| As a common disease in andrology,the incidence of oligoasthenozoospermia is on the rise.Because the pathological mechanism of oligoasthenozoospermia has not been fully clarified,western medicine mainly adopts the cause-based treatment.Traditional Chinese medicine pays attention to the comprehensive conditioning of macroscopic and microscopic and can exert its unique advantages where western medicine cannot reach.This study assessed the quality of systematic reviews of traditional Chinese medicine to treat oligoasthenozoospermia through a systematic review overview.Zuogui Wan,as a prescription for the treatment of oligoasthenospermia,has an excellent clinical effect,but the mechanism of action remains to be clarified.This study explored the efficacy and possible mechanism of the Zuogui Wan in treating oligoasthenozoospermia through both clinical and experimental aspects.Objective:The objectives of this study were three:1 To assess the quality of systematic reviews of Chinese medicine to treat oligoasthenozoospermia.2 To explore the effects of Zuogui Wan on semen parameters,sperm DFI and TCM syndrome scores.At the same time,we explored the correlation of age,BMI,and semen parameters with DFI.3 To explore the effect and possible mechanism of Zuogui Wan on the structure and function of spermatogenic cells.Method:1 Overview of the systematic review:This study screened from the database a systematic review or meta-analysis(SR/MA)of traditional Chinese medicine alone or in combination with western medicine in treating oligoasthenospermia.In this study,AMSTAR 2 and GRADE were used to evaluate the methodological quality and evidence level of SR/MA.The overall response rate,spouse pregnancy rate,the quality of semen,and sperm DFI were used as outcome indicators.This study assessed the quality of systematic reviews of traditional Chinese medicine alone or in combination with western medicine and summarized the evidence for their impact on fertility-related outcomes in oligoasthenozoospermia.2 Clinical research:In this study,patients with kidney-yin deficiency type oligoasthenozoospermia and abnormal sperm DFI were used as the observation objects,and Zuogui Wan was the intervention drug.This study adopts a multi-center,self-controlled before-and-after approach,and is expected to conduct a 90-day intervention in the 100 patients.The observation indicators of this study were semen routine parameters,DFI and TCM syndrome score.Patients enrolled in this study were visited every 30 days and 3-6 months after the end of treatment.3 Experimental study:In this study,spermatogonia(GC-1 spg)were used as the observation object,and the cells were randomly divided into 4 groups:blank group(C),model group(M),levocarnitine group(LEV),and Zuogui Wan Group(ZGW).The spermatogenic disorder cell model was prepared using triptolide(GTW)-containing serum.The LEV and ZGW groups were intervened by adding LEV and ZGW-containing serum based on the former.In this study,flow cytometry was used to observe cell apoptosis and cell cycle,and an electron microscope was used to monitor changes in cell ultrastructure.In this study,Western blot was used to detect the protein expression levels of PI3K,p-AKT,and Bcl-2 in each group of cells,and RT-PCR was used to detect the mRNA expression levels of PI3K,p-AKT,and Bcl-2 in each group of cells.Result:1 The results of the overview of the systematic review showed that in terms of efficacy,there is moderate-quality evidence that Chinese herbal decoctions are more effective than western medicine alone.Low-quality evidence suggests that adding traditional Chinese medicine to Western medicine can improve efficacy.There is very low-quality evidence that Chinese patent medicines are more effective than Western medicines.In terms of spouse pregnancy rate,there is moderate-quality evidence that adding kidney-tonifying traditional Chinese medicine to western medicine can help improve the pregnancy rate of the patient’s spouse.Low-quality evidence suggests that traditional Chinese medicine treatment is more effective in increasing the pregnancy rate of a patient’s spouse than Western medicine.The addition of Qilin Pills based on western medicine helps to improve the pregnancy rate of the patient’s spouse.There is very low-quality evidence that Chinese patent medicines are more effective in improving pregnancy rates than western medicines.In terms of sperm concentration,low-and very-low-quality evidence suggests that Chinese medicine is more effective in increasing sperm concentration than Western medicine.Based on western medicine,supplemented by traditional Chinese medicine,treatment helps to improve sperm