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1.
Yamaguchi M Morooka Y Misawa H Tsurusaki Y Nakajima R 《Journal of cellular biochemistry》2002,86(3):520-529
Rats were generated by pronuclear injection of the transgene with a cDNA construct encoding rat regucalcin that is a regulatory protein of Ca2+ signaling. Transgenic (TG) founders were fertile, transmitted the transgene at the expected frequency, and bred to homozygote. Western analysis of the cytosol prepared from the tissue of TG female rats (5-week-old) showed a remarkable expression of regucalcin (3.3 kDa) protein in the liver, kidney cortex, heart, lung, stomach, brain, spleen, muscle, colon, and duodenum. Regucalcin expression of TG male rats was seen in the liver, kidney cortex, heart, and lung. In wild-type (wt) male and female rats, regucalcin was mainly present in the liver and kidney cortex. Regucalcin inhibited protein phosphatase activity in rat kidney cortex cytosol and activated Ca2+-ATPase activity in rat heart muscle microsomes. The suppressive effect of regucalcin on protein phosphatase activity was significantly enhanced in the cytosol of kidney cortex of TG male and female rats as compared with those of wt rats. Likewise, heart muscle microsomal Ca2+-ATPase activity was significantly enhanced in TG rats. The changes in their enzyme's activities in TG rats were completely abolished in the presence of anti-regucalcin monoclonal antibody (100 ng/ml) in the enzyme reaction mixture. Moreover, the body weight of TG female rats was significantly lowered as compared with that of wt rats. Serum inorganic phosphorus concentration was significantly increased in TG male and female rats, while serum calcium, glucose, triglyceride, free cholesterol, albumin, and urea nitrogen concentrations were not significantly altered in TG rats. Regucalcin TG rats should be a useful model to define a regulatory role of endogenous regucalcin in the tissues in vivo. 相似文献
2.
ATPase activity in rat heart sarcoplasmic reticulum was stimulated in a concentration-dependent manner by both Ca2+ and Mg2+ in the complete absence of the other cation. Increasing concentrations of Mg2+ produced an apparent inhibition of the Ca2+-dependent ATP hydrolysis. CDTA (trans-1,2-diaminocyclohexane-N,N,N,N-tetraacetate) had no effect on these responses. The results indicate the presence of a low affinity non-specific divalent cation-stimulated ATPase in rat heart sarcoplasmic reticulum. However, sarcoplasmic reticulum vesicles transported Ca2+ with a high affinity (K0.5 Ca2+ = 0.41 M) suggesting the presence of a high affinity Ca2+-transporting ATPase. Calmodulin did not stimulate rat heart sarcoplasmic reticulum ATPase activity over a range of Ca2+ and Mg2+ concentrations and failed to stimulate membrane phosphorylation and Ca2+ transport into sarcoplasmic reticulum vesicles. Calmodulin antagonists trifluoperazine and compound 48180 did not affect the ATPase activity. Catalytic subunit of cAMP-dependent protein kinase was also ineffective in stimulating the ATPase activity. These results suggest the presence of an ATPase activity in rat heart sarcoplasmic reticulum with different properties from the high affinity Ca2+-pumping ATPase previously characterized in dog heart and other species.Abbreviations cAMP
adenosine 3,5-monophosphate
- CaM
calmodulin
- CDTA
trans-1,2-diaminocyclohexane-N,N,N,N-tetraacetate
- EDTA
ethylene-diaminetetraacetate
- EGTA
ethylene glycol bis(-aminoethyl ether)-N,N,N,N-tetraacetate
- PLB
phospholamban
- SR
sarcoplasmic reticulum
- TFP
trifluoperazine 相似文献
3.
Eduardo M. R. Reis Carolyn W. Slayman Sergio Verjovski-Almeida 《Bioscience reports》1996,16(2):107-113
In recent years, expression of rabbit sarcoplasmic reticulum (SR) Ca2+-ATPase in heterologous systems has been a widely used strategy to study altered enzymes generated by site-directed mutagenesis. Various eukaryotic expression systems have been tested, all of them yielding comparable amounts of recombinant protein. However, the relatively low yield of recombinant protein obtained so far suggests that novel purification techniques will be required to allow further characterization of this enzyme based on direct ligand-binding measurements. 相似文献
4.
