Targeting TCA cycle metabolites by small molecule inhibitors ameliorate LPS induced immune tolerance in Macrophages through epigenetic mechanisms

Abstract

Abstract Background Severe and chronic infections, such as sepsis and Tuberculosis, result in macrophage immune tolerance in order to prevent pathologic inflammation. Immune tolerance is believed to be a result of metabolic dysregulation, especially of the TCA cycle. Metabolites play a critical role in regulating epigenetic mechanism, but many aspects remain poorly understood. Methods An in-vitro LPS tolerance model was used to evaluate if manipulating the TCA cycle could mitigate immune tolerance. Intracellular signaling, DNA methylation, and nuclear localization were measured using RPPA, Illumina EPIC, and confocal microscopy, respectively. Results Metabolic inhibitors of glycolysis (metformin), the TCA cycle (BAY 1436032), and mTOR (rapamycin and everolimus) blocked LPS-induced immune tolerance. LPS treatment changed the methylome with hypermethylation of key genes of the TCA cycle including PDP2, NDUFS8, TRAP1 and PDK2 and lead to increased nuclear localization of the TCA enzymes, which could be ameliorated with inhibitors. Increased levels of key mediators of the TCA by LPS treatment including p-mTOR, pAKT, LDHA, IDH1, LC3A and epigenetic regulator p300 was significantly downregulated by pre-treatment with metformin, everolimus and BAY 1436032. BAY 1436032 also significantly reduced the LPS induced upregulation of epigenetic enzymes including histone deacetylases (HDAC) 3,4, and EZH2. Conclusion Previous studies demonstrated that LPS induces epigenetic-mediated immune tolerance. Here we show this can be mitigated by inhibiting glycolysis, mTOR pathway and TCA cycle. This could have clinical implications in treatment of sepsis and Tuberculosis among other diseases. Supported by NIAID K23 AI141681-02