SERCA Cys⁶⁷⁴sulphonylation and inhibition of L-type Ca²⁺influx contribute to cardiac dysfunction in endotoxemic mice, independent of cGMP synthesis

Abstract

The goal of this study was to identify the cellular mechanisms responsible for cardiac dysfunction in endotoxemic mice. We aimed to differentiate the roles of cGMP [produced by soluble guanylyl cyclase (sGC)] versus oxidative posttranslational modifications of Ca(2+) transporters. C57BL/6 mice [wild-type (WT) mice] were administered lipopolysaccharide (LPS; 25 μg/g ip) and euthanized 12 h later. Cardiomyocyte sarcomere shortening and Ca(2+) transients (ΔCai) were depressed in LPS-challenged mice versus baseline. The time constant of Ca(2+) decay (τCa) was prolonged, and sarcoplasmic reticulum Ca(2+) load (CaSR) was depressed in LPS-challenged mice (vs. baseline), indicating decreased activity of sarco(endo)plasmic Ca(2+)-ATPase (SERCA). L-type Ca(2+) channel current (ICa,L) was also decreased after LPS challenge, whereas Na(+)/Ca(2+) exchange activity, ryanodine receptors leak flux, or myofilament sensitivity for Ca(2+) were unchanged. All Ca(2+)-handling abnormalities induced by LPS (the decrease in sarcomere shortening, ΔCai, CaSR, ICa,L, and τCa prolongation) were more pronounced in mice deficient in the sGC main isoform (sGCα1(-/-) mice) versus WT mice. LPS did not alter the protein expression of SERCA and phospholamban in either genotype. After LPS, phospholamban phosphorylation at Ser(16) and Thr(17) was unchanged in WT mice and was increased in sGCα1(-/-) mice. LPS caused sulphonylation of SERCA Cys(674) (as measured immunohistochemically and supported by iodoacetamide labeling), which was greater in sGCα1(-/-) versus WT mice. Taken together, these results suggest that cardiac Ca(2+) dysregulation in endotoxemic mice is mediated by a decrease in L-type Ca(2+) channel function and oxidative posttranslational modifications of SERCA Cys(674), with the latter (at least) being opposed by sGC-released cGMP.

Lipopolysaccharide and cytokines inhibit rat cardiomyocyte contractility in vitro

“…This is similar to the decrease in cardiac contractility that is commonly seen in animal models of SIC (22, 27, 29–31) and is consistent with the notion that cardiac dysfunction in sepsis is due to the effects of circulating mediators.…”

“…In previous experiments, this method gave identical results when compared to measuring the L-type Ca 2+ current by patch-clamp (22). …”

“…This is an important concern when one attempts to determine whether inflammatory mediators exert inhibitory effects on cardiac contractile function. Obviously, any mediators that would inhibit contractility by inhibiting SERCA (22, 26), or RyR (26, 27), for example, would not affect the contractile function of neonatal cells, and thus be reported as false negatives.…”

“…A decrease in ΔCa i is also the predominant mechanism underlying cardiac depression in many animal models of SIC (22, 27, 29–31) indicating that Cytomix incubation of ARVM does constitute a representative in vitro model for SIC. In contrast, the decrease in myofilament sensitivity, which has been reported to underlie exclusively (i.e.…”

Septic cardiomyopathy in rat LPS-induced endotoxemia: relative contribution of cellular diastolic Ca2+ removal pathways, myofibrillar biomechanics properties and action of the cardiotonic drug levosimendan

“…This is in contrast with studies in endotoxemic mice (LPS 25 lg/g, assessed after 12 h), where posttranslational modifications of SERCA Cys(674) and depressed SR Ca 2? load were found, indicative of decreased SERCA activity [15]. However, data were derived from Ca 2?…”