摘要
目的运用超高效液相色谱-四极杆-静电场轨道阱串联质谱(UHPLC-QE-MS)技术分析橘荔散结丸的药物成分,围绕核因子-κB(NF-κB)信号通路探讨橘荔散结丸诱导子宫肌瘤细胞凋亡的炎症机制。方法①采用UHPLC-QE-MS技术分析橘荔散结丸化学成分。②培养原代人子宫肌瘤细胞,免疫组化法进行鉴定。实验分为5组,空白组常规培养,基质组加入0.05%DMSO培养,米非司酮组加入1×10-5 mol/L米非司酮含药液培养,橘荔散结丸高、低浓度组分别加入2 mg/mL和1 mg/mL橘荔散结丸含药液培养。采用流式细胞术检测子宫肌瘤细胞凋亡率,采用Western blot法检测子宫肌瘤细胞中IKKα、IKBα、p-IKBα、p65、肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)蛋白表达情况,采用qPCR技术检测子宫肌瘤细胞中IKKα、IKBα、p65、TNF-α、IL-6 mRNA表达量。结果鉴别橘荔散结丸主要化学成分91个,冻干粉与中成药间共有22个。米非司酮组和橘荔散结丸高、低浓度组子宫肌瘤细胞早期凋亡率和晚期凋亡率均明显高于空白组和基质组(P均<0.05),且米非司酮组、橘荔散结丸高浓度组早期凋亡率和晚期凋亡率均明显高于橘荔散结丸低浓度组(P均<0.05),橘荔散结丸高浓度组早期凋亡率明显高于米非司酮组(P<0.05)。橘荔散结丸高浓度组和米非司酮组子宫肌瘤细胞中IKKα、p-IKBα、p65、TNF-α蛋白相对表达量均明显高于空白组和橘荔散结丸低浓度组(P均<0.05),且橘荔散结丸高浓度组和米非司酮组子宫肌瘤细胞中IL-6蛋白相对表达量均明显高于空白组(P均<0.05);橘荔散结丸高浓度组子宫肌瘤细胞中p-IKBα、p65、IL-6蛋白相对表达量均明显低于米非司酮组(P均<0.05)。米非司酮组、橘荔散结丸高浓度组、橘荔散结丸低浓度组子宫肌瘤细胞中IKKα、IKBα、p65、IL-6 mRNA相对表达量和米非司酮组、橘荔散结丸高浓度组TNF-αmRNA相对表达量均明显高于空白组(P均
Objective It is to analyze the drug components of Juli Sanjie Pill(JSP)by ultra-high performance liquid chromatography-quadrupole-electrostatic orbitrap mass spectrometry(UHPLC-QE-MS),and to investigate the inflammatory mechanism of JSP in inducing uterine fibroid cell apoptosis via nuclear factorκB(NF-κB)signaling pathway.Methods①The chemical components of JSP were analyzed by UHPLC-QE-MS.②Primary human uterine fibroid cells were cultured and identified by immunohistochemical method.The experiment was divided into 5 groups,the blank group was cultured routinely,the substrate group was cultured by adding 0.05%DMSO,the mifepristone group was cultured by adding 1×10-5 mol/L mifepristone-containing solution,and the JSP high and low concentration groups were cultured by adding 2 mg/mL and 1 mg/mL JSP-containing solution,respectively.The apoptosis rate was detected by flow cytometry.The expressions of IKKα,IKBα,p-IKBα,p65,TNF-αand IL-6 in uterine fibroid cells were detected by Western blot,and the mRNA expressions of IKKα,IKBα,p65,TNF-αand IL-6 were detected by qPCR.Results There were 91 main chemical components of JSP,22 in lyophilized powder and Chinese patent medicine.The early and late apoptotic rates of uterine fibroid cells in JSP high and low concentration groups and mifepristone group were all significantly higher than those in the blank group and substrate group(all P<0.05),and the early and late apoptotic rates of uterine fibroid cells in JSP high concentration group and mifepristone group were all significantly higher than those in the JSP low concentration group(all P<0.05),the rates in the high concentration group were significantly higher than those in the mifepristone group(P<0.05).The relative expressions of IKKα,p-IKBα,p65,and TNF-αproteins in uterine fibroid cells of the JSP high concentration group and mifepristone group were significantly higher than those of the blank group and JSP low concentration group(all P<0.05),and the relative expressions of IL-6 protein in uterine fibroid
作者
谢卓庭
王乃平
程晓榆
关永格
李坤寅
XIE Zhuoting;WANG Naiping;CHENG Xiaoyu;GUAN Yongge;LI Kunyin(Dongguan Hospital of Guangzhou University of Chinese Medicine,Dongguan 523000,Guangdong,China;The Third Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangzhou 510000,Guangdong,China)
出处
《现代中西医结合杂志》
CAS
2023年第17期2350-2356,2364,共8页
Modern Journal of Integrated Traditional Chinese and Western Medicine
基金
广东省自然科学基金项目(2020A1515010600)
广东省中管局项目(20203003)
东莞市社会发展科技项目(高水平医院建设专项)(20231800903501)。
