par-4与2型糖尿病胰岛功能障碍关系及其机制研究

发布时间：2019-04-29 10:16

【摘要】：目的:通过检测2型糖尿病患者血清par-4(Prostate Apoptosis Response-4,前列腺凋亡反应因子-4)浓度及干扰高糖高脂模拟糖尿病环境中胰岛细胞par-4的表达,观察并初步探讨Par-4对2型糖尿病胰岛β细胞凋亡和分泌功能的影响及其机制。方法:收集2型糖尿病患者组(T2DM组,n=30)及健康对照者组(N组,n=30)的临床基本资料;采集样本空腹静脉血液,采用ELISA法检测血清par-4浓度。培养NOD小鼠胰岛β细胞株NIT-1细胞,随机分为正常对照组、Par-4干扰组、高糖高脂组、高糖高脂+Par-4干扰组,采用ELISA(n=6)法检测各组par-4分泌浓度和葡萄糖刺激后的胰岛素分泌浓度;TUNEL法(n=8)检测各组细胞凋亡率;MTT法(n=10)检测各组细胞增殖能力;Western blot(n=3)检测各组细胞par-4和GRP78(Glucose Regulated Protein 78,葡萄糖糖调节蛋白78)表达水平。结果:2型糖尿病组和健康对照组两组患者临床基本资料无明显差异,2型糖尿病组患者血浆FBG(fasting blood-glucose空腹血糖)、HbA1c(Hemoglobin A1C糖化血红蛋白)和TG(Triglyceride,甘油三酯,3.73±3.44mmol/L)明显高于正常对照组,HDL(high density lipoprotein高密度脂蛋白,1.23±0.40mmol/L)、CP(C-peptide,C肽,1.29±0.41 nmol/L)及HOMA-β指数(48.9±27.08%)低于正常对照组,差异均有统计学意义(P0.05),而两者间胰岛素水平无明显统计学差异(P0.05),T2DM组患者血清par-4水平(17.44±3.65ng/ml)明显高于N组(14.08±2.94 ng/ml),差异有统计学意义(P0.05);相关分析显示在2型糖尿病组,血清par-4分泌水平与HOMA-β指数(r=-0.455,p=0.012)和C肽(r=-0.393,P=0.032)之间存在负相关关系,HOMA-β指数和C肽之间存在正相关关系(r=0.497,P=0.005)。与正常对照组(C组,凋亡率:3.14±1.08%)比较,par-4干扰组(I组)胰岛β细胞凋亡率(4.32±0.79%)、par-4浓度(67.6±16.32 ng/ml)、胰岛素分泌功能(6.65±0.36 m IU/L)和细胞增殖功能(0.70±0.14)均无明显差异(P0.05);与C组比较,高糖高脂组(D组)胰岛β细胞凋亡率(33.82±3.15%)、par-4分泌浓度(388.18±18.43 ng/ml)、par-4和GRP78表达明显增加,胰岛素分泌浓度(3.59±0.20 m IU/L)和胰岛细胞增殖功能(0.36±0.10)明显减少,,差异均有统计学意义(P0.05),高糖高脂+par-4干扰组(T组)胰岛素分泌功能(4.48±0.25 mIU/L)、细胞增殖功能(0.50±0.17)降低,par-4分泌浓度(164.58±16.62 ng/ml)升高,差异均有统计学意义(P0.05);与D组比较,T组细胞凋亡率(18.34±2.11%)、par-4分泌浓度、par-4和GRP78蛋白表达下降,胰岛素分泌浓度和细胞增殖功能升高,差异均有统计学意义(P0.05)。结论:2型糖尿病患者较之正常组胰岛细胞功能降低,血清par-4浓度明显升高且与HOMA-β指数负相关,干扰par-4表达可以但不能完全改善高糖高脂诱导的胰岛β细胞凋亡和功能障碍,其机制可能与降低了β细胞内质网应激(endoplasmic reticulum stress,ERS)水平有关。
[Abstract]:Objective: to detect the concentration of prostate apoptosis response factor-4 (par-4 (Prostate Apoptosis Response-4,-4) in serum of patients with type 2 diabetes mellitus and to interfere with the expression of islet cell par-4 in diabetic environment with high glucose and high fat. To observe and explore the effect of Par-4 on the apoptosis and secretion of islet 尾 cells in type 2 diabetes mellitus and its mechanism. Methods: the clinical data of patients with type 2 diabetes mellitus (T2DM group, n = 30) and healthy controls (group N, n = 30) were collected, fasting venous blood samples were collected and serum par-4 concentration was measured by ELISA method. NOD mice islet 尾 cell line NIT-1 cells were cultured and randomly divided into normal control group, Par-4 interference group, high glucose and high fat Par-4 interference group. The par-4 secretion concentration and insulin secretion concentration after glucose stimulation were measured by ELISA (n = 6) method. The apoptosis rate was measured by TUNEL assay, the proliferation ability of the cells was measured by MTT assay, the expression levels of par-4 and GRP78 (Glucose Regulated Protein 78 and glucose regulatory protein 78 were detected by MTT assay. Results: there was no significant difference between type 2 diabetes mellitus group and healthy control group in clinical basic data. Plasma FBG (fasting blood-glucose fasting blood glucose, HbA1c (Hemoglobin A1C glycosylated hemoglobin) and TG (Triglyceride, triglyceride in type 2 diabetes mellitus group were not significantly different from those in type 2 diabetes mellitus group. 