The Effect Of Methamphetamine On Function Of Erection,HPT Axis And HPA Axis In Male Animal Models

[Background]Substance abuse,which can cause serious damage to health of abusers and threaten the social public security,has become a global social problem.The related drug control and rehabilitation work has become a major problem and arduous task that global governments all need to actively face at this stage.The emergence of new synthetic drugs represented by Methamphetamine(MA,commonly known as "ICE")has furtherly exacerbated the spread of substance abuse.Epidemiological studies at home and abroad have found that the age composition of MA abusers has an obvious younger trend.Part One:Effect of MA on erection of rats[Objective]With MA exposure model of male rats,HIF-lα/A2bR/PI3K/PKBα/eNOS/cGMP signaling pathway and COX-2/PTGIS/cAMP signaling pathway were used to explore the effects and possible mechanisms of MA exposure on penile erectile function in acute phase,chronic phase and after abstinence.[Methods]1.50 male rats were randomly assigned to 1 of 5 groups:control(acute control,chronic control),MA(acute MA,chronic MA)and abstinence.Rats in MA and abstinence groups received intraperitoneal injection of MA solution regularly;in the abstinence group,the MA was ceased in the last 10 days;the rats in the control groups received intraperitoneal injection of the same volume of normal saline simultaneously.2.The penile erectile function was evaluated regularly by observing spontaneous erection and BR.3.ELISA:the levels of 8-OHdG in serum;the levels of NOS,cGMP and cAMP in penile cavernous tissue.4.RT qPCR was used to detect mRNA expression levels of HIF-lα,A2bR,eNOS,COX-2 and PTGIS in penile cavernous tissue;Western blot was used to detect protein expression levels of HIF-1α,A2bR,PI3K,p-PKBα,p-eNOS,COX-2 and PTGIS.[Results]1.Male rat models of MA acute exposure,chronic exposure and abstinence were successfully constructed.2.Erectile function:compared with the acute control group,the erectile function of the acute MA group was significantly enhanced;compared with the chronic control group,the erectile function of the chronic MA group was significantly reduced,and the erectile function of the abstinence group was not significantly different.3.HIF-1α/A2bR/PI3K/PKBα/eNOS/cGMP signal pathway:(1)Compared with the acute control group,the levels of 8-OHdG,NOS and cGMP detected by ELISA in acute MA group were significantly higher,and the mRNA expression levels of HIF-lα,A2bR and eNOS were significantly increased,and the protein expression levels of HIF-lα,A2bR,PI3K,p-PKBa and p-eNOS were significantly increased.(2)Compared with the chronic control group,the levels of 8-OHdG,NOS and cGMP detected by ELISA in chronic MA group were decreased significantly,the mRNA expression levels of A2bR and eNOS were significantly decreased,and the protein expression levels of A2bR,PI3K,p-PKBa and p-eNOS were significantly decreased,but no significant difference of HIF-1α mRNA and protein expression.(3)Compared with the chronic control group,there was no significant difference in levels of 8-OHdG,NOS and cGMP detected by ELISA in abstinence group,no significant difference in mRNA expression levels of HIF-lα,A2bR and eNOS,and no significant difference in the protein expression levels of HIF-1α,A2bR,PI3K,p-PKBα and p-eNOS.4.COX-2/PTGIS/cAMP signal pathway(1)Compared with the acute control group,the mRNA and protein expression levels of COX-2 and PTGIS were significantly higher in the acute MA group,and the cAMP level detected by ELISA was significantly higher.(2)Compared with the chronic control group,the mRNA and protein expression levels of COX-2 and PTGIS were significantly lower in the chronic MA group,and the cAMP level detected by ELISA was significantly lower.(3)Compared with the chronic control group,there was no significant difference in cAMP level detected by ELISA in the abstinence group,and no significant difference in mRNA and protein expression levels of COX-2 and PTGIS.[Conclusions]1.Rat models of acute and chronic MA exposure and withdrawal can be established by intraperitoneal injection,and can be used for research of erectile function.2.Erectile function is enhanced after acute exposure and impaired after chronic exposure,but it can be restored after withdrawal.3.MA exposure may affect the erectile function of rats through the HIF-1α/A2bR/PI3K/PKBα/ENOS/cGMP signal pathway:acute MA exposure can increase oxidative stress and serum 8-OHdG level,and up-regulate the expression of HIF1α/A2bR/PI3K/PKBα/eNOS/cGMP signal pathway to enhance erectile function;chronic MA exposure can increase oxidative stress and serum 8-OHdG level,although there was no significant change in the expression of HIF-1,but downstream A2bR/PI3K/PKBα/eNOS/cGMP signal pathway of HIF-la was down-regulated and inhibited erectile function;after withdrawal,oxidative stress disappeared and the expression of HIF-lα/A2bR/PI3K/PKBα/eNOS/cGMP signal pathway and erectile function returned to the control level.4.MA exposure may affect the erectile function of rats through the COX-2/PTGIS/cAMP signal pathway:acute MA exposure can up-regulate the expression of COX-2/PTGIS/cAMP signal pathway and enhance erectile function;chronic MA exposure can down-regulate the expression of COX-2/PTGIS/cAMP signal pathway and inhibit erectile function;after withdrawal,the expression of COX-2/PTGIS/cAMP signal pathway and erectile function returned to the control level.Part Two:Effect of chronic MA exposure on HPT axis and HPA axis function in non-human primates[Objective]By constructing an animal model of non-human primates after chronic exposure to MA,to explore the effects and possible damage of chronic exposure to MA on the function of HPT axis and HPA axis from the perspective of endocrine hormone level and target organ molecular biology.[Methods]1.10 adult male monkeys were divided into 2 groups:control group(normal saline)and MA group(normal saline solution of MA).Normal saline or normal saline solution of MA were injected at a gradually increasing dose for 4 weeks,and then maintained for 24 weeks.2.Collect blood samples and test the levels of thyroid and adrenal related hormones every week.3.RT qPCR and Western blot were used to detect:the expression levels of Bcl-2,Bax and TSH-R in thyroid;the expression levels of Bcl-2,Bax and ACTH-R in adrenal cortex.[Results]1.A non-human primate model of chronic MA exposure was successfully constructed.2.The levels of T4,FT4,T3,TSH,ACTH and cortisol in MA group were increased significantly,but there was no significant difference in the levels of FT3,TGAb,TPOAb and aldosterone.3.Most thyroid glands in MA group showed obvious disordered structure and different sizes of acini,accompanied by lymphocyte infiltration and local fibrosis.4.In the thyroid tissue of MA group,the expression levels of Bcl-2 and TSH-R were decreased significantly,and the expression level of Bax was increased significantly.5.In the adrenal cortex of MA group,the expression levels of Bcl-2 and ACTH-R were decreased significantly,and the expression level of Bax was increased significantly.[Conclusions]1.The animal model of chronic MA exposure in non-human primates can be successfully constructed by intravenous injection.2.In the non-human primate model,long-term MA abuse can change function of HPT axis and HPA axis.3.In the non-human primate model,long-term MA abuse can cause obvious histological and microstructure disorder in thyroid tissue,but there is no similar change in adrenal tissue.4.In the non-human primate model,long-term MA abuse can reduce the expression of anti-apoptotic gene Bcl-2 and increase the expression of pro-apoptotic gene Bax in thyroid and adrenal cortex,which may induce apoptosis or micro injury of thyroid and adrenal cortex cells,and finally lead to the function change of HPT axis and HPA axis.