Tumor necrosis factor receptor associated factor 3 regulated the signaling pathway of STING/IFN-I for improving the lupus nephritis related podocyte injury and understanding its mechanism

Objective  To explore the effect and mechanism of TRAF3 on IgG-induced apoptosis and inflammation of human podocyte cell (HPC).

Methods  HPC cells were cultured in vitro and induced with purified IgG from lupus nephritis (LN) patients to create an inflammatory environment for 24h. The cells were divided into six groups of control, IgG-LN, IgG-LN plus siRNA, IgG-LN plus si-TRAF3, IgG-LN plus STING inhibitor H-151 (1 μmol/L) and IgG-LN plus si-TRAF3 and STING activator ADU-S100 (10 μmol/L). CCK-8 assay was utilized for detecting HPC cell viability; Flow cytometry for examining cell apoptosis; real-time polymerase chain reaction (RT-PCR) for measuring IFN-α mRNA level in HPC cells in each group; enzyme-linked immunosorbent assay (ELISA) for measuring the levels of interferon-alpha (IFN-α), interleukin-6 (IL-6), interleukin-1 beta (IL-1β) and tumor necrosis factor-alpha (TNF-α); Western blot for detecting the expressions of TRAF3 and STING protein; co-inmunoprecipitation (Co-IP) for verifying the interaction between TRAF3 and STING.

Results  TRAF3 silencing inhibited IgG-induced cell apoptosis and secretion of inflammatory factors (P<0.01). TRAF3 inhibited IgG-induced activation of STING/IFN-I pathway through conjugating with STING (P<0.01). Blunting of STING/IFN-I pathway improved IgG-induced cell apoptosis and secretion of inflammatory factors (P<0.01). Activation of STING/IFN-I pathway reversed the relieving effect of TRAF3 silencing on cell apoptosis and inflammation.

Conclusions  RAF3 silencing may mute STING/IFN-I activation and alleviate serum IgG induced cell apoptosis and inflammatory responses in LN patients.