Smart-phone based electrocardiogram wavelet decomposition and neural network classification

This paper discusses ECG classification after parametrizing the ECG waveforms in the wavelet domain. The aim of the work is to develop an accurate classification algorithm that can be used to diagnose cardiac beat abnormalities detected using a mobile platform such as smart-phones. Continuous time recurrent neural network classifiers are considered for this task. Records from the European ST-T Database are decomposed in the wavelet domain using discrete wavelet transform (DWT) filter banks and the resulting DWT coefficients are filtered and used as inputs for training the neural network classifier. Advantages of the proposed methodology are the reduced memory requirement for the signals which is of relevance to mobile applications as well as an improvement in the ability of the neural network in its generalization ability due to the more parsimonious representation of the signal to its inputs.

Cardiac protein kinases: the cardiomyocyte kinome and differential kinase expression in human failing hearts

Aims. Protein kinases are potential therapeutic targets for heart failure, but most studies of cardiac protein kinases derive from other systems, an approach that fails to account for specific kinases expressed in the heart and the contractile cardiomyocytes. We aimed to define the cardiomyocyte kinome (i.e. the protein kinases expressed in cardiomyocytes) and identify kinases with altered expression in human failing hearts. Methods and Results. Expression profiling (Affymetrix microarrays) detected >400 protein kinase mRNAs in rat neonatal ventricular myocytes (NVMs) and/or adult ventricular myocytes (AVMs), 32 and 93 of which were significantly upregulated or downregulated (>2-fold), respectively, in AVMs. Data for AGC family members were validated by qPCR. Proteomics analysis identified >180 cardiomyocyte protein kinases, with high relative expression of mitogen-activated protein kinase cascades and other known cardiomyocyte kinases (e.g. CAMKs, cAMP-dependent protein kinase). Other kinases are poorly-investigated (e.g. Slk, Stk24, Oxsr1). Expression of Akt1/2/3, BRaf, ERK1/2, Map2k1, Map3k8, Map4k4, MST1/3, p38-MAPK, PKCδ, Pkn2, Ripk1/2, Tnni3k and Zak was confirmed by immunoblotting. Relative to total protein, Map3k8 and Tnni3k were upregulated in AVMs vs NVMs. Microarray data for human hearts demonstrated variation in kinome expression that may influence responses to kinase inhibitor therapies. Furthermore, some kinases were upregulated (e.g. NRK, JAK2, STK38L) or downregulated (e.g. MAP2K1, IRAK1, STK40) in human failing hearts. Conclusions. This characterization of the spectrum of kinases expressed in cardiomyocytes and the heart (cardiomyocyte and cardiac kinomes) identified novel kinases, some of which are differentially expressed in failing human hearts and could serve as potential therapeutic targets.

Dystrophin abnormalities in polymyositis and dermatomyositis.

The expression of dystrophin in muscle biopsies from nine cases of polymyositis, ten cases of juvenile dermatomyositis and three adults with dermatomyositis was studied by Western blot analysis and immunocytochemistry. Five antibodies corresponding to different N- and C-terminal regions of the dystrophin gene were used. Sixteen of the 22 cases (73%) showed an abnormality in the expression of dystrophin on Western blot analysis, either with a reduced molecular weight protein or a reduced amount. Immunostaining was abnormal in 11 out of 19 cases (58%) and showed varying degrees of discontinuity or loss of sarcolemmal staining. Immunolabelling of these areas with antibodies to beta-spectrin was normal implying that the changes were not caused by a loss of the sarcolemma. These results show that secondary changes in the expression of dystrophin can occur in the absence of an abnormality in the corresponding gene and that dystrophin cannot be used in isolation as a diagnostic marker for muscular dystrophy.

Endothelin-1 and fibroblast growth factors stimulate the mitogen-activated protein kinase signaling cascade in cardiac myocytes. The potential role of the cascade in the integration of two signaling pathways leading to myocyte hypertrophy.

