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目的观察3T3-L1前体脂肪细胞诱导分化过程中Visfatin蛋白分泌水平的变化,探讨胰岛素对脂肪细胞Visfatin分泌水平的调节作用。方法体外培养3T3-L1前体脂肪细胞,采用1-甲基-3-异丁基黄嘌呤(MIX)、胰岛素、地塞米松方案诱导3T3-L1前体脂肪细胞分化为成熟脂肪细胞,诱导分化过程中采用油红O染料鉴定并观察细胞分化情况。在诱导3T3-L1前体脂肪细胞分化成熟的不同时段(第0、4、6、8天)收集细胞培养上清液;并以100nmol/L胰岛素干预诱导分化第6、8天的成熟脂肪细胞,设未干预组为对照组,采用酶联免疫吸附法(ELISA)检测细胞培养上清液中Visfatin的蛋白水平。结果未诱导分化的3T3-L1前体脂肪细胞仅分泌低水平Visfatin蛋白[(29.0±2.4)μg/L],诱导分化后随3T3-L1脂肪细胞逐渐分化成熟,脂肪细胞培养上清液中Visfatin的蛋白水平逐渐升高,至分化第8天成熟脂肪细胞Visfatin分泌达到高峰[(48.0±3.4)μg/L]。以100nmol/L胰岛素干预第6天和第8天分化成熟的脂肪细胞后,细胞培养上清液Visfatin的蛋白水平均较对照组显著升高[(57.9±5.7)μg/Lvs(45.7±3.8)μg/L;(61.0±4.2)μg/Lvs(48.0±3.4)μg/LPa<0.05]。结论Visfatin蛋白分泌与脂肪细胞成熟度有关,胰岛素对脂肪细胞Visfatin的分泌可能具有正性调节作用。
Objective To observe the changes of Visfatin protein secretion during 3T3-L1 preadipocyte differentiation and to investigate the regulatory effect of insulin on visfatin secretion of adipocytes. Methods 3T3-L1 precursor adipocytes were cultured in vitro. 3T3-L1 preadipocytes were induced to differentiate into mature adipocytes by 1-methyl-3-isobutylxanthine (MIX), insulin and dexamethasone Oil red O dye was used in the process to identify and observe cell differentiation. The cell culture supernatants were harvested at different time points (day 0, day 4, day 6, day 8) to induce the differentiation and maturation of 3T3-L1 precursor adipocytes. The mature adipocytes were induced to differentiate on the 6th and 8th days with 100nmol / L insulin . The non-intervention group was set as the control group. The protein level of Visfatin in the cell culture supernatant was detected by enzyme-linked immunosorbent assay (ELISA). Results The 3T3-L1 preadipocytes which did not induce differentiation secreted only low levels of Visfatin protein ([(29.0 ± 2.4) μg / L], and gradually differentiated and matured with 3T3-L1 adipocytes after differentiation. Visfatin (48.0 ± 3.4) μg / L] in mature adipocytes on the 8th day after differentiation. The protein level of Visfatin in cell culture supernatant was significantly higher than that in control group (57.9 ± 5.7) μg / Lvs (45.7 ± 3.8) after differentiation of mature adipocytes on day 6 and day 8 with 100 nmol / L insulin. μg / L; (61.0 ± 4.2) μg / Lvs (48.0 ± 3.4) μg / LPa <0.05]. Conclusion Visfatin protein secretion is related to adipocyte maturation. Insulin may have a positive regulatory effect on visfatin secretion in adipocytes.