猫爪草皂苷对结肠癌LoVo细胞凋亡和细胞内Ca^(2+)浓度的影响

Effect of Radix Ranunculus Ternate Saponins on Apoptosis and the Free Ca^(2+) Concentration in Human Colon Carcinoma LoVo Cells

目的:探讨猫爪草皂苷(Radix ranunculus ternate saponins,RRTS)对人结肠癌LoVo细胞增殖和凋亡的调控作用及其机制。方法:按猫爪草皂苷浓度分为1、10、100、500μg·ml-14个处理组和对照组(未加猫爪草皂苷),处理LoVo细胞不同时间后,倒置显微镜观察细胞形态;采用MTT法检测细胞增殖的抑制情况;流式细胞仪(FCM)进行细胞周期分析并计算凋亡率;激光共聚焦(LCSM)测定皂苷对LoVo胞内Ca2+浓度的影响。结果:与对照组比较,猫爪草皂苷处理组可使细胞明显变圆、体积缩小、脱壁细胞增多;对LoVo细胞的增殖抑制作用增强,随着浓度的增加凋亡率增加,可看出细胞阻滞在G1期,随着给药浓度的增加,细胞凋亡峰(AP峰)越来越明显。作用于LoVo细胞4 h后,胞内Ca2+明显上升,4 h、8 h时略有下降,但仍维持在较高水平(P<0.001)。结论:猫爪草皂苷具有明显抑制人结肠癌LoVo细胞增殖和诱导凋亡作用,引起凋亡的途径可能与Ca2+的增加有关。

Objective： To explore the regulatory effect of radix ranunculus temate saponins （RRTS） on proliferation and apoptosis in human colon carcinoma LoVo cells. Methods： The LoVo cells were divided into 4 treatment groups（ 1, 10, 100, 500 ug· ml^-1 RRTS） and blank control group. The morphological alterations of LoVo cells after treated for different time were observed by inverse microscopy, the inhibition of cell proliferation was detected by MTT assay, the cell cycle phase was analyzed and the apoptosis was inspected by flow cytometry; LCSM （laser confocal scanning microscopy） was used to observe the affection of RRTS on the free Ca^2＋ concentration of the cells. Results： After being treated with RRTS, LoVo cells grew round and small obviously and were against the wall under the inverse microscopel with the increasing of the concentration of RRTS and the elongation of the treated time, the LoVo cell growth was inhibited, the inhibition of proliferation of LoVo cells increased in a time - and dose-dependent manner （ P 〈 0. 001 ）; the apoptotic rates of the dose-dependent treated groups increased significantly vs the control＇s （P〈 0. 001 ）; cell cycle was blocked in G1 phase, and typical sub-diploid peaks were found by FCM analysis at different dosage groups; LCSM shows that Ca^2＋ efflux increases obviously after treated by RRTS for 4 h. Although it decreased a bit after 8 h, Ca^2＋ efflux still maintained at a high level after 12 h （P 〈 0. 001 ）. Conclusion： RRTS could significantly inhibit the growth of LoVo cells, and induce the apoptosis through Ca^2＋ efflux increase, probably.