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纸质出版日期:2021,
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鸢尾素通过自噬对棕榈酸诱导的INS-1细胞功能和凋亡的影响[J]. 重庆医科大学学报, 2021,46(8):908-914.
Effect of irisin on function and apoptosis of INS-1 cells induced by palmitic acid through autophagy[J]. Journal of Chongqing Medical University, 2021,46(8):908-914.
鸢尾素通过自噬对棕榈酸诱导的INS-1细胞功能和凋亡的影响[J]. 重庆医科大学学报, 2021,46(8):908-914. DOI: 10.13406/j.cnki.cyxb.002642.
Effect of irisin on function and apoptosis of INS-1 cells induced by palmitic acid through autophagy[J]. Journal of Chongqing Medical University, 2021,46(8):908-914. DOI: 10.13406/j.cnki.cyxb.002642.
目的: 探讨鸢尾素对棕榈酸(palmitic acid,PA)诱导的胰岛β细胞功能和凋亡的影响。方法: 培养INS-1细胞,用不同浓度梯度的鸢尾素干预INS-1细胞,根据蛋白结果筛选出最佳鸢尾素干预浓度为1 500 ng/mL。然后将INS-1细胞分为正常组、PA组、PA+1 500 ng/mL鸢尾素组、PA+1 500 ng/mL鸢尾素+雷帕霉素组、PA+1 500 ng/mL鸢尾素+3-MA组。用3.3 mmol/L和16.7 mmol/L的葡萄糖浓度刺激检测干预后胰岛素的分泌水平,Western blot检测自噬相关蛋白p-mTOR、mTOR、p62、LC3Ⅱ、LC3Ⅰ的蛋白水平,用mGFP-GFP-LC3腺病毒转染观察自噬流水平,流式细胞仪检测细胞凋亡水平。结果采用SPSS 20.0软件进行统计分析,计量资料以均数±标准差(±s)表示,多组间差异比较采用单因素方差分析及Turkey HSD检验。结果: ①PA组LC3Ⅱ/Ⅰ的比值较正常组增高(0.745±0.035 vs.1.015±0.012,P=0.000),随着鸢尾素干预浓度增加,LC3Ⅱ/Ⅰ的水平较PA组逐渐升高且有统计学意义(I1组1.465±0.028,I2组2.269±0.063,I3组1.858±0.074,I4组2.355±0.074,均P=0.000),鸢尾素浓度为1 500 ng/mL时LC3Ⅱ/Ⅰ的水平最高。②PA组INS-1细胞上清液中胰岛素水平在3.3 mmol/L和16.7 mmol/L葡萄糖刺激下较正常组均下降[(0.447±0.067)ng/mL vs.(0.138±0.012)ng/mL,P=0.000;(0.625±0.041)ng/mL vs.(0.245±0.043)ng/mL,P=0.000],鸢尾素干预后胰岛素水平较PA组均升高[(0.138±0.012)ng/mL vs.(0.322±0.041)ng/mL,P=0.001;(0.245±0.043)ng/mL vs.(0.414±0.036)ng/mL,P=0.005]。③与正常组相比,PA干预细胞后,自噬体数量增加(2.33±1.53 vs.10.33±1.53,P=0.000),而自噬溶酶体数量无明显变化(2.33±1.53 vs.3.33±1.53,P=0.939),鸢尾素干预后,自噬体数量和自噬溶酶体数量较PA组均升高(10.33±1.53 vs.15.67±1.53,P=0.007;3.33±1.53 vs.12.00±2.00,P=0.001)。④与正常组相比,PA干预后细胞凋亡率增加[(11.417±1.297)%vs.(46.507±0.525)%,P=0.000],鸢尾素干预后凋亡率降低[(46.507±0.525)%vs.(33.767±1.083)%,P=0.000]。结论: 鸢尾素通过自噬改善PA诱导的INS-1细胞分泌功能和凋亡。
Objective: To investigate the effect of irisin on the function and apoptosis of pancreatic β cells induced by palmitic acid (PA). Methods: Firstly
INS-1 cells were cultivated and treated with different concentration gradients of irisin. The optimal concentration of 1 500 ng/mL was selected according to the protein results of Western blot. Secondly
INS-1 cells were divided into the normal group
the PA group
the palmitate acid+1 500 ng/mL irisin group
the palmitate acid+1 500 ng/mL irisin+rapamycin group and the palmitate acid+1 500 ng/mL irisin+3-MA group. The glucose-stimulated insulin secretion (GSIS) test (with glucose levels of 3.3 mmol/L and 16.7 mmol/L) was performed to determine the insulin level in each group. Western blot was used to determine the levels of mamlian target of rapamycin (mTOR)
p-mTOR
p62
LC3Ⅱand LC3Ⅰ; mRFP-GFP-LC3 and flow cytometry were used to determine the autophagic flow and cell apoptosis
respectively. SPSS 20.0 was used to conduct data analysis. Results were presented as the mean±SD
one-way analysis of variance followed by Turkey HSD was used to calculate differences among the various study groups. Results: ①Compared with the normal group
the ratio of LC3Ⅱ/Ⅰin the PA group was higher (0.745±0.035 vs.1.015±0.012
P=0.000). With the increase of irisin concentration
the level of LC3Ⅱ/Ⅰwas gradually increased and had statistical significance (I1: 1.465±0.028
I2: 2.269±0.063
I3: 1.858±0.074
I4: 2.355±0.074; P=0.000). When irisin concentration was 1 500 ng/mL
the level of LC3Ⅱ/Ⅰwas the highest.②GSIS levels in the INS-1 cell supernatant in the PA group was significantly reduced for stimulation of 3.3 mmol/L and 16.7 mmol/L of glucose when compared with the normal group [(0.447±0.067) ng/mL vs. (0.138±0.012) ng/mL
P=0.000; (0.625±0.041) ng/mL vs. (0.245±0.043) ng/mL
P=0.000]
while in the irisin treated group were increased when compared with the PA group [(0.138±0.012) ng/mL vs. (0.322±0.041) ng/mL
P=0.001; (0.245±0.043) ng/mL vs. (0.414±0.036) ng/mL
P=0.005].③The number of autophagosome in the PA group was significantly higher than that in the normal group (2.33±1.53 vs.10.33±1.53
P=0.000)
but there was no obvious change in the number of autophagolysosome (2.33±1.53 vs.3.33±1.53
P=0.939). In the irisin group
the autolysosome number and autolysosome number was increased when compared with PA group (10.33±1.53 vs.15.67±1.53
P=0.007; 3.33±1.53 vs.12.00±2.00
P=0.001). Treated with PA for 24 hours
the apoptosis rate of INS-1 cells was increased when compared with the normal group [(11.417±1.297) %vs. (46.507±0.525) %
P=0.000]
while the irisin-treated group had a significantly reduced apoptosis rate when compared with the PA group [(46.507±0.525) %vs. (33.767±1.083) %
P=0.000]. Conclusion: Irisin ameliorates apoptosis and improves secretion function of INS-1 cells induced by PA through autophagy.
鸢尾素自噬棕榈酸胰岛β细胞功能凋亡
irisinautophagypalmitic acidislet βcell functionapoptosis
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