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Published on: May 27, 2015
Endocytosis‒Mediated Invasion and Pathogenicity of Streptococcus agalactiae in Rat Cardiomyocyte (H9C2)
Sharma Pooja1, Muthuirulan Pushpanathan1, Paramasamy Gunasekaran1
1Department of Genetics, School of Biological Sciences, Madurai Kamaraj University, Madurai, 625 021, India.
Insights
Streptococcus agalactiae invades cardiovascular cells via cytoskeleton-dependent endocytosis. Lipoteichoic acid (LTA) on S. agalactiae is crucial for this internalization and subsequent host cell damage, impacting cardiovascular disease patients.
Area of Science:
- * Microbiology and Molecular Pathogenesis
- * Cardiovascular Disease Research
- * Cell Biology
Background:
- * Streptococcus agalactiae infections pose a significant mortality risk for cardiovascular disease (CVD) patients, particularly those with prosthetic heart valves.
- * The precise mechanisms by which S. agalactiae contributes to CVD pathogenesis remain incompletely understood.
- * Understanding S. agalactiae-host cell interactions is critical for developing targeted therapeutic strategies.
Purpose of the Study:
- * To elucidate the pathogenic mechanisms of Streptococcus agalactiae in rat cardiomyocytes (H9C2 cells).
- * To investigate the role of bacterial internalization and host cell cytoskeleton in S. agalactiae pathogenesis.
- * To determine the contribution of lipoteichoic acid (LTA) to S. agalactiae-induced cytotoxicity and internalization.
Main Methods:
- * Utilized H9C2 cells to study S. agalactiae uptake, employing chemical inhibitors (genistein, chlorpromazine, nocodazole, cytochalasin B) to probe cellular uptake pathways.
- * Analyzed gene expression of cytoskeleton components (Acta1, Tubb5) using RT-PCR and RT-qPCR.
- * Assessed host cell viability and bacterial internalization, and investigated the role of lipoteichoic acid (LTA) by treating bacteria with benzyl penicillin and extracting LTA.
Main Results:
- * Both live and dead S. agalactiae cells were internalized by H9C2 cells, dependent on functional actin and microtubule cytoskeleton.
- * S. agalactiae infection led to increased expression of cytoskeleton genes and induced cytotoxicity in H9C2 cells.
- * Treatment with benzyl penicillin reduced S. agalactiae internalization and cytotoxicity, linked to LTA removal; purified LTA demonstrated dose-dependent cytotoxicity.
Conclusions:
- * Streptococcus agalactiae internalizes into cardiomyocytes via energy-dependent endocytic processes requiring a functional host cytoskeleton.
- * The lipoteichoic acid (LTA) of S. agalactiae is a key virulence factor mediating bacterial internalization and inducing host cell cytotoxicity.
- * These findings provide critical insights into S. agalactiae pathogenesis in the context of cardiovascular disease.
Abstract:
Streptococcus agalactiae infection causes high mortality in cardiovascular disease (CVD) patients, especially in case of setting prosthetic valve during cardiac surgery. However, the pathogenesis mechanism of S. agalactiae associate with CVD has not been well studied. Here, we have demonstrated the pathogenicity of S. agalactiae in rat cardiomyocytes (H9C2). Interestingly, both live and dead cells of S. agalactiae were uptaken by H9C2 cells. To further dissect the process of S. agalactiae internalization, we chemically inhibited discrete parts of cellular uptake system in H9C2 cells using genistein, chlorpromazine, nocodazole and cytochalasin B. Chemical inhibition of microtubule and actin formation by nocodazole and cytochalasin B impaired S. agalactiae internalization into H9C2 cells. Consistently, reverse‒ transcription PCR (RT‒PCR) and quantitative real time‒PCR (RT-qPCR) analyses also detected higher levels of transcripts for cytoskeleton forming genes, Acta1 and Tubb5 in S. agalactiae‒infected H9C2 cells, suggesting the requirement of functional cytoskeleton in pathogenesis. Host survival assay demonstrated that S. agalactiae internalization induced cytotoxicity in H9C2 cells. S. agalactiae cells grown with benzyl penicillin reduced its ability to internalize and induce cytotoxicity in H9C2 cells, which could be attributed with the removal of surface lipoteichoic acid (LTA) from S. agalactiae. Further, the LTA extracted from S. agalactiae also exhibited dose‒dependent cytotoxicity in H9C2 cells. Taken together, our data suggest that S. agalactiae cells internalized H9C2 cells through energy‒dependent endocytic processes and the LTA of S. agalactiae play major role in host cell internalization and cytotoxicity induction.
