American journal of physiology. Heart and circulatory physiology, 2003
The aim of this study was to determine whether changes in protein content and/or gene expression ... more The aim of this study was to determine whether changes in protein content and/or gene expression of Na+-K+-ATPase subunits underlie its decreased enzyme activity during ischemia and reperfusion. We measured protein and mRNA subunit levels in isolated rat hearts subjected to 30 min of ischemia and 30 min of reperfusion (I/R). The effect of ischemic preconditioning (IP), induced by three cycles of ischemia and reperfusion (10 min each), was also assessed on the molecular changes in Na+-K+-ATPase subunit composition due to I/R. I/R reduced the protein levels of the alpha2-, alpha3-, beta1-, and beta2-isoforms by 71%, 85%, 27%, and 65%, respectively, whereas the alpha1-isoform was decreased by <15%. A similar reduction in mRNA levels also occurred for the isoforms of Na+-K+-ATPase. IP attenuated the reduction in protein levels of Na+-K+-ATPase alpha2-, alpha3-, and beta2-isoforms induced by I/R, without affecting the alpha1- and beta1-isoforms. Furthermore, IP prevented the reduction...
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
The gap junction protein connexin-43 (Cx43) exists mainly in the phosphorylated state in the norm... more The gap junction protein connexin-43 (Cx43) exists mainly in the phosphorylated state in the normal heart, while ischemia induces dephosphorylation. Phosphatase(s) involved in cardiac Cx43 dephosphorylation have not as yet been identified. We examined the acute effects of ischemia on the dephosphorylation of the gap junction protein connexin-43 in isolated adult cardiomyocytes and isolated perfused hearts. In addition we tested the effectiveness of protein phosphatase 1 and 2A (PP1/2A) inhibitors in preventing Cx43 dephosphorylation. In both models, significant accumulation of the 41 kDa non-phosphorylated Cx43, accompanied by decreased relative levels of the 43–46 kDa phosphorylated Cx43, was observed at 30 min of ischemia. Okadaic acid decreased ischemia-induced Cx43 dephosphorylation; it also decreased the accumulation of non-phosphorylated Cx43 at the intercalated discs of myocytes in the whole heart. Calyculin A, but not fostriecin, also decreased ischemia-induced Cx43 dephosphorylation in isolated cardiomyocytes. It is concluded that isolated adult myocytes respond to ischemia in a manner similar to whole hearts and that ischemia-induced dephosphorylation of Cx43 is mediated, at least in part, by PP1-like phosphatase(s).
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
Etomoxir, an inhibitor of mitochondrial carnitine palmitoyltransferase-1, is known to attenuate t... more Etomoxir, an inhibitor of mitochondrial carnitine palmitoyltransferase-1, is known to attenuate the changes in myosin isoforms and sarcoplasmic reticular function that occur in diabetic rat hearts. In the present study, we tested the hypothesis that etomoxir also prevents the diabetes-induced depression of sarcolemmal (SL) Na+-K+ ATPase activity by differentially affecting its α and β-subunit levels. Streptozotocin-induced diabetes was associated with a decreased in α2-, α3-subunit levels, whereas the α1-, and β1-subunits were unchanged. Treatment of diabetic rats for 4 weeks with etomoxir (8 mg/kg/day) increased the α1-subunit levels, but failed to prevent the decrease in α2- and α3-subunit levels. In euglycemic control rats, etomoxir increased the α1-subunit protein level per g heart weight, but did not alter the α2-, α3- and β1-subunit levels. The large decrease in Na+-K+ ATPase activity per g heart weight in diabetic rats was prevented by etomoxir, which suggests that the increased α1-subunit levels seen with this drug compensated for the decreased α2- and α3-subunit levels. The SL yield was also increased by etomoxir in euglycemic rats in proportion to the higher α1-subunit level, which resulted in an unchanged α1-content when expressed per mg SL protein; however, the α2- and β1-subunit levels were reduced (p 2- and α3-subunit levels of diabetic rats were associated with reduced mRNA abundance. However, no increase in α1-subunit mRNA abundance was seen in the etomoxir treated rats, which suggests that possibly post-transcriptional mechanisms are occurring in these hearts.
