Biomedical subjects

Tushi Singal

Publications and source records attributed to Tushi Singal.

5 recordsLinked to original sources

PubMed2006

Norepinephrine-induced changes in gene expression of phospholipase C in cardiomyocytes.

Previously, we have reported that norepinephrine (NE)-mediated cardiac hypertrophy may occur due to stimulation of alpha1-adrenoceptors and phospholipase C (PLC) activity. Since the signal transduction mechanisms involving PLC isozymes in cardiomyocytes are not well established, the present study was conducted to test the hypothesis that stimulation of cardiac PLC activity by NE increases the gene expression for PLC isozymes via a PKC and ERK 1/2-dependent pathway. For this purpose, mRNA levels for PLC beta1, beta3, gamma1, and delta1 isozymes were determined in isolated adult rat cardiomyocytes upon incubation in the absence and presence of NE. The NE-induced increases in PLC isozyme mRNA levels were not only attenuated by prazosin, an inhibitor of alpha1-adrenergic receptor, but also by U73122, an inhibitor of PLC activity. Alterations in NE-induced PLC gene expression by both prazosin and U73122 were associated with inhibition of PLC activity. The inhibition of NE-stimulated PLC gene expression by bisindolylmaleimide, a PKC inhibitor, and PD98059, an ERK1/2 inhibitor, indicated that PKC-MAPK signaling may be involved in this signal transduction pathway. The observed NE-induced changes in gene expression in the presence of different inhibitors were associated with corresponding changes in the protein content. Furthermore, significant increases in mRNA levels and protein contents for all PLC isozymes were found in cardiomyocytes treated with phorbol 12-myristate 13-acetate, a PKC activator. These data indicate that PLC isozymes may regulate their own gene expression through a PKC and ERK 1/2-dependent pathway in a cycle of events associated with the cardiomyocyte hypertrophic response.

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PubMed2006

Inhibition of phospholipase C-gamma 1 augments the decrease in cardiomyocyte viability by H2O2.

The present study was conducted to examine the role of a major cardiac phospholipase C (PLC) isozyme, PLC-gamma 1, in cardiomyocytes during oxidative stress. Left ventricular cardiomyocytes were isolated by collagenase digestion from adult male Sprague-Dawley rats (250-300 g) and treated with 20, 50, and 100 microM H2O2 for 15 min. A concentration-dependent (up to 50 microM) increase in the mRNA level and membrane protein content of PLC-gamma 1 was observed with H2O2 treatment. Furthermore, PLC-gamma 1 was activated in response to H2O2, as revealed by an increase in the phosphorylation of its tyrosine residues. There was a marked increase in the phosphorylation of the antiapoptotic protein Bcl-2 by H2O2; this change was attenuated by a PLC inhibitor, U-73122. Although both protein kinase C (PKC)-delta and -epsilon protein contents were increased in the cardiomyocyte membrane fraction in response to H2O2, PKC-epsilon activation, unlike PKC-delta, was attenuated by U-73122 (2 microM). Inhibition of PKC-epsilon with inhibitory peptide (0.1 microM) prevented Bcl-2 phosphorylation. Moreover, different concentrations (0.05, 0.1, and 0.2 microM) of this peptide augmented the decrease in cardiomyocyte viability in response to H2O2. In addition, a decrease in cardiomyocyte viability, as assessed by trypan blue exclusion, due to H2O2 was also seen when cells were pretreated with U-73122 and was as a result of increased apoptosis. It is therefore suggested that PLC-gamma 1 may play a role in cardiomyocyte survival during oxidative stress via PKC-epsilon and phosphorylation of Bcl-2.

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PubMed2005

Differential changes in phospholipase D and phosphatidate phosphohydrolase activities in ischemia-reperfusion of rat heart.

