Gope, A. | Search bioRxiv

bioRxiv2022

Glycyrrhizin, an inhibitor of HMGB1 induces autolysosomal degradation function and inhibits H. pylori infection

Helicobacter pylori a key agent for causing gastric complications is linked with peptic ulcer, gastritis, and in severe cases gastric cancer. In response to infection, host cells stimulate autophagy to maintain cellular homeostasis. However, H. pylori have evolved the ability to usurp the hosts autophagic machinery. High mobility group box1 (HMGB1), an alarmin molecule is a regulator of autophagy and its expression is augmented in gastric cancer and many other cancers. Therefore, this study aims to explore the role of glycyrrhizin (a known inhibitor of HMGB1) in autophagy during H. pylori infection. Human gastric cancer (AGS) cells were infected with H. pylori SS1 strain and further treatment was done with glycyrrhizin. Western blot was used to examine the expression levels of autophagy proteins. Autophagy and lysosomal activity were monitored by immunofluorescence. We have performed knockdown of HMGB1 to verify the effect of glycyrrhizin by siRNA transfection method. H. pylori-infection in vivo C56BL/6 mice model was established and the effect of glycyrrhizin treatment was studied. We found that the autophagy-lysosomal pathway was impaired due to a significant increase in lysosomal membrane permeabilization during H. pylori infection in AGS cells. Subsequently, glycyrrhizin treatment restored the lysosomal membrane integrity, accompanied by an increase in cathepsin B activity and reduction of ROS and inflammatory cytokine IL-8. The recovered lysosomal function enhanced autolysosome formation and concomitantly attenuated the intracellular H. pylori growth by eliminating the pathogenic niche from gastric cells. Additionally, glycyrrhizin treatment inhibited inflammation and improved gastric tissue damages in mice.

microbiology↗

bioRxiv2021

Macrophage cell lines and murine infection by Salmonella Typhi L-form bacteria

Antibiotic resistance of pathogenic bacteria has emerged as a major threat to public health worldwide. While stable resistance due to the acquisition of genomic mutations or plasmids carrying antibiotic-resistance genes is well-established, much less is known about the temporary and reversible resistance induced by antibiotic treatment, such as the one due to treatment with bacterial cell-wall inhibiting antibiotics like ampicillin. Typically, ampicillin concentration in the blood and other tissues gradually increases over time after initiation of the treatment. As a result, the bacterial population is exposed to a concentration gradient of ampicillin. This is different from in vitro drug testing where the organism is exposed to fixed drug concentrations from the beginning till the end. To mimic the mode of antibiotic exposure of microorganisms in the tissues, we cultured the wild type, ampicillin-sensitive Salmonella Typhi Ty2 strain (S. Typhi Ty2) in the presence of increasing concentrations of ampicillin over a period of 14 days. This resulted in the development of a strain that exhibited several features of the so-called L-form of bacteria, such as the absence of cell wall, altered shape and slower growth rate compared with the parental strain. Studies on the pathogenesis of S. Typhi L-form showed efficient infection of the murine and human macrophage cell lines. More importantly, S. Typhi L-form was also able to establish infection in a mouse model to the extent comparable to its parental strain. These results suggested that L-form generation following initiation of antibiotic treatment could lead to drug escape of S. Typhi and direct spread to new cells (macrophages), which sustain the infection. Oral infection by the L-form bacteria underscores the potential of rapid disease transmission through faeco-oral route, highlighting the need for new approaches to decrease the reservoir of infection.

microbiology↗