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
Previous studies have shown that the renin–angiotensin system (RAS) is activated in diabetes and ... more Previous studies have shown that the renin–angiotensin system (RAS) is activated in diabetes and this may contribute to the subcellular remodelling and heart dysfunction in this disease. Therefore, we examined the effects of RAS blockade by enalapril, an angiotensin-converting enzyme inhibitor, and losartan, an angiotensin receptor AT1 antagonist, on cardiac function, myofibrillar and myosin ATPase activity as well as myosin heavy chain (MHC) isozyme expression in diabetic hearts. Diabetes was induced in rats by a single injection of streptozotocin (65 mg/kg; i.v.) and these animals were treated with and without enalapril (10 mg/kg/day; oral) or losartan (20 mg/kg/day; oral) for 8 weeks. Enalapril or losartan prevented the depressions in left ventricular rate of pressure development, rate of pressure decay and ventricular weight seen in diabetic animals. Both drugs also attenuated the decrease in myofibrillar Ca2+-ATPase, Mg2+-ATPase and myosin ATPase activity seen in diabetic rats. The diabetes-induced increase in β-MHC content and gene expression as well as the decrease in α-MHC content and mRNA levels were also prevented by enalapril and losartan. These results suggest the occurrence of myofibrillar remodelling in diabetic cardiomyopathy and provide evidence that the beneficial effects of RAS blockade in diabetes may be associated with attenuation of myofibrillar remodelling in the heart. (Mol Cell Biochem 261: 271–278, 2004)
Canadian Journal of Physiology and Pharmacology, 2006
The present study determined whether changes in the activity and isoforms of protein kinase C (PK... more The present study determined whether changes in the activity and isoforms of protein kinase C (PKC) are associated with cardiac hypertrophy and heart failure owing to volume overload induced by aortocaval shunt (AVS) in rats. A significant increase in Ca2+-dependent and Ca2+-independent PKC activities in the homogenate and particulate fractions, unlike the cystolic fraction, of the hypertrophied left ventricle (LV) were evident at 2 and 4 weeks after inducing the AVS. This increase coincided with increases in PKC-alpha and PKC-zeta contents at 2 week and increases in PKC-alpha, PKC-beta1, PKC-beta2, and PKC-zeta contents at 4 weeks in the hypertrophied LV. By 8 and 16 weeks of AVS, PKC activity and content were unchanged in the failing LV. On the other hand, no increase in the PKC activity or isoform content in the hypertrophied right ventricle (RV) was observed during the 16 weeks of AVS. The content of G alpha q was increased in the LV at 2 weeks but then decreased at 16 weeks, whereas G alpha q content was increased in RV at 2 and 4 weeks. Our data suggest that an increase in PKC isoform content neither plays an important role during the development of cardiac hypertrophy nor participates in the phase leading to heart failure owing to volume overload.
To determine the sequence of alterations in cardiac sarcolemmal (SL) Na+-Ca2+ exchange, Na+-K+ AT... more To determine the sequence of alterations in cardiac sarcolemmal (SL) Na+-Ca2+ exchange, Na+-K+ ATPase and Ca2+-transport activities during the development of diabetes, rats were made diabetic by an intravenous injection of 65 mg/kg alloxan. SL membranes were prepared from control and experimental hearts 1-12 weeks after induction of diabetes. A separate group of 4 week diabetic animals were injected with insulin (3 U/day) for an additional 4 weeks. Both Na+-K+ ATPase and Ca2+-stimulated ATPase activities were depressed as early as 10 days after alloxan administration; Mg2+ ATPase activity was not depressed throughout the experimental periods. Both Na+-Ca2+ exchange and ATP-dependent Ca2+-uptake activities were depressed in diabetic hearts 2 weeks after diabetes induction. These defects in SL Na+-K+ ATPase and Ca-transport activities were normalized upon treatment of diabetic animals with insulin. Northern blot analysis was employed to compare the relative mRNA abundances of α--subunit of Na+-K+ ATPase and Na+-Ca2+ exchanger in diabetic ventricular tissue vs. control samples. At 6 weeks after alloxan administration, a significant depression of the Na+-K+ ATPase α-- subunit mRNA was noted in diabetic heart. A significant increase in the Na+-Ca2+ exchanger mRNA abundance was observed at 3 weeks which returned to control by 5 weeks. The results from the alloxan-rat model of diabetes support the view that SL membrane abnormalities in Na+-K+ ATPase, Na+Ca2+ exchange and Ca2+-pump activities may lead to the occurrence of intracellular Ca2+ overload during the development of diabetic cardiomyopathy but these defects may not be the consequence of depressed expression of genes specific for those SL proteins.
