龙血竭有效部位对胎鼠皮肤成纤维细胞生长的作用

Growth of Fetal Mouse Skin Fibroblasts Stimulated by Effect Locus of Resina Draconis

目的：探讨龙血竭不同有效部位对体外培养的胎鼠皮肤成纤维细胞生长及分泌的影响。方法：孕龄12．5～14．5d昆明小鼠胚胎，无菌条件下将躯干部皮肤制成匀浆，接种于25m^2塑料培养瓶，体积分数10％FBS的高糖DMEM培养基培养传代，P=3代细胞以8×10^6个／L接种于96孔板，细胞饥饿24h后进行实验。采用石油醚、氯仿、乙酸乙酯、正丁醇、乙醇等极性递增溶剂依次萃取龙血竭中的各种有效部位，均选用0．125g／L、0．25g／L、0．5g/L、1g／L、2g/L、4g/L五个浓度梯庹作用于96孔板中的细胞（DMSO助溶），24h后MTT法检测不同条件下细胞的生长曲线，免疫组化检测细胞分泌Ln、Ⅰ型胶原的变化。10％FBS的高糖DMEM培养条件为空白对照，1％DMSO的高糖DMEM培养条件为实验对照。结果：龙血竭乙酸乙酯有效部位在低浓度对胎鼠成纤维细胞的生长有轻度抑制作用，随浓度增加促生长作用增强，1g/L龙血竭乙酸乙酯有效部位对胎鼠皮肤成纤维细胞的促生长作用最佳，并促进In、Ⅰ型胶原的分泌。龙血竭其它有效部位对胎鼠成纤维细胞均无刺激生长的作用。结论：龙血竭乙酸乙酯有效部位能促进胎鼠皮肤成纤维细胞的生长和分泌。

Objectives： To investigate the growth of fetal mouse skin fibroblasts stimulated by effect locus of Resina Draconis .Methods： The fetal mouse were executed at 12.5 - 14.5d. The skin of trtmk was homogeneted and transplanted into 25cm^2 culture flask in germ free condition. The tissue was cultured with high-glucose DMEM media containing 10% FBSNF3 fibroblasts were transplanted into 96 shadow mask with 8×10^6/L. These cells were tested being starved 24h later. Resina Dracon/s were extracted by ligarine, chloroform, acetoacetate, n - butanol and ethyl alcohol to get various kinds of effect locus. Those cells in 96 shadow mask were tested by 5 concentration gradients, 0.125g/L, 0.25g/L, 0.5g/L, 1g/L, 2g/L, 4g/L.The growth curves got by MTT.Ln and type Ⅰ collagen protein were tested by immuno histochemistry. The blank control was HG-DMEM containing 10% FBS. The experimental control was HG-DMEM containing 10% FBS and 1% DMSO.Results： The fetal mouse skin fibroblasts were slightly inhibited by the effect locus extracted by acetoacetate of Resina Dracon/s at low concentration. Then the cells were proliferated with higher concentration. The best concentration was 1g/L.Ln and type Ⅰ collagen protein were excreted more.The other effect locus of Resina Draconis couldn＇ t stimulate the cells growth. Conclusions： The effect locus extracted by acetoacetate of Resina Draconis can stimulate the growth and secretion of fetal mouse skin fibroblasts.