血竭素高氯酸盐对成纤维细胞增殖与成纤维细胞生长因子mRNA表达的影响

Effects of dracorhodin perchiorate on proliferation and fibroblast growth factor mRNA expression of dermal fibroblasts

目的观察血竭素高氯酸盐（Dp）对人皮肤成纤维细胞（Fh）生物学特性的影响，探讨其促进创面愈合的机制。方法分离培养人正常Fb，将Dp以不同浓度（0．063、0．125、0．250、0．625、1．250、2．500mg／L）分别加入培养液，采用噻唑蓝（MTT）比色法检测Dp在不同浓度以及不同时间点对体外培养的Fb增殖作用的影响。通过流式细胞仪，实时荧光定量聚合酶链式反应（real-time PCR）分别检测最适浓度培养条件下Fh的细胞周期变化以及成纤维细胞生长因子（FGF）mRNA的合成表达。结果Dp在0．625—1．250mg／L浓度范围内，其吸光度值（0．237±0．012、0．243±0．017）均高于对照组（0．208±0．011）差异有统计学意义（t值分别为2．11、2．23，P〈0．05），此浓度范围可促进Fb增殖，且呈剂量依赖性，在浓度为1．250mg／L时（A值0．243±0．017），促增殖作用最为显著（t=2．23，P〈0．01），流式细胞仪结果显示，在浓度为1．250mg／L时，Dp可明显促进Fh通过G，／S及s／G2期限制点，S期及G2／M期细胞与对照组比较明显增多，G0／G1期细胞与对照组比较明显减少（t值分别为4．32、7．53、3．27，P〈0．05）。细胞因子mRNA表达测定中，1．250mg／LDp组与对照组比较表达明显上调，差异亦有统计学意义（t=1．48，P〈0．05）。结论Dp能显著促进Fb增殖，加快Fb周期进程，同时可促进FGF的mRNA表达，可能与血竭促进创面愈合的机制有关。

Objective To observe the effects of dracorhodin perchlorate （Dp） on the biological characteristics of human dermal fibroblasts in vitro, and explore the possible therapeutic mechanism on inducing wound healing. Methods Human normal fibroblasts （Fb） were isolated and cultured, and different concentrations of Dp were added into the culture medium. By using methyl thiazol tetrazolium （MTF） colorimetric assay, the effects of Dp at different concentrations and different time points in vitro on proliferation of Fb were analyzed. The changes in the cell cycle of Fb and cytokine expression of fibroblast growth factor （FGF） mRNA synthesis were detected in optimal concentration by flow cytometry and real-time fluo-rescence quantitative polymerase chain reaction （real-time PCR）. Results Dp in the 0. 625-1. 250 mg/L concentration range could promote Fb proliferation in a dose-dependent manner, and the most significant effect on the proliferation （ t = 2. 11,2. 23, P 〈 0. 01 ） was obtained at a concentration of 1. 250 mg/L. 1. 250 mg/L Dp could promote Fb to pass through GJS and S/G2 phase restriction point, the number of cells in S phase and GJM phase was increased significantly compared with the control group, and number of cells in G0/G1 phase was reduced significantly as compared with the control group （ t = 4. 32, 7. 53, 3.27 ,P 〈 0. 05）. As compared with the control group, the cytokine mRNA expression was significantly increased in Fb treated with 1. 250 mg/L Dp （ t = 1.48, P 〈 0. 05 ）. Conclusion Dp can significantly pro-mote the proliferation of Fb, accelerate the process of Fb cycle, and promote the mRNA expression of FGF at the same time, which might be contributed to the mechanisms of Dp promoting wound healing.