巴戟散水提物协同调控凋亡通路促进梅花鹿耳缘成纤维细胞增殖OA

[Objective]This study investigated the effect of spray-dried powder of Bajisan on the proliferation of auricular marginal skin fibroblasts in Cervus nippon and its regulatory effect on the expression of apoptosis-related genes.[Method]Twenty-four adult Cervus nippon were randomly divided into control group and treatment group.The treatment group(500 mg·kg-1)was fed a diet containing Water Extract of Bajisan daily for one month.Auricular marginal skin tissue was collected after treatment completion.Primary fibroblasts were isolated and cultured using the explant attachment method.Effect of Water Extract of Bajisan on cell proliferation activity and apoptosis-related gene expression were analyzed through cell morphology observation,MTT assay for cell viability,growth curve plotting,and qPCR technology.[Result]The results showed that cells from both groups exhibited typical fibroblast morphological characteristics.The cell growth curves exhibited an'S'shape.The proliferation rate of the treatment group was significantly higher than that of the control group(P<0.05).The peak cell density in the treatment group occurred on day 4 post-seeding,earlier than in the control group(day 6).MTT assay revealed that at 72 h post-seeding for P2 and P3 generation cells,the proliferation activity in the treatment group was 1.97 times and 1.68 times higher,respectively,compared to the control group(P<0.05).qPCR analysis demonstrated that,compared to the control group,the treatment group significantly upregulated the relative mRNA expression level of the anti-apoptotic gene Bcl-2(P<0.01)and downregulated the relative mRNA expression level of the pro-apoptotic gene Bax(P<0.05),resulting in a significantly increased Bcl-2/Bax ratio(P<0.05).Furthermore,the treatment group significantly inhibited the relative mRNA expression levels of key genes in the intrinsic(mitochondrial)apoptotic pathway(Caspase-9 and Apaf1;P<0.01),as well as key genes in the extrinsic(death receptor)apoptotic pathway(Fas,TNFRSF1A and DR5)and the apoptosis execution molecules(Caspase-3 and Caspase-8;P<0.05).These results indicate that Water Extract of Bajisan can promote the proliferation of auricular marginal skin fibroblasts in Cervus nipponand significantly inhibit cell apoptosis.Its mechanism involves the synergistic regulation of both intrinsic and extrinsic apoptotic pathways,manifested by upregulating anti-apoptotic gene expression,downregulating pro-apoptotic gene expression,and inhibiting the caspase cascade reaction.[Conclusion]This research provides technical support for the construction of germplasm resource banks of endangered deer species and also offers a new perspective for the study of the mechanism of action of traditional Chinese medicine compound prescriptions in the field of cell biology.