Slit2 Protects Hearts Against Ischemia-Reperfusion Injury by Inhibiting Inflammatory Responses and Maintaining Myofilament Contractile Properties

Abstract

Background: The secreted glycoprotein Slit2, previously known as an axon guidance cue, has recently been found to protect tissues in pathological conditions; however, it is unknown whether Slit2 functions in cardiac ischemia-reperfusion (IR) injury. Methods: Langendorff-perfused isolated hearts from Slit2-overexpressing (Slit2-Tg) mice and C57BL/6J mice (background strain) were subjected to 20 min of global ischemia followed by 40 min of reperfusion. We compared Slit2-Tg with C57BL/6J mice in terms of left ventricular function and infarct size of post-IR hearts along with tissue histological and biochemical assessments (mRNA and protein expression, phosphorylation status, and myofilament contractile properties). Results: Slit2 played cardioprotective roles in maintaining contractile function and reducing infarct size in post-IR hearts. IR increased the expression of the Slit2 receptor Robo4 and the membrane receptor Slamf7, but these increases were suppressed by Slit2 overexpression post IR. This suppression was associated with inhibition of the nuclear translocation of NFκB p65 and reductions in IL-1β and IL-18 release into perfusates. Furthermore, Slit2 overexpression attenuated the increases in myofilamentassociated PKCs and phosphorylation of cTnI at Ser43 in the post-IR myocardium. The myofilament calcium sensitivity and actomyosin MgATPase activity were preserved in the post-IR Slit2 myocardium. Conclusion: Our work demonstrates that Slit2 inhibits inflammatory responses and maintains myofilament contractile properties, thus contributing, at least in part, to the prevention of structural and functional damage during IR.

A Porcine Model of Heart Failure With Preserved Ejection Fraction Induced by Chronic Pressure Overload Characterized by Cardiac Fibrosis and Remodeling

“…Hamdani et al ( 23 ) further found that the phosphorylation levels of several titin sites were decreased, accompanied by corresponding changes in the expression levels of protein kinases and phosphatases. Our previous study revealed that myofilament phosphorylation is sensitive to all cardiac stresses ( 19 , 21 ). However, myofilament phosphorylation is quite dynamic during cardiac stress.…”

“…RNA sequencing was used to profile gene expression following DAC treatment as previously described ( 19 ). Briefly, total RNA was extracted using TRIzol reagent (cat.…”

“…Myofilament phosphorylation levels were analyzed to examine their responses to chronic pressure overload. Myofilaments were extracted as previously described, with modifications ( 19 – 21 ). The hearts were homogenized in an ice-cold cell lysis buffer comprising 60 mM KCl, 30 mM imidazole (pH 7.0), 2 mM MgCl 2 , and proteinase inhibitors, and then centrifuged at 12,000 × g at 4°C for 15 min.…”

“…Myofilament phosphorylation levels were analyzed to examine their responses to chronic pressure overload. Myofilaments were extracted as previously described, with modifications (19)(20)(21).…”

Cardioprotective Effects of n-3 Polyunsaturated Fatty Acids: Orchestration of mRNA Expression, Protein Phosphorylation, and Lipid Metabolism in Pressure Overload Hearts

“…Pathological examinations were conducted to determine the cardiac structure, fibrosis, vessel morphology, and cardiomyocyte size as previously described ( 15 , 22 ). Masson's trichrome staining was performed to identify collagen fibers and muscle tissue, whereas wheat germ agglutinin was used to visualize cardiomyocyte plasma membranes and calculate the cardiomyocyte size.…”