| Objective To study the effect and mechanism of Shenggu Zaizao Pill (SGZZP) on MLO-Y4 osteoid cells by inhibiting TNF-α-regulated programmed necrosis. Methods CCK8 assay was used to detect and select the optimal concentration of serum containing SGZZP (10%, 20%, 40%, 60%, 100%) for TNF-α-induced MLO-Y4 cell proliferation in vitro. The effect of the optimal concentration of SGZZP on the proliferation of MLO-Y4 cells in vitro under different intervention conditions was detected with CCK8 assay. Flow cytometry was used to detect the apoptosis of MLO-Y4 osteoid cells under various conditions. The mRNA of RIP1, RIP3, MLKL and Caspase 8 associated with programmed necrosis was detected with real-time quantitative PCR. The protein levels of RIP1, RIP3, MLKL, and Caspase 8 associated with programmed necrosis in MLO-Y4 cells were detected with Western blotting. Results Compared to the the blank group, SGZZP group promoted the proliferation rate of MLO-Y4 cells most obviously when the dose concentration was 40%. Compared to the model group, SGZZP promoted the proliferation of MLO-Y4 cells, and inhibited the apoptosis of MLO-Y4 cells. Compared to the blank group, TNF-α promoted mRNA levels of RIP1, RIP3, MLKL, and Caspase 8. Compared to the model group, SGZZP inhibited the protein expressions of RIP1, RIP3, MLKL, and Caspase 8. Conclusion SGZZP may regulate the axis of RIP1/RIP3-MLKL signaling pathway by inhibiting the expression of TNF-α. It regulates the programmed necrosis RIP1/RIP3-MLKL signaling pathway by inhibiting TNF-α, thereby reducing the intracellular RIP1, RIP3, and MLKL and promoting the proliferation of MLO-Y4 cells. |