Monday, May 6, 2024
5/6/2024
PROJECT SUMMARY: It has been well recognized that individuals with type 2 diabetes (T2DM) are increasingly susceptible to Mycobacterium tuberculosis (Mtb) infection. The tripeptide, glutathione (GSH) protects all cells against oxidizing agents, free radicals and reactive oxygen intermediates (ROI), either directly or through enzymatic action of GSH peroxidases and GSH-transferases. Interestingly, mycobacteria do not synthesize GSH. Rather, they produce mycothiol in millimolar amounts. We observed that the virulent laboratory strain of Mtb, H37Rv is sensitive to physiological concentrations of GSH (5mM) when grown in vitro. We also found that enhancing the levels of GSH in human macrophages resulted in inhibition in the growth of intracellular Mtb. Thus, GSH has direct antimycobacterial activity, functioning as an effector molecule in innate defense against Mtb infection. These results unfold a novel and potentially important innate defense mechanism adopted by human macrophages to control Mtb infection. We also reported that GSH in combination with cytokines such as IL-2 and IL-12 enhances the functional activity of natural killer cells to inhibit the growth of Mtb inside human monocytes. We then demonstrated that GSH activates the functions of T lymphocytes to control Mtb infection inside human monocytes. These results indicate that GSH inhibits the growth of Mtb by both direct antimycobacterial effects as well as by enhancing the functions of immune cells. Finally, we demonstrated that the levels of GSH are significantly compromised in the plasma, monocytes and red blood cells (RBCs) derived from individuals with T2DM and this decrease is due diminished levels of GSH de novo synthesis enzymes and due to increased production of free radicals. These results led us to hypothesize that GSH- deficiency in the lung parenchyma can impair the formation of granuloma and dampen the granulomatous effector responses against Mtb infection, thereby favoring bacterial replication and pathogenesis. We will test our hypothesis by performing in vivo infection studies in GSH-deficient diabetic mice (db/db) and in glutamate cysteine ligase modifier subunit knock-out (GCLM-/-) mice. SPECFIC AIMS: SPCIFIC AIM 1: To determine the effects of GSH deficiency in the lung parenchyma in altering the innate and adaptive immune responses against Mtb infection. SPCIFIC AIM 2: To assess the effects of GSH restoration in enhancing the granulomatous responses against Mtb infection. SPCIFIC AIM 3: To determine the synergistic effects of liposomal GSH in conjunction with suboptimal concentration of Rifampicin in clearing Mtb infection. Studies proposed in this application will enable us to characterize the underlying mechanisms by which GSH deficiency in the lung parenchyma leads to altered immune responses in the granulomas resulting in increased susceptibility to Mtb infection and tap the potential use of GSH as a possible immunomodulatory agent. The proposed research will therefore provide compelling evidence on whether appropriate GSH-enhancing agents can be used to prevent the development of active TB in individuals with T2DM.
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