目的:通过体内外实验观察固本畅枢法对肥胖糖尿病大鼠糖脂代谢的影响及胰岛β细胞MIN6凋亡的干预作用。方法:体内实验:高脂高糖喂养+小剂量链脲佐菌素(streptozocin,STZ)一次性腹腔注射复制肥胖糖尿病大鼠模型,连续灌胃给药4周,观察大...目的:通过体内外实验观察固本畅枢法对肥胖糖尿病大鼠糖脂代谢的影响及胰岛β细胞MIN6凋亡的干预作用。方法:体内实验:高脂高糖喂养+小剂量链脲佐菌素(streptozocin,STZ)一次性腹腔注射复制肥胖糖尿病大鼠模型,连续灌胃给药4周,观察大鼠饮水、摄食、尿量及一般情况,检测模型大鼠的空腹血糖(fasting blood glucose,FBG)、空腹胰岛素(fasting insulin,FINS)、甘油三酯(triglyceride,TG)、总胆固醇(cholesterol,TC)、低密度脂蛋白胆固醇(low density lipoprotein,LDL-C)、高密度脂蛋、白胆固醇(high density lipoprotein,HDL-C)水平,观察胰腺、肝脏、肌肉、脂肪组织病理,采用TUNEL-DAPI共染色法检测各组大鼠胰腺β细胞凋亡情况,了解细胞凋亡形态学特征;体外实验:培养小鼠胰岛β细胞MIN6,采用流式细胞术定量分析MIN6细胞凋亡情况。结果:体内实验:固本畅枢法能减少糖尿病模型大鼠的饮水、摄食、尿量(P<0.01),差异有统计学意义;能降低FBG、FINS、TC、TG,升高HDL水平(P<0.05,P<0.01),差异有统计学意义;可修复胰腺、肝脏、肌肉、脂肪组织病理结构,使其形态更规则;能改善模型组大鼠胰腺β细胞凋亡时的细胞形态(P<0.01),差异有统计学意义);体外实验:固本畅枢法能减少模型组MIN6细胞的凋亡细胞数量。结论:固本畅枢法改善糖脂代谢紊乱,其作用机制与修复组织病理、抑制胰岛β细胞凋亡有关。展开更多
This study assessed the effects of leukemia-related protein 16 (LRP16) on the regulation of pancreatic functions in mouse insulinoma (MIN6) cells. Cells with down-regulated expression of LRP16 were obtained by a shRNA...This study assessed the effects of leukemia-related protein 16 (LRP16) on the regulation of pancreatic functions in mouse insulinoma (MIN6) cells. Cells with down-regulated expression of LRP16 were obtained by a shRNA interference strategy. Insulin content and glucose-stimulated insulin secretion (GSIS) were examined by radioimmunoassay. Western blotting was applied to detect protein expression. Glucose-stimulated sub-cellular localization of PDX-1 was immunocytochemically determined. Cell proliferation and apoptosis were detected by flow cytometry. Our results showed that LRP16 regulated insulin content in MIN6 cells by controlling expression of insulin and insulin transcription factors. LRP16 gene silence in MIN6 cells led to reduced cell proliferation and increased apoptosis. The observation of phosphorylation of serine-473 Akt and the localization of PDX-1 to the nucleus under glucose-stimulation exhibited that LRP16 was a component mediating Akt signaling in MIN6 cells. These results suggest that LRP16 plays a key role in maintaining pancreatic β-cell functions and may help us to understand the protective effects of estrogen on the functions of pancreatic β-cells.展开更多
Background Pancreatic β cells are susceptible to fatty acid-induced apoptosis. The 17β-estradiol （E2） protects pancreatic βcells from apoptosis, mediated by the estrogen receptor-a （ERa）. The mRNA level and pro...Background Pancreatic β cells are susceptible to fatty acid-induced apoptosis. The 17β-estradiol （E2） protects pancreatic βcells from apoptosis, mediated by the estrogen receptor-a （ERa）. The mRNA level and promoter activity of leukemia-related protein （LRP） 16 were significantly increased by E2 in E R-a and LRP 16 was a co-activator of ER-a. The aim of the study was to assess the effects of LRP16 on fatty acid-induced apoptosis in MIN6 cells. Methods Cells with over-expressing LRP16 were obtained by lipidosome transfection. Insulin content and glucose-stimulated insulin secretion （GSIS） were examined by radioimmunoassay. Western blotting was applied to detect protein expression. Apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） and flow cytometry. The forkhead boxO1 （FoxO1） subcellular localization was determined by immunocytochemical analysis. Results MIN6-LRP16 cells with overexpression of LRP16 were successfully established, and protein expression of LRP16 was 2.29-fold of that of control cells （MIN6-3.1, P 〈0.05）. Insulin content and GSIS in MIN6-LRP16 were substantially increased compared with those in control cells. When cells were stimulated with glucose, increased phosphorylation of extracellular signal-regulated kinase （ERK） 1/2 and serine-threonine kinase （Akt） were observed in MIN6-LRP16. When cells were under palmitate pressure, the TUNEL-positive rate in MIN6-LRP16 was （17.0±0.5）%, while it in MIN6-3.1 was （22.0±0.4）%. In palmitate-treated cells, attenuated Akt phosphorylation was observed, but the attenuation in Akt activity was partially restored in MIN6-LRP16 cells. Meanwhile, nuclear localization of FoxO1 in MIN6-LRP16 was apparently reduced compared with that in control cells. Conclusions LRP16 regulated insulin content and GSIS in MIN6 cells by ERK1/2 and Akt activated way. Meanwhile, LRP16 overexpression protected MIN6 cells from fatty acid-induced apoptosis by partially restoring Akt phosphorylati展开更多
Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", c...Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", commonly known as ‘Diabetes mellitus’ in clinics.Previous studies have shown that SGY-P could reduce insulin resistance and repair damaged pancreas in db/db mice, but the underlying mechanisms were unclear. Here, we investigated whether treatment with SGY-P could protect pancreatic β-cells from apoptosis and uncovered the underlying mechanisms. db/db mice were used to observe the hypoglycemic and islet protective effect in vivo. Apoptosis was induced in mouse insulinoma 6(MIN6) cells by palmitate, following which the cells were treated with SGY-P for elucidating the anti-apoptotic mechanism in vitro. Cell viability and nuclear morphology were detected by CCK-8 assay and Hoechst 33258 staining. The expression levels of apoptosis-, endoplasmic reticulum(ER) stress-, and autophagy-related proteins were measured by western blot. The results showed that SGY-P reduced fasting blood glucose, pancreatic pathological changes, and islet β-cell apoptosis in db/db mice. Palmitate-induced apoptosis in MIN6 cells was decreased by SGY-P treatment. Hence, SGY-P therapy exhibited a protective effect on pancreatic β-cells by decreasing the expression of cleaved caspase-3, cleaved PARP and Bax, and increasing Bcl-2 by suppressing ER stress(Bip/XBP1/IRE1α/CHOP/Caspase-12) and autophagy(LC3/p62/Atg5) pathways.2/Atg5 pathways.展开更多
The sweet taste receptors comprised of T1r2 and T1r3, sense glucose concentrations in the gastrointestine. While hyperglycemia was reported to decrease the T1R2 and T1R3 tanscript levels in healthy subjects, no change...The sweet taste receptors comprised of T1r2 and T1r3, sense glucose concentrations in the gastrointestine. While hyperglycemia was reported to decrease the T1R2 and T1R3 tanscript levels in healthy subjects, no change was observed in type 2 diabetes patients. We investigated which glucose level and nutrients affect those transcript levels in MIN 6 and primary cultured taste buds cells using quantitative Reverse Trancription Polymerase Chain Reaction. High glucose diminished T1r2 transcript levels in MIN 6 and primary cultured taste buds cells. Resveratrol and its analogue augmented transcript levels of T1r1 and T1r2 above normal levels in MIN 6 cells in the medium with 25 mM glucose. Adenine, but not guanine, augmented T1r2 transcript levels of MIN 6 cells in the medium with 25 mM glucose. These results imply that nutrients in meals could affect sweet taste sensitivity by modulating T1r2 transcript levels in response to blood glucose levels.展开更多
BACKGROUND Type 2 diabetes(T2 D) is characterized by insufficient insulin secretion caused by defective pancreatic β-cell function or insulin resistance,resulting in an increase in blood glucose.However,the mechanism...BACKGROUND Type 2 diabetes(T2 D) is characterized by insufficient insulin secretion caused by defective pancreatic β-cell function or insulin resistance,resulting in an increase in blood glucose.However,the mechanism involved in this lack of insulin secretion is unclear.The level of vascular endothelial growth factor B(VEGF-B) is significantly increased in T2 D patients.The inactivation of VEGF-B could restore