丙烯酰胺对大鼠睾丸组织PCNA、EGFR蛋白表达的影响及机制研究
发布时间:2018-06-01 14:07
本文选题:丙烯酰胺 + 睾丸 ; 参考:《重庆医科大学》2013年硕士论文
【摘要】:目的 研究丙烯酰胺(acrylamide,AA)对大鼠睾丸组织的亚急性毒性作用,及其对增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)和表皮生长因子受体(epidermal growth factor receptor,,EGFR)蛋白表达的影响,初步探讨丙烯酰胺生殖毒性的可能作用机制。 方法 将32只5~6周龄雄性SD大鼠随机分为4组,每组8只,分别为空白对照组,4mg/kg、12mg/kg和36mg/kg剂量丙烯酰胺染毒组,连续灌胃染毒30天。处理结束后,心脏取血,化学发光法检测血清中睾酮(testosterone,T)和雌二醇(estradiol,E2)的含量,取双侧睾丸组织,HE染色观察睾丸组织病理学变化,TUNEL检测睾丸细胞凋亡情况,RT-PCR检测睾丸组织PCNA、EGFR mRNA的表达,免疫组化和Westernblot检测睾丸组织PCNA、EGFR蛋白的表达。 结果 与对照组比较,12、36mg/kg染毒组大鼠体重明显下降(P0.05),36mg/kg染毒组大鼠睾丸脏器系数升高(P0.05);HE染色显示,12、36mg/kg染毒组大鼠睾丸组织出现了明显的病理形态学改变,具体表现为生精细胞减少、生精上皮萎缩、管腔出现空泡等;TUNEL显示12、36mg/kg染毒组睾丸凋亡细胞显著增多(P0.05);化学发光法显示,各染毒组大鼠血清T浓度呈剂量依赖型下降(P0.05),12、36mg/kg染毒组大鼠血清E2浓度下降(P0.05);RT-PCR显示,12、36mg/kg染毒组大鼠睾丸组织PCNA和EGFR的mRNA表达均明显降低(P0.05);免疫组化和Western blot显示,12、36mg/kg染毒组大鼠睾丸组织PCNA的蛋白表达明显降低(P0.05),各染毒组大鼠睾丸组织EGFR的蛋白表达明显降低(P0.05),差异具有统计学意义。 结论 一定剂量的丙烯酰胺染毒可导致大鼠睾丸组织发生病理学变化,凋亡细胞增多。PCNA、EGFR蛋白在大鼠睾丸组织中的表达减少,在一定程度上可抑制生精细胞DNA的合成,打乱睾丸生精细胞的分裂、增殖与凋亡之间的动态平衡,扰乱性激素水平,最终导致大鼠不同程度的生殖毒性,为丙烯酰胺生殖毒性的机制研究提供实验依据。
[Abstract]:Purpose To study the subacute toxicity of acrylamide acrylic acid (AA) to testis of rats and its effect on the expression of proliferating cell nuclear antigen-PCNA (PCNA) and epidermal growth factor receptor (EGFR) protein of proliferating cell nuclear antigen (PCNA) and epidermal growth factor receptor (EGFR). To explore the possible mechanism of reproductive toxicity of acrylamide. Method Thirty-two 5-week-old male SD rats were randomly divided into 4 groups (n = 8 in each group). The rats in the control group (n = 8) were treated with 4 mg 路kg ~ (-1) of 36mg/kg and 12 mg / kg / kg of 36mg/kg respectively, and the rats were given intragastric administration for 30 days. After the treatment, the blood was taken from the heart, and the contents of testosterone Tand estradiolone E2) in serum were detected by chemiluminescence method. The histopathological changes of testis were observed by HE staining in bilateral testis. The apoptosis of testicular cells was detected by Tunel. The expression of PCNA- EGFR mRNA in testicular tissues was detected by RT-PCR, and the expression of PCNA- EGFR protein was detected by immunohistochemistry and Westernblot. Result Compared with the control group, the weight of the rats in the 125mg / kg group decreased significantly (P 0.05 mg / kg) and the testicular organ coefficient increased. The HE staining showed that the testicular tissues of the rats exposed to 123mg / kg P0.05mg / kg showed obvious pathological morphology changes, and the spermatocyte decreased. Testicular apoptotic cells were significantly increased in 12 ~ 36mg / kg group with atrophy of spermatogenic epithelium and vacuole in the lumen, and showed by chemiluminescent method. Serum T concentration decreased in a dose-dependent manner in each exposure group (P 0.05 mg / kg). Serum E _ 2 concentration decreased in the rats of each group. RT-PCR showed that the mRNA expression of PCNA and EGFR in testis of rats exposed to 12 ~ 36mg / kg was significantly lower than that of the control group, and the expression of mRNA in testis was significantly decreased by immunohistochemistry and Western blot, and the results of Western blot showed that the rats were exposed to 120.36 mg / kg of P0. 05 mg / kg. The expression of PCNA protein in testis of the rats in the control group decreased significantly, while the expression of EGFR protein in the testis of each exposed group was significantly lower than that in the control group (P 0.05). Conclusion A certain dose of acrylamide could induce pathological changes in rat testis, decrease the expression of DNA protein in testis and inhibit the synthesis of DNA in spermatogenic cells. Disrupting the division of testicular spermatogenic cells, the dynamic balance between proliferation and apoptosis, and disturbing the level of sex hormones, eventually lead to different degrees of reproductive toxicity in rats, which provides experimental basis for the study of the mechanism of acrylamide reproductive toxicity.
【学位授予单位】:重庆医科大学
【学位级别】:硕士
【学位授予年份】:2013
【分类号】:R114
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