concentration.There is very low-quality evidence that Chinese patent medicine is superior to western medicine in improving sperm concentration.In terms of total sperm motility,there is low-quality evidence that Chinese medicine supplemented with Western medicine can improve total sperm motility.There is very low-quality evidence that Chinese medicine improves total sperm motility than Western medicine.Low-quality evidence suggests that herbal medicines are more effective than vitamins in improving sperm DFI.2 The clinical study results showed that a total of 91 patients completed the treatment and visit.After treatment,the liquefaction status of the semen samples of the enrolled patients improved significantly.The mean value of semen volume was more remarkable than before treatment,but there was no statistical difference(P>0.05).The concentration and the total number of sperm were significantly increased(P<0.01).Sperm forward motility(PR)and total motility(PR+NP)were significantly increased(P<0.01).Sperm DFI,especially at lower levels,was significantly decreased(P<0.01).In terms of TCM syndrome score,compared with before treatment,the total score of TCM syndrome and individual TCM syndrome score of patients after treatment were significantly improved(P<0.01).The correlation coefficient between patient age and DFI was r=0.059,P=0.576>0.05,and the correlation coefficient between BMI and DFI was r=0.036,P=0.738>0.05.There was no correlation between the patient’s age,BMI,and DFI.The correlation coefficient between sperm PR value and DFI was r=-0.331,P=0.001<0.05,indicating we can initially construct a weak correlation between sperm PR and DFI and a linear regression model.The results show that R2=26.5%,and the model interpretation is not reasonable.D-W=1.939,the data are independent of each other.ANOVA analysis showed F=31.808,P=0.000<0.05,and the regression model was DFI=45.212-0.72×PR.The correlation coefficient of total sperm motility and DFI was r=-0.285,P=0.006<0.05,indicating a weak correlation between total sperm motility and DFI.We could initially construct a linear regression model between total sperm motility and DFI.The results show that R2=14.9%,the model interpretation is not reasonable;D-W=1.984,which meets the independence condition.ANOVA analysis showed F=15.567,P=0.000<0.05,and the regression model was DFI=41.611-0.426×(PR+NP).The correlation coefficient between sperm concentration and DFI was r=-0.199,P=0.059>0.05,and the correlation coefficient between total sperm and DFI was r=-0.175,P=0.098>0.05,indicating that there was no correlation between sperm concentration and total number and DFI.3 The experimental results showed that:(1)In terms of cell apoptosis,after the intervention of the C,M,LEV,and ZGW groups,respectively,compared with the C group,the apoptosis ratio of the M group and LEV group was significantly increased,and the ZGW group was significantly increased.There was no statistical difference between the groups(P=0.719).Compared with the M group,the apoptosis ratio of the LEV group and the ZGW group was significantly reduced,indicating that both LEV and ZGW can reduce the apoptosis of cells.Compared with the LEV group,apoptosis was lower in the ZGW group.(2)In terms of the cell cycle,compared with group C,the proportion of cells in the G0/G1 phase in group M was significantly decreased,there was no statistical difference in the LEV group,and the proportion of cells in the ZGW group was significantly decreased.Compared with the M group,the proportion of cells in the G0/G1 phase in the LEV and ZGW groups increased.Compared with the LEV group,the proportion of cells in the G0/G1 phase in the ZGW group was increased.Compared with group C,the percentage of the S phase was significantly higher in group M and LEV,and there was no statistical difference in group ZGW.Compared with the M group,the S-phase percentage was significantly lower in the LEV and ZGW groups.Compared with the LEV group,the percentage of the S phase was significantly lower in the ZGW group.Compared with the C group,the percentage of spermatogenic cells in the G2/M phase of the M,LEV,and ZGW groups were significantly decreased.Compared with the M group,the percentage of spermatogenic cells in the G2/M phase in the LEV and ZGW groups significantly increased.Compared with the LEV group,there was no statistical difference in the ZGW group(P=0.538).