角质细胞生长因子对老龄鼠胸腺细胞增殖作用的研究

Proliferative effect of keratinocyte growth factor on thymocyte

目的观察角质细胞生长因子(KGF)通过恢复胸腺上皮细胞(TEC)数量促进老龄鼠胸腺细胞增殖的作用。方法①体内实验:BALB/C小鼠10只,随机分为KGF处理组和对照组,每组各5只,连续7d分别给予皮下注射重组人KGF(5mg·kg-1.d-1)和生理盐水,4周后检测小鼠胸腺和脾组织的细胞总数,流式细胞仪分别检测胸腺组织中CD4单阳性(CD4SP)、CD8单阳性(CD8SP)、CD4CD8双阳性(DP)、CD4CD8双阴性(DN)以及脾组织中CD4SP、CD8SP各T细胞亚群的比例;②体外实验:鼠胸腺上皮来源的1C6细胞,分别经含不同浓度(25、50、100、200、400、800ng/ml)的重组人KGF和不含重组人KGF的完全培养液(对照组)作用24h,采用噻唑蓝(MTT)法测定细胞增殖率。结果细胞计数和流式细胞仪检测显示,与对照组相比,KGF处理组平均胸腺细胞总数和CD4SP、CD8SP、DP、DN各T细胞亚群的平均细胞数以及KGF处理组平均脾细胞总数和CD4SP、CD8SP各T细胞亚群的平均细胞数均有明显增加(均P<0.05)。MTT法检测显示,在低浓度(<200ng/ml)时,1C6细胞增殖率随KGF浓度的增加不断升高;当KGF浓度为200ng/ml时,1C6细胞增殖率达到最高值;而在高浓度(>200ng/ml)时,1C6细胞增殖率随KGF浓度的增加又逐渐下降;与对照组比较,浓度为100、200、400ng/ml的重组人KGF对1C6细胞具有明显的促增殖作用(均P<0.05)。结论KGF可通过恢复老龄鼠TEC数量以及促进胸腺细胞生成和增加成熟T细胞向外周输出而增强机体的免疫力。

Objective To observe keratinocyte growth factor （KGF） to enhance the proliferation of thymus cells through restoring the number of thymic epithelial cells （TEC） in aged mice. Methods （1）In vivo： BALB/C mice were randomly divided into two groups （KGF-treated group and control group, n = 10, 5 for each ）.They were treated with recombinant human KGF （5 mg·kg^-1·d^-1 ） and saline for 7 consecutive days of subcutaneous injection and the absolute cell number of thymus and spleen were detected 4 weeks later. The ratios of CD4 single positive （CD4SP）, CD8 single positive （CD8SP）, CD4CD8 double positive （DP） and CD4CD8 double negative （DN） T cell subsets in thymus were detected by flow cytometry and so were CD4SP and CD8SP T cell subsets in spleen; （2）In vitro： 1C6 cells originated from mouse thymic epithelial cells were cultured in complete culture medium with different concentrations （25, 50, 100, 200, 400, 800 ng/ml） of recombinant human KGF and without recombinant human KGF （control group） for 24 h, and cell proliferation rate was detected by MTT assay. Results Cell counting and flow cytometry show that the mean number of absolute cell and CD4SP, CD8SP, DE DN T cell subsets of thymus and the mean number of absolute cell and CD4SP, CD8SP T cell subsets of spleen increase significantly in KGF group as compared with the control group （P 〈 0.05）. MTT assay shows that when KGF concentration is less than 200 ng/ml, the rate of cell proliferation increases as the concentration rase, when KGF concentration is 200 ng/ml, the rate reaches the peak value and when KGF concentration is more than 200 ng/ml, the rate decreases gradually as the concentration declined. Proliferation effect on 1C6 cells is obvious when the concentration of recombinant human KGF are 100, 200 and 400 ng/ml （P 〈 0.05）, as compared with the control group respectively. Conclusion KGF could enhance body immune function through restoring the number of TEC, promoting the cytopoiesis of thymus and increasing the peripheral output of mature T cells in aged mice.