This study examined the status of sarcolemmal Na+/K+-ATPase activity in rat heart under condition... more This study examined the status of sarcolemmal Na+/K+-ATPase activity in rat heart under conditions of Ca2+-paradox to explore the existence of a relationship between changes in Na+/K+-pump function and myocardial Na+ as well as K+ content. One min of reperfusion with Ca2+ after 5 min of Ca2+-free perfusion reduced Na+/K+-ATPase activity in the isolated heart by 53% while Mg2+-ATPase, another sarcolemmal bound enzyme, retained 74% of its control activity. These changes in sarcolemmal ATPase activities were dependent on the duration and Ca2+ concentration of the initial perfusion and subsequent reperfusion periods; however, the Na+/K+-ATPase activity was consistently more depressed than Mg2+-ATPase activity under all conditions. The depression in both enzyme activities was associated with a reduction in Vmax without any changes in Km values. Low Na+ perfusion and hypothermia, which protect the isolated heart from the Ca2+-paradox, also prevented reperfusion-induced enzyme alterations. A significant relationship emerged upon comparison of the changes in myocardial Na+ and K+ content to Na+/K+-ATPase activity under identical conditions. At least 60% of the control enzyme activity was necessary to maintain normal cation gradients. Depression of the Na+/K+-ATPase activity by 60-65% resulted in a marked increase and decrease in intracellular Na+ and K+ content, respectively. These results suggest that changes in myocardial Na+ and K+ content during Ca2+-paradox are related to activity of the Na+/K+-pump; the impaired Na+/K+-ATPase activity may lead to augmentation of Ca2+-overload via an enhancement of the Na+/Ca2+-exchange system.
Previous studies have shown that cardiac Na+ -K+ ATPase activity in the UM-X7.1 hamster strain is... more Previous studies have shown that cardiac Na+ -K+ ATPase activity in the UM-X7.1 hamster strain is decreased at an early stage of genetic cardiomyopathy and remains depressed; however, the mechanism for this decrease is unknown. The objective of the present study was to assess whether changes in the expression of cardiac Na+-K+ ATPase subunits in control and UM-X7.1 cardiomyopathic hamsters are associated with alterations in the enzyme activity. Accordingly, we examined sarcolemmal Na+-K+ ATPase activity as well as protein content and mRNA levels for the alpha1, alpha2, alpha3 and beta1-subunit of the Na+-K+ ATPase in 250-day-old UM-X7.1 and age-matched, control Syrian hamsters; this age corresponds to the severe stage of heart failure in the UM-X7.1 hamster. Na+-K+ ATPase activity in UM-X7.1 hearts was decreased compared to controls (9.0 +/- 0.8 versus 5.6 +/- 0.8 micromol Pi/mg protein/hr). Western blot analysis revealed that the protein content of Na+-K+ ATPase alpha1- and beta1-subunits were increased to 164 +/- 27% and 146 +/- 22% in UM-X7.1 hearts respectively, whereas that of the alpha2- and alpha3-subunits were decreased to 82 +/- 5% and 69 +/- 11% of control values. The results of Northern blot analysis for mRNA levels were consistent with the protein levels; mRNA levels for the alpha1- and beta1-subunits in UM-X7.1 hearts were elevated to 165 +/- 14% and 151 +/- 10%, but the alpha2-subunit was decreased to 60 +/- 8% of the control value. We were unable to detect mRNA for the alpha3-subunit in either UM-X7. 1 or control hearts. These data suggest that the marked depression of Na+-K+ ATPase activity in UM-X7.1 cardiomyopathic hearts may be due to changes in the expression of subunits for this enzyme.
