| 1. |
- Afonso, Georgia, et al.
(författare)
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Critical parameters in blood processing for T-cell assays: Validation on ELISpot and tetramer platforms
- 2010
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Ingår i: Journal of Immunological Methods. - : Elsevier BV. - 1872-7905 .- 0022-1759. ; 359:1-2, s. 28-36
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Tidskriftsartikel (refereegranskat)abstract
- Assays detecting antigen (Ag)-specific T-cell responses in immune-mediated processes are increasingly employed to understand disease pathogenesis and "immune staging". The quantity and quality of starting peripheral blood mononuclear cell (PBMC) preparations are important factors in the performance of such assays. We therefore compared final PBMC yield and function by modifying parameters at the blood drawing, storage and processing steps. While drawing blood in vacuum-driven tubes or syringes and separating PBMCs on density gradients using standard or membrane (Leucosep (R)) tubes made no difference, storing tubes for 18 h without any agitation led to PBMC preparations contaminated with granulocytes and decreased interferon (IFN)-gamma enzyme-linked immunospot (ELISpot) responses. Even agitated blood showed a trend towards reduced ELISpot responses and increased human leukocyte Ag (HLA) multimer readouts when stored for 18 h compared to 3 h. These changes were reduced by diluting blood prior to storage. Washing PBMCs with media containing 10% human serum increased PBMC yields by 40.5%, without affecting ELISpot responses and multimer counts. However, washes with >10% human serum decreased multimer counts, with no additional improvement in PBMC yields. These findings may be relevant for optimizing and harmonizing PBMC processing procedures for T-cell assays. (C) 2010 Elsevier B.V. All rights reserved.
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| 2. |
- Agardh, Carl-David, et al.
(författare)
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Clinical evidence for the safety of GAD65 immunomodulation in adult-onset autoimmune diabetes.
- 2005
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Ingår i: Journal of Diabetes and its Complications. - : Elsevier BV. - 1873-460X .- 1056-8727. ; 19:4, s. 238-246
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Tidskriftsartikel (refereegranskat)abstract
- The purpose of this Phase II study was to evaluate if alum-formulated human recombinant GAD65 is safe and does not compromise beta cell function. The study was conducted as a randomized, double blind, placebo-controlled, dose-escalation clinical trial in a total of 47 Latent Autoimmune Diabetes in Adults (LADA) patients who received either placebo or 4, 20, 100, or 500 μg Diamyd subcutaneously at Weeks 1 and 4. Safety evaluations, including neurology, beta cell function tests, diabetes status assessment, hematology, biochemistry, and cellular and humoral immunological markers, were repeatedly assessed over 24 weeks. None of the patients had significant study-related adverse events (AE). Fasting c-peptide levels at 24 weeks were increased compared with placebo (P=.0015) in the 20 μg but not in the other dose groups. In addition, both fasting (P=.0081) and stimulated (P=.0236) c-peptide levels increased from baseline to 24 weeks in the 20 μg dose group. GADA log levels clearly increased (P=.0002) in response to 500 μg Diamyd. The CD4+CD25+/CD4+CD25− cell ratio increased (P=.0128) at 24 weeks in the 20 μg group. No sudden increase in HbA1c or plasma glucose or decrease in beta cell function was observed in any of the dose groups. These positive findings for clinical safety further support the clinical development of Diamyd as a therapeutic to prevent autoimmune diabetes.
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| 3. |
- Andersson, Cecilia K, et al.
(författare)
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Glucose tolerance and beta-cell function in islet autoantibody-positive children recruited to a secondary prevention study.
- 2013
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Ingår i: Pediatric Diabetes. - : Hindawi Limited. - 1399-543X .- 1399-5448. ; 14:5, s. 341-349
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Tidskriftsartikel (refereegranskat)abstract
- AIMS: Children with type 1 diabetes (T1D) risk and islet autoantibodies are recruited to a secondary prevention study. The aims were to determine metabolic control in relation to human leukocyte antigen (HLA) genetic risk and islet autoantibodies in prepubertal children. METHODS: In 47 healthy children with GADA and at least one additional islet autoantibody, intravenous glucose tolerance test (IvGTT) and oral glucose tolerance test (OGTT) were performed 8-65 d apart. Hemoglobin A1c, plasma glucose as well as serum insulin and C-peptide were determined at fasting and during IvGTT and OGTT. RESULTS: All children aged median 5.1 (4.0-9.2) yr had autoantibodies to two to six of the beta-cell antigens GAD65, insulin, IA-2, and the three amino acid position 325 variants of the ZnT8 transporter. In total, 20/47 children showed impaired glucose metabolism. Decreased (≤30 μU/mL insulin) first-phase insulin response (FPIR) was found in 14/20 children while 11/20 had impaired glucose tolerance in the OGTT. Five children had both impaired glucose tolerance and FPIR ≤30 μU/mL insulin. Number and levels of autoantibodies were not associated with glucose metabolism, except for an increased frequency (p = 0.03) and level (p = 0.01) of ZnT8QA in children with impaired glucose metabolism. Among the children with impaired glucose metabolism, 13/20 had HLA-DQ2/8, compared to 9/27 of the children with normal glucose metabolism (p = 0.03). CONCLUSION: Secondary prevention studies in children with islet autoantibodies are complicated by variability in baseline glucose metabolism. Evaluation of metabolic control with both IvGTT and OGTT is critical and should be taken into account before randomization. All currently available autoantibody tests should be analyzed, including ZnT8QA.