3.73 卤3.44 mmol/L) was significantly higher than that in the normal control group (1.230.40 mmol/L), CP (C peptide, 1.230.40 mmol/L), CP (C peptide). 1.29 卤0.41 nmol/L and HOMA- 尾 index (48.9 卤27.08%) were significantly lower than those in the normal control group (P0.05), but there was no significant difference in insulin level between the two groups (P0.05). The level of serum par-4 in T2DM group (17.44 卤3.65 ng/ml) was significantly higher than that in N group (14.08 卤2.94 ng/ml), the difference was statistically significant (P0.05). The correlation analysis showed that there was a negative correlation between serum par-4 secretion and HOMA- 尾 index (r = 0.455,0.012) and C peptide (r = 0.393, P = 0.032) in type 2 diabetes mellitus. There was a positive correlation between HOMA- 尾 index and C peptide (r = 0.497, P < 0.005). Compared with normal control group (C group, apoptosis rate: 3.14 卤1.08%), 尾-cell apoptosis rate (4.32 卤0.79%) and par-4 concentration (67.6 卤16.32 ng/ml) in par-4 interference group (I group) were significantly higher than those in normal control group (C group, 3.14 卤1.08%). There was no significant difference in insulin secretion function (6.65 卤0.36 m IU/L) and cell proliferation function (0.70 卤0.14) (P0.05). Compared with group C, the apoptosis rate (33.82 卤3.15%), the secretion concentration of par-4 (388.18 卤18.43 ng/ml), the expression of par-4 and GRP78 in high glucose group (D group) were significantly higher than those in group C (P < 0.05). Insulin secretion concentration (3. 59 卤0. 20 m IU/L) and islet cell proliferation function (0. 36 卤0. 10) were significantly decreased (P 0. 05). The insulin secretion function (4.48 卤0.25 mIU/L), the cell proliferation function (0.50 卤0.17) and the par-4 secretion concentration (164.58 卤16.62 ng/ml) in the high glucose and high fat par-4 interference group (T group) were lower than those in the control group (164.58 卤16.62 ng/ml). The difference was statistically significant (P0.05); Compared with D group, apoptosis rate (18.34 卤2.11%), par-4 secretion concentration, par-4 and GRP78 protein expression decreased, insulin secretion concentration and cell proliferation function increased in T group (P0.05). Conclusion: the function of islet cells in type 2 diabetic patients is lower than that in the normal group. The serum par-4 concentration is significantly higher and negatively correlated with the HOMA- 尾 index in patients with type 2 diabetes mellitus. Interfering with the expression of par-4 could but not completely improve the apoptosis and dysfunction of 尾-cell induced by high glucose and high fat. The mechanism may be related to the decrease of the level of (endoplasmic reticulum stress,ERS induced by 尾-cell endoplasmic reticulum stress.
【学位授予单位】：第三军医大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R587.1

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