Diets rich in omega-3 polyunsaturated fatty acids are associated with decreased incidences of car... more Diets rich in omega-3 polyunsaturated fatty acids are associated with decreased incidences of cardiovascular disease. The extent of incorporation and distribution of these beneficial fats into body tissues is uncertain. Rabbits were fed regular rabbit chow or a diet containing 10% ground flaxseed that is highly enriched with the omega-3 polyunsaturated fatty acid alpha-linolenic acid (ALA). The high-flaxseed diet resulted in an incorporation of ALA in all tissues, but mostly in the heart and liver with little in the brain. Docosahexaenoic and eicosapentaenoic acid levels were also selectively increased in some tissues, and the effects were not as large as ALA. Arachidonic acid and the ratio of omega-6/omega-3 fatty acids were decreased in all tissues obtained from the flax-supplemented group. Consumption of dietary flaxseed appears to be an effective means to increase ALA content in body tissues, but the degree will depend upon the tissues examined.
Journal of Molecular and Cellular Cardiology, 2002
Prostaglandins, Leukotrienes and Essential Fatty Acids, Volume 34, Issue 7, Pages A18, July 2002,... more Prostaglandins, Leukotrienes and Essential Fatty Acids, Volume 34, Issue 7, Pages A18, July 2002, Authors:Bradley P. Ander; Anna R. Weber; Penelope Rampersad; James SC Gilchrist; Grant N. Pierce; Anton Lukas.
American journal of physiology. Heart and circulatory physiology, 2003
The aim of this study was to determine whether changes in protein content and/or gene expression ... more The aim of this study was to determine whether changes in protein content and/or gene expression of Na+-K+-ATPase subunits underlie its decreased enzyme activity during ischemia and reperfusion. We measured protein and mRNA subunit levels in isolated rat hearts subjected to 30 min of ischemia and 30 min of reperfusion (I/R). The effect of ischemic preconditioning (IP), induced by three cycles of ischemia and reperfusion (10 min each), was also assessed on the molecular changes in Na+-K+-ATPase subunit composition due to I/R. I/R reduced the protein levels of the alpha2-, alpha3-, beta1-, and beta2-isoforms by 71%, 85%, 27%, and 65%, respectively, whereas the alpha1-isoform was decreased by <15%. A similar reduction in mRNA levels also occurred for the isoforms of Na+-K+-ATPase. IP attenuated the reduction in protein levels of Na+-K+-ATPase alpha2-, alpha3-, and beta2-isoforms induced by I/R, without affecting the alpha1- and beta1-isoforms. Furthermore, IP prevented the reduction...
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
The gap junction protein connexin-43 (Cx43) exists mainly in the phosphorylated state in the norm... more The gap junction protein connexin-43 (Cx43) exists mainly in the phosphorylated state in the normal heart, while ischemia induces dephosphorylation. Phosphatase(s) involved in cardiac Cx43 dephosphorylation have not as yet been identified. We examined the acute effects of ischemia on the dephosphorylation of the gap junction protein connexin-43 in isolated adult cardiomyocytes and isolated perfused hearts. In addition we tested the effectiveness of protein phosphatase 1 and 2A (PP1/2A) inhibitors in preventing Cx43 dephosphorylation. In both models, significant accumulation of the 41 kDa non-phosphorylated Cx43, accompanied by decreased relative levels of the 43–46 kDa phosphorylated Cx43, was observed at 30 min of ischemia. Okadaic acid decreased ischemia-induced Cx43 dephosphorylation; it also decreased the accumulation of non-phosphorylated Cx43 at the intercalated discs of myocytes in the whole heart. Calyculin A, but not fostriecin, also decreased ischemia-induced Cx43 dephosphorylation in isolated cardiomyocytes. It is concluded that isolated adult myocytes respond to ischemia in a manner similar to whole hearts and that ischemia-induced dephosphorylation of Cx43 is mediated, at least in part, by PP1-like phosphatase(s).
