| Objective By observing the effects of Shu-Mu Brain-Kidney acupuncture points and Distant-approximal acupuncture points electro-acupuncture therapy on rats with oligo-weak spermatozoospermia model,the effects of two different matching points on the improvement of reproductive function of rats with hypothalamic-pituitary-testicular axis dysfunction and related mechanisms were explored.Methods1.Animals and grouping: 60 SPF-grade 8-week-old SD male rats were randomly divided into six groups: control group(C),model group(M),acupuncture method with Shu-Mu Brain-Kidney point(S),acupuncture method with distant-approximal point(D),positive drug group(P),and non meridian,each group of 10 animals.2.Modeling and Intervention: The C group was given saline gavage daily,and the remaining groups were modeled with 20 mg/100 g of adenine gavage for 28 days.After successful modeling,electroacupuncture intervention was performed according to two different acupuncture point methods,once a day for 30 mins each time.The first one is the Shu-Mu Brain-Kidney method,in which the 3 lines of the frontal area,Kidney Shu,Jingmen and Taixi are selected;the second one is the distant-approximal method,in which Zhongji,Guanyuan,Sansanli and Sanyinjiao are selected.10 m L/kg of levocannidine oral solution is given to the P group by gavage,and non-acupuncture points 1,2 and 3 are given to the N group.The operation was the same as that of the electroacupuncture group,and each group was intervened for 28 consecutive days.3.After 28 days of intervention,both kidneys and testes of rats in each group were weighed and the organ coefficients were calculated;epididymal spermatozoa were taken to detect their number,density,viability and vitality;HE staining was performed to observe the morphological changes of testicular pathology;ELISA was performed to detect serum sex hormones testosterone(T),gonadotropin-releasing hormone(Gn RH),follicle-stimulating hormone(FSH),luteinizing hormone(LH),estradiol(E2),luteinizing hormone(E2),and luteinizing hormone(LH).estradiol(E2),inhibin B(INHB),and prolactin(PRL);Western Blot detected the expression levels of TGF-β,P-smad2,and P-smad3 in testicular and hypothalamic tissues.The experimental data were analyzed by SPSS26.0 statistical software.Results1.The rats in group C were in good spirits,responsive to external stimuli,normal diet and urine and stool,while rats in group M were depressed and tired,unresponsive,with reduced diet,yellow hair,bowed back and curled up,unformed stool and large amount of urine,which were basically consistent with the Chinese medicine model of "kidney yang deficiency".Compared with the M group,the rats in each treatment group gradually improved their mental status,their fur color gradually regained its luster,their body weight increased,their stools gradually formed,and their urine volume relatively decreased.2.Histopathological changes of rat testes: HE staining showed that the walls of the spermatogenic tubules in group C were round and thick,and spermatogenic cells at all levels were visible in the lumen structure,and the interstitial structure was clear and intact;in group M,the spermatogenic tubules were atrophied and degenerated,and the walls of the tubules became thin,and the spermatogenic cells at all levels were obviously reduced.Compared with group M,testicular spermatogenic epithelial cells in groups S and D were restored,but still loosely arranged,and the number of spermatogenic cells at all levels was obviously restored;testicular tissues in group P were relatively improved,spermatogenic tubules were relatively full,and the structure tended to be intact;testicular varicose seminiferous tubules in group N had different degrees of atrophy and deformation,and the spermatogenic cells were disordered at different levels.3.Analysis of rat organ coefficients: adenine modeling caused damage to the kidneys of rats,and the left and right kidney coefficients of rats in the M group were higher than those in the C group(p<0.05);both acupuncture methods had significant restorative effects on the kidneys of rats,and the left and right kidney coefficients of rats in both groups were significantly lower than those in the M group(p<0.05),while there was no significant difference between the P group and the M group(p >0.05),indicating that the two acupuncture treatments The improvement effect on renal coefficient was better than that of levocannabinoids;there was no significant difference in renal coefficients after