Group A streptococcus (GAS) causes a particularly severe form of septic shock termed Streptococcal Toxic Shock Syndrome (StrepTSS) and mortalities range from 30-70% despite modern medical practices. Marked hypotension and early multi-organ failure are distinctive features and recently evidence of severely impaired cardiac function has been documented in some patients with StrepTSS and these patients have the highest mortality. Our preliminary results suggest that extremely small amounts of streptolysin O, a potent pore-forming toxin produced by GAS, causes rapid and profound changes (1-5 min) in the ability of individual heart muscle cells to contract in response to electrical stimuli. These changes were directly correlated with marked impairment of calcium handling by these cells. We hypothesize that SLO facilitates calcium entry into the cardiac cell from the external environment via its pore-forming ability. In addition, calcium is not adequately removed from the inside of the cardiac myocyte despite the presence of elaborate pathways for this purpose. This suggests that the ATP-driven enzyme that removes calcium from the cytoplasm is adversely affected by SLO. Matrix metalloproteinases (MMP) are a family of zinc-requiring enzymes that break down structural proteins such as collagen in various types of tissue including the heart. Hearts from experimental animals with GAS bacteremia express high levels of only MMP-13. While MMPs-2 and -9 have been clearly implicated in acute Gram negative septic cardiomyopathy and in the cardiac extracellular remodeling under various stresses such as hypertension and heart attacks, no such evidence exists for MMP-13. Yet disassembled collagen scaffolding cannot function efficiently as structural support for myofibrils. Thus, MMP-13, induced by bacterial virulence factors such as SLO, may uniquely drive acute cardiac dysfunction in StrepTSS. Thus, the most likely hypothesis regarding cardiac dysfunction in StrepTSS is that early in the course of infection, SLO has direct effects on heart function and that later in the course of infection, cardiomyocytes, infiltrating inflammatory cells and/or cardiac fibroblasts produce MMP-13 that further contributes to the deterioration of cardiac myocyte function. Thus the first specific aim i to determine how SLO causes an increase in intracellular Ca2+ in cardiac myocytes and a decrease in calcium reuptake into storage pools. Studies will utilize single isolated murine cardiac myocytes to measure the dynamics of intracellular Ca2+ transients in response to SLO and to a unique SLO construct that can bind cells but cannot form pores.
The second aim i s to determine what drives upregulation of cardiac MMP-13 in hearts of animals with GAS bacteremia and to elucidate the functional consequences of this upregulation. For this, whole hearts and isolated cardiac cells from both wild-type mice and those deficient in MMP-13 will be studied. MMP-13 gene expression, protein production and functional enzymatic activity will be measured by PCR, ELISA, and zymography, respectively. Mechanical functioning of individual cardiomyocytes will be assessed with an IonOptix system designed for this purpose. The long-term objective is to provide a molecular basis for cardiac dysfunction in severe infections caused by organisms that produce toxins homologous to SLO such that new targeted therapies can be developed to limit infection severity and improve survival.
Severe group A streptococcal soft tissue infections have been described in civilians of all ages and in veterans in VA hospitals and nursing homes. Mortality and morbidity of this infection remain high, especially in elderly individuals, despite appropriate antibiotic treatment and intensive care measures. Survivors frequently require several months of acute care hospitalization and rehabilitation due to consequences of shock and cardiac failure and to the loss of muscle groups or entire extremities as a result of life-saving surgical debridement. This proposal will investigate the mechanisms underlying the cardiomyopathy associated with StrepTSS. Results of these studies will define a series of potential interventions to treat, preven and attenuate the severity of illness at distinct molecular levels. Consequently, these experimental results could be translated into novel therapeutic strategies to improve the management of patients with StrepTSS, thereby shortening hospital stay, reducing morbidity and improving outcome.