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
Etomoxir, an inhibitor of mitochondrial carnitine palmitoyltransferase-1, is known to attenuate t... more Etomoxir, an inhibitor of mitochondrial carnitine palmitoyltransferase-1, is known to attenuate the changes in myosin isoforms and sarcoplasmic reticular function that occur in diabetic rat hearts. In the present study, we tested the hypothesis that etomoxir also prevents the diabetes-induced depression of sarcolemmal (SL) Na+-K+ ATPase activity by differentially affecting its α and β-subunit levels. Streptozotocin-induced diabetes was associated with a decreased in α2-, α3-subunit levels, whereas the α1-, and β1-subunits were unchanged. Treatment of diabetic rats for 4 weeks with etomoxir (8 mg/kg/day) increased the α1-subunit levels, but failed to prevent the decrease in α2- and α3-subunit levels. In euglycemic control rats, etomoxir increased the α1-subunit protein level per g heart weight, but did not alter the α2-, α3- and β1-subunit levels. The large decrease in Na+-K+ ATPase activity per g heart weight in diabetic rats was prevented by etomoxir, which suggests that the increased α1-subunit levels seen with this drug compensated for the decreased α2- and α3-subunit levels. The SL yield was also increased by etomoxir in euglycemic rats in proportion to the higher α1-subunit level, which resulted in an unchanged α1-content when expressed per mg SL protein; however, the α2- and β1-subunit levels were reduced (p 2- and α3-subunit levels of diabetic rats were associated with reduced mRNA abundance. However, no increase in α1-subunit mRNA abundance was seen in the etomoxir treated rats, which suggests that possibly post-transcriptional mechanisms are occurring in these hearts.
The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during... more The aim of this study was to assess whether depression of cardiac Na+,K(+)-ATPase activity during ischemia/reperfusion (I/R) is associated with alterations in Na+,K(+)-ATPase isoforms, and if oxidative stress participates in these I/R-induced changes. Na+,K(+)-ATPase alpha1, alpha2, alpha3, beta1, beta2, and beta3 isoform contents were measured in isolated rat hearts subjected to I/R (30 min of global ischemia followed by 60 min of reperfusion) in the presence or absence of superoxide dismutase plus catalase (SOD+CAT). Effects of oxidative stress on Na+,K(+)-ATPase isoforms were also examined by perfusing the hearts for 20 min with 300 microM hydrogen peroxide or 2 mM xanthine plus 0.03 U/ml xanthine oxidase (XXO). I/R significantly reduced the protein levels of all alpha and beta isoforms. Treatment of I/R hearts with SOD+CAT preserved the levels of alpha2, alpha3, beta1, beta2, and beta3 isoforms, but not that of the alpha1 isoform. Perfusion of hearts with hydrogen peroxide and XXO depressed all Na+,K(+)-ATPase alpha and beta isoforms, except for alpha1. These results indicate that the I/R-induced decrease in Na+,K(+)-ATPase may be due to changes in Na+,K(+)-ATPase isoform expression and that oxidative stress plays a role in this alteration. Antioxidant treatment attenuated the I/R-induced changes in expression of all isoforms except alpha1, which appears to be more resistant to oxidative stress.
Previous studies have shown that the renin–angiotensin system (RAS) is activated in diabetes and ... more Previous studies have shown that the renin–angiotensin system (RAS) is activated in diabetes and this may contribute to the subcellular remodelling and heart dysfunction in this disease. Therefore, we examined the effects of RAS blockade by enalapril, an angiotensin-converting enzyme inhibitor, and losartan, an angiotensin receptor AT1 antagonist, on cardiac function, myofibrillar and myosin ATPase activity as well as myosin heavy chain (MHC) isozyme expression in diabetic hearts. Diabetes was induced in rats by a single injection of streptozotocin (65 mg/kg; i.v.) and these animals were treated with and without enalapril (10 mg/kg/day; oral) or losartan (20 mg/kg/day; oral) for 8 weeks. Enalapril or losartan prevented the depressions in left ventricular rate of pressure development, rate of pressure decay and ventricular weight seen in diabetic animals. Both drugs also attenuated the decrease in myofibrillar Ca2+-ATPase, Mg2+-ATPase and myosin ATPase activity seen in diabetic rats. The diabetes-induced increase in β-MHC content and gene expression as well as the decrease in α-MHC content and mRNA levels were also prevented by enalapril and losartan. These results suggest the occurrence of myofibrillar remodelling in diabetic cardiomyopathy and provide evidence that the beneficial effects of RAS blockade in diabetes may be associated with attenuation of myofibrillar remodelling in the heart. (Mol Cell Biochem 261: 271–278, 2004)
Canadian Journal of Physiology and Pharmacology, 2006
