静脉注射神经生长因子基因修饰的骨髓基质干细胞对辐射诱导小鼠肝细胞凋亡的影响

Intravenous injection of nerve growth factor gene-modified bone marrow stromal stem cells on the apoptosis of the mouse hepatic cells induced by radiation

背景:研究发现神经生长因子能减少神经细胞和心肌细胞的凋亡。目的:假设神经生长因子基因修饰的骨髓基质干细胞对辐照诱导小鼠肝细胞凋亡具有保护作用,拟验证其可能性。设计、时间及地点:随机对照动物实验,于2007-01/08在广东医学院实验动物中心实验室完成。材料:昆明小鼠120只,随机分细胞注射组40只、单纯照射组40只和正常对照组40只。方法:单纯照射组采用60Coγ射线照射小鼠,吸收剂量率188.2cGy/min,细胞注射组照射前2d静脉给予神经生长因子基因修饰的骨髓基质干细胞5×1010L-1;正常对照组不辐射损伤。观察小鼠30d存活率和死亡动物存活时间;用原位末端标记法和流式细胞仪观察受照小鼠肝细胞凋亡及p53表达,再用Westernblot进一步测定p53的表达水平。主要观察指标:①受照射小鼠30d存活率。②受照射小鼠肝细胞凋亡情况。③受照射小鼠肝细胞p53表达。结果:细胞注射组30d存活率高于单纯照射组。单纯照射组小鼠大量肝细胞发生凋亡,呈灶性。细胞注射组小鼠肝细胞凋亡少于单纯照射组,凋亡细胞呈散在分布。细胞注射组肝细胞p53标记的荧光强度和细胞比例明显低于单纯照射组(P<0.05)。细胞注射组p53带灰度值低于单纯照射组(P<0.05)。结论:神经生长因子基因修饰的骨髓基质干细胞能明显抑制受照小鼠肝细胞凋亡及抑制肝细胞p53的表达。

BACKGROUD： Studies indicate that nerve growth factor （NGF） can decrease the apoptosis of nerve cells and myocardial cells. OBJECTIVE： To suppose that NGF gene-modified bone marrow stromal stem cells have protective effect on the apoptosis of mouse hepatic cells induced by radiation, and verify the possibility. DESIGN, TIME AND SETTING： The randomized controlled animal experiment was done at the Central Laboratory of Experimental Animals, Guangdong Medical College, between January 2007 and August 2007. MATERIALS： 120 Kunming mice were randomly divided into cell injection group, simple irradiation group and normal control group, with 40 mice in each group. METHODS： Mice in the simple irradiation group were radiated with ^60Coγ -irradiation, the absorbed dose rate was 188.2 cGy/min; mice in the cell injection group were intravenously injected 5 × 10^10/L NGF gene-modified bone marrow stromal stem cells at 2 days before irradiation; mice in the normal control group were not irradiated. The 30-day survival rate of mice and the average life span of dead mice post-irradiation were observed. FACS was used to detect the apoptosis and p53 expression of hepatic cells from irradiated mice, and the expression level of p53 was further examined with western blotting. MAIN OUTCOME MEASURES： The 30-day survival rate of irradiated mice; apoptosis condition of irradiated mouse hepatic cells; the expression level of p53 of irradiated mice. RESULTS： The 30-day survival rate of mice in the cell injection group was higher than that in the simple irradiation group. Apoptosis occurred in large quantity of hepatic cells of mice in the simple irradiation group and apoptosis cells presented focus distribution. Hepatic cell apoptosis of mice in the cell injection group was less than that in the simple irradiation group, and apoptosis cells presented diffused distribution. The fluorescence intensity of hepatic cell p53 marker and the cell ratio in the cell injection group were significantly lower than that in the simple irradiation group （P 〈 0.05）. The gray value of p53 band in the cell injection group was lower than that in the simple irradiation group （P 〈 0.05）. CONCLUSION： NGF gene-modified bone marrow stromal stem cells can inhibit the apoptosis of mouse hepatic cells induced by radiation and the p53 expression of hepatic cells.