雷公藤内酯醇及雷公藤内酯醇聚合物胶束对HT29细胞增殖及凋亡的影响

Antitumor effect of triptolide and triptolide loaded polymeric micelles in HT29 cell

目的研究雷公藤内酯醇(Triptolide TP)和雷公藤内酯醇聚合物胶束(Triptolide loaded poly-meric micelles,TP-PM)对人结肠癌HT29细胞增殖及凋亡的作用及机制,评价聚合物胶束作为雷公藤内酯醇纳米载体的可行性。方法用含不同浓度的TP和TP-PM(1-100ng/mL)培养液孵育HT29细胞24h、48h和72h后,利用WST-1、BrdU试剂盒测定细胞的存活率、增殖率;通过测定LDH活性判断细胞膜完整性;利用流式细胞仪测定细胞死亡、存活和凋亡率;并通过Caspase-Glo3/7定量试剂盒测定细胞caspase3/7的表达情况。结果 TP和TP-PM对HT29细胞活性和细胞增殖的抑制作用显著,呈明显的剂量、时间依赖性,TP-PM对HT29的抑制作用优于TP(P<0.05)。TP和TP-PM对HT29细胞膜的破坏轻微。FDA-PI双染结果显示,两组中FDA阳性细胞均呈现时间、剂量依赖性的下降(P<0.01);PI阳性细胞率呈现时间、剂量依赖性的上升(P<0.01);FDA、PI双阴性细胞也明显上升,但不呈时间、剂量依赖关系。Caspase测定结果显示TP和TP-PM均可引起Caspase3/7活性增高,TP-PM组引起的凋亡作用强于TP组(P<0.05)。结论 TP和TP-PM可以通过诱导凋亡的方式抑制肿瘤细胞生长和增殖,且TP-PM的作用强于TP,聚合物胶束是一种极具前景的载药体系。

Objective To investigate the antitumor mechanism and compare the antitumor effect of triptolide （TP） and triptolide loaded polymeric micelles （TP-PM） in HT29 cell,and evaluate the feasibility of polymeric micelle as triptolide nanocarrier.Methods HT29 cells were incubated with TP and TP-PM for 24h,48h and 72h,then,cell viability was measured by WST-1 assay; cell membrane integrity was measured by LDH assay,and cell proliferation was measured by BrdU assay.Additionally,cell survival and apoptosis was measured by FDA-PI staining,and caspase 3/7 activity was measured by Caspase-Glo 3/7 assay kit.Results HT29 cell viability and proliferation was apparently inhibited by the TP and TP-PM,the inhibitions showed a time-dependent and dose-dependent manner,and the inhibitory effect of TP-PM was better than TP （P0.05）.Both TP and TP-PM caused only slight HT 29 cell membrane damage.FDA-PI staining results indicated that FDA-positive cell showed a time-dependent and dose-dependent down trend,while PI-positive cell show a time-dependent and dose-dependent ascending trend.FDA,PI double negative cell also increased more,however,it neither showed a time-dependent nor dose-dependent effect.HT29 cell caspase 3/7 activity increased after incubated with TP and TP-PM,and TP-PM induced a higher apoptosis than TP （P0.05）.Conclusions Both TP and TP-PM can potently inhibit HT29 cell growth and proliferation via induction of apoptosis,TP-PM shows a better inhibitory effect than TP.Polymeric micelle is a promising drug carrier.