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      Gene Expression Profiles of Beta-Cell Enriched Tissue Obtained by Laser Capture Microdissection from Subjects with Type 2 Diabetes

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          Abstract

          Background

          Changes in gene expression in pancreatic beta-cells from type 2 diabetes (T2D) should provide insights into their abnormal insulin secretion and turnover.

          Methodology/Principal Findings

          Frozen sections were obtained from cadaver pancreases of 10 control and 10 T2D human subjects. Beta-cell enriched samples were obtained by laser capture microdissection (LCM). RNA was extracted, amplified and subjected to microarray analysis. Further analysis was performed with DNA-Chip Analyzer (dChip) and Gene Set Enrichment Analysis (GSEA) software. There were changes in expression of genes linked to glucotoxicity. Evidence of oxidative stress was provided by upregulation of several metallothionein genes. There were few changes in the major genes associated with cell cycle, apoptosis or endoplasmic reticulum stress. There was differential expression of genes associated with pancreatic regeneration, most notably upregulation of members of the regenerating islet gene (REG) family and metalloproteinase 7 (MMP7). Some of the genes found in GWAS studies to be related to T2D were also found to be differentially expressed. IGF2BP2, TSPAN8, and HNF1B (TCF2) were upregulated while JAZF1 and SLC30A8 were downregulated.

          Conclusions/Significance

          This study made possible by LCM has identified many novel changes in gene expression that enhance understanding of the pathogenesis of T2D.

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          Most cited references67

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          Model-based analysis of oligonucleotide arrays: expression index computation and outlier detection.

          Recent advances in cDNA and oligonucleotide DNA arrays have made it possible to measure the abundance of mRNA transcripts for many genes simultaneously. The analysis of such experiments is nontrivial because of large data size and many levels of variation introduced at different stages of the experiments. The analysis is further complicated by the large differences that may exist among different probes used to interrogate the same gene. However, an attractive feature of high-density oligonucleotide arrays such as those produced by photolithography and inkjet technology is the standardization of chip manufacturing and hybridization process. As a result, probe-specific biases, although significant, are highly reproducible and predictable, and their adverse effect can be reduced by proper modeling and analysis methods. Here, we propose a statistical model for the probe-level data, and develop model-based estimates for gene expression indexes. We also present model-based methods for identifying and handling cross-hybridizing probes and contaminating array regions. Applications of these results will be presented elsewhere.
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            Function and biological roles of the Dickkopf family of Wnt modulators.

            C Niehrs (2006)
            Dickkopf (Dkk) genes comprise an evolutionary conserved small gene family of four members (Dkk1-4) and a unique Dkk3-related gene, Dkkl1 (soggy). They encode secreted proteins that typically antagonize Wnt/beta-catenin signaling, by inhibiting the Wnt coreceptors Lrp5 and 6. Additionally, Dkks are high affinity ligands for the transmembrane proteins Kremen1 and 2, which also modulate Wnt signaling. Dkks play an important role in vertebrate development, where they locally inhibit Wnt regulated processes such as antero-posterior axial patterning, limb development, somitogenesis and eye formation. In the adult, Dkks are implicated in bone formation and bone disease, cancer and Alzheimer's disease.
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              Five stages of evolving beta-cell dysfunction during progression to diabetes.

              This article proposes five stages in the progression of diabetes, each of which is characterized by different changes in beta-cell mass, phenotype, and function. Stage 1 is compensation: insulin secretion increases to maintain normoglycemia in the face of insulin resistance and/or decreasing beta-cell mass. This stage is characterized by maintenance of differentiated function with intact acute glucose-stimulated insulin secretion (GSIS). Stage 2 occurs when glucose levels start to rise, reaching approximately 5.0-6.5 mmol/l; this is a stable state of beta-cell adaptation with loss of beta-cell mass and disruption of function as evidenced by diminished GSIS and beta-cell dedifferentiation. Stage 3 is a transient unstable period of early decompensation in which glucose levels rise relatively rapidly to the frank diabetes of stage 4, which is characterized as stable decompensation with more severe beta-cell dedifferentiation. Finally, stage 5 is characterized by severe decompensation representing a profound reduction in beta-cell mass with progression to ketosis. Movement across stages 1-4 can be in either direction. For example, individuals with treated type 2 diabetes can move from stage 4 to stage 1 or stage 2. For type 1 diabetes, as remission develops, progression from stage 4 to stage 2 is typically found. Delineation of these stages provides insight into the pathophysiology of both progression and remission of diabetes.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2010
                13 July 2010
                : 5
                : 7
                : e11499
                Affiliations
                [1 ]Section on Islet Transplantation and Cell Biology, Research Division, Joslin Diabetes Center and the Department of Medicine, Harvard Medical School, Boston, Massachusetts, United States of America
                [2 ]Molecular Pathology Unit, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America
                [3 ]Section of Endocrinology and Metabolism of Organ Transplantation, Department of Endocrinology and Metabolism, University of Pisa, Pisa, Italy
                Uppsala University, Sweden
                Author notes

                Conceived and designed the experiments: LM SBW PM GCW. Performed the experiments: LM JT SD DCS. Analyzed the data: LM JT AS SBW GCW. Contributed reagents/materials/analysis tools: PM. Wrote the paper: LM GCW.

                Article
                10-PONE-RA-17289R1
                10.1371/journal.pone.0011499
                2903480
                20644627
                0a0dd125-8f6e-4e12-bf36-cdcb2daed140
                Marselli et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                History
                : 10 March 2010
                : 6 June 2010
                Page count
                Pages: 13
                Categories
                Research Article
                Molecular Biology
                Cell Biology/Gene Expression
                Diabetes and Endocrinology/Type 2 Diabetes

                Uncategorized
                Uncategorized

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