insulin sensitivity in db/db mice by reducing fatty acid accumulation.It is speculated that VEGF-B is related to pancreatic β-cell dysfunction and is an important factor affecting β-cell secretion of insulin.As an in vitro model of normal pancreatic β-cells,the MIN6 cell line can be used to analyze the mechanism of insulin secretion and related biological effects.AIM To study the role of VEGF-B in the insulin secretion signaling pathway in MIN6 cells and explore the effect of VEGF-B on blood glucose regulation.METHODS The MIN6 mouse pancreatic islet β-cell line was used as the model system.By administering exogenous VEGF-B protein or knocking down VEGF-B expression in MIN6 cells,we examined the effects of VEGF-B on insulin secretion,Ca2+ and cyclic adenosine monophosphate(cAMP) levels,and the insulin secretion signaling pathway.RESULTS Exogenous VEGF-B inhibited the secretion of insulin and simultaneously reduced the levels of Ca2+ and cAMP in MIN6 cells.Exogenous VEGF-B also reduced the expression of phospholipase C gamma 1(PLCγ1),phosphatidylinositol 3-kinase(PI3 K),serine/threonine kinase(AKT),and other proteins in the insulin secretion pathway.Upon knockdown of VEGF-B,MIN6 cells exhibited increased insulin secretion and Ca2+ and cAMP levels and upregulated expression of PLCγ1,PI3 K,AKT,and other proteins.CONCLUSION VEGF-B can regulate insulin secretion by modulating the levels of Ca2+ and cAMP.VEGF-B involvement in insulin secretion is related to the expression of PLCγ1,PI3 K,AKT,and other signaling proteins.These results provide theoretical support and an experimental basis for the study of VEG展开更多
文摘本研究主要探讨D-柠檬烯对衣霉素(Tm)诱导胰腺MIN6细胞损伤的保护作用。设置溶剂对照组、Tm组、4-苯基丁酸(PBA)组和不同浓度的D-柠檬烯组,用Griess试剂检测Tm刺激胰岛β细胞产生的NO水平;Annexin V-FITC/PI双染法流式检测细胞凋亡; RT-PCR法检测mRNA的表达水平; Western Blot法检测相关蛋白的表达。结果显示,D-柠檬烯组与Tm组相比,NO产生量减少、细胞凋亡率降低、内质网应激相关基因sXbp1和CHOP的mRNA表达量显著降低、p-eIF2α的蛋白表达量显著下降。结果表明D-柠檬烯对衣霉素(Tm)诱导的胰腺MIN6细胞损伤具有显著的保护作用。
文摘目的:通过体内外实验观察固本畅枢法对肥胖糖尿病大鼠糖脂代谢的影响及胰岛β细胞MIN6凋亡的干预作用。方法:体内实验:高脂高糖喂养+小剂量链脲佐菌素(streptozocin,STZ)一次性腹腔注射复制肥胖糖尿病大鼠模型,连续灌胃给药4周,观察大鼠饮水、摄食、尿量及一般情况,检测模型大鼠的空腹血糖(fasting blood glucose,FBG)、空腹胰岛素(fasting insulin,FINS)、甘油三酯(triglyceride,TG)、总胆固醇(cholesterol,TC)、低密度脂蛋白胆固醇(low density lipoprotein,LDL-C)、高密度脂蛋、白胆固醇(high density lipoprotein,HDL-C)水平,观察胰腺、肝脏、肌肉、脂肪组织病理,采用TUNEL-DAPI共染色法检测各组大鼠胰腺β细胞凋亡情况,了解细胞凋亡形态学特征;体外实验:培养小鼠胰岛β细胞MIN6,采用流式细胞术定量分析MIN6细胞凋亡情况。结果:体内实验:固本畅枢法能减少糖尿病模型大鼠的饮水、摄食、尿量(P<0.01),差异有统计学意义;能降低FBG、FINS、TC、TG,升高HDL水平(P<0.05,P<0.01),差异有统计学意义;可修复胰腺、肝脏、肌肉、脂肪组织病理结构,使其形态更规则;能改善模型组大鼠胰腺β细胞凋亡时的细胞形态(P<0.01),差异有统计学意义);体外实验:固本畅枢法能减少模型组MIN6细胞的凋亡细胞数量。结论:固本畅枢法改善糖脂代谢紊乱,其作用机制与修复组织病理、抑制胰岛β细胞凋亡有关。
基金supported by grants from the National Na-ture Science Foundation of China(No.30771042and30971127)Major National Basic Research Develop-ment Program of China(Program973)(2006CB503903)
文摘This study assessed the effects of leukemia-related protein 16 (LRP16) on the regulation of pancreatic functions in mouse insulinoma (MIN6) cells. Cells with down-regulated expression of LRP16 were obtained by a shRNA interference strategy. Insulin content and glucose-stimulated insulin secretion (GSIS) were examined by radioimmunoassay. Western blotting was applied to detect protein expression. Glucose-stimulated sub-cellular localization of PDX-1 was immunocytochemically determined. Cell proliferation and apoptosis were detected by flow cytometry. Our results showed that LRP16 regulated insulin content in MIN6 cells by controlling expression of insulin and insulin transcription factors. LRP16 gene silence in MIN6 cells led to reduced cell proliferation and increased apoptosis. The observation of phosphorylation of serine-473 Akt and the localization of PDX-1 to the nucleus under glucose-stimulation exhibited that LRP16 was a component mediating Akt signaling in MIN6 cells. These results suggest that LRP16 plays a key role in maintaining pancreatic β-cell functions and may help us to understand the protective effects of estrogen on the functions of pancreatic β-cells.
