睾丸组织移植对内分泌衰退小鼠学习记忆损害的改善作用

Improving effect of testicular tissue graft on impaired learning and memory in endocrine recession mice

目的研究睾丸组织移植对内分泌衰退小鼠学习记忆损害的改善作用，并探讨其可能的作用机制。方法将40只昆明种雄性小鼠用随机数字表法分为假手术组、去势组、雄激素组和移植组，每组10只。切除小鼠双侧睾丸，以建立去势小鼠模型；移植组将同种小鼠皋丸组织块植入去势小鼠背部皮下肌肉层；雄激素组小鼠灌胃给予雄激素，剂量为30mg／kg，治疗3个月；其他组给予等量生理盐水。90d后，ELISA法测量各组小鼠内分泌水平；Morris水迷宫观察小鼠行为学变化；HE染色观察海马CA1区形态学改变；蛋白免疫印迹检测小鼠脑内雄激素受体和凋亡相关蛋白，即B细胞淋巴瘤／白血病-2（Bcb2）、Bcl-2相关X蛋白（Bax）、半胱氨酸天冬氨酸蛋白酶3（caspase-3）和半胱氨酸天冬氨酸蛋白酶9（caspase-9）的表达。结果假手术组小鼠血清睾丸酮为（50．50±11．41）nmol／L，去势组为（15．50±2．84）nmol／L，雄激素组为（41．504-10．01）nmol／L，移植组为（38．00±9．78）nmol／L。与去势组相比较，移植组小鼠血清睾丸酮水平较高（P〈0．01）；移植组小鼠逃逸潜伏期缩短，游泳路程明显减少，穿越平台次数大幅度增加（P〈0．01）；移植组海马CAl区神经元形态结构有所改善，且Bax、半胱氨酸天冬氨酸蛋白酶3（caspase-3）和半胱氨酸天冬氨酸蛋白酶（caspase-9）蛋白表达水平明显减少（P〈0．01），雄激素受体和Bcl-2蛋白表达量显著上升（P〈0．01）。结论睾丸组织移植可能是通过分泌内源性睾丸酮，结合脑内雄激素受体，调控神经元凋亡相关基因的表达，上调Bcl-2蛋白水平，降低Bax、caspase-3和caspase-9蛋白的表达量，抑制大脑神经元凋亡，进而改善学习记忆能力。

Objective To study the improving effect of testicNar tissue graft on impaired learning and memory in endocrine recession mice, and to explore the possible mechanisms. Method Forty Kunming male mice were randomly divided into four groups by random number table; sham operation group, castration group, androgen （30 mg/kg） control group and testicular tissue graft group. The bilateral testes of mice were cut off to establish castrated mouse model. After modeling, the back subcutaneous muscle layers of castrated mice were implanted on the aUogeneic testicular tissue blocks in graft group. Androgen group mice were intragastrically administrated with androgen for three months, and other two groups of mice were given an equal volume of normal saline. At the end of treatment, the levels of serum testosterone in each group of mice were measured by ELISA. Leaming and memory abilities of the mice were assessed by Morris water maze behavioral test. Morphological changes in the hippocampal CA1 region were observed through hematoxylin-eosin staining （HE）. The expression levels of AR and apoptosis-related proteins （Bcl-2, Bax, caspase-3 and caspase-9） in the brain were detected by Western blotting. Result As compared with castration group, the levels of serum testosterone in graft group were increased significantly （P〈0. 01 ）, learning and memory abilities were greatly strengthened （P 〈0. 01）, the morphological structures in hippocampal CA1 neurons were improved, and the expression levels of Bax, caspase-3 and caspase-9 were significantly reduced （P〈0. 01）, while the expression levels of AR and Bcl-2 had obvious enhancements （P〈0. 01）. Conclusion Testicular tissue graft can improve learning and memory ability probably by secreting endogenous testosterone which will bind with androgen receptor in the brain, then regulating the expression of neuronal apoptosiS related genes, increasing Bcl-2 protein level and reducing the expression of Box, caspase-3 and caspase-9 protein, and finally inhibiting neuronal apoptosis.