京尼平对脂多糖诱导的KGN细胞株炎性反应的影响研究

Effect of genipin on lipopolysaccharide induced inflammation response of KGN cell line

目的研究不同质量浓度京尼平对脂多糖诱导的人卵巢颗粒细胞系(KGN细胞株)炎性反应的影响。方法2018年9月,用脂多糖诱导KGN细胞株炎性反应,24h后分别加入不同质量浓度(0、0.05、0.10、0.20、0.30mmol/L)京尼平,设置空白对照组(添加相应剂量溶剂二甲基亚砜的DMEM/F12培养基)、脂多糖组和脂多糖+0.05、0.10、0.20、0.30mmol/L京尼平组。采用CCK-8法检测KGN细胞株的活性;采用酶联免疫吸附试验检测各组上清液中雌激素水平,以及白细胞介素-4(IL-4)、IL-5、IL-1β、IL-6的表达;蛋白质印迹法检测各组17α羟化酶(CYP17A1)、芳香化酶(CYP19A1)、线粒体解偶联蛋白2(UCP2)、核因子κB(NF-κB)蛋白表达。结果与空白对照组比较,高质量浓度(≥0.10mmol/L)京尼平对KGN细胞株生长具有明显抑制作用,差异均有统计学意义(P<0.05);低质量浓度(≤0.05mmol/L)京尼平对KGN细胞株生长无明显影响,与空白对照组比较,差异均无统计学意义(P>0.05)。脂多糖诱导的KGN细胞株IL-4、IL-5、IL-1β、IL-6表达,以及UCP2、NF-κB蛋白表达均明显增加,差异均有统计学意义(P<0.05);脂多糖诱导的KGN细胞株用高质量浓度京尼平干预后IL-4、IL-5、IL-1β、IL-6表达,以及NF-κB蛋白表达均明显降低,差异均有统计学意义(P<0.05);UCP2、CYP17A1、CYP19A1蛋白表达变化均不明显,差异均无统计学意义(P>0.05);各组雌激素水平比较,差异均无统计学意义(P>0.05)。结论脂多糖能诱导KGN细胞株发生明显的炎性反应,京尼平对炎性反应具有明显的抑制作用,并伴随NF-κB蛋白表达的下降而更明显。而京尼平在炎性状态下可能不影响雌激素的分泌。

Objective To study the effect of different concentrations of genipin on lipopolysaccharide(LPS)induced inflammatory response of human ovarian granulosa cell line(KGN cell line).Methods The inflammatory response of KGN cell line was induced by LPS.After 24 h,the different mass concentrations of genipin(0,0.05,0.10,0.20,0.30 mmol/L)were added respectively.The blank control group(DMEM/F12 culture medium adding corresponding dose solvent dimethyl sulfoxide),LPS group and LPS+0.05,0.10,0.20,0.30 mmol/L genipin groups were set.The activity of KGN cell line was detected by CCK-8.The estrogen level and expression of IL-4,IL-5,IL-1βand IL-6 in supernatant were detected by enzyme-linked immunosorbent assay.The expressions of CYP17 A1,CYP19 A1,UCP2 and NF-κB protein were detected by Western blot.Results Compared with blank control group,the high mass concentration of genipin(≥0.1 mmol/L)had significant inhibitory effect on the growth of KGN cell line,and the difference was statistically significant(P<0.05);the low mass concentration of genipin(≤0.05 mmol/L)had no significant influence on the growth of KGN cell line,compared with the control group,the difference had no statistical significance(P>0.05).LPS induced the IL-4,IL-5,IL-1βand IL-6 expression of KGN cell line,and significant increase of UCP2 and NF-κB protein expression,and the differences were statistically significant(P<0.05).In LPS induced KGN cell line,after the high mass concentration of genipin intervention,the IL-4,IL-5,IL-1βand IL-6 expression and NF-κB protein expression were significantly decreased,and the differences were statistically significant(P<0.05).The changes of UCP2,CYP17 A1,CYP19 A1 protein expression all were unobvious,and the differences were not statistically significant(P>0.05).The estrogen level had no statistical difference among various groups(P>0.05).Conclusion LPS can induce the KGN cell line to develop significant inflammatory response,genipin has obvious inhibiting effect on the inflammatory response,which is more obvious with the decrease of NF-κB protein expression.But in the inflammatory state,genipin may not affect the secretion of estrogen.