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| 4. |
- Barchetta, Ilaria, et al.
(författare)
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Epigenetic changes induced by maternal factors during fetal life : Implication for type 1 diabetes
- 2021
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Ingår i: Genes. - : MDPI AG. - 2073-4425. ; 12:6
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Forskningsöversikt (refereegranskat)abstract
- Organ-specific autoimmune diseases, such as type 1 diabetes, are believed to result from T-cell-mediated damage of the target tissue. The immune-mediated tissue injury, in turn, is known to depend on complex interactions between genetic and environmental factors. Nevertheless, the mechanisms whereby environmental factors contribute to the pathogenesis of autoimmune diseases remain elusive and represent a major untapped target to develop novel strategies for disease prevention. Given the impact of the early environment on the developing immune system, epigenetic changes induced by maternal factors during fetal life have been linked to a likelihood of developing an autoimmune disease later in life. In humans, DNA methylation is the epigenetic mechanism most extensively investigated. This review provides an overview of the critical role of DNA methylation changes induced by prenatal maternal conditions contributing to the increased risk of immune-mediated diseases on the offspring, with a particular focus on T1D. A deeper understanding of epigenetic alterations induced by environmental stressors during fetal life may be pivotal for developing targeted prevention strategies of type 1 diabetes by modifying the maternal environment.
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| 5. |
- Behnan, Jinan, et al.
(författare)
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Recruited brain tumor-derived mesenchymal stem cells contribute to brain tumor progression.
- 2014
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Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 32:5, s. 1110-1123
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Tidskriftsartikel (refereegranskat)abstract
- The identity of the cells that contribute to brain tumor structure and progression remains unclear. Mesenchymal stem cells (MSCs) have recently been isolated from normal mouse brain. Here, we report the infiltration of MSC-like cells into the GL261 murine glioma model. These Brain Tumor derived Mesenchymal Stem Cells (BT-MSCs) are defined with the phenotype (Lin-Sca-1+CD9+CD44+CD166+/-) and have multipotent differentiation capacity. We show that the infiltration of BT-MSCs correlates to tumor progression; furthermore, BT-MSCs increased the proliferation rate of GL261 cells in vitro. For the first time, we report that the majority of GL261 cells expressed mesenchymal phenotype under both adherent and sphere culture conditions in vitro, and that the non-MSC population is non-tumorigenic in vivo. Although the GL261 cell line expressed mesenchymal phenotype markers in vitro, most BT-MSCs are recruited cells from host origin in both wild type GL261 inoculated into GFP-transgenic mice and GL261-GFP cells inoculated into wild type mice. We show the expression of chemokine receptors CXCR4 and CXCR6 on different recruited cell populations. In vivo, the GL261 cells change marker profile, and acquire a phenotype that is more similar to cells growing in sphere culture conditions. Finally, we identify a BT-MSC population in human glioblastoma that is CD44+CD9+CD166+ both in freshly isolated and culture-expanded cells. Our data indicate that cells with MSC-like phenotype infiltrate into the tumor stroma and play an important role in tumor cell growth in vitro and in vivo. Thus we suggest that targeting BT-MSCs could be a possible strategy for treating glioblastoma patients. Stem Cells 2013.
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| 6. |
- Bekris, L. M., et al.
