Ifn-γ-mediated HMGB1Effect On The Proliferation Of Mouse Mesangial Cells And Its Possible Mechanism

Objective: In this study, mouse mesangial cells and RNAi techniquewere used to explore whether HMGB1mediates IFN-γ-induced mesangial cellproliferation and the possible mechanism.Systemic lupus erythematosus (SLE) is a common spontaneous systemicautoimmune disease, and affects the skin and multiple organs. Abouttwo-thirds of the patients with systemic lupus erythematosus associated withlupus nephritis (LN) occurs, and the degree of kidney damage is to determinethe prognosis of the main indicators. At same time, the LN is also a majorcause of death of LN. But in the present study, causes of LN is not very clear.Recent evidence has revealed that many cytokines are likely involved in lupusnephritis and the unbalance between proinflammatory and anti-inflammatorycytokines plays an important role in the immune response. High mobilitygroup protein (HMGB1) was originally found as a nuclear protein andwidely involved in all aspects of the cell, such as transcription, replication.HMGB1is either released by activated cells (such as activatedmacrophages/monocytes, and erythroleukemia cells) or passively released bynecrotic or damaged cells into the extracellular milieu. Our previousexperiments revealed that the level of HMGB1mRNA and protein in theperipheral blood of patients with lupus nephritis increased, which meaned thatHMGB1is one of the important inflammatory cytokines in the pathogenesisof lupus nephritis, and related to the mesangial cell proliferation. However,the possible mechanism how HMGB1effect on cell proliferation is stilluncertain. Methods: IFN-γ was first used to stimulate mouse mesangialcells(MMC), Brdu technique was used to detect the proliferation of MMC.Furthermore, MMC was transfected with sh-RNA vector aimed to HMGB1toelucidate whether HMGB1was involved in IFN-γ-induced MMC proliferation.Finally, a mouse mesangial cell line was used to determine the effect ofHMGB1on the cell proliferation and p-Akt expression, and whether sh-RNAvector aimed to Akt inhibite the proliferation of MMC cells.Results:1IFN-γ induced the proliferation of MMCCompared with the control group, the proliferation level of MMCstimulated by IFN-γ increased from4h, it reached to the peak at8h.2The sh-HMGB1and sh-AKT vector effectively knocked down theexpression of HMGB1and AKT protein and mRNA in MMC cells.RT-PCR and westernblot showed that targeted HMGB1and AKT siRNAcould knock down cellular HMGB1and AKT expression; the non-silencingcontrol siRNA had no impact on the expression of HMGB1and AKT.3The sh-HMGB1vector effectively prevented IFN-γ-induced cellproliferation in MMC cells.Compared with the control group, IFN-γ induced cell proliferation of MMCcells. However, the level of cell proliferation decreased following transfectionof IFN-γ-stimulated MMC cells with the sh-HMGB1vector.4The expression of p-AKT protein increased after stimulated by HMGB1.Compared with control group, the expression of p-AKT significantlyincreased at45mins after stimulated by HMGB1, but there was no effect onAKT protein.5The sh-AKT vector effectively prevented HMGB1-induced cell proliferationin MMC cells.Compared with the control group, HMGB1induced the cell proliferationof MMC. In addition, the sh-AKT vector decreased the level ofHMGB1-induced cell proliferation. Conclusions:1IFN-γ could promote the proliferation level of MMC by increasing theexpression of HMGB1.2HMGB1could improve proliferation level of MMC partly by activating thePI3K-AKT signal pathway.