Objective

2

To investigate the effect of

Yishen Tongluo Formula

（益肾通络方， YSTL） on sperm DNA damage and the possible mechanism.

Methods

2

Forty ICR mice were randomly divided into control group， model group， YSTL low-dose （YSTLL）， medium-dose （YSTLM） and high-dose （YSTLH） group， with 8 mice in each group. Except for the control group， mice were given 100 mg/（kg·d） of Benzo（a）pyrene （BaP） to replicate the sperm DNA damage model.The three YSTL groups were modeled in the morning， while received the decoction of YSTL at the dose of 5.95， 11.89 and 23.78 g/（kg·d） by gavage respectively in YSTLL， YSTLM and YSTLH groups； the control group and model group received 0.1 ml/10 g of normal saline by gavage； the administration lasted for 8 weeks in all gorups. The sperm DNA fragmentation index （DFI） was detected. Transcriptome and proteome analysis were performed according to DFI results. Trend analysis， GO and KEGG analysis were performed based on the two omics results. The target genes of YSTL were screened out by nine quadrant method and verified by Western blotting.

Results

2

The DFI of the model group was significantly higher than that of the control group （

P

＜0.01）. Compared to that in the model group， the DFI significantly decreased in the YSTLH group （

P

＜0.01）， while there was not significant difference in the YSTHL and YSTHM groups （

P

＞0.05）. Therefore， the transcriptome and proteome analysis were performed in the control group， model group， and YSTLH group. A total of 1272 differential expressed genes （DEGs） and 1309 differential expressed proteins （DEPs） were screened out. Eight trend types were screened out； as profile 2 was the gene/protein which was down-regulated in the model group and restored after the intervention of YSTH， and profile 5 was the one up-regulated in the model group and restored after the intervention of YSTH. Profile 2 and 5 were selected for further analysis. GO analysis showed that development process， intracellular part and molecular function regulation ranked first in the biological processes， cellular components and molecular function categories， respectively. The top five KEGG-enriched pathways of genes of profile 2 and profile 5 were platinum drug resistance， osteoclast differentiation， various cell apoptosis， apoptosis， and nonalcoholic fatty liver disease， while proteins were mainly enriched in systemic lupus erythematosus， nitrogen metabolism， protein digestion and absorption， African trypanosomiasis and B cell receptor signaling pathway. X-ray cross complementation repair gene 1 （Xrcc1）， cytochrome b5 reductase 3 （Cyb5r3）， hemopexin 1 （Hebp1）， and serine protease 45 （Prss45） were screened out by the nine-quadrant method， and significant decreases were found of Xrcc1 and Prss45 protein expression in the testis of the model group than the control group through Western blotting verification （

P

＜0.05）. The expression of Xrcc1 in the YSTLH group was significantly up-regulated compared to the model group （

P

＜0.01）.

Conclusion

2

YSTL can repair sperm DNA damage caused by BaP exposure. The mechanism may be related to the pathways of drug resistance and apoptosis， as well as Xrcc1.