Maximally effective concentrations of endothelin-1 (ET-1), acidic FGF (aFGF), or 12-O-tetradecanoylphorbol-13-acetate (TPA) activated mitogen-activated protein kinase (MAPK) by 3-4-fold in crude extracts of myocytes cultured from neonatal rat heart ventricles. Maximal activation was achieved after 5 min. Thereafter, MAPK activity stimulated by ET-1 or aFGF declined to control values within 1-2 h, whereas activation by TPA was more sustained. Two peaks of MAPK activity (a 42- and a 44-kDa MAPK) were resolved in cells exposed to ET-1 or aFGF by fast protein liquid chromatography on a Mono Q column. One major and one minor peak of MAPK kinase (MAPKK) was stimulated by ET-1 or aFGF. Cardiac myocytes expressed protein kinase C (PKC)-alpha, -delta, -epsilon and -zeta as shown immunoblotting. Exposure to 1 microM TPA for 24 h down-regulated PKC-alpha, -delta, and -epsilon, but not PKC-zeta. This maneuver wholly abolished the activation of MAPK on re-exposure to TPA but did not affect the response to aFGF. The effect of ET-1 was partially down-regulated. ET-1 stimulated phospho[3H]inositide hydrolysis 18-fold, whereas aFGF stimulated by only 30%. Agonists which initially utilize dissimilar signaling pathways may therefore converge at the level of MAPKK/MAPK and this may be relevant to the hypertrophic response of the heart.

Expression of protein kinase C isoforms during cardiac ventricular development.

The expression of protein kinase C (PKC) isoforms (PKC-alpha, PKC-beta 1, PKC-delta, PKC-epsilon, and PKC-zeta) was studied by immunoblotting in whole ventricles of rat hearts during postnatal development (1-26 days) and in the adult. PKC-alpha, PKC-beta 1, PKC-delta, PKC-epsilon, and PKC-zeta were detected in ventricles of 1-day-old rats, although PKC-alpha and PKC-beta 1 were only barely detectable. All isoforms were rapidly downregulated during development, with abundances relative to total protein declining in the adult to < 25% of 1-day-old values. PKC-beta 1 was not detectable in adult ventricles. The specific activity of PKC was also downregulated. The rat ventricular myocyte becomes amitotic soon after birth but continues to grow, increasing its protein content 40- to 50-fold between the neonate and the 300-g adult. An important question is thus whether the amount of PKC per myocyte is downregulated. With the use of isolated cells, immunoblotting showed that the contents per myocyte of PKC-alpha and PKC-epsilon increased approximately 10-fold between the neonatal and adult stages. In rat ventricles, the rank of association with the particulate fraction was PKC-delta > PKC-epsilon > PKC-zeta. Association of these isoforms with the particulate fraction was less in the adult than in the neonate. In primary cultures of ventricular myocytes prepared from neonatal rat hearts, 1 microM 12-O-tetradecanoylphorbol-13-acetate (TPA) elicited translocation of PKC-alpha, PKC-delta, and PKC-epsilon from the soluble to the particulate fraction in < 1 min, after which time no further translocation was observed. Prolonged exposure (16 h) of myocytes to 1 microM TPA caused essentially complete downregulation of these isoforms, although downregulation of PKC-epsilon was slower than for PKC-delta. In contrast, PKC-zeta was neither translocated nor downregulated by 1 microM TPA. Immunoblotting of human ventricular samples also revealed downregulation of PKC relative to total protein during fetal/postnatal development.

Adrenergic receptor stimulation of the mitogen-activated protein kinase cascade and cardiac hypertrophy.