基金This work was supported by a grant from the National Natural Science Foundation of China （No. 30771042）.
文摘Background Pancreatic β cells are susceptible to fatty acid-induced apoptosis. The 17β-estradiol （E2） protects pancreatic βcells from apoptosis, mediated by the estrogen receptor-a （ERa）. The mRNA level and promoter activity of leukemia-related protein （LRP） 16 were significantly increased by E2 in E R-a and LRP 16 was a co-activator of ER-a. The aim of the study was to assess the effects of LRP16 on fatty acid-induced apoptosis in MIN6 cells. Methods Cells with over-expressing LRP16 were obtained by lipidosome transfection. Insulin content and glucose-stimulated insulin secretion （GSIS） were examined by radioimmunoassay. Western blotting was applied to detect protein expression. Apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling （TUNEL） and flow cytometry. The forkhead boxO1 （FoxO1） subcellular localization was determined by immunocytochemical analysis. Results MIN6-LRP16 cells with overexpression of LRP16 were successfully established, and protein expression of LRP16 was 2.29-fold of that of control cells （MIN6-3.1, P 〈0.05）. Insulin content and GSIS in MIN6-LRP16 were substantially increased compared with those in control cells. When cells were stimulated with glucose, increased phosphorylation of extracellular signal-regulated kinase （ERK） 1/2 and serine-threonine kinase （Akt） were observed in MIN6-LRP16. When cells were under palmitate pressure, the TUNEL-positive rate in MIN6-LRP16 was （17.0±0.5）%, while it in MIN6-3.1 was （22.0±0.4）%. In palmitate-treated cells, attenuated Akt phosphorylation was observed, but the attenuation in Akt activity was partially restored in MIN6-LRP16 cells. Meanwhile, nuclear localization of FoxO1 in MIN6-LRP16 was apparently reduced compared with that in control cells. Conclusions LRP16 regulated insulin content and GSIS in MIN6 cells by ERK1/2 and Akt activated way. Meanwhile, LRP16 overexpression protected MIN6 cells from fatty acid-induced apoptosis by partially restoring Akt phosphorylati
基金Chinese Medicine Research Program of Hunan Province(No.201999)。
文摘Sangguayin preparation(SGY-P) is refined from the traditional Chinese medicinal compound Sangguayin, which"clears heat and promotes fluid" and "tonifies kidney and spleen" for "Xiaoke", commonly known as ‘Diabetes mellitus’ in clinics.Previous studies have shown that SGY-P could reduce insulin resistance and repair damaged pancreas in db/db mice, but the underlying mechanisms were unclear. Here, we investigated whether treatment with SGY-P could protect pancreatic β-cells from apoptosis and uncovered the underlying mechanisms. db/db mice were used to observe the hypoglycemic and islet protective effect in vivo. Apoptosis was induced in mouse insulinoma 6(MIN6) cells by palmitate, following which the cells were treated with SGY-P for elucidating the anti-apoptotic mechanism in vitro. Cell viability and nuclear morphology were detected by CCK-8 assay and Hoechst 33258 staining. The expression levels of apoptosis-, endoplasmic reticulum(ER) stress-, and autophagy-related proteins were measured by western blot. The results showed that SGY-P reduced fasting blood glucose, pancreatic pathological changes, and islet β-cell apoptosis in db/db mice. Palmitate-induced apoptosis in MIN6 cells was decreased by SGY-P treatment. Hence, SGY-P therapy exhibited a protective effect on pancreatic β-cells by decreasing the expression of cleaved caspase-3, cleaved PARP and Bax, and increasing Bcl-2 by suppressing ER stress(Bip/XBP1/IRE1α/CHOP/Caspase-12) and autophagy(LC3/p62/Atg5) pathways.2/Atg5 pathways.
