摘要
目的基于JAK2/STAT1/3信号通路探讨当归补血汤含药血清对肿瘤坏死因子α(TNF-α)诱导的人类风湿关节炎成纤维样滑膜细胞(HFLS-RA)增殖、凋亡的影响及机制。方法将Wistar大鼠随机分为空白组、雷公藤多苷组(9.45 mg·kg^(-1))及当归补血汤高、中、低剂量组(15、7.5、3.75 g·kg^(-1)),连续灌胃给药7 d,制备大鼠空白血清及含药血清。将HFLS-RA细胞随机分为6组,即空白组(10%空白组大鼠血清培养基)、模型组(20 ng·mL^(-1)TNF-α+10%空白组大鼠血清培养基)、雷公藤多苷组(20 ng·mL^(-1)TNF-α+10%雷公藤多苷组大鼠血清培养基),以及当归补血汤含药血清低、中、高剂量组(20 ng·mL^(-1)TNF-α+10%当归补血汤低、中、高剂量组大鼠血清培养基);按照分组设置加入终质量浓度为20 ng·mL^(-1)的TNF-α,1 h后加入相应含药(或空白)血清处理24 h。采用CCK-8法检测HFLS-RA细胞增殖活性;流式细胞术(Annexin V-FITC/PI双染法)检测HFLS-RA细胞凋亡情况;RT-PCR法检测细胞中Bax、Bcl-2、caspase-3 mRNA表达;ELISA法检测细胞上清液中白细胞介素6(IL-6)、白细胞介素15(IL-15)水平;Western Blot法检测细胞中Bax、Bcl-2、caspase-3、p-JAK2、p-STAT3、p-STAT1蛋白表达水平。结果与空白组比较,模型组HFLS-RA细胞的增殖活性显著升高(P<0.01),细胞凋亡率明显下降(P<0.05),IL-6、IL-15水平显著升高(P<0.01);Bax、caspase-3 mRNA及蛋白表达水平显著降低(P<0.01),而Bcl-2 mRNA及蛋白表达水平显著升高(P<0.01),Bax/Bcl-2比值显著降低(P<0.01);细胞p-JAK2、p-STAT1、p-STAT3蛋白表达水平显著升高(P<0.01)。与模型组比较,雷公藤多苷组及当归补血汤含药血清各剂量组HFLS-RA细胞的增殖活性均显著降低(P<0.01),细胞凋亡率显著升高(P<0.05,P<0.01),IL-6、IL-15水平均显著降低(P<0.01);Bax、caspase-3mRNA及蛋白表达水平显著升高(P<0.01),而Bcl-2 mRNA及蛋白表达水平显著降低(P<0.05,P<0.01),Bax/Bcl-2比值显著升高(P<0.01);当�
Objective To explore the effects and mechanisms of Danggui Buxue Decoction-containing serum on the proliferation and apoptosis of human fibroblast-like synoviocyte in rheumatoid arthritis(HFLS-RA)induced by TNF-α based on JAK2/STAT1/3 signaling pathway.Methods Wistar rats were randomly divided into the blank group,the tripterygium glycosides group(9.45 mg·kg^(-1))and the Danggui Buxue Decoction high-,medium-and low-dose groups(15,7.5 and 3.75 g·kg^(-1)).All the groups were administered by gavage for 7 consecutive days,and the blank serum and drug-containing serum were prepared.HFLS-RA cells were randomly divided into6 groups,i.e.blank group(10%serum medium in blank rats),model group(20 ng·mL^(-1)TNF-α+10%serum medium in blank rats),tripterygium glycosides group(20 ng·mL^(-1)TNF-α+10%tripterygium glycosides in rat serum medium),and Danggui Buxue Decoction-containing serum low-,medium-and high-dose groups(20 ng·mL^(-1)TNF-α+10%Danggui Buxue Decoction-containing serum low-,medium-and high-dose in rat serum medium).TNF-α was added at a final mass concentration of 20 ng·mL^(-1)according to the group setting,and the corresponding drug-containing(or blank)serum was added after 1 hour.The proliferative activity of HFLS-RA cell was detected by CCK-8;the apoptosis of HFLS-RA cells was detected by flow cytometry(Annexin V-FITC/PI double staining);the mRNA expressions of BAX,Bcl-2 and caspase-3 in the cell supernatant were detected by RT-PCR;interleukin 6(IL-6)and interleukin 15(IL-15)levels in cell supernatant were detected by ELISA;protein expressions of Bax,Bcl-2,caspase-3,p-JAK2,p-STAT3,p-STAT1 in HFLS-RA cells were detected by Western Blot.Results Compared with the blank group,the proliferative activity of HFLS-RA cells in the model group was significantly increased(P<0.01),apoptosis rate was significantly decreased(P<0.05),IL-6 and IL-15levels were significantly increased(P<0.01);mRNA and protein levels of Bax and caspase-3 were significantly decreased(P<0.01),while mRNA and protein levels of Bcl-2 were sign
作者
赵莹
王爽
潘志
汪少琼
王颖航
ZHAO Ying;WANG Shuang;PAN Zhi;WANG Shaoqiong;WANG Yinghang(Changchun University of Chinese Medicine,Changchun 130117 Jilin,China;The Affiliated Hospital of Changchun University of Chinese Medicine,Changchun 130117 Jilin,China)
出处
《中药新药与临床药理》
CAS
CSCD
北大核心
2023年第7期870-877,共8页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
国家自然科学基金项目(82074324)
吉林省自然科学基金项目(20210101235JC)。
