Over the past decades, drug-resistant Mycobacterium tuberculosis strains have presented a significant challenge, with inadequate diagnosis of tuberculosis (TB) cases. Here, we report the draft whole-genome sequence of drug-resistant M. tuberculosis strain S7, which was isolated from a patient from Tripura, India, who was diagnosed with pulmonary TB.
A diet derived agent Curcumin (Diferuloylmethane), demonstrated its clinical application in inflammation, infection and cancer conditions. Nevertheless, its impact on the proteome of epithelial cells of non-small cell lung carcinoma (NSCLC) is yet to be explored. We employed a stable isotope labeling method for cell culture (SILAC) based relative quantitative proteomics and informatics analysis to comprehend global proteome change in A549 cells treated with curcumin and/or Lipopolysaccharide (LPS). Pretreated A549 cells were infected with Mycobacterium tuberculosis H37Rv strain to monitor bacterial load. With exposure to curcumin and LPS, out of the 1492 identified proteins, 305 and 346 proteins showed deregulation respectively. The expression of BID and AIFM1 mitochondrial proteins which play critical role in apoptotic pathway were deregulated in curcumin treated cells. Higher mitochondria intensity was observed in curcumin treated A549 cells than LPS treatment. Simultaneous treatment of curcumin and LPS neutralized the effect of LPS. Curcumin and/or LPS pretreated A549 cells infected with H37Rv, showed successful bacterial internalization. LPS treated A549 cells after infection showed increased bacterial load than curcumin compared to non-treated infected cells. However, simultaneous treatment of curcumin and LPS neutralized the effect of LPS. This study generated molecular evidence to deepen our understanding of the anti-inflammatory role of curcumin and may be useful to identify molecular targets for drug discovery.
In vitro studies involving cell lines or primary cells, provide critical insights into their physiology under normal and perturbed conditions like cancer and infection. Given that there are multiple sources of carbon, nitrogen, and other nutrients available in routinely used standard media (such as DMEM, RPMI), it is vital to quantify their contribution to cellular metabolism. 13C based Isotopic tracers of the media components can be used to kinetically track their oxidation by the cell systems such as Human Lung Carcinoma (A549) cells. In this study, a universally labelled glucose tracer ([13C6]glucose) was used to quantify its metabolic contribution that provided further insights into the central carbon metabolism of A549 cells. Gas chromatography and mass spectrometry (GC-MS) based mass isotopomer analysis (average 13C) of methanolic extracts (glycerol: 5.46±3.53 % and lactate: 74.4±2.65 %), amino acids derived from acid hydrolysates of protein (Serine: 4.51±0.21 %, Glycine: 2.44±0.31 %, Alanine: 24.56±0.59 %, Glutamate: 8.81±0.85 %, Proline: 6.96±0.53 % and Aspartate: 10.72±0.95 %) and the metabolites of the culture filtrate (glycerol: 43.14±1.45 % and lactate: 81.67±0.91 %), allowed to capture the relative contribution of glucose. We observed the Warburg effect and a significant amount of lactate contributed from glucose, was released to the media. 13C glycerol of glucogenic origin was kinetically released to the culture filtrate and might be playing a critical role in metabolic reprogramming of A549 cells. Part of the protein biomass contributed from amino acids were of glucogenic origin. Besides, the workflow adopted for 13C analysis and derived average 13C of each metabolite provided a standard methodology that could be useful in defining the metabolic phenotypes of cells in normal and perturbed conditions. Understanding precisely the altered cellular metabolism to meet the biomass demand under a range of physiological conditions, kinetically, may identify pathways for targeted and effective therapeutic interventions.
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