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The Effect of NaAsO2 on Insulin Synthesis and Secretion of Pancreatic Islet β-cells

Author: WangRui
Tutor: MuXiaoLing
School: Shihezi University
Course: Human Anatomy and Embryology
Keywords: Sodium arsenite NIT-1 cells Insulin Apoptosis Reactive oxygen species
CLC: R587.1
Type: Master's thesis
Year: 2010
Downloads: 16
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Abstract


Objective: To observe the sodium arsenite (NaAsO2) the synthesis and secretion of pancreatic beta cells NIT-1 insulin (Insulin). And to further explore its possible mechanism. Methods: Cultured NIT-1 cells with different concentrations of NaAsO2 (0,1,2,4,8,16 and 32μmol / L) for different times (24h and 48h), using the MTT cell proliferation activity. According to the results of MTT select the appropriate concentration NaAsO2 (0, 1, 4, and 8μmol / L) effector cells at different times (24h and 48h), measured by enzyme-linked immunosorbent assay (ELISA) with different concentrations of glucose (of 5.5,16.5 mmol / L) stimulated Insulin secretion; RT-PCR method to detect the mRNA expression of Insulin; Annexin V / PI double staining flow cytometry apoptosis of cells in each group; confocal laser the microscope fluorescence staining cells reactive oxygen species (ROS) changes . The role of the NaAsO2 (0,4,8 μmol / L) for 48h, fluorescence immunohistochemistry to detect cell islets - duodenal homeobox domain protein (PDX-1) translocation; Western-blot detection cell p -JNK, PDX-1 expression. Results: (1) NaAsO2 NIT-1 cell proliferation activity: when NaAsO2 concentration is greater than 4 micromol / L inhibited cell proliferation, inhibition of cell proliferation with NaAsO2 concentration increase and extension of time to increase. (2) NaAsO2 (1,4, and 8μmol / L) NIT-1 cells Insulin secretion and mRNA expression: role 24h 8μmol / L of different concentrations of glucose-stimulated secretion of Insulin and Insulin mRNA expression were reduced compared with the control group difference was statistically significant; role 48h micromol / L group and 8μmol / L different concentrations of glucose-stimulated secretion of Insulin and Insulin mRNA expression were reduced compared with control group difference was statistically significant, but 8μmol / L group Insulin secretion stimulated by high glucose (16.5 mmol / L) and base (5.5 mmol / L) Insulin secretion is no longer a significant difference. (3) NaAsO2 NIT-1 cell apoptosis: of NaAsO2 apoptosis in a dose-and time-dependent, the total rate of apoptosis with the increase in the concentration of NaAsO2 and the extension of time increased. When the NaAsO2 concentration greater than 4 micromol / L promote apoptosis was statistically significant (p lt; 0.05) (4) NaAsO2 ROS levels in NIT-1 cells: In addition to the control group in each group of ROS expression, intracellular ROS levels increase and the extension of time with NaAsO2 concentration increased. The (5) NaAsO2 the NIT-1 cells PDX-1 translocation: NaAsO2 inhibition of PDX-1 nuclear localization signal, caused by PDX-1 translocation to the cytoplasm. (6) NaAsO2 p-JNK NIT-1 cells, PDX-1 protein expression: p-JNK protein expression increased PDX-1 protein decreased, compared with control group differences were statistically significant. Conclusion: NaAsO2 can reduce Insulin biosynthesis and secretion by pancreatic beta cells. Potential mechanism may be: (1) NaAsO2 can be generated through the induction of oxidative stress caused by intracellular reactive oxygen species (ROS) levels, and then through the activation of the JNK pathway induces apoptosis in pancreatic beta cells. (2) NaAsO2 reduced PDX-1 protein expression and activation of the JNK pathways lead to inhibition of PDX-1 nuclear localization Insulin biosynthesis and secretion dysfunction. Arsenic NIT-1 cells secretion of Insulin and Insulin gene expression and the role of time, dose-related Insulin synthesis and secretion disorders may be arsenic affect islet cell function and thus lead to one of the mechanisms of diabetes.

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CLC: > Medicine, health > Internal Medicine > Endocrine diseases and metabolic diseases > Islet disease > Diabetes
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