摘要
目的:探究加味二妙颗粒有效成分盐酸药根碱清除人乳头瘤病毒(human papilloma virus,HPV)的分子作用机制。方法:利用生物信息学数据库筛查宫颈鳞状细胞癌组织中差异表达基因并对其进行生存分析。体外验证实验利用慢病毒载体构建稳定过表达HPV16 E6/E7蛋白的人角质形成细胞系HaCaT,用盐酸药根碱0.001、0.05、0.2、1、5μmol/L处理HaCaT-HPV16E6/E7细胞系,采用Cell Counting Kit-8(CCK-8)检测盐酸药根碱对HaCaT-HPV16 E6/E7细胞的影响,采用实时荧光定量聚合酶链式反应(Real-time PCR)检测盐酸药根碱对HaCaT-HPV16 E6/E7细胞中相关细胞因子白介素-1β(IL-1β)、白介素-8(IL-8)、白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)的影响,蛋白免疫印迹(Western blot)法检测C2TA蛋白及NF-κB通路相关蛋白的表达,探讨盐酸药根碱在人乳头瘤病毒蛋白E6、E7逃避免疫识别的作用机制。结果:通过数据库筛查得知C2TA与宫颈鳞状细胞癌密切相关,其低表达影响患者的生存情况。体外实验验证得知,与对照组比较,盐酸药根碱给药组能够显著抑制HaCaT-HPV16 E6/E7组中的细胞增殖能力(P <0.01)。盐酸药根碱能恢复HaCaT-HPV16 E6/E7细胞中C2TA的蛋白表达,并参与NF-κB通路激活及下游促炎细胞因子释放(P <0.01)。结论:加味二妙颗粒的有效成分盐酸药根碱可有效增强上皮细胞对人乳头瘤病毒的敏感性,其作用机制可能C2TA表达及NF-κB通路激活导致的细胞因子释放有关。
Objective: To investigate the molecular mechanism of Jatrorrhizine Hydrochloride, an effective component of modified Ermiao Granule, in eliminating human papillomavirus(HPV). Methods: Differential genes of cervical squamous epithelium carcinoma(CSEC) was screened and their existence were analyzed by using databases of biological information. Lentivirus was used to construct a stable and overexpressing HaCaT cell line HPV16 E6/E7. HaCaT-HPV16 E6/E7 cells were treated with Jatrorrhizine Hydrochloride at the doses of 0. 001 μmol/L, 0. 05 μmol/L, 0. 2 μmol/L, 1 μmol/L and 5 μmol/L respectively. The effect of Jatrorrhizine Hydrochloride on HaCaT-HPV16 E6/E7 cells was detected by CCK-8 assay. The effect of Jatrorrhizine Hydrochloride on the levels of IL-1β, IL-8, IL-6 and TNF-α was estimated by using Real-time q-PCR. The expressions of C2TA and NF-κB pathway-related proteins were detected by Western blot to explore the action mechanism of Jatrorrhizine Hydrochloride in the immune escape of HPV E6 and HPV E7. Results: C2TA was closely associated with CSEC through database screening. The low depression of C2TA influenced the survival status of the patients. In vitro experiment showed that compared with that in the control group, the cell proliferation of HaCaT-HPV16 E6/E7 was significantly inhibited in the Jatrorrhizine Hydrochloride groups(P<0.01). Administration of Jatrorrhizine Hydrochloride could rescue the expression of C2TA and participate the activation of NF-κB signaling pathway and the release of down-stream proinflammatory cytokines(P<0.01). Conclusion: Jatrorrhizine Hydrochloride can effectively enhance the sensitivity of epithelial cells to HPV, and its action mechanism may be related to C2TA expression and cytokine release caused by activation of NF-κB pathway.
作者
戴任金铭
时晨
李秀
商婷婷
任青玲
DAI Renjinming;SHI Chen;LI Xiu;SHANG Tingting;REN Qingling(Jiangsu Province Hospital of Chinese Medicine Affiliated to Nanjing University of Chinese Medicine,Nanjing 210000,China;TCM Pediatric Institute of Nanjing University of Chinese Medicine,Nanjing 210000,China)
出处
《中医药信息》
2023年第1期1-8,共8页
Information on Traditional Chinese Medicine
基金
国家自然科学基金面上项目(8207150903)
江苏省中医妇产生殖临床医学创新中心项目(ZX202102)
江苏省科研创新与实践计划科研创新类项目(KYCX21_1680)。