(3)In terms of the ultrastructure of cells,compared with group C,group M had significant changes in the cell membrane,matrix,organelles,and nucleus.In terms of the cell membrane,many cells were ruptured,and pseudopodia and bulges appeared around the membrane.In terms of the matrix,the main change is that the density of the matrix is reduced,and the overall structure is evacuated.In terms of organelles,mitochondria were swollen and deformed,and mitochondrial membranes even disappeared.Rough endoplasmic reticulum expansion was evident with degranulation.Golgi body hypertrophy with dilation.In the nucleus,the main changes are severe deformation and depression,with the nuclear membrane exposed.Compared with the M group,the cell state of the LEV group and the ZGW group was significantly better,and the damage to the cell membrane,organ,and nucleus was recovered.(4)In terms of protein and mRNA expression,compared with group C,the expression of PI3K in group M and LEV was significantly decreased(P<0.01),and the expression of PI3K in the ZGW group was not statistically different from that in group C(P=0.243).Compared with the M group,there was no significant difference in the expression of PI3K in the LEV group(P=0.071),and the expression of PI3K in the ZGW group was significantly increased(P=0.002<0.01).Compared with the LEV group,the expression of PI3K in the ZGW group was significantly increased(P=0.039<0.05).Compared with group C,the expression levels of p-AKT in group M,LEV,and ZGW were significantly decreased(P<0.01).Compared with the M group,the expression levels of p-AKT in the LEV and ZGW groups were significantly increased(P<0.01).Compared with the LEV group,the expression of p-AKT in the ZGW group was significantly increased(P=0.015<0.05).Compared with group C,the expression of Bcl-2 in group M,LEV,and ZGW was significantly decreased(P<0.01).Compared with group M,the expression of Bcl-2 in the LEV and ZGW groups was significantly increased(P=0.002,0.001<0.01).Compared with the LEV group,the expression of Bcl-2 in the ZGW group was significantly increased(P=0.002<0.01).Compared with group C,the mRNA expression of PI3K in group M,LEV,and ZGW was significantly decreased(P<0.01).Compared with the M group,the mRNA expression of PI3K in the LEV and ZGW groups was significantly increased(P=0.003,0.001<0.01).Compared with the LEV group,the expression of PI3K in the ZGW group was significantly increased(P=0.009<0.01).Compared with group C,the mRNA expression of p-AKT in group M,LEV,and ZGW was significantly decreased(P<0.01).Compared with the M group,the mRNA expression of p-AKT in the LEV and ZGW groups was significantly increased(P=0.004,0.002<0.01).Compared with the LEV group,the mRNA expression of p-AKT in the ZGW group was significantly increased(P=0.026<0.05).Compared with the C group,the mRNA expression of Bcl-2 in the M group and the LEV group was significantly decreased(P<0.01),but there was no statistical difference in the ZGW group(P=0.848).The mRNA expression of Bcl-2 in the group was significantly increased(P=0.005,0.003<0.01).Compared with the LEV group,the mRNA expression of Bcl-2 in the ZGW group was significantly increased(P=0.008<0.01).Conclusion:1 The results of an overview of the systematic review showed that Chinese medicine alone or in combination with western medicine was more effective in improving fertility outcomes and semen quality than western medicine alone.Overall,the level of evidence in the meta-analysis of TCM treatment of oligoasthenozoospermia is low,and the design of the original study needs to be strengthened.Sperm DFI is also an important indicator for assessing male fertility and needs more attention.2 The clinical trials showed that Zuogui Wan can significantly improve sperm motility,concentration,and the total number of oligoasthenospermia patients in terms of semen routine parameters but does not considerably improve semen volume.In sperm DFI,Zuogui Wan can reduce sperm DFI in patients,especially for patients with mild abnormal DFI levels.The improvement is pronounced.In terms of TCM syndrome scores,Zuogui Wan also performed well,and each visit had a lower score than the previous one.In terms of correlations,the level of sperm DFI was not correlated with age,BMI,sperm concentration,and the total number of included patients.Still,it was negatively associated with forwarding motility and total motility.3 The results of cell experiments showed that ZGW can reduce the damage caused by GTW,repair the ultrastructure of cells,and maintain their normal cell functions.ZGW can promote cell proliferation and reduce apoptosis,which may be achieved by regulating the cell cycle and the expression of PI3K/AKT-related proteins and mRNAs.This study provided a solid clinical basis for Zuogui Wan in treating oligoasthenozoospermia and explores its specific mechanism of improving spermatogenesis at the cellular level. |
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