文摘The sweet taste receptors comprised of T1r2 and T1r3, sense glucose concentrations in the gastrointestine. While hyperglycemia was reported to decrease the T1R2 and T1R3 tanscript levels in healthy subjects, no change was observed in type 2 diabetes patients. We investigated which glucose level and nutrients affect those transcript levels in MIN 6 and primary cultured taste buds cells using quantitative Reverse Trancription Polymerase Chain Reaction. High glucose diminished T1r2 transcript levels in MIN 6 and primary cultured taste buds cells. Resveratrol and its analogue augmented transcript levels of T1r1 and T1r2 above normal levels in MIN 6 cells in the medium with 25 mM glucose. Adenine, but not guanine, augmented T1r2 transcript levels of MIN 6 cells in the medium with 25 mM glucose. These results imply that nutrients in meals could affect sweet taste sensitivity by modulating T1r2 transcript levels in response to blood glucose levels.
基金Supported by National Natural Science Foundation of China,No.31771284National Natural Science Foundation of China Youth Project,No.31702024+1 种基金Major Basic Research Project of Shandong Provincial Natural Science Foundation,No.ZR2019ZD27Shandong Province Higher Educational Science and Technology Plan Project,No.J17KA258。
文摘BACKGROUND Type 2 diabetes(T2 D) is characterized by insufficient insulin secretion caused by defective pancreatic β-cell function or insulin resistance,resulting in an increase in blood glucose.However,the mechanism involved in this lack of insulin secretion is unclear.The level of vascular endothelial growth factor B(VEGF-B) is significantly increased in T2 D patients.The inactivation of VEGF-B could restore insulin sensitivity in db/db mice by reducing fatty acid accumulation.It is speculated that VEGF-B is related to pancreatic β-cell dysfunction and is an important factor affecting β-cell secretion of insulin.As an in vitro model of normal pancreatic β-cells,the MIN6 cell line can be used to analyze the mechanism of insulin secretion and related biological effects.AIM To study the role of VEGF-B in the insulin secretion signaling pathway in MIN6 cells and explore the effect of VEGF-B on blood glucose regulation.METHODS The MIN6 mouse pancreatic islet β-cell line was used as the model system.By administering exogenous VEGF-B protein or knocking down VEGF-B expression in MIN6 cells,we examined the effects of VEGF-B on insulin secretion,Ca2+ and cyclic adenosine monophosphate(cAMP) levels,and the insulin secretion signaling pathway.RESULTS Exogenous VEGF-B inhibited the secretion of insulin and simultaneously reduced the levels of Ca2+ and cAMP in MIN6 cells.Exogenous VEGF-B also reduced the expression of phospholipase C gamma 1(PLCγ1),phosphatidylinositol 3-kinase(PI3 K),serine/threonine kinase(AKT),and other proteins in the insulin secretion pathway.Upon knockdown of VEGF-B,MIN6 cells exhibited increased insulin secretion and Ca2+ and cAMP levels and upregulated expression of PLCγ1,PI3 K,AKT,and other proteins.CONCLUSION VEGF-B can regulate insulin secretion by modulating the levels of Ca2+ and cAMP.VEGF-B involvement in insulin secretion is related to the expression of PLCγ1,PI3 K,AKT,and other signaling proteins.These results provide theoretical support and an experimental basis for the study of VEG
