| 1. |
- Akerman, Ildem, et al.
(författare)
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Human Pancreatic β Cell lncRNAs Control Cell-Specific Regulatory Networks
- 2017
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Ingår i: Cell Metabolism. - : Elsevier BV. - 1550-4131. ; 25:2, s. 400-411
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Tidskriftsartikel (refereegranskat)abstract
- Recent studies have uncovered thousands of long non-coding RNAs (lncRNAs) in human pancreatic β cells. β cell lncRNAs are often cell type specific and exhibit dynamic regulation during differentiation or upon changing glucose concentrations. Although these features hint at a role of lncRNAs in β cell gene regulation and diabetes, the function of β cell lncRNAs remains largely unknown. In this study, we investigated the function of β cell-specific lncRNAs and transcription factors using transcript knockdowns and co-expression network analysis. This revealed lncRNAs that function in concert with transcription factors to regulate β cell-specific transcriptional networks. We further demonstrate that the lncRNA PLUTO affects local 3D chromatin structure and transcription of PDX1, encoding a key β cell transcription factor, and that both PLUTO and PDX1 are downregulated in islets from donors with type 2 diabetes or impaired glucose tolerance. These results implicate lncRNAs in the regulation of β cell-specific transcription factor networks.
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| 2. |
- Atla, Goutham, et al.
(författare)
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Genetic regulation of RNA splicing in human pancreatic islets
- 2022
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Ingår i: Genome Biology. - : Springer Science and Business Media LLC. - 1474-760X. ; 23, s. 1-28
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundNon-coding genetic variants that influence gene transcription in pancreatic islets play a major role in the susceptibility to type 2 diabetes (T2D), and likely also contribute to type 1 diabetes (T1D) risk. For many loci, however, the mechanisms through which non-coding variants influence diabetes susceptibility are unknown.ResultsWe examine splicing QTLs (sQTLs) in pancreatic islets from 399 human donors and observe that common genetic variation has a widespread influence on the splicing of genes with established roles in islet biology and diabetes. In parallel, we profile expression QTLs (eQTLs) and use transcriptome-wide association as well as genetic co-localization studies to assign islet sQTLs or eQTLs to T2D and T1D susceptibility signals, many of which lack candidate effector genes. This analysis reveals biologically plausible mechanisms, including the association of T2D with an sQTL that creates a nonsense isoform in ERO1B, a regulator of ER-stress and proinsulin biosynthesis. The expanded list of T2D risk effector genes reveals overrepresented pathways, including regulators of G-protein-mediated cAMP production. The analysis of sQTLs also reveals candidate effector genes for T1D susceptibility such as DCLRE1B, a senescence regulator, and lncRNA MEG3.ConclusionsThese data expose widespread effects of common genetic variants on RNA splicing in pancreatic islets. The results support a role for splicing variation in diabetes susceptibility, and offer a new set of genetic targets with potential therapeutic benefit.
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| 3. |
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| 4. |
- Brandhorst, Daniel, et al.
(författare)
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Multicenter Assessment of Animal-free Collagenase AF-1 for Human Islet Isolation
- 2017
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Ingår i: Cell Transplantation. - : Sage Publications. - 0963-6897 .- 1555-3892. ; 26:10, s. 1688-1693
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Tidskriftsartikel (refereegranskat)abstract
- Animal-free (AF) SERVA Collagenase AF-1 and Neutral Protease (NP) AF GMP Grade have recently become available for human islet isolation. This report describes the initial experiences of 3 different islet transplant centers. Thirty-four human pancreases were digested using 1 vial of the 6 different lots of Collagenase AF-1 (2,000-2,583 PZ-U/vial) supplemented with 4 different lots of NP AF in a range of 50 to 160 DMC-U per pancreas. Isolation, culture, and quality assessment were performed using standard techniques as previously described. All data are presented as mean +/- standard error of the mean (SEM). Variability of pancreas weight was associated with a wide range of collagenase and NP activities, ranging from 12.7 to 46.6 PZ-U/g (26.0 +/- 1.5 PZ-U/g) and 0.4 to 3.0 DMC-U/g (1.5 +/- 0.1 DMC-U/g), respectively. Postpurification islet yield was 296,494 +/- 33,620 islet equivalents (IEQ) equivalent to 3,274 +/- 450 IEQ/g with a purity of 55.9% +/- 3.2%. Quality assessment performed after 2 to 4 d of culture demonstrated a viability of 88.1% +/- 1.5% and a stimulation index of 3.7 +/- 0.7. Eighteen of the 34 preparations were transplanted into type 1 diabetic patients equivalent to a transplantation rate of 52.9%. Six preparations, which were infused into patients as first transplant, could be analyzed and increased the fasting C-peptide level from 0.11 +/- 0.08 pretransplant to 1.23 +/- 0.24 and 2.27 +/- 0.31 ng/mL 3 and 6 mo posttransplant (P < 0.05), respectively. Insulin requirements were simultaneously reduced at the same time from 39.2 +/- 3.8 IU/d before transplantation to 10.8 +/- 4.1 and 4.0 +/- 2.3 IU/d, after 3 and 6 mo posttransplant (P < 0.05), respectively. This study demonstrates the efficiency of AF SERVA Collagenase AF-1 and NP AF for clinical islet isolation and transplantation. The new plant-based production process makes these products a safe new option for the islet field.