The present study determined whether changes in the activity and isoforms of protein kinase C (PK... more The present study determined whether changes in the activity and isoforms of protein kinase C (PKC) are associated with cardiac hypertrophy and heart failure owing to volume overload induced by aortocaval shunt (AVS) in rats. A significant increase in Ca2+-dependent and Ca2+-independent PKC activities in the homogenate and particulate fractions, unlike the cystolic fraction, of the hypertrophied left ventricle (LV) were evident at 2 and 4 weeks after inducing the AVS. This increase coincided with increases in PKC-alpha and PKC-zeta contents at 2 week and increases in PKC-alpha, PKC-beta1, PKC-beta2, and PKC-zeta contents at 4 weeks in the hypertrophied LV. By 8 and 16 weeks of AVS, PKC activity and content were unchanged in the failing LV. On the other hand, no increase in the PKC activity or isoform content in the hypertrophied right ventricle (RV) was observed during the 16 weeks of AVS. The content of G alpha q was increased in the LV at 2 weeks but then decreased at 16 weeks, whereas G alpha q content was increased in RV at 2 and 4 weeks. Our data suggest that an increase in PKC isoform content neither plays an important role during the development of cardiac hypertrophy nor participates in the phase leading to heart failure owing to volume overload.
To determine the sequence of alterations in cardiac sarcolemmal (SL) Na+-Ca2+ exchange, Na+-K+ AT... more To determine the sequence of alterations in cardiac sarcolemmal (SL) Na+-Ca2+ exchange, Na+-K+ ATPase and Ca2+-transport activities during the development of diabetes, rats were made diabetic by an intravenous injection of 65 mg/kg alloxan. SL membranes were prepared from control and experimental hearts 1-12 weeks after induction of diabetes. A separate group of 4 week diabetic animals were injected with insulin (3 U/day) for an additional 4 weeks. Both Na+-K+ ATPase and Ca2+-stimulated ATPase activities were depressed as early as 10 days after alloxan administration; Mg2+ ATPase activity was not depressed throughout the experimental periods. Both Na+-Ca2+ exchange and ATP-dependent Ca2+-uptake activities were depressed in diabetic hearts 2 weeks after diabetes induction. These defects in SL Na+-K+ ATPase and Ca-transport activities were normalized upon treatment of diabetic animals with insulin. Northern blot analysis was employed to compare the relative mRNA abundances of α--subunit of Na+-K+ ATPase and Na+-Ca2+ exchanger in diabetic ventricular tissue vs. control samples. At 6 weeks after alloxan administration, a significant depression of the Na+-K+ ATPase α-- subunit mRNA was noted in diabetic heart. A significant increase in the Na+-Ca2+ exchanger mRNA abundance was observed at 3 weeks which returned to control by 5 weeks. The results from the alloxan-rat model of diabetes support the view that SL membrane abnormalities in Na+-K+ ATPase, Na+Ca2+ exchange and Ca2+-pump activities may lead to the occurrence of intracellular Ca2+ overload during the development of diabetic cardiomyopathy but these defects may not be the consequence of depressed expression of genes specific for those SL proteins.
This study examined the status of sarcolemmal Na+/K+-ATPase activity in rat heart under condition... more This study examined the status of sarcolemmal Na+/K+-ATPase activity in rat heart under conditions of Ca2+-paradox to explore the existence of a relationship between changes in Na+/K+-pump function and myocardial Na+ as well as K+ content. One min of reperfusion with Ca2+ after 5 min of Ca2+-free perfusion reduced Na+/K+-ATPase activity in the isolated heart by 53% while Mg2+-ATPase, another sarcolemmal bound enzyme, retained 74% of its control activity. These changes in sarcolemmal ATPase activities were dependent on the duration and Ca2+ concentration of the initial perfusion and subsequent reperfusion periods; however, the Na+/K+-ATPase activity was consistently more depressed than Mg2+-ATPase activity under all conditions. The depression in both enzyme activities was associated with a reduction in Vmax without any changes in Km values. Low Na+ perfusion and hypothermia, which protect the isolated heart from the Ca2+-paradox, also prevented reperfusion-induced enzyme alterations. A significant relationship emerged upon comparison of the changes in myocardial Na+ and K+ content to Na+/K+-ATPase activity under identical conditions. At least 60% of the control enzyme activity was necessary to maintain normal cation gradients. Depression of the Na+/K+-ATPase activity by 60-65% resulted in a marked increase and decrease in intracellular Na+ and K+ content, respectively. These results suggest that changes in myocardial Na+ and K+ content during Ca2+-paradox are related to activity of the Na+/K+-pump; the impaired Na+/K+-ATPase activity may lead to augmentation of Ca2+-overload via an enhancement of the Na+/Ca2+-exchange system.
