No abstract

Dominant mutations in sarcomere protein genes cause hypertrophic cardiomyopathy, an inherited human disorder with increased ventricular wall thickness, myocyte hypertrophy, and disarray. To understand the early consequences of mutant sarcomere proteins, we have studied mice (designated αMHC 403/+ ) bearing an Arg403Gln missense mutation in the α cardiac myosin heavy chain. We demonstrate that Ca 2+ is reduced in the sarcoplasmic reticulum of αMHC 403/+ mice, and levels of the sarcoplasmic reticulum Ca 2+ -binding protein calsequestrin are diminished in advance of changes in cardiac histology or morphology. Further evidence for dysregulation of sarcoplasmic reticulum Ca 2+ in these animals is seen in their decreased expression of the ryanodine receptor Ca 2+ -release channel and its associated membrane proteins and in an increase in ryanodine receptor phosphorylation. Early administration of the L-type Ca 2+ channel inhibitor diltiazem restores normal levels of these sarcoplasmic reticular proteins and prevents the development of pathology in αMHC 403/+ mice. We conclude that disruption of sarcoplasmic reticulum Ca 2+ homeostasis is an important early event in the pathogenesis of this disorder and suggest that the use of Ca 2+ channel blockers in advance of established clinical disease could prevent hypertrophic cardiomyopathy caused by sarcomere protein gene mutations. cytoplasm, sarcoplasmic reticulum (SR), and sarcomereaccount for excitation-contraction coupling. Depolarization triggers entry of small amounts of Ca 2+ through the L-type Ca 2+ channels located on the cell membrane, which in turn prompts SR Ca 2+ release by cardiac ryanodine receptors (RyR's), a process termed calcium-induced Ca 2+ release. The resulting rapid rise in cytosolic levels fosters Ca 2+ -troponin-C interactions and triggers sarcomere contraction. Activation of the ATP-dependent calcium pump (SERCA) recycles cytosolic Ca 2+ into the SR to restore sarcomere relaxation. To understand the mechanism by which calcium dysregulation occurs in αMHC 403/+ myocytes, we studied Ca 2+ -binding proteins in specific myocyte compartments and monitored the hypertrophic response to the Arg403Gln missense mutation. We report that changes in Ca 2+ -binding protein levels occur in advance of disease, and we demonstrate that restoration of these protein levels by the L-type Ca 2+ channel inhibitor diltiazem prevents clinical expression of hypertrophic cardiomyopathy in αMHC 403/+ mice. MethodsMice. αMHC 403/+ mice were generated as described (13) and were bred and maintained on the 129/SvEv genetic background. Selected mice were treated with diltiazem added to their drinking water (450 mg/l) corresponding to 1.8 mg of diltiazem per day. Short-term studies involved treatment of mice with either enalapril, atenolol, or fludrocortisone added to their drinking water to achieve a dose of 25 mg/kg/d. All mice were maintained according to protocols approved by the Institutional Animal Care and Use Committee of Harvard Medical School.RNA and protein analyses. Nor...

No abstract

Dominant mutations in sarcomere protein genes cause hypertrophic cardiomyopathy, an inherited human disorder with increased ventricular wall thickness, myocyte hypertrophy, and disarray. To understand the early consequences of mutant sarcomere proteins, we have studied mice (designated αMHC 403/+ ) bearing an Arg403Gln missense mutation in the α cardiac myosin heavy chain. We demonstrate that Ca 2+ is reduced in the sarcoplasmic reticulum of αMHC 403/+ mice, and levels of the sarcoplasmic reticulum Ca 2+ -binding protein calsequestrin are diminished in advance of changes in cardiac histology or morphology. Further evidence for dysregulation of sarcoplasmic reticulum Ca 2+ in these animals is seen in their decreased expression of the ryanodine receptor Ca 2+ -release channel and its associated membrane proteins and in an increase in ryanodine receptor phosphorylation. Early administration of the L-type Ca 2+ channel inhibitor diltiazem restores normal levels of these sarcoplasmic reticular proteins and prevents the development of pathology in αMHC 403/+ mice. We conclude that disruption of sarcoplasmic reticulum Ca 2+ homeostasis is an important early event in the pathogenesis of this disorder and suggest that the use of Ca 2+ channel blockers in advance of established clinical disease could prevent hypertrophic cardiomyopathy caused by sarcomere protein gene mutations. cytoplasm, sarcoplasmic reticulum (SR), and sarcomereaccount for excitation-contraction coupling. Depolarization triggers entry of small amounts of Ca 2+ through the L-type Ca 2+ channels located on the cell membrane, which in turn prompts SR Ca 2+ release by cardiac ryanodine receptors (RyR's), a process termed calcium-induced Ca 2+ release. The resulting rapid rise in cytosolic levels fosters Ca 2+ -troponin-C interactions and triggers sarcomere contraction. Activation of the ATP-dependent calcium pump (SERCA) recycles cytosolic Ca 2+ into the SR to restore sarcomere relaxation. To understand the mechanism by which calcium dysregulation occurs in αMHC 403/+ myocytes, we studied Ca 2+ -binding proteins in specific myocyte compartments and monitored the hypertrophic response to the Arg403Gln missense mutation. We report that changes in Ca 2+ -binding protein levels occur in advance of disease, and we demonstrate that restoration of these protein levels by the L-type Ca 2+ channel inhibitor diltiazem prevents clinical expression of hypertrophic cardiomyopathy in αMHC 403/+ mice. MethodsMice. αMHC 403/+ mice were generated as described (13) and were bred and maintained on the 129/SvEv genetic background. Selected mice were treated with diltiazem added to their drinking water (450 mg/l) corresponding to 1.8 mg of diltiazem per day. Short-term studies involved treatment of mice with either enalapril, atenolol, or fludrocortisone added to their drinking water to achieve a dose of 25 mg/kg/d. All mice were maintained according to protocols approved by the Institutional Animal Care and Use Committee of Harvard Medical School.RNA and protein analyses. Nor...