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| 5. |
- Hårdstedt, Maria
(författare)
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Studies of Innate and Adaptive Immunity in Islet Transplantation
- 2014
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Clinical islet transplantation is today an established alternative treatment for a selected group of type 1 diabetes patients. The predominant technique for transplantation is infusion of islets in the liver via the portal vein. Obstacles to advancing islet transplantation include limited engraftment resulting from an immediate blood-mediated inflammatory reaction (IBMIR), a life-long need for immunosuppression and the shortage of organs available.In this thesis, innate and adaptive immunity were explored in allogeneic and xenogeneic settings, with the long-term goal of preventing islet graft destruction. Methods for studying immune responses to islets in blood and engrafted islets in liver tissue (intragraft gene expression) were developed and refined. The innate response to human islets and exocrine tissue in ABO-compatible blood was characterized up to 48 h using a novel whole-blood model. Physiological changes in the blood during incubations were explored and adjusted to allow prolonged experiments. Increased production of chemokines targeting CXCR1/2, CCR2 and CXCR3 was observed, accompanied by massive intra-islet neutrophil infiltration. Notably, endocrine and exocrine tissue triggered a similarly strong innate immune response.Two studies of adult porcine islet transplantation to non-human primates (NHPs) were performed. Expression of immune response genes induced in liver tissue of non-immunosuppressed NHPs (≤72 h) was evaluated after porcine islet transplantation. Up-regulation of CXCR3 mRNA, together with IP-10, Mig, MIP-1α, RANTES, MCP-1 and cytotoxic effector molecule transcripts, was associated with T-cell and macrophage infiltration at 48-72 h. Long-term survival (>100 days) of adult porcine islets in a NHP model was later demonstrated using T-cell-based immunosuppression, including co-stimulatory blockade (anti-CD154 mAb). Graft failure was associated with increased levels of circulating, indirectly activated T cells, non-Gal pig-specific IgG and gene transcripts of inflammatory cytokines. Microarray analysis of the response to inflammatory cytokines in cultured porcine islets identified genes involved in cell death, immune responses and oxidative stress; this gene pattern coincided with physiological changes (decrease in insulin and ATP content).In summary, allogeneic whole-blood experiments and xenogeneic in vivo studies underscored the importance of preventing early inflammation and cell-recruitment to avoid islet graft loss in islet transplantation. Long-term survival of porcine islets in NHPs was shown to be feasible using T-cell-directed immunosuppression, including anti-CD154 mAb.
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| 6. |
- Miguel-Escalada, Irene, et al.
(författare)
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Human pancreatic islet three-dimensional chromatin architecture provides insights into the genetics of type 2 diabetes
- 2019
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Ingår i: Nature Genetics. - : Springer Science and Business Media LLC. - 1061-4036 .- 1546-1718. ; 51:7, s. 1137-1148
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Tidskriftsartikel (refereegranskat)abstract
- Genetic studies promise to provide insight into the molecular mechanisms underlying type 2 diabetes (T2D). Variants associated with T2D are often located in tissue-specific enhancer clusters or super-enhancers. So far, such domains have been defined through clustering of enhancers in linear genome maps rather than in three-dimensional (3D) space. Furthermore, their target genes are often unknown. We have created promoter capture Hi-C maps in human pancreatic islets. This linked diabetes-associated enhancers to their target genes, often located hundreds of kilobases away. It also revealed >1,300 groups of islet enhancers, super-enhancers and active promoters that form 3D hubs, some of which show coordinated glucose-dependent activity. We demonstrate that genetic variation in hubs impacts insulin secretion heritability, and show that hub annotations can be used for polygenic scores that predict T2D risk driven by islet regulatory variants. Human islet 3D chromatin architecture, therefore, provides a framework for interpretation of T2D genome-wide association study (GWAS) signals.
