| Aspongopus is a kind of traditional Chinese medicine with the function of warming kidney and strengthening yang.It is often used to treat kidney deficiency,impotence and other diseases,namely erectile dysfunction.Erectile dysfunction is now a disease of great concern in the world.Many middle-aged and old men are troubled by it and cannot be cured.Clinical treatment of erectile dysfunction mainly through oral sildenafil treatment,and a small amount of supplementary testosterone therapy,but most of these methods can only play a short-term therapeutic effect,can not be cured completely.Aspongopus,as a traditional Chinese medicine for strengthening yang,has rarely been reported on the mechanism of treatment of erectile dysfunction,which hinders the development and utilization of Aspongopus resources.The Chinese Pharmacopoeia(2015 edition)records that Aspongopus have the functions of regulating qi and relieving pain,warming the midst and promoting yang.They are used for fear of cold,bloating pain,liver and stomach pain,impotence due to kidney deficiency,soreness of waist and knee.As a traditional Chinese medicine for strengthening yang,there are few reports on the mechanism of the treatment of erectile dysfunction,which hinders the development and utilization of the resources of the insect.In this study,the anti-fatigue ability of extract of Aspongopus against fatigue,warming kidney and strengthening Yang was studied in vivo and in vitro by means of normal mice,Yang-deficiency mice,Yang-deficiency rats and Leydig cell model of testis,and the material basis and mechanism of Aspongopus warming kidney and strengthening yang were explored.OBJECTIVE: To confirm the warming kidney and strengthening yang effect of Aspongopus,locate the main active parts of Aspongopus,explore the pharmacological effects of similar products of Aspongopus,and analyze the possible mechanism of Aspongopus.METHODS:(1)The serum lactic acid content,blood urea nitrogen content and exhaustive swimming time of normal mice were tested by anti-fatigue test;the mating ability,testosterone content in serum,organ coefficient and HE staining histomorphology of testicular tissue were observed in the model of kidney-yang deficiency induced by hydrocortisone.(2)By dividing the total extracts of Aspongopus into petroleum ether parts,water-soluble parts and alcohol-precipitated parts,the mating ability of mice was observed and testosterone content was detected.(3)Collect the common analogues of Aspongopus on the market,such as cyclopelta parva distant,Black worm,Stink bug,kidney-yang deficiency mice induced by hydrocortisone,observe the mating ability,and detect the content of testosterone in serum.(4)Isolation and purification of rat Leydig cells,ELISA was used to detect the effects of different analogues and extracts of Aspongopus on testosterone secretion of Leydig cells.RT-PCR was used to detect the expression levels of St AR,3β-HSD and CYP11a1 in cells.(5)Kidney-yang deficiency rats were induced by subcutaneous injection of hydrocortisone.Mating ability,organ coefficient,serum testosterone,luteinizing hormone and follicle stimulating hormone content and HE staining histomorphology of testis were observed.Proteomics analysis mainly involved in biological processes and signaling pathways.Western blotting analysis of St AR,3β-HSD,CYP17a1 protein expression.RESULTS:(1)By analyzing the anti-fatigue experiment results of the extract of Aspongopus on mice,the dosage group of Aspongopus(7g/kg)could reduce the content of lactic acid and urea nitrogen in the serum of normal mice,enhance the exhaustive swimming time,and improve the anti-fatigue ability.The experiment also showed that the extract of Aspongopus could improve the mating ability of mice with kidney-yang deficiency caused by hydrocortisone.Mating ability of Yang deficiency mice restored the coefficient of gonad organs and testosterone level of Yang deficiency mice,and had protective effect on testicular tissue damage.(2)The effects of different parts of the extract on the mating ability of normal mice were studied.It was found that the alcohol-precipitated part of the extract had obvious yang-strengthening effect.It could shorten the capture latency and mating latency of mice,increase the capture times and mating times of mice,thereby improving the mating ability and the content of testosterone in serum.(3)The effects of analogues of Aspongopus on the mating ability of mice with kidney-yang deficiency induced by hydrocortisone were studied.It was found that the two insects,cyclopelta parva distant,Black worm,could improve the mating ability of mice and restore the testosterone level,but the insects,Stink bug,had no obvious effect.(4)To analyze the effects of extracts of the insect and its analogues on testosterone secretion in Leydig cells of testis.Aspongopus,cyclopelta parva distant and Black worm all have the effect of promoting testosterone secretion,while the effect of total water extract is obviously stronger than that of 50% alcohol extract.The total extract of water was precipitated by 80% ethanol.The alcohol precipitation part could promote the secretion of testosterone,and the high polar component might be the active ingredient for tonifying kidney and strengthening yang.By analyzing the effect of alcohol precipitation site on the expression of related genes in c AMP/PKA signaling pathway,we found that alcohol precipitation site could promote the synthesis of testosterone by promoting the expression of St AR and other genes,thus promoting the secretion of testosterone.(5)By studying the effect of extract of Aspongopus on mating ability of rats with kidney-yang deficiency,we can see that extract of Aspongopus can restore the weight loss of gonad organs caused by hydrocortisone and repair the pathological damage of testicular tissue;Aspongopus can regulate the secretion of testosterone by regulating hypothalamus-pituitary-gonadal axis hormone pathway;Aspongopus extract group can improve mating ability of rats.By shortening the catching latency and mating latency,increasing the catching times and mating times,the catching percentage and mating percentage were increased.Especially,the effect of mid-dose is the best,which has extremely significant difference.Proteomic analysis revealed that the possible signaling pathways of the aphrogenic effect of the insect include steroids,lipid metabolism,spermatogenesis,protein transport and other pathways,as well as the coagulation mechanism.The results of proteomics were verified by Western blotting.Testosterone was regulated by up-regulating the expression of St AR,3β-HSD,CYP17a1 protein and regulating the c AMP/PKA signaling pathway.CONCLUSION: The extract of Aspongopus has anti-fatigue,warming kidney and strengthening yang effects on normal mice,Yang-deficiency mice and Yang-deficiency rats.According to the model of Leydig cells of testis,it is speculated that the possible action site is the high polar component in the alcohol precipitation site,and that the testosterone secretion may be regulated by c AMP-PKA signaling pathway and hypothalamus-pituitary-gonad axis pathway in order to warm the kidney and strengthen the yang. |
PDF Full Text Request