The Effects Of Shengmaisan On The Lifespan Of Isp-1 And Mev-1 Mutant C.elegans And It’s Mechanism

Shengmaisan（SMS）is one of the most classical formula for treating both qi and yin deficiency in the traditional Chinese medicine.It is clinically used to treat cardiovascular diseases,diabetes and other diseases.Research shows that SMS has the pharmacological activity of anti-oxidation,anti-tumor,improving learning and memory ability and cognitive performance,it suggests that SMS has potential in anti-aging application.The previous work of SMS in our lab found that SMS significantly affected the lifespan of Caenorhabditis elegans with different genetic backgrounds,and even played the opposite role.In this study,we will base on the previous evidence obtained in our lab and use the isp-1 C.elegans with mitochondrial respiratory chain complexⅢiron-sulfur cluster protein subunit gene mutation and mev-1 worm with mitochondrial respiratory chain complex II succinate dehydrogenase subunit basic gene mutation to study the mechanism of SMS playing an opposite role on the lifespan of these two C.elegans.Our results will provide scientific data to understand the reason why SMS exhibits the opposite effect in C.elegans under different genetic backgrounds.SMS was used to treat isp-1 and mev-1 mutant nematodes,and then the tested animals in each group were subjected to transcriptome sequencing in our work.The results showed that contrast to the wild-type N2,SMS had a similar gene expression pattern for isp-1 and mev-1 mutant C.elegans.The GO and KEGG enrichment analysis of differential gene showed that after SMS treatment they enriched in some biological processes such as metabolic process,oxidation-reduction process,innate immune response,proteolysis,fatty acid degradation and etc.The oxidation-reduction process is common to all treatment groups of C.elegans.Therefore,it is necessary to further study the effect of SMS on the redox homeostasis of C.elegans.We chose H₂DCF-DA fluorescent dye to directly measure the change of ROS level in C.elegans before and after SMS treatment,and MitoSOX^TM Red was used to evaluate the effect of SMS on mitochondrial superoxide anion levels.The results showed that ROS level in isp-1 mutant C.elegans firstly increased and then decreased during the treament,and the superoxide anion level always significantly reduced.Whereas the levels of ROS and superoxide anion of mev-1 mutant C.elegans increased.ROS and superoxide anion in wild-type N2 varied similarly to isp-1 C.elegans.The levels of SOD-1,SOD-3 and GST-4 significantly increase in C.elegans after SMS treatment,but HSP-16.2 only significantly increased after heat shock.Further more,we examined the rate of oxygen consumption,ATP content and the mitochondrial fission and fusion after SMS treatment.The results showed that SMS increased the oxygen consumption rate and the ATP content of wild-type and mutant C.elegans,and partially alleviated the mitochondrial fragmented after isp-1 or mev-1RNAi.The results of this study suggest that SMS can increase ROS levels,promote the protective response,regulate mitochondrial function,exert antioxidant effect and ultimately affect worm lifespan.Our results support that SMS exerting a dual role on lifespan depends on whether the net redox level is within some certain threshold in an organism.The results of this study will provide data for deeply understanding the mechanism of effect of SMS on lifespan in mitochondrial dysfunction C.elegans,and will provide new clues for the potential clinical applications of SMS.