This observation suggests that FKBP12

This observation suggests that FKBP12.6-mediated signaling is linked to physiological, rather than pathological hypertrophy,24,25consistent with FKBP12.6-deficient mice never displaying heart failure, a common end stage for the pathological hypertrophic heart. To determine whether FKBP12.6-deficient males also develop exercise-induced cardiac arrhythmia and sudden death in addition to hypertrophy, we compared conscious ECG parameters in these mutant mice and male littermates (56 month old) (Fig. are able Rabbit Polyclonal to GRAK to rescue abnormal cardiac hypertrophy and abnormal calcium release in FKBP12.6-deficient mice. == Introduction == Ca2+release from intracellular stores to the cytoplasm is regulated by calcium release channels, such as ryanodine receptors (RyR) in cardiac, skeletal, and smooth muscle cells1and inositol 1,4,5-trisphosphate receptor in many nonmuscle cell types.2RyR1 and RyR2 are required for the maintenance of skeletal and cardiac muscle calcium homeostasis and excitationcontraction coupling, respectively. The activation and inactivation of RyR is regulated by a number of channel modulators,1including FKBP12.6, calmodulin, protein LY341495 kinase A, and Ca2+/calmodulin dependent kinase (CaMKII). FK506 binding proteins (FKBPs) are immunophilins that bind to the immunosuppressive drugs FK506 and rapamycin.2FKBP12 (also known asfkbp1a) and FKBP12.6 (also known asfkbp1b) are cytoplasmic proteins that share 85% amino acid identity and a similar tissue distribution. Although FKBP12 is highly expressed in cardiomyocytes, FKBP12.6 is the predominant isoform associated with RyR2.1,35Despite their similarities in protein structure, FKBP12 and FKBP12.6 have distinct roles for normal cardiac function. These differences are clearly illustrated in mice with targeted deletion of FKBP12 or FKBP12.6. Mice homozygous for a null mutation in FKBP12 are embryonic lethal due to aberrant cardiac development.6In contrast, FKBP12.6-deficient mice display normal cardiac development and are viable. However, once mature, adult FKBP12.6-deficient mice display abnormal cardiac physiology including either enlarged hearts7or exercise-induced cardiac arrhythmias and sudden death.8Furthermore, our recent study demonstrated that FKBP12 is a key regulator for cardiac voltage-gated sodium channel.9 RyR is a large tetrameric calcium release channel and is composed of four identical subunits (560 kDa). The majority of channel modulators interact with N-terminal regulatory foot region.1FKBP12.6 has been shown to regulate RyR2 closure via its interaction with RyR2.1014Two independent mouse strains deficient in FKBP12.6 had distinctively different abnormal cardiac phenotypes. The first strain (129SvEv/C57 hybrid or 129SvEv inbred background) displayed sex-specific adult cardiac hypertrophy.7The second strain (DBA/lacJ inbred background) displayed stress/exercise-induced cardiac sudden death.8Interestingly, intracellular calcium release appeared to be altered in both FKBP12.6-deficient mouse models, suggesting that secondary genetic factors may contribute to the final pathogenic outcome. However, there are still many unanswered questions regarding the abnormal cardiac phenotypes in FKBP12.6-deficient mice. It has not been tested whether FKBP12.6-deficient mice in 129SvEv/C57 background can develop exercise-induced cardiac sudden death or whether the cardiac functional defects seen in FKBP12.6-deficient mice (in both strains) are a direct consequence of abnormal calcium handling or an indirect consequence of other altered physiological function(s), such as hypertension7or an altered immune system.15Resolving these questions will help us to further understand the physiological role of FKBP12.6 in heart function and heart failure. In this report, we determined the physiological impact of cardiomyocyte-restricted overexpression of FKBP12.6 on cardiac function and intracellular Ca2+release in cardiomyocytes. We demonstrated that MHC-FKBP12.6 mice were able to rescue abnormal cardiac hypertrophy and Ca2+release phenotype in FKBP12.6-deficient mice. These data provide further insight into the role of FKBP12.6 in Ca2+release and cardiac hypertrophy. == Materials and Methods == == Generation of MHC-FKBP12.6 Transgenic and MHC-FKBP12.6/FKBP12.6Deficient Mice == FKBP12.6-deficient mice were generated as described previously7and were maintained in 129SvEv/C57BL backgrounds. To generate MHC-FKBP12.6 transgenic mice, an -myosin heavy chain (MHC) promoter was used to drive a human FKBP12.6 cDNA (coding region), followed by an SV40 early region transcription terminator/polyadenylation site. A myc epitope tag was added in frame to the 5 end of FKBP12.6 cDNA. The myc-epitope tag allowed us to distinguish transgenic FKBP12.6 from endogenous mouse FKBP12.6 and FKBP12. MHC-FKBP12.6 mice were generated as previously described.10To generate mice in which FKBP12.6 was only LY341495 expressed in the myocardium (i.e., FKBP12.6 was not present in any other tissue), we first cross-bred MHC-FKBP12.6+mice to FKBP12.6 homozygous mutants (FKBP12.6/) to generate mice LY341495 positive for the transgene (i.e., MHC-FKBP12.6) and heterozygous for the FKBP12.6 knock-out allele (FKBP12.6+/). MHC-FKBP12.6+: FKBP12.6+/were further cross-bred to FKBP12.6/mice to generate MHC-FKBP12.6+: FKBP12.6/mice. == Morphological, Histological, and Echocardiogram Analyses == Cardiac structure and function of mice were also assessed by histological analysis and echocardiograph (ECG) as previously described.16In brief, the heart weight to body weight ratio (mg/g) was used as a measure of cardiac hypertrophy. Cardiac tissue LY341495 samples were fixed in 10% formalin in phosphate-buffered saline (pH 7.4), paraffin embedded, and sectioned (8-m thickness). Sections were further processed and stained.