大鼠原代心肌细胞 z-bo 2020-12-10 374

　　Objective To explore the role of miR-199a in primary rat cardiomyocytes induced by LPS.

　　 Method The primary rat cardiomyocytes were divided into control group (NC), LPS group, LPS+miR-199a mimic group, and LPS+miR-199a inhibitor group. In this experiment, the CCK8 method was used to explore the effect of miR-199a on the cell viability of primary rat myocardium. Subsequently, ELISA was used to detect the release of miR-199a on LPS-induced inflammatory factors such as TNF-α and IL-1β in primary rat cardiomyocytes. Flow cytometry was used to detect the apoptotic protein changes of miR-199a for LPS-induced primary rat cardiomyocytes. Use Western blot experiments to study the expression of miR-199a miR-199a mimics or inhibitors, and to study the expression of miR-199a on LPS-induced p-p65, p65, p-iκBa, iκBa and apoptotic proteins in primary rat cardiomyocytes Happening. Results The CCK8 method found that compared with the NC group, LPS inhibited the vitality of rat primary myocardial cells (P<0.01), but after treatment with the analog, the primary myocardial vitality induced by LPS was more severely inhibited (P<0. 05). At the same time, compared with the mimic group, the primary cardiomyocyte viability of rats in the miR-199a inhibitor group was inhibited and decreased (P<0.01). The ELISA results showed that compared with the NC group, LPS induced the release of TNF-α and IL-1β (P<0.05). Compared with the LPS group, the miR-199a mimic group aggravated the release of TNF-α (P<0.01) and IL-1β (P<0.05), while the miR-199a inhibitor group TNF-α (P<0.01) <0.01) and IL-1β (P<0.05) release decreased. The results of flow cytometry and Western blot showed that miR-199a mimics at 50 nmol/L can increase the expression of caspase3 and bax and down-regulate the expression of bcl2 to exacerbate the apoptosis of primary rat cardiomyocytes induced by LPS (P<0. 05). Further testing of the NF-κΒ pathway found that LPS can activate the NF-κΒ pathway of primary rat cardiomyocytes through the up-regulation of p-iκBa and p-p65 expression. However, compared with the LPS group, p-p65 and p-p65 and p-p65 were treated with the mimic. The up-regulation of p-iκBa is more obvious. At the same time, after blocking miR-199a, the expression of p-p65 and p-iκBa was down-regulated compared with the mimic group. These findings indicate that treatment with mimetics can aggravate the activation of NF-κB signaling pathways in LPS-induced myocarditis in vivo. Further use of NF-κB pathway inhibitors found that miR-199a mimics did not aggravate the inhibition of LPS on the viability of primary rat cardiomyocytes, which further suggested that miR-199a may aggravate cardiomyocyte damage through the NF-κB pathway.

　　 Conclusion miR-199a can aggravate the damage of cardiomyocytes. This damage is mainly achieved through the NF-κB signaling pathway.