Kazuya Yamagata

T130I mutation in HNF-4α gene is a loss-of-function mutation in hepatocytes and is associated with late-onset Type 2 diabetes mellitus in Japanese subjects

Aims/hypothesisMutations in hepatocyte nuclear factor (HNF)-4α gene cause a form of maturity-onset diabetes of the young (MODY1). The T130I mutation is a rare missense mutation, which affects a conserved amino acid in a DNA binding domain. This mutation can be found in the general population, so this variant alone does not cause MODY. However, its significance in the development of late-onset Type 2 diabetes is not known.MethodsWe screened 423 unrelated Japanese patients with late-onset Type 2 diabetes and 354 unrelated non-diabetic control subjects for the T130I mutation in the HNF-4α gene. The transactivation ability of T130I-HNF-4α was assessed using reporter gene assay.ResultsThe frequency of the T130I mutation was higher in Type 2 diabetic patients (p=0.015, odds ratio 4.3, 95%CI 1.24–14.98) than control subjects. The serum HDL-cholesterol concentration was lower in Type 2 diabetic patients with the T130I mutation compared with those without this mutation (p=0.006). Reporter gene analysis showed that T130I-HNF-4α transcriptional activity was not impaired compared with wild-type HNF-4α in Hela and MIN6 cells, but it was reduced in HepG2 and primary cultured mouse hepatocytes (27–78% of wild type, p<0.05).Conclusion/interpretationOur findings suggest that T130I-HNF-4α is a loss-of-function mutation in hepatocytes and that this mutation is associated with late-onset Type 2 diabetes in Japanese subjects. The T130I mutation in the HNF-4α gene might be involved in the development of Type 2 diabetes in the Japanese population.

Analysis of expression profiles of islet-associated transcription and growth factors during β-cell neogenesis from duct cells in partially duct-ligated mice

Introduction &bgr;-cell neogenesis from pancreatic duct cells has been reported to occur in duct-ligated rat. Nevertheless, detailed process of this phenomenon has not been clarified. Aims and Methodology To clarify the mechanism of &bgr;-cell neogenesis, a partial pancreatic duct ligation mouse model was created. Proliferation of duct cells, &bgr;-cell neogenesis, and expression of transcription factors and differentiation/growth factors were studied by immunohistochemistry, cDNA array, and RT-PCR methods. Results In the duct-ligated portion of the pancreas, newly formed islet-like cell clusters (ICCs) were observed arising from the ducts on day 7 and afterward. Transcription factors, such as pancreatic and duodenal homeobox gene-1 (PDX-1), paired box factor 6 (Pax6), islet1 and Nkx2.2-positive cells, and protein gene product 9.5 (PGP9.5) were also induced in duct lining cells. By cDNA microarray analysis, expression of insulin-like growth factor-1 (IGF-1) and transforming growth factor &bgr;1 (TGF-&bgr;1) were above control levels on day 5, and RT-PCR showed an increase from day 5 to day 28. IGF-1 and activin A–positive cells were detected in ducts. In addition, expression of betacellulin (BTC), heparin-binding epidermal growth factor–like growth factor (HB-EGF), and TGF-&agr; were also increased from day 3 or 5. Conclusion These findings suggest that &bgr;-cell or endocrine precursors are localized among duct lining cells. Induction of several islet cell–associated transcription factors and differentiation and/or growth factors may play important roles during &bgr;-cell neogenesis in this model.

Expression profile of MODY3/HNF-1α protein in the developing mouse pancreas

HeadingAbstractn Aims/hypothesis. One subtype of MODY (MODY3) results from the heterozygous mutation of a hepatocyte nuclear factor (HNF)-1α. The pattern of HNF-1α expression in the normal pancreas has not been determined. This study aimed to clarify the profile of HNF-1α protein expression in the developing mouse pancreas.n Methods. Double immunofluorescence staining was carried out for HNF-1α and pancreatic hormones or transcription factors (PDX-1, Pax6, Isl1, and Nkx2.2). The expression of these transcription factors was also studied in the beta cells of HNF-1α mutant mice.n Results. HNF-1α was expressed by both endocrine and exocrine cells of the pancreas. Double immunofluorescence staining showed that HNF-1α was expressed in the nuclei of alpha cells, beta cells, delta cells, and pancreatic polypeptide (PP) cells. HNF-1α was first detected in most pancreatic epithelial cells on embryonic day 10.5 (E10.5), and hormone-positive endocrine cells and amylase-positive cells expressed HNF-1α on E15.5. Most of the Pax6-, Isl1-, or PDX-1-positive cells showed co-expression of HNF-1α. However, HNF-1α immunoreactivity was not observed in 36.0% of Nkx2.2-positive cells. Expression of Nkx2.2, Isl1 and Pax6 seemed to be normal in the beta cells of transgenic mice with dominant negative overexpression of HNF-1α. Expression of PDX-1 did not change in the beta cells of pre-diabetic HNF-1α (–/–) mice, but expression was markedly decreased in the diabetic stage.n Conclusion/interpretation. HNF-1α is expressed by both endocrine cells and exocrine cells of the pancreas from the foetal stage along with other transcription factors, so HNF-1α might play a role during development.