紫草素对Cx32蛋白组成的细胞缝隙连接功能的影响

The effect of Shikonin on the function of gap junction composed of Cx32

目的探究紫草素对宫颈癌Hela细胞缝隙连接通讯功能(gap junctional intercellular communication,GJIC)和缝隙连接蛋白32(Cx32蛋白)表达水平的影响。方法将细胞分成两组培养,加多西环素为诱导组,不加多西环素为非诱导组,采用CCK-8(Cell Counting Kit-8)法检测不同浓度紫草素对Hela细胞毒性的影响;采用细胞接种荧光示踪法(parachute assay)观察不同浓度紫草素对缝隙连接(gap junction,GJ)功能的影响;采用蛋白免疫印迹法(western-blot)研究紫草素在影响GJ功能浓度范围内对Cx32蛋白表达的影响。结果紫草素在0~4μmol/L浓度时对非诱导组Hela细胞无毒性作用;紫草素在0~3μmol/L浓度时对诱导组Hela细胞无毒性作用;诱导组细胞在紫草素1~3μmol/L浓度时荧光传递量会随浓度增加;紫草素(1~3μmol/L)能浓度依赖性增强GJ功能及Cx32蛋白表达量。结论紫草素能够增强Cx32组成的细胞GJ功能,这种增强作用可能与其增加Cx32蛋白表达水平有关。

Objective To investigate the effect of shikonin on gap junctional intercellular communication （GJIC） and connexin 32 （Cx32） expression of cervical cancer Hela cells. Methods The cultured Hela cells were divided into two groups, with one group being induced with doxycycline and the other without any treatment. CCK-8 （ Cell Counting Kit-8 ） method was used to evaluate the effects of different concentrations of shikonin on Hela cells; the effects of different concentrations of shikonin on gap junction were monitored by cell inoculation with fluorescent tracer; variation of Cx32 expression was evaluated by Western blotting. Results Shikonin was shown to have no significant cytotoxicity on Hela in control group when the concentration ranged from 0 to 4 mol/L; Shikonin was displayed to have no obvious toxicity on Hela cells in induced group when the concentration was around 0-3 mol/L; Shikonin was presented to be able to increase the amount of fluorescence transfer flow in induced cells when concentration arrived at 1 - 3 mol/L. Conclusion Shikonin was shown to be capable of enhancing the GJ function of Cx32 cells, which may be related to the increased expression of Cx32 protein.