Temporal and mutation-specific alterations in Ca²⁺ homeostasis differentially determine the progression of cTnT-related cardiomyopathies in murine models

Abstract

Naturally occurring mutations in cardiac troponin T (cTnT) result in a clinical subset of familial hypertrophic cardiomyopathy. To determine the mechanistic links between thin-filament mutations and cardiovascular phenotypes, we have generated and characterized several transgenic mouse models carrying cTnT mutations. We address two central questions regarding the previously observed changes in myocellular mechanics and Ca(2+) homeostasis: 1) are they characteristic of all severe cTnT mutations, and 2) are they primary (early) or secondary (late) components of the myocellular response? Adult left ventricular myocytes were isolated from 2- and 6-mo-old transgenic mice carrying missense mutations at residue 92, flanking the TNT1 NH(2)-terminal tail domain. Results from R92L and R92W myocytes showed mutation-specific alterations in contraction and relaxation indexes at 2 mo with improvements by 6 mo. Alterations in Ca(2+) kinetics remained consistent with mechanical data in which R92L and R92W exhibited severe diastolic impairments at the early time point that improved with increasing age. A normal regulation of Ca(2+) kinetics in the context of an altered baseline cTnI phosphorylation suggested a pathogenic mechanism at the myofilament level taking precedence for R92L. The quantitation of Ca(2+)-handling proteins in R92W mice revealed a synergistic compensatory mechanism involving an increased Ser16 and Thr17 phosphorylation of phospholamban, contributing to the temporal onset of improved cellular mechanics and Ca(2+) homeostasis. Therefore, independent cTnT mutations in the TNT1 domain result in primary mutation-specific effects and a differential temporal onset of altered myocellular mechanics, Ca(2+) kinetics, and Ca(2+) homeostasis, complex mechanisms which may contribute to the clinical variability in cTnT-related familial hypertrophic cardiomyopathy mutations.

Disease Pathways and Novel Therapeutic Targets in Hypertrophic Cardiomyopathy

“…The exceptions to this poison peptide basis for HCM are those MYBPC3 mutations that reduce the amount of full-length protein, resulting in insufficient protein for normal function (haploinsufficiency). 18 20 likely contribute to many aspects of HCM. In a transgenic troponin T model of HCM, the burden of ventricular arrhythmias correlated with the degree of Ca 2ϩ sensitization resulting from the mutation.…”

“…20 Moreover, studies in HCM patients with severe hypertrophy and impaired contractile reserve emphasize the putative role of SERCA2 mRNA downregulation, potentially altering Ca 2ϩ handling. 33 A logical extension of such observations is the targeting of SERCA2a, PLN, and related proteins in HCM.…”

Hypertrophic Cardiomyopathy: Genetic Alterations, Pathogenesis and Pathophysiology

“…Экспериментальные модели ГКМП выявляют изменение содержания внутриклеточного Са 2+ , включая снижение его уровня в саркоплазмати-ческом ретикулуме и увеличение концентрации кати-онов в диастолу [23,28] В последние годы накоплено множество научных доказательств, подтверждающих, что мутации белков саркомеров, в частности тонких филаментов (напри-мер, мутации кардиального тропонина Т), в действи-тельности повышают чувствительность контрактиль-ных элементов к Са 2+ и усиливают силу сокращения [29,30]. Биофизический анализ мутированных бел-ков саркомера, являющихся "гиперконтрактиль-ными", с завышенными энергетическими затратами для продукции силы сокращения, привел исследова-телей к более специфической гипотезе: молекулярная основа ГКМП представлена клеточным энергетиче-ским дефицитом, в результате неэкономной функции саркомеров [30,31].…”

“…При этом белки миозина, содержащие мутации, типичные для ГКМП, обусловливают уве-личение АТФ-азной активности, генерированной силы сокращения и усиление скольжения актиновых филаментов [14]. Мутация кардиального тропонина 2 (TNNT2) оказывает аналогичный эффект: она сопро-вождается активацией АТФ-азы и усилением силы сокращения саркомеров [23].…”

Rescue of Familial Hypertrophic Cardiomyopathy by Altering Sarcomeric Exposure and Response to Calcium

“…These variable clinical phentoypes strongly suggest that a specific FHC mutation in a contractile protein seemingly can be modified by environmental factors and modifier genes. For example, pathological symptoms associated with FHC mutations in the troponin T gene appear to be minimal; however, most of these patients die of sudden cardiac arrest (18). Patients with FHC mutations in the α-tropomyosin (α-Tm) gene exhibit a wide phenotypic range of symptoms.…”