摘要
为探究热应激对猪睾丸Spo11、DNA双链断裂分子标记γH2AX和DNA重组修复分子标记Rad51表达和定位的影响,笔者构建了环境热应激(37℃环境温度,每日加热3 h,连续7 d)和睾丸局部热应激(42℃,局部阴囊加热1 h)的猪动物模型,未处理组猪睾丸作为对照。通过免疫组织化学染色、Western-blot和qRT-PCR等方法检测Spo11、Rad51和γH2AX在猪睾丸中的表达和定位。免疫组化结果显示,对照组生精小管内可见各级生精细胞,Spo11和Rad51主要定位于初级精母细胞的细胞核,γH2AX免疫阳性染色见于初级精母细胞的细胞核。与对照组相比,环境热应激组初级精母细胞的细胞核Spo11和γH2AX免疫阳性着色无明显差异,Rad51免疫阳性着色较弱;局部热应激组初级精母细胞的细胞核Spo11、Rad51和γH2AX免疫阳性着色较强,细胞定位未发生变化。Western-blot和qRT-PCR结果显示,与对照组相比,环境热应激组Spo11蛋白、m RNA和γH2AX蛋白水平均无显著差异,Rad51蛋白和m RNA表达水平均显著降低(P<0.01);局部热应激组Spo11蛋白和m RNA表达水平均显著增加(P<0.05),Rad51蛋白、m RNA和γH2AX蛋白表达水平均显著降低(P<0.01)。结果表明,环境热应激对睾丸Spo11和γH2AX蛋白表达水平无显著影响,下调Rad51表达水平,提示DNA双链断裂后修复受损;局部热应激上调初级精母细胞Spo11、Rad51和γH2AX表达水平,提示DNA双链断裂增加。
s:The study aimed to investigate the effects of heat stress on the expression and localization of Spo11,γH2AX as the DNA double-strand break molecular marker and Rad51 as the DNA recombination repair molecular marker in testis of boar.In present study,environmental heating at 37℃,3 h per day for 7 days and local scrotal heating at 42℃for 1 h boar models were constructed.Testis of boars without heat stress as control.The expression and localization of Spo11,Rad51 andγH2AX in heat stress testis tissues of boar were detected by immunohistochemical staining,qRT-PCR and Western-blot.The immunohistochemical results showed that various types of spermatogenic cells were observed in the spermatogenic tubules of the control group,Spo11 and Rad51 were mainly localized to the nuclei of primary spermatocytes,andγH2AX immunopositive staining was observed in the nuclei of primary spermatocytes.Compared with the control group,the immunopositive staining of Spo11 andγH2AX were not signif icantly different,and the weaker immunopositive staining of Rad51 was observed in the nuclei of primary spermatocytes of environmental heat stress group,the stronger Spo11,Rad51 andγH2AX immunopositive staining were observed in the nucleus of primary spermatocytes of local heat stress group,cellular localization was not significantly different.Western-blot and qRT-PCR results showed compared with the control group,the levels of Spo11 protein,mRNA andγH2AX protein were not significantly different and the expression levels of Rad51 protein and mRNA were significantly reduced(P<0.01)in the environmental heat stress group.The expression levels of Spo11 protein and mRNA were significantly increased(P<0.05)and the expression levels of Rad51 protein,mRNA andγH2AX protein were significantly reduced(P<0.01)in the local heat stress group.In conclusion,environmental heat stress had no significant effect on the expression levels of Spo11 andγH2AX proteins in testis,and the expression level of Rad51 was down-regulated suggested that impaired repair after DNA double strand break,local heat stress upregulated the expression levels of Spo11,Rad51 andγH2AX in primary spermatocytes indicated that increased DNA double-strand break.
作者
李东晓
李佳容
赵子惠
常倩雯
白涛
范小瑞
贺俊平
LI Dong-xiao;LI Jia-rong;ZHAO Zi-hui;CHANG Qian-wen;BAI Tao;FAN Xiao-rui;HE Jun-ping(College of Veterinary Medicine,Shanxi Agricultural University,Taigu 030801,China)
出处
《中国兽医科学》
CAS
CSCD
北大核心
2023年第9期1200-1206,共7页
Chinese Veterinary Science
基金
国家自然科学基金项目(31470124)
山西省优秀博士来晋工作奖励资金科研项目(SXBYKY2022067)。
