Genotype-Dependent and Independent Calcium Signaling Dysregulation in Human Hypertrophic Cardiomyopathy
Background—Aberrant calcium signaling may contribute to arrhythmias and adverse remodeling in hypertrophic cardiomyopathy (HCM). Mutations in sarcomere genes may distinctly alter calcium handling pathways.
Methods—We analyzed gene expression, protein levels, and functional assays for calcium regulatory pathways in human HCM surgical samples with (n=25) and without (n=10) sarcomere mutations compared with control hearts (n=8).
Results—Gene expression and protein levels for calsequestrin, L-type calcium channel, sodium-calcium exchanger, phospholamban (PLN), calcineurin, and calcium/calmodulin-dependent protein kinase type II (CaMKII) were similar in HCM compared to controls. CaMKII protein abundance was increased only in sarcomere-mutation HCM (p<0.001). The CaMKII target, pT17-PLN, was 5.5-fold increased only in sarcomere-mutation HCM (p=0.01), as was auto-phosphorylated CaMKII (p<0.01) suggestive of constitutive activation. Calcineurin (PPP3CB) mRNA was not increased, nor was RCAN1 mRNA level, indicating lack of calcineurin activation. Further, MEF2 and NFAT transcription factor activity was not increased in HCM, suggesting that calcineurin pathway activation is not an upstream cause of increased CAMKII protein abundance or activation. SERCA2A mRNA transcript levels were reduced in HCM regardless of genotype, as was SERCA2/PLN protein ratio (45% reduced, p=0.03). 45Ca SERCA uptake assay showed reduced uptake velocity in HCM regardless of genotype (p=0.01). The cardiac ryanodine receptor (RyR2) was not altered in transcript, protein, or phosphorylated (pS2808, pS2814) protein abundance, and [3H]ryanodine binding was not different in HCM, consistent with no major modification of RyR2.
Conclusions—Human HCM demonstrates calcium mishandling through both genotype-specific and common pathways. Post-translational activation of the CaMKII pathway is specific to sarcomere mutation-HCM, while SERCA2 abundance and SR Ca uptake are depressed in both sarcomere mutation-positive and negative HCM.
- hypertrophic cardiomyopathy
- calcium/calmodulin-dependent protein kinase II
- calcium regulation
- Received October 26, 2015.
- Revision received July 17, 2016.
- Accepted August 5, 2016.