Phospholipase D (PLD2) produces phosphatidic acid (PA), which is converted to 1,2 diacylglycerol (DAG) by phosphatidate phosphohydrolase (PAP2). Since PA and DAG regulate Ca(2+) movements, we examined PLD2 and PAP2 in the sarcolemma (SL) and sarcoplasmic reticular (SR) membranes from hearts subjected to ischemia and reperfusion (I-R). Although SL and SR PLD2 activities were unaltered after 30 min ischemia, 5 min reperfusion resulted in a 36% increase in SL PLD2 activity, whereas 30 min reperfusion resulted in a 30% decrease in SL PLD2 activity, as compared to the control value. SR PLD2 activity was decreased (39%) after 5 min reperfusion, but returned to control levels after 30 min reperfusion. Ischemia for 60 min resulted in depressed SL and SR PLD2 activities, characterized with reduced V(max) and increased K(m) values, which were not reversed during reperfusion. Although the SL PAP2 activity was decreased (31%) during ischemia and at 30 min reperfusion (28%), the SR PAP2 activity was unchanged after 30 min ischemia, but was decreased after 5 min reperfusion (25%) and almost completely recovered after 30 min reperfusion. A 60 min period of ischemia followed by reperfusion caused an irreversible depression of SL and SR PAP2 activities. Our results indicate that I-R induced cardiac dysfunction is associated with subcellular changes in PLD2 and PAP2 activities.

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PubMed2004

Phospholipase C may be involved in norepinephrine-induced cardiac hypertrophy.

Cardiac hypertrophy is characterized by increased cardiomyocyte size, mRNA levels for atrial natriuretic factor (ANF), and protein synthesis. Although activation of the phosphoinositide-specific phospholipase C (PLC) leads to the generation of diacylglycerol (DAG) and inositol 1,4,5-trisphosphate, the involvement of PLC in hypertrophic response remains to be fully understood. The present study was therefore undertaken to examine if the inhibition of PLC activity is associated with a decrease in ANF expression and protein synthesis in cardiomyocytes, due to norepinephrine (NE), a known hypertrophic agent. NE resulted in an increase in ANF gene expression and protein synthesis in adult rat cardiomyocytes, these effects of NE were attenuated by a PLC inhibitor, U73122. The NE-induced increase in ANF gene expression and protein synthesis was also inhibited by an alpha-adrenoceptor blocker, prazosin. Both U73122 and prazosin depressed the NE-induced increase in DAG production in cardiomyocytes. These results indicate that the alpha-adrenoceptor mediated PLC activation may be involved in the process of NE-induced cardiac hypertrophy.

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PubMedDate not supplied

Expression of phospholipase D isozymes in scar and viable tissue in congestive heart failure due to myocardial infarction.

The phospholipase D (PLD) associated with the cardiac sarcolemmal (SL) membrane hydrolyses phosphatidylcholine to produce phosphatidic acid, an important phospholipid signaling molecule known to influence cardiac function. The present study was undertaken to examine PLD isozyme mRNA expression, protein contents and activities in congestive heart failure (CHF) subsequent to myocardial infarction (MI). MI was induced in rats by occlusion of the left anterior descending coronary artery. At 8 weeks after the surgical procedure, hemodynamic assessment revealed that these experimental rats were at a moderate stage of CHF. Semi-quantitative reverse transcriptase-polymerase chain reaction revealed that PLD1 and PLD2 mRNA amounts were unchanged in viable left ventricular (LV) tissue of the failing heart. Furthermore, this technique demonstrated the presence of PLD1 and PLD2 mRNA in the scar tissue. While SL PLD1 and PLD2 protein contents were elevated in the viable LV tissue of the failing heart, SL PLD1 activity was significantly decreased, whereas SL PLD2 activity was significantly increased. On the other hand, although PLD1 protein was undetectable, PLD2 protein and activity were detected in the scar tissue. Our findings suggest that differential changes in PLD isozymes may contribute to the pathophysiology of CHF and may also be involved in the processes of scar remodeling.

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