—Troponin I is a subunit of the thin filament–associated troponin-tropomyosin complex involved in calcium regulation of skeletal and cardiac muscle contraction. We deleted the cardiac isoform of troponin I by using gene targeting in murine embryonic stem cells to determine the developmental and physiological effects of the absence of this regulatory protein. Mice lacking cardiac troponin I were born healthy, with normal heart and body weight, because a fetal troponin I isoform (identical to slow skeletal troponin I) compensated for the absence of cardiac troponin I. Compensation was only temporary, however, as 15 days after birth slow skeletal troponin I expression began a steady decline, giving rise to a troponin I deficiency. Mice died of acute heart failure on day 18, demonstrating that some form of troponin I is required for normal cardiac function and survival. Ventricular myocytes isolated from these troponin I–depleted hearts displayed shortened sarcomeres and elevated resting tension measured under relaxing conditions and had a reduced myofilament Ca sensitivity under activating conditions. The results show that (1) developmental downregulation of slow skeletal troponin I occurs even in the absence of cardiac troponin I and (2) the resultant troponin I depletion alters specific mechanical properties of myocardium and can lead to a lethal form of acute heart failure.

Several cardiac troponin I (cTnI) mutations are associated with restrictive cardiomyopathy (RCM) in humans. We have created transgenic mice (cTnI193His mice) that express the corresponding human RCM R192H mutation. Phenotype of this RCM animal model includes restrictive ventricles, biatrial enlargement and sudden cardiac death, which are similar to those observed in RCM patients carrying the same cTnI mutation. In the present study, we modified the overall cTnI in cardiac muscle by crossing cTnI193His mice with transgenic mice expressing an N-terminal truncated cTnI (cTnI-ND) that enhances relaxation. Protein analyses determined that wild type cTnI was replaced by cTnI-ND in the heart of double transgenic mice (Double TG), which express only cTnI-ND and cTnI R193H in cardiac myocytes. The presence of cTnI-ND effectively rescued the lethal phenotype of RCM mice by reducing the mortality rate. Cardiac function was significantly improved in Double TG mice when measured by echocardiography. The hypersensitivity to Ca2+ and the prolonged relaxation of RCM cTnI193His cardiac myocytes were completely reversed by the presence of cTnI-ND in RCM hearts. The results demonstrate that myofibril hypersensitivity to Ca2+ is a key mechanism that causes impaired relaxation in RCM cTnI mutant hearts and Ca2+ desensitization by cTnI-ND can correct diastolic dysfunction and rescue the RCM phenotypes, suggesting that Ca2+ desensitization in myofibrils is a therapeutic option for treatment of diastolic dysfunction without interventions directed at the systemic β-adrenergic-PKA pathways.

BackgroundPrenatal alcohol exposure may cause cardiac development defects, however, the underlying mechanisms are not yet clear. In the present study we have investigated the roles of histone modification by curcumin on alcohol induced fetal cardiac abnormalities during the development.Methods and resultsQ-PCR and Western blot results showed that alcohol exposure increased gene and active forms of caspase-3 and caspase-8, while decreased gene and protein of bcl-2. ChIP assay results showed that, alcohol exposure increased the acetylation of histone H3K9 near the promoter region of caspase-3 and caspase-8, and decreased the acetylation of histone H3K9 near the promoter region of bcl-2. TUNEL assay data revealed that alcohol exposure increased the apoptosis levels in the embryonic hearts. In vitro experiments demonstrated that curcumin treatment could reverse the up-regulation of active forms of caspase-3 and caspase-8, and down-regulation of bcl-2 induced by alcohol treatment. In addition, curcumin also corrected the high level of histone H3K9 acetylation induced by alcohol. Moreover, the high apoptosis level induced by alcohol was reversed after curcumin treatment in cardiac cells.ConclusionsThese findings indicate that histone modification may play an important role in mediating alcohol induced fetal cardiac apoptosis, possibly through the up-regulation of H3K9 acetylation near the promoter regions of apoptotic genes. Curcumin treatment may correct alcohol-mediated fetal cardiac apoptosis, suggesting that curcumin may play a protective role against alcohol abuse caused cardiac damage during pregnancy.

Protein kinase C (PKC) is the target for a number of tumor promoters. The mechanism underlying the promoting effects of bile acids in colorectal cancer is not understood. We report that sodium deoxycholate (DOC) triggered activation of PKC in physiological conditions. The biphasic effects of DOC upon PKC activation were Ca(2+)-stimulated and did not require phosphatidylserine (PtdSer) as phospholipid co-factor. The optimal rate of activation was obtained at 0.4 mM DOC and reached approximately half the maximal rate of activation obtained in the presence of PtdSer. Similarly to PtdSer, DOC supported diacylglycerol- as well as phorbol-ester-mediated PKC activation. The reciprocal effects of PtdSer and DOC upon PKC in either 0.5 mM CaCl2 or 0.5 mM EGTA suggest that DOC interacts with the phospholipid-binding domain to elicit PKC activation. DOC-supported enzyme activation exhibited substrate specificity different from that of PtdSer-supported enzyme activation. All tested primary and secondary bile acids activated PKC to various extents, with DOC being the most potent. We suggest that amphipathic bile acids acting in a PtdSer-like manner provide the hydrophobic environment required for PKC activation. Treatment of 32P-labeled platelets and colonic cells HT29 Cl.19A with DOC enhanced the phosphorylation of endogenous substrates for PKC. Colonic cells responsive at 50 microM DOC, appeared to be 10-fold more sensitive than platelets. We suggest that direct or indirect activation of PKC by bile acids may account for the promoting effects of these non-phorbol-ester-type tumor promoters.