The role of myeloid cells such as neutrophils in providing host defense to microbial infections is well- established; however, the contribution of monocytes/macrophages (M?) to the pathophysiology of bacterial endophthalmitis is less clear. Our preliminary studies revealed that M? depletion results in increased inflammatory mediators at the resolution phase, suggesting their involvement in the resolution of endophthalmitis. The M? perform multiple tasks, including sensing pathogens, tissue repair, and, in response to host-derived mediators, they differentiate into distinct functional phenotypes; a feature termed plasticity. The classically activated M? (M1) produce inflammatory cytokines and nitric oxide, contributing to host tissue damage. Conversely, the alternatively activated M? (M2) mediate tissue repair through the elimination of damaged cells/tissue and the production of anti-inflammatory molecules to resolve inflammation. Therefore, understanding the mechanisms governing the phenotypic switch of M? can be utilized to develop novel therapeutic strategies. Our transcriptome and metabolomics analyses of the bacteria-infected retina directed us to the identification of adenosine monophosphate-activated protein kinase (AMPK), a metabolic gene, which modulates the infiltrating myeloid cell phenotype in endophthalmitis. We discovered that mice with global deletion (knockout) of AMPK?1 (KO) developed severe endophthalmitis and pathology compared to wild type (WT) mice. M? lacking AMPK?1 maintained a low metabolic state, even in the hyper-inflammatory state. To precisely examine the role of AMPK in myeloid cells, we induced endophthalmitis in myeloid cell specific KO of AMPK?1 (LysM-KO) and observed that LysM-KO displayed exacerbated inflammation and reduced retinal function compared to WT mice, suggesting an essential role of AMPK in myeloid cells in the pathogenesis of bacterial endophthalmitis. Building on these findings, we propose to test our central hypothesis that AMPK exerts protective effects in bacterial endophthalmitis by modulating the polarization of infiltrating monocytes/M? to promote inflammation resolution and that metabolic reprograming is an underlying mechanism of the monocytes/M? phenotype switch. To test our hypothesis, in Aim 1, we will investigate the mechanisms underlying reduced AMPK activity in bacterial endophthalmitis by examining the modification of LKB1 via nitrosylation or chemical adduct formation.
Aim 2 tests the hypothesis that AMPK?1 ablation enhances the activation state of myeloid cells and maintains their proinflammatory (M1) state during the resolution phase of the disease.
In Aim 3, we will decipher the bioenergetic events, regulated by AMPK in M?, that polarize and maintain their pro-inflammatory nature. The anticipated results of this study will demonstrate that defective AMPK activity in myeloid cells, mainly in monocytes/M?? impacts the resolution of endophthalmitis via regulation of cellular metabolism. Also, it may provide novel therapeutic targets for the development of anti- inflammatory therapies for endophthalmitis and other microbial infections.
This study will use a mouse model of bacterial endophthalmitis and cultured monocytes/macrophages to study the role of a metabolic gene AMP-activated protein kinase (AMPK) in regulating innate responses in endophthalmitis, a devastating complication of ocular trauma and/or surgery. In light of an aging U.S. population and the increasing prevalence of antibiotic-resistant bacterial pathogens causing ocular infections, this study is of paramount importance, as it may lead to the development of new anti-inflammatory therapeutics for the prevention and/or treatment of bacterial endophthalmitis.
| Singh, Pawan Kumar; Khatri, Indu; Jha, Alokkumar et al. (2018) Determination of system level alterations in host transcriptome due to Zika virus (ZIKV) Infection in retinal pigment epithelium. Sci Rep 8:11209 |
| Guest, John M; Singh, Pawan Kumar; Revankar, Sanjay G et al. (2018) Isavuconazole for Treatment of Experimental Fungal Endophthalmitis Caused by Aspergillus fumigatus. Antimicrob Agents Chemother 62: |
