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目的 应用体外培养海人藻酸 (KA)兴奋性毒素损伤的细胞模型 ,研究 GDNF对损伤的背根节神经元的不同作用 ,为进一步研究 GDNF对神经元作用机制 ,表达产生及检测方法提供思路 .方法 采用原代培养的方法 ,用 N1无血清分离培养背根节神经元 ,再用 5μm ol· L- 1 KA作用 6～ 8h ,加上含有 GDNF的无血清培养基 ,继续培养 2 4h,然后用MTT法检测反映细胞的活性 ,胎盘蓝染色记数、细胞总蛋白测定以及形态学观察突起的生长状况 .结果 1细胞活性 A值 :GDNF+KA(0 .0 32 8± 0 .0 0 6 ) ,KA(0 .0 2 85± 0 .0 0 75 ) ,GDNF (0 .0 398± 0 .0 0 2 ) ,Blank(0 .0 41± 0 .0 0 2 ) (P<0 .0 5 ) ;2活细胞数相应为 GDNF+KA (6 0± 4.4) ,KA(35 .7±2 .2 ) ,GDNF (5 9.3± 3.6 ) ,Blank (5 7.7± 2 .9) ;3细胞的总蛋白分别为 GDNF+KA (70 .3± 9.2 ) (P<0 .0 1) ,KA (49.0± 3.7) ,GDNF (75 .0± 7.3) ,Blank (6 8.0± 5 .5 ) (P<0 .0 1) ;4突起长度 :实验组与对照组不明显 ,GDNF组与空白对照组不明显 .结论 在正常无血清体外培养情况下 GDNF对DRG神经元作用不明显 ,在 KA兴奋性毒素损伤后 ,对细胞的活性、存活及总蛋白合成有明显的保护作用 ,但对突起的生长则没有明显的促进作用 .
OBJECTIVE: To study the different effects of GDNF on injured DRG neurons in vitro by culturing cell models of excitotoxicity of kainic acid (KA) in vitro and to provide ideas for further study on the mechanism, expression and detection of GDNF on neurons. Methods Primary culture was used to culture dorsal root ganglion neurons with serum-free N1, and then treated with 5μmol · L-1 KA for 6 ~ 8h. Serum-free medium containing GDNF was added to continue culture for 24 hours. Then MTT assay was used to detect the activity of the cells, the number of placental blue staining, the determination of the total cellular protein and morphological observation of the growth of the cells.Results1 Cell activity A value: GDNF + KA (0.032 8 ± 0 .0 0 6), KA (0. 0285 ± 0. 0 0 75), GDNF (0. 398 ± 0. 0 0 2), Blank 0 0 1 0 0 0 2 0 (P 0. The number of viable cells was GDNF + KA (60 ± 4.4), KA (35.7 ± 2.2), GDNF (5 9.3 ± 3.6), Blank (5 7.7 ± 2.9) The total protein of cells were GDNF + KA (70.3 ± 9.2) (P <0.01), KA (49.0 ± 3.7), GDNF (75.0 ± 7.3), Blank (6 8.0 ± 5.5) (P <0 01); 4 Protrusion length: the experimental group and the control group was not obvious, GDNF group The blank control group was not obvious.Conclusion GDNF has no obvious effect on DRG neurons in normal serum-free in vitro culture, and has obvious protective effect on the activity, survival and total protein synthesis of KA after excitotoxicity of KA, Prominent growth did not significantly promote the role.