Phenylephrine and noradrenaline (alpha-adrenergic agonism) or isoprenaline (beta-adrenergic agonism) stimulated protein synthesis rates, increased the activity of the atrial natriuretic factor gene promoter and activated mitogen-activated protein kinase (MAPK). The EC50 for MAPK activation by noradrenaline was 2-4 microM and that for isoprenaline was 0.2-0.3 microM. Maximal activation of MAPK by isoprenaline was inhibited by the beta-adrenergic antagonist, propranolol, whereas the activation by noradrenaline was inhibited by the alpha1-adrenergic antagonist, prazosin. FPLC on a Mono-Q column separated two peaks of MAPK (p42MAPK and p44MAPK) and two peaks of MAPK-activating activity (MEK) activated by isoprenaline or noradrenaline. Prolonged phorbol ester exposure partially down-regulated the activation of MAPK by noradrenaline but not by isoprenaline. This implies a role for protein kinase C in MAPK activation by noradrenaline but not isoprenaline. A role for cyclic AMP in activation of the MAPK pathway was eliminated when other agonists that elevate cyclic AMP in the cardiac myocyte did not activate MAPK. In contrast, MAPK was activated by exposure to ionomycin, Bay K8644 or thapsigargin that elevate intracellular Ca2+. Furthermore, depletion of extracellular Ca2+ concentrations with bis-(o-aminophenoxy)ethane-NNN'N'-tetra-acetic acid (BAPTA) or blocking of the L-type Ca2+ channel with nifepidine or verapamil inhibited the response to isoprenaline without inhibiting the responses to noradrenaline. We conclude that alpha- and beta-adrenergic agonists can activate the MEK/MAPK pathway in the heart by different signalling pathways. Elevation of intracellular Ca2+ rather than cyclic AMP appears important in the activation of MAPK by isoprenaline in the cardiac myocyte.

Stimulation of the p38 mitogen-activated protein kinase pathway in neonatal rat ventricular myocytes by the G protein-coupled receptor agonists, endothelin-1 and phenylephrine: a role in cardiac myocyte hypertrophy?

We examined the activation of the p38 mitogen-activated protein kinase (p38-MAPK) pathway by the G protein-coupled receptor agonists, endothelin-1 and phenylephrine in primary cultures of cardiac myocytes from neonatal rat hearts. Both agonists increased the phosphorylation (activation) of p38-MAPK by approximately 12-fold. A p38-MAPK substrate, MAPK-activated protein kinase 2 (MAPKAPK2), was activated approximately fourfold and 10 microM SB203580, a p38-MAPK inhibitor, abolished this activation. Phosphorylation of the MAPKAPK2 substrate, heat shock protein 25/27, was also increased. Using selective inhibitors, activation of the p38-MAPK pathway by endothelin-1 was shown to involve protein kinase C but not Gi/Go nor the extracellularly responsive kinase (ERK) pathway. SB203580 failed to inhibit the morphological changes associated with cardiac myocyte hypertrophy induced by endothelin-1 or phenylephrine between 4 and 24 h. However, it decreased the myofibrillar organization and cell profile at 48 h. In contrast, inhibition of the ERK cascade with PD98059 prevented the increase in myofibrillar organization but not cell profile. These data are not consistent with a role for the p38-MAPK pathway in the immediate induction of the morphological changes of hypertrophy but suggest that it may be necessary over a longer period to maintain the response.

The p38-MAPK inhibitor, SB203580, inhibits cardiac stress-activated protein kinases/c-Jun N-terminal kinases (SAPKs/JNKs).

SB203580 is a recognised inhibitor of p38-MAPKs. Here, we investigated the effects of SB203580 on cardiac SAPKs/JNKs. The IC50 for inhibition of p38-MAPK stimulation of MAPKAPK2 was approximately 0.07 microM, whereas that for total SAPK/JNK activity was 3-10 microM. SB203580 did not inhibit immunoprecipitated JNK1 isoforms. Three peaks of SAPK/JNK activity were separated by anion exchange chromatography, eluting in the isocratic wash (44 kDa), and at 0.08 M (46 and 52 kDa) and 0.15 M NaCl (54 kDa). SB203580 (10 microM) completely inhibited the 0.15 M NaCl activity and partially inhibited the 0.08 M NaCl activity. Since JNK1 antibodies immunoprecipitate the 46 kDa activity, this indicates that SB203580 selectively inhibits 52 and 54 kDa SAPKs/JNKs.