Previous studies have shown that cardiac Na+ -K+ ATPase activity in the UM-X7.1 hamster strain is... more Previous studies have shown that cardiac Na+ -K+ ATPase activity in the UM-X7.1 hamster strain is decreased at an early stage of genetic cardiomyopathy and remains depressed; however, the mechanism for this decrease is unknown. The objective of the present study was to assess whether changes in the expression of cardiac Na+-K+ ATPase subunits in control and UM-X7.1 cardiomyopathic hamsters are associated with alterations in the enzyme activity. Accordingly, we examined sarcolemmal Na+-K+ ATPase activity as well as protein content and mRNA levels for the alpha1, alpha2, alpha3 and beta1-subunit of the Na+-K+ ATPase in 250-day-old UM-X7.1 and age-matched, control Syrian hamsters; this age corresponds to the severe stage of heart failure in the UM-X7.1 hamster. Na+-K+ ATPase activity in UM-X7.1 hearts was decreased compared to controls (9.0 +/- 0.8 versus 5.6 +/- 0.8 micromol Pi/mg protein/hr). Western blot analysis revealed that the protein content of Na+-K+ ATPase alpha1- and beta1-subunits were increased to 164 +/- 27% and 146 +/- 22% in UM-X7.1 hearts respectively, whereas that of the alpha2- and alpha3-subunits were decreased to 82 +/- 5% and 69 +/- 11% of control values. The results of Northern blot analysis for mRNA levels were consistent with the protein levels; mRNA levels for the alpha1- and beta1-subunits in UM-X7.1 hearts were elevated to 165 +/- 14% and 151 +/- 10%, but the alpha2-subunit was decreased to 60 +/- 8% of the control value. We were unable to detect mRNA for the alpha3-subunit in either UM-X7. 1 or control hearts. These data suggest that the marked depression of Na+-K+ ATPase activity in UM-X7.1 cardiomyopathic hearts may be due to changes in the expression of subunits for this enzyme.
Diets rich in omega-3 polyunsaturated fatty acids are associated with decreased incidences of car... more Diets rich in omega-3 polyunsaturated fatty acids are associated with decreased incidences of cardiovascular disease. The extent of incorporation and distribution of these beneficial fats into body tissues is uncertain. Rabbits were fed regular rabbit chow or a diet containing 10% ground flaxseed that is highly enriched with the omega-3 polyunsaturated fatty acid alpha-linolenic acid (ALA). The high-flaxseed diet resulted in an incorporation of ALA in all tissues, but mostly in the heart and liver with little in the brain. Docosahexaenoic and eicosapentaenoic acid levels were also selectively increased in some tissues, and the effects were not as large as ALA. Arachidonic acid and the ratio of omega-6/omega-3 fatty acids were decreased in all tissues obtained from the flax-supplemented group. Consumption of dietary flaxseed appears to be an effective means to increase ALA content in body tissues, but the degree will depend upon the tissues examined.
Journal of Molecular and Cellular Cardiology, 2002
Prostaglandins, Leukotrienes and Essential Fatty Acids, Volume 34, Issue 7, Pages A18, July 2002,... more Prostaglandins, Leukotrienes and Essential Fatty Acids, Volume 34, Issue 7, Pages A18, July 2002, Authors:Bradley P. Ander; Anna R. Weber; Penelope Rampersad; James SC Gilchrist; Grant N. Pierce; Anton Lukas.
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