Roles Of P38MAPK And JNK Signalings On Testicular Mitochondrial Oxidative Stress In Hypothyroid Male Rats

Objectives:The aim of this study was to construct hypothyroid model induced by propylthiouracil?PTU?,and then explore the possible molecular mechanism of p38mitogen-activated protein kinase?p38MAPK?and c-Jun NH2-terminal kinase?JNK?signaling on testiscular mitochondrial oxidative stress in hypothyroid male rats to provide a new theoretical basis for studying the mechanism of male reproductive damage in hypothyroid rats.Methods:?1?Establish the hypothyroid model of rats.According to body weight?240-270g?,30 Wistar male rats were divided into control group?C group,NS,0 mg/kg b.w PTU?,Low-dose group?L group,0.1 mg/kg b.w PTU?and high-dose group?H group,10 mg/kg b.w PTU?by intragastric administration.Body weights of all rats were measured once every three days,and they were killed after 60 days.?2?The initial and final body weights were recorded on the first day and day of sacrifice.At the same time,the testes weights were recorded on the day of sacrifice and then were homogenized rapidly.?3?Blood samples were collected to measure the levels of serum triiodothyronine?T3?,thyroxine?T4?and thyroid stimulating hormone?TSH?.?4?The activities of superoxide dismutase?SOD?,catalase?CAT?and Ca²⁺-ATPase in testiscular mitochondria were determined with spectrophotometric assay.?5?Real-time quantitative polymerase chain reaction?RT-qPCR?were used to quantify the relative mRNA expressions of androgen receptor?AR?,p38MAPK and JNK in testiscular mitochondria.?6?Western blot were performed to detected the protein expressions of AR,p38MAPK,JNK as well as phosphorylated p38MAPK?p-p38MAPK?and phosphorylated JNK?p-JNK?in testiscular mitochondria.Results:?1?The detection of thyroid hormone level in serum.Compared with C and L groups,levels of serum T3 and T4 were significantly decreased?p<0.05?,while the TSH level was significantly increased?p<0.05?.?2?The analysis of general toxicity showed that significant decreases of body weights were observed in 30 and 60 days as well as testis weights in 60 days,whereas a significant increase of the relative testes weights was found in the H group compared to the C and L groups?p<0.05?.?3?The activity of Ca²⁺-ATPase in testiscular mitochondria was significantly decreased in H and L groups compared with C group?p<0.05?.?4?After the intragastric administration for 60 d,the activity of SOD in testiscular mitochondria was significantly decreased in the H group compared to C group?p<0.05?and the activity of CAT was significantly decreased in H and L groups compared with C group?p<0.01?.?5?The relative mRNA expressions of AR and p38MAPK in testiscular mitochondria in the H group were higher than those in the C group?p<0.05?,whereas there was no significant difference of the relative JNK mRNA expression in each group?p>0.05?.?6?Compared to C and L groups,the relative protein expression of AR in testiscular mitochondria was significantly decreased in H group?p<0.05?.The relative protein expressions of p-p38MAPK and p-JNK in testiscular mitochondria were significantly increased in H group compared with C and L groups?p<0.01and p<0.05 respectively?.However,there were no significant differences of the p38MAPK and JNK expressions in each group?p>0.05?.Conclusions:The hypothyroid model of male rats was established induced by PTU in this study.The results of present study indicates that the possible molecular mechanisms of male reproductive damage induced by hypothyroidism could be related to changes of AR expression and Ca²⁺-ATPase activity,induction of oxidative stress and activations of p38MAPK and JNK signaling in the testicular mitochondria.This study provides a new theoretical basis for studying the mechanism of male reproductive damage in hypothyroid rats.