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. 2021 Mar 29;2021:6670495. doi: 10.1155/2021/6670495

Figure 7.

Figure 7

The function of ATP6V0D2 is independent of the V-ATPase activity that induces Notch1 signaling. (a) Mannose-siATP6V0D2 conjugated vector was injected into the tail vein before IR operation, and then, the protein levels of NICD and Hes1 of macrophages were detected at 6 h after IR by western blot.(b) Injection of siATP6V0D2 (iv) or siNotch1 + siATP6V0D2 4 h before IR surgery, siCtrl as control. Then, detected the level of ALT and AST in serum at IR 6 h by ELISA. (c) Relative expression of IL1β, MCP-1, IL10, and Arg1 in liver macrophages quantitied by qPCR. (d) BMDMs were transfected with siATP6V0D2 or siCtrl for 24 h, then stimulated with LPS for 0 h or 4 h, harvested the cells to detect the protein levels of NICD and Hes1 by western blot. (e) Injection of siATP6V0D2 (iv) or siNotch1 + siATP6V0D2 4 h before IR surgery, siCtrl as control. Then, detected the mRNA levels of NLRP3 and IL18 at 6 h by RT-qPCR (representative of 4-6 experiments). (f) Macrophages at IR6h which pretreated with siATP6V0D2 or siATP6V0D2 + siNotch1 for 4 h or additional rapamycin (5 mg/kg, ip) for 1 h were harvested in vitro. The protein levels of NLRP3 and cle-IL1β in Mφs were detected by western blot (representative of three experiments).