紫杉醇-壳聚糖缓释膜对胆管成纤维细胞增殖的抑制作用

Inhibitation of paclitaxel-chitosan sustain film on biliary fibroblasts cell proliferation

目的探讨紫杉醇-壳聚糖缓释膜(PTX-Chitosan Sustain film,PTX-CSF)对胆管成纤维细胞增殖和凋亡的影响。方法采用组织块法进行成纤维细胞的原代单细胞培养。将实验分为5组:未处理组,单纯PTX处理组(250nM)和低、中、高壳聚糖PTX缓释膜处理组(100nM、250nM、500nM)。使用噻唑蓝(methyl thiazolyl tetrazolium,MTT)比色法检测细胞的增殖情况,划痕实验检测紫杉醇对TGF-β1诱导细胞迁移的抑制作用,流式细胞术(flow cytometry,FCM)检测细胞的凋亡及细胞周期。结果紫杉醇-壳聚糖缓释膜能有效抑制胆管成纤维细胞的增殖,具有剂量和时间依赖性。其抑制作明显强于单纯的紫杉醇用药,且随着时间的延长,优势逐渐体现。紫杉醇-壳聚糖缓释膜能有效抑制TGF-β1对胆管成纤维细胞的促迁移作用,较单纯PTX组具有更长的时间效应。72h时,紫杉醇-壳聚糖缓释膜组成纤维细胞的凋亡率显著高于单纯PTX组和对照组(P<0.05),后2者之间无统计学差异。72h时,紫杉醇-缓释膜组和单纯PTX组的G2/M期细胞比例均较未处理组显著增加,2者相比,前者也明显高于后者,差异均具有统计学意义(P<0.05)。结论紫杉醇-壳聚糖缓释膜具有较强的细胞毒性作用及明显的缓释效果,可以维持较长时间的有效浓度,从而较单纯紫杉醇用药更能持续地抑制胆管成纤维的增殖。

Objective To explore the effect of Paclitaxel-Chitosan Sustain film on growth, apoptosis and cell cycle of biliary fibroblasts cells. Methods Human biliary fibroblasts cells were cultured and treated with PTX-CSF and naked PTX,separately, untreated cells as blank control. The experiment was divided into five groups:untreated group, simple PTX-treated group (250nM) and low, medium and high chitosan sustained-release film PTX-treated group (100 nM, 250 nM, 500 nM). The proliferations of cells were determined by MTT assay. The apoptosis and cell cycle of cells were detected by FCM. Results The proliferation of biliary fibroblasts cells was inhibited by PTX-CSF with time-dependent and dose-dependent, and the inhibiting effect was more obvious than naked PTX treatment as the time went on. Meanwhile, PTX-CSF could inhibit the magration of bile duct fibroblasts induced by TGF-β1,and had longer effect than naked PTX. After 72 h, the apoptosis rate of cells treated with PTX-CSF was significantly higher than cells treated with naked PTX or untreated cells(P<0.05), the difference between naked PTX or untreated cells was not significant. Compared with untreated cells, the proportion of G 2/M in cells treated with PTX or PTX-CSF were significantly increased, and the former was sinificantly higher than the latter(P<0.05). Conclusion Compared with naked PTX, PTX-CSF has strong cytotoxic effects and obviously sustained-release effect. The effective concentration can be maintain for a long time by PTX-CSF, and it could be as the novel drug delivery system to continuously inhibit proliferation of bile duct fibroblasts.