulatory element harboring rs880890 exhibits high enhancer activity and shows significant allelic bias. The A allele was favored over the G allele, particularly under shear stress conditions, mediated through alterations in the HSF1 (heat shock factor 1) motif and binding. CRISPR deletion of rs880890 enhancer resulted in downregulation of CALCRL expression, whereas HSF1 knockdown resulted in a significant decrease in rs880890-enhancer activity and CALCRL expression. A significant decrease in HSF1 binding to the enhancer region in endothelial cells was observed under disturbed flow compared with unidirectional flow. CALCRL knockdown and variant perturbation experiments indicated the role of CALCRL in mediating eNOS (endothelial nitric oxide synthase), APLN (apelin), angiopoietin, prostaglandins, and EDN1 (endothelin-1) signaling pathways leading to a decrease in cell proliferation, tube formation, and NO production.CONCLUSIONS:Overall, our results demonstrate the existence of an endothelial-specific HSF (heat shock factor)-regulated transcriptional enhancer that mediates CALCRL expression. A better understanding of CALCRL gene regulation and the role of single-nucleotide polymorphisms in the modulation of CALCRL expression could provide important steps toward understanding the genetic regulation of shear stress signaling responses.
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