Effect of Hemidesmus indicus R.Br. root extract against Salmonella enterica serovar Typhimurium-induced apoptosis in murine macrophage cell line (P388D1).

ABSTRACT

Background & objectives: Previous studies on natural products had mainly dealt with their antimicrobial activity and studies on the interference of these bioactive compounds with host-bacterial interaction is limited. The present study was undertaken to investigate the effect of the sterols and fatty acids present in the chloroform fraction of crude methanol extract of Hemidesmus indicus root (CHI) on Salmonella enterica serovar Typhimurium (S. Typhimurium) mediated apoptosis in a murine macrophage cell line (P388D1). Methods: Bacterial sensitivity test was carried out with different concentrations of CHI and the optimum dose was fixed as 100 μg/ml for CHI, which was safe on host cells as the CD50 (50% of cell death) dose of CHI was determined to be 500 μg/ml in the P388D1 cell line. Results: The CHI-treated bacteria had negligible cytotoxicity and were less potent to invade and proliferate intracellularly. Murine macrophages infected with wild bacteria, stained with Hoechst 33258, had swollen and damaged morphology with characteristic apoptotic bodies whereas macrophages infected with treated bacteria had comparative normal architecture. Immunofluorescence and transmission electron micrographs both confirmed that CHI-treated bacteria were defective and smaller than the wild bacteria. Ultrastructures of P388D1 cells infected with wild bacteria showed many ingested bacteria and characteristic Salmonella-containing vacuoles (SCV). Some cells had condensed or fragmented nuclei with swollen mitochondria, whereas most of the cells infected with treated bacteria were normal in morphology and a few had internalized bacteria, but the typical bacteria laden SCV was not observed in cells infected with CHI-treated S. Typhimurium. Interpretation & conclusions: Our results showed that the choloroform fraction of H. indicus root blocked the cytotoxic activity of S. Typhimurium in a macrophage cell line. More studies need to be done to elaborate and confirm our findings.