摘要
目的探讨蠲饮泄肺方对脂多糖(LPS)诱导的气道黏液高分泌的抑制作用及其机制。方法培养人支气管上皮16HBE细胞,分别用脂多糖作用6 h、0.005%浓度的蠲饮泄肺方相应处理细胞,并对细胞进行SOCS1-siRNA转染,荧光定量PCR技术(RTPCR)、Western blot法检测转染效果。根据不同的处理措施,分为空白对照组、试验1组(仅脂多糖刺激)、试验2组(脂多糖刺激+SOCS1-siRNA转染)、试验3组(脂多糖刺激+蠲饮泄肺方)、试验4组(脂多糖刺激+SOCS1-siRNA转染+蠲饮泄肺方)。Western blot法分别检测各组细胞裂解液中SOCS1、MUC5AC、磷酸化JAK1和STAT1表达量,同时检测JAK1、STAT1总蛋白,以甘油醛-3-磷酸脱氢酶(GADPH)蛋白表达量为内参。结果 RT-PCR、Western blot法检测SOCS1-siRNA转染效果明确。与空白对照组比较,试验1组细胞内SOCS1表达被抑制,MUC5AC、JAK1和STAT1磷酸化水平增高(P<0.001);试验2组细胞内SOCS1表达被抑制及MUC5AC、JAK1和STAT1磷酸化水平增高更明显(P<0.001);试验3组相比试验1组,细胞内SOCS1表达增高,MUC5AC、JAK1和STAT1磷酸化水平下降(P<0.001);试验4组相比试验2组,细胞内SOCS1表达增高,MUC5AC、JAK1和STAT1磷酸化水平下降(P<0.001)。使用蠲饮泄肺方后能改善脂多糖和SOCS1-siRNA对细胞内SOCS1表达的抑制,降低JAK1和STAT1磷酸化水平及MUC5AC的表达。结论蠲饮泄肺方能抑制脂多糖诱导的MUC5AC高表达,从而抑制气道黏液高分泌,与SOCS1抑制JAK1/STAT1信号通路、产生负反馈调节、降低磷酸化水平有关。
Objective To investigate the inhibitory effects of Juanyin Xiefei Formula on airway mucus hypersecretion induced by lipopolysaccharide （LPS） and its mechanism. Methods The human bronchial epithelial cells 16HBE were cultured. The cells were treated with lipopolysaccharide for 6 hours and/or 0.005% Juanyin Xiefei Formula, and the cells were transfected with suppressor of cytokine signaling 1 （SOCS1）-siRNA. The transfection effect was detected by RT-PCR and Western blot. According to the different treatment measures, the cells were divided into the blank control group, experiment 1 group （only lipopolysaccharide stimulation）, experiment 2 group （lipopolysaccharide stimulation plus SOCS1-siRNA transfection） , experiment 3 group （lipopolysaccharide stimulation plus Juanyin Xiefei Formula） , experiment 4 group （lipopolysaccharide stimulation plus SOCS1-siRNA transfeetion plus Juanyin Xiefei Formula）. The expressions of SOCS1, MUC5AC, phosphorylated JAK1 and STAT1 in the cell lysate of each group were detected by Western blot, and the total proteins of JAK1 and STAT1 was detected, with the protein expression of glyceraldehyde- 3- phosphate dehydrogenase （GADPH） as the internal parameter. R^ults The effect of SOCSI-siRNA transfection was obvious detected by RT-PCR and Western blot. Compared with the blank control group, the expression of SOCS1 in the experiment 1 group was inhibited, the expression of MUC5AC and the phosphorylation levels of JAK1 and STAT1 were increased （ P〈0.001 ）. The inhibition on SOCS1 expression and the increase of MUC5AC expression, JAK1 and STAT1 phosphorylation in the experiment 2 group were more significant （P〈0.001）. Compared with the experiment 1 group, the SOCSI expression was increased, the MUC5AC expression and the phosphorylation levels of JAKI and STATI were decreased in the experiment 3 group （ P〈0.001 ）. Compared with the experiment 2 group, the SOCS1 expression was increased, and the MUC5AC expression and the phosphorylation levels of JAKI and STAT1 were decreased in the experiment 4 group （P〈O.001）. After the treatmem of Juanyin Xiefei Formula, the inhibition of lipopo｜ysaccharide and SOCS1-siRNA on SOCSI expression was improved, the phosphorylation levels of JAK 1 and STAT1 and the expression of MUC5AC were decreased. Conclusion Juanyin Xiefei Formu[a can inhibit the high expression of MUC5AC induced by lipopolysaccharide, and then inhibit the airway mucus hypersecretion, which is related to SOCS1 inhibition on JAK1/STAT1 signaling pathway, negative feedback regulation and reduction of phosphorylation level.
作者
钟秀君
顾文燕
徐威
汤杰
杨佩兰
王庆其
ZHONG Xiujun;GU Wenyan;XU Wei;TANG Jie;YANG Peilan;WANG Qingqi(Department of Respiration,Yueyang Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China;Yueyang Clinical Medical School,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China;School of Basic Medical Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)
出处
《上海中医药杂志》
2018年第9期69-73,共5页
Shanghai Journal of Traditional Chinese Medicine
基金
上海市卫计委2017年度中医优势病种培育项目(zybz-2017007)
上海中医药大学预算内(自然科学类)资助项目(2016YSN51)