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| 7. |
- Nano, Rita, et al.
(författare)
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Heterogeneity of Human Pancreatic Islet Isolation Around Europe : Results of a Survey Study
- 2020
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Ingår i: Transplantation. - : LIPPINCOTT WILLIAMS & WILKINS. - 0041-1337 .- 1534-6080. ; 104:1, s. 190-196
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Tidskriftsartikel (refereegranskat)abstract
- Background: Europe is currently the most active region in the field of pancreatic islet transplantation, and many of the leading groups are actually achieving similar good outcomes. Further collaborative advances in the field require the standardization of islet cell product isolation processes, and this work aimed to identify differences in the human pancreatic islet isolation processes within European countries.Methods: A web-based questionnaire about critical steps, including donor selection, pancreas processing, pancreas perfusion and digestion, islet counting and culture, islet quality evaluation, microbiological evaluation, and release criteria of the product, was completed by isolation facilities participating at the Ninth International European Pancreas and Islet Transplant Association (EPITA) Workshop on Islet-Beta Cell Replacement in Milan.Results: Eleven islet isolation facilities completed the questionnaire. The facilities reported 445 and 53 islet isolations per year over the last 3 years from deceased organ donors and pancreatectomized patients, respectively. This activity resulted in 120 and 40 infusions per year in allograft and autograft recipients, respectively. Differences among facilities emerged in donor selection (age, cold ischemia time, intensive care unit length, amylase concentration), pancreas procurement, isolation procedures (brand and concentration of collagenase, additive, maximum acceptable digestion time), quality evaluation, and release criteria for transplantation (glucose-stimulated insulin secretion tests, islet numbers, and purity). Moreover, even when a high concordance about the relevance of one parameter was evident, thresholds for the acceptance were different among facilities.Conclusions: The result highlighted the presence of a heterogeneity in the islet cell product process and product release criteria.
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| 8. |
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| 9. |
- Nano, Rita, et al.
(författare)
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Islets for Research : Nothing Is Perfect, but We Can Do Better
- 2019
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Ingår i: Diabetes. - : AMER DIABETES ASSOC. - 0012-1797 .- 1939-327X. ; 68:8, s. 1541-1543
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Tidskriftsartikel (refereegranskat)abstract
- In December 2018, Diabetes and Diabetologia began requiring authors of papers reporting data obtained from studies on human islets to report critical characteristics of the human islets used for research. The islet community was asked to provide feedback on it. Here is the contribution by the European Consortium for Islet Transplantation.
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| 10. |
- Olerud, Johan, 1977-
(författare)
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Role of Thrombospondin-1 in Endogenous and Transplanted Pancreatic Islets
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Type 1 diabetes mellitus is a severe life-long disease with a pronounced risk of developing secondary complications. One way to avoid the latter is to restore the fine tuning of blood glucose homeostasis by transplantation of pancreatic islets. However, isolated islets need to be properly engrafted and to re-establish a vascular network in order to regain function. Earlier studies have shown that pancreatic islets experimentally transplanted to e.g. the liver or the kidney become poorly revascularized. In the present thesis, mice deficient of the angiostatic factor thrombospondin-1 (TSP-1) were found to have an impaired beta-cell function. Development of this beta-cell dysfunction was prevented by treatment of TSP-1 deficient mice from birth with the TGFbeta-1 activating sequence of TSP-1. TSP-1 in islets was predominantly expressed in the endothelial cells. Isolated islet endothelial cells was observed to have a low proliferatory and migratory capacity towards angiogenic stimuli, but this could be reversed by neutralizing antibodies to the angiostatic factors alpha1-antitrypsin, endostatin or TSP-1. Transient downregulation of TSP-1 expression in mouse islet cells prior to transplantation improved graft revascularization, blood perfusion, oxygenation and function when evaluated one-month post-transplantation. The same result was achieved when islets or recipients of islets were pre-treated with the hormone prolactin one-month post-transplantation. The present study illustrates the importance of the angiostatic factor TSP-1 for islet beta-cell function and engraftment of islets following transplantation. Interference with TSP-1 can possibly be used to improve the results of clinical islet transplantation.
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