the two needle interventions(p >0.05).The absence of significant changes in rat testicular coefficients may be related to the short modeling time,which did not cover the whole rat spermatogenic cycle.4.Comparison of rat semen quality: adenine modeling led to a decrease in rat semen quality,which was significantly lower in group M than in group C(p<0.05),indicating that the modeling of oligozoospermia was successful;the Shu-Mu Brain-Kidney method,distant-approximal acupuncture point method and levocannabinoid had some degree of recovery on rat semen quality,which was higher in groups S,D and P than in group M(p <0.05);sperm density in group S The sperm density in group S was higher than that in group D and group P(p<0.05),indicating that the restoration of sperm density was best with the use of Shu-Mu Brain-Kidney and the matching points.5.Comparison of sex hormone levels of rats in each group: adenosine modelling had some effect on the sex hormone levels of rats,and the levels of T,Gn RH and INHB in group M were lower than those in group C(p<0.05),while the levels of FSH,LH,PRL and E2 were higher than those in group C(p<0.05).Both acupuncture and levocannidine promoted T,Gn RH,INHB secretion in rats a nd higher than group M(p<0.05),and positive feedback regulated hypothalamic-pituita ry-testicular axis and promoted testicular spermatogenic function.It also reduced the a bnormal secretion of FSH,LH,and E2 and lower than the M group(p<0.05),and the distant-approximal acupuncture method and levocarnitine reduced PRL and lower than the M group(p<0.05),with negative feedback regulating the hypothalamic-pituitary-t esticular axis to achieve normal sex hormone levels and maintain gonadal axis balanc e.Compared with levocarnitine,the distant-approximal acupuncture point method was more effective in promoting T secretion(p < 0.05),and the Shu-Mu Brain-Kidney matching acupuncture point method was more effective in reducing abnormally high Gn RH and INHB(p < 0.05).The regulation of hormone levels by the distant-approximal acupuncture points method was better than that by the Shu-Mu Brain-Kidney acupuncture points method.PRL and LH in group D were lower than those in group S(p < 0.05),and the distant-approximal acupuncture points method reduced the abnormally increased hormone levels and regulated gonadal axis function;Gn RH and INHB were higher than those in group S(p < 0.05),and distant-approximal acupuncture points method enhanced the pulse signal and regulated reproductive function.6.Changes in expression of TGF-β /smads signaling pathway-related proteins in hypothalamus and testis: TGF-β/smads signaling pathway conduction was abnormal after successful modeling,and TGF-β,p-smad2,and p-smad3 in testis and hypothalamus were significantly higher in group M than in group C(p < 0.05).After intervention by different methods,TGF-β,p-smad2,and p-smad3 in testis and hypothalamus tissues were lower in each treatment group than in the M group(p < 0.05).There was no significant difference between TGF-β in testis and hypothalamus in group S and group D compared with group P(p > 0.05).Comparison of the two acupuncture methods revealed that p-smad3 in testicular tissue was lower in group S than in group D(p < 0.05),indicating that the intervention effect of the Shu-Mu Brain-Kidney matching acupuncture method on abnormal phosphorylation of smad3 was more pronounced than that of thedistant-approximal acupuncture method.There was no significant difference between the two acupuncture methods in hypothalamic tissues(p > 0.05).Conclusion1.Both the Shu-Mu Brain-Kidney Acupuncture Point Method and the Distant-approximal Acupuncture Point Method had significant improvement and regulation effects on organ coefficients,semen quality and sex hormone levels in rats.Among them,the effect of the Shu-Mu Brain-Kidney Point Method on improving sperm density was better,and the effect of the Distant-approximal Point Method on regulating hormone levels was more obvious,reflecting the advantages of different acupuncture methods in treating oligozoospermia.2.The possible mechanism of the intervention of the Shu-Mu Brain-Kidney Acupuncture Point Method and the Distant-approximal Acupuncture Point Method in rats with oligozoospermia is to influence the testicular spermatogenesis process through the common TGF-β/smads signaling pathway in the hypothalamic-pituitary-testicular axis,thus achieving the improvement of oligozoospermia. |
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