Ryanodine Receptor Calcium Leak in Circulating B-Lymphocytes as a Biomarker in Heart Failure

Abstract

Background: Advances in congestive heart failure (CHF) management depend on biomarkers for monitoring disease progression and therapeutic response. During systole, intracellular Ca²⁺ is released from the sarcoplasmic reticulum into the cytoplasm through type-2 ryanodine receptor/Ca²⁺ release channels. In CHF, chronically elevated circulating catecholamine levels cause pathological remodeling of type-2 ryanodine receptor/Ca²⁺ release channels resulting in diastolic sarcoplasmic reticulum Ca²⁺ leak and decreased myocardial contractility. Similarly, skeletal muscle contraction requires sarcoplasmic reticulum Ca²⁺ release through type-1 ryanodine receptors (RyR1), and chronically elevated catecholamine levels in CHF cause RyR1-mediated sarcoplasmic reticulum Ca²⁺ leak, contributing to myopathy and weakness. Circulating B-lymphocytes express RyR1 and catecholamine-responsive signaling cascades, making them a potential surrogate for defects in intracellular Ca²⁺ handling because of leaky RyR channels in CHF.

Methods: Whole blood was collected from patients with CHF, CHF following left-ventricular assist device implant, and controls. Blood was also collected from mice with ischemic CHF, ischemic CHF+S107 (a drug that specifically reduces RyR channel Ca²⁺ leak), and wild-type controls. Channel macromolecular complex was assessed by immunostaining RyR1 immunoprecipitated from lymphocyte-enriched preparations. RyR1 Ca²⁺ leak was assessed using flow cytometry to measure Ca²⁺ fluorescence in B-lymphocytes in the absence and presence of RyR1 agonists that empty RyR1 Ca²⁺ stores within the endoplasmic reticulum.

Results: Circulating B-lymphocytes from humans and mice with CHF exhibited remodeled RyR1 and decreased endoplasmic reticulum Ca²⁺ stores, consistent with chronic intracellular Ca²⁺ leak. This Ca²⁺ leak correlated with circulating catecholamine levels. The intracellular Ca²⁺ leak was significantly reduced in mice treated with the Rycal S107. Patients with CHF treated with left-ventricular assist devices exhibited a heterogeneous response.

Conclusions: In CHF, B-lymphocytes exhibit remodeled leaky RyR1 channels and decreased endoplasmic reticulum Ca²⁺ stores consistent with chronic intracellular Ca²⁺ leak. RyR1-mediated Ca²⁺ leak in B-lymphocytes assessed using flow cytometry provides a surrogate measure of intracellular Ca²⁺ handling and systemic sympathetic burden, presenting a novel biomarker for monitoring response to pharmacological and mechanical CHF therapy.

Keywords: biomarker; calcium; heart failure; ion channels.