(författare)
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GAD65 autoantibody epitopes in adult patients with latent autoimmune diabetes following GAD65 vaccination
- 2007
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Ingår i: Diabetic Medicine. - : Wiley. - 1464-5491 .- 0742-3071. ; 24:5, s. 521-526
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Tidskriftsartikel (refereegranskat)abstract
- Aims Subcutaneous injection of recombinant human GAD65 (rhGAD65) in patients with latent autoimmune diabetes in adults (LADA) correlates with an increase in C-peptide levels. In this study we analysed the effect of rhGAD65 administration on the GAD65-specific autoimmune response. Methods Longitudinal serum samples obtained from LADA patients (n = 47) who received 4, 20, 100 or 500 mu g alum-formulated rhGAD65 or placebo by subcutaneous injection twice (4 weeks apart) were analysed for their epitope recognition using GAD65-specific recombinant Fab and GAD65/67 fusion proteins. Results Overall, minor changes in the epitope pattern were observed using either approach. Only in the 500-mu g dosage group was an increase in GAD65Ab level associated with a significant increase in the binding to a conformational epitope located at the middle part of GAD65. Conclusions Our data suggest that the apparent beneficial effects of 20 mu g alum-formulated recombinant human GAD65 is not explained by changes in the GAD65Ab epitope pattern.
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| 7. |
- Bergman, Marie-Louise, et al.
(författare)
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CTLA-4-/- mice display T cell-apoptosis resistance resembling that ascribed to autoimmune-prone non-obese diabetic (NOD) mice
- 2001
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Ingår i: Journal of Autoimmunity. - : Elsevier. - 0896-8411 .- 1095-9157. ; 16:2, s. 105-113
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Tidskriftsartikel (refereegranskat)abstract
- The genes conferring susceptibility to autoimmune (insulin-dependent) diabetes mellitus (IDDM) are, in most cases, not defined. Among the loci so far identified as associated with murine IDDM (Idd1-19), only the nature of Idd1 has been assessed. Here we show that thymocytes and peripheral lymphocytes of the non-obese diabetic (NOD) mouse are relatively resistant to apoptosis induced by gamma-irradiation. By linkage analysis of F2 progeny mice, we map this trait to a locus on chromosome 1 containing the Idd5 diabetes susceptibility region. By the use of congenic mice, we confirm the linkage data and map this locus to a 6 cM region on proximal chromosome 1. Ctla4, being localized in this chromosomal region and mediating crucial functions in T cell biology, is a logical candidate gene in the Idd5 susceptibility region. In line with this, we demonstrate that T cells from Ctla4(-/-)deficient mice show a similar resistance to gamma-irradiation-induced apoptosis as observed in the NOD mice. This reinforces the notion that CTLA-4 contributes to the pathogenesis of autoimmune diabetes.
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| 8. |
- Beyan, Huriya, et al.
(författare)
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Guthrie card methylomics identifies temporally stable epialleles that are present at birth in humans
- 2012
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1549-5469 .- 1088-9051. ; 22:11, s. 2138-2145
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Tidskriftsartikel (refereegranskat)abstract
- A major concern in common disease epigenomics is distinguishing causal from consequential epigenetic variation. One means of addressing this issue is to identify the temporal origins of epigenetic variants via longitudinal analyses. However, prospective birth-cohort studies are expensive and time consuming. Here, we report DNA methylomics of archived Guthrie cards for the retrospective longitudinal analyses of in-utero-derived DNA methylation variation. We first validate two methodologies for generating comprehensive DNA methylomes from Guthrie cards. Then, using an integrated epigenomic/genomic analysis of Guthrie cards and follow-up samplings, we identify interindividual DNA methylation variation that is present both at birth and 3 yr later. These findings suggest that disease-relevant epigenetic variation could be detected at birth, i.e., before overt clinical disease. Guthrie card methylomics offers a potentially powerful and cost-effective strategy for studying the dynamics of interindividual epigenomic variation in a range of common human diseases.
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| 9. |
- Bolmeson, Caroline, et al.
(författare)
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Differences in islet-enriched miRNAs in healthy and glucose intolerant human subjects.
- 2011
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 1090-2104 .- 0006-291X. ; Dec, s. 16-22
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Tidskriftsartikel (refereegranskat)abstract
- Many microRNAs (miRNAs) are known to be cell-type specific and are implicated in development of diseases. We investigated the global expression pattern of miRNAs in human pancreatic islets compared to liver and skeletal muscle, using bead-based technology and quantitative RT-PCR. In addition to the known islet-specific miR-375, we also found enrichment of miR-127-3p, miR-184, miR-195 and miR-493∗ in the pancreatic islets. The expression of miR-375, miR-127-3p, miR-184 and the liver-enriched miR-122 were positively correlated to insulin biosynthesis, while the expression of miR-127-3p and miR-184 were negatively correlated to glucose-stimulated insulin secretion (GSIS). These correlations were absent in islets of glucose intolerant donors (HbA1c⩾6.1). We suggest the presence of an islet-specific miRNA network, which consists of at least miR-375, miR-127-3p and miR-184, potentially involved in insulin secretion. Our results provide new insight into miRNA-mediated regulation of insulin secretion in healthy and glucose intolerant subjects.
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| 10. |
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