白细胞介素-2对大鼠心肌Ca2+ATPase和Na+ /K+ATPase的影响 总被引:3,自引:0,他引:3
为了探讨IL-2对心肌细胞内钙影响的可能机制,用光学法检测心肌肌浆网Ca^2 ATPase的活性,以及细胞膜Ca^2 ATPase和Na^ /K^ ATPase的活性。结果:(1)用IL-2(10、40、200、800U/ml)灌流心脏后,其肌浆网Ca^2 ATPase的活性随IL-2浓度的升高而增强;(2)在ATP浓度为0.1-4mmol/L时,Ca^2 ATPase的活性随ATP浓度的升庙则增强,由IL-2(200U/ml)灌流后的心脏获得肌浆网(SR),其Ca^2 ATPase的活性对ATP的反应强于对照组;(3)在[Ca^2 ]为1-40μmol/L时,心脏SR Ca^2 ATPase的活性随[Ca^2 ]增加而增强,而IL-2灌流心脏后分离的SR,其Ca^2 ATPase活性在[Ca^2 ]升高时没有明显改变;(4)用nor-BNI(10nmol/L)预处理5min后,IL-2(200U/ml)灌流后不再使SR Ca^2 ATPase的活性增强;(5)用PTX(5mg/L)预处理后,IL-2对SR Ca^2 ATPase的影响减弱;(6)用磷脂酶C(PLC)抑制剂U73122(5μmol/L)处理后,IL-2不再使SR Ca^2 ATPase活性增高;(7)用IL-2直接处理从正常大鼠分离的SR后,对SR Ca^2 ATPase活性无明显影响;(8)IL-2灌流后,对心肌细胞膜Ca^2 ATPase和Na^ /K^ ATPase活性没有显著。上述结果表明,IL-2灌流心脏后使心肌肌浆网Ca^2 ATPase的活性增加,心肌细胞膜上的κ-阿片受体及其下游的G蛋白和PLC介导了IL-2的作用。尽管IL-2提高SR Ca^2 ATPase对ATP的反应性,但却抑制SR Ca^2 ATPase对钙离子的敏感性。IL-2对心肌细胞膜Ca^2 ATPase和Na^ /K^ ATPase的活性无明显影响。 相似文献
5.
Regucalcin increases Ca2+-ATPase activity in the mitochondria of brain tissues of normal and transgenic rats 总被引:1,自引:0,他引:1
The role of regucalcin, which is a regulatory protein in intracellular signaling, in the regulation of Ca(2+)-ATPase activity in the mitochondria of brain tissues was investigated. The addition of regucalcin (10(-10) to 10(-8) M), which is a physiologic concentration in rat brain tissues, into the enzyme reaction mixture containing 25 microM calcium chloride caused a significant increase in Ca(2+)-ATPase activity, while it did not significantly change in Mg(2+)-ATPase activity. The effect of regucalcin (10(-9) M) in increasing mitochondrial Ca(2+)-ATPase activity was completely inhibited in the presence of ruthenium red (10(-7) M) or lanthanum chloride (10(-7) M), both of which are inhibitors of mitochondrial uniporter activity. Whether the effect of regucalcin is modulated in the presence of calmodulin or dibutyryl cyclic AMP (DcAMP) was examined. The effect of regucalcin (10(-9) M) in increasing Ca(2+)-ATPase activity was not significantly enhanced in the presence of calmodulin (2.5 microg/ml) which significantly increased the enzyme activity. DcAMP (10(-6) to 10(-4) M) did not have a significant effect on Ca(2+)-ATPase activity. The effect of regucalcin (10(-9) M) in increasing Ca(2+)-ATPase activity was not seen in the presence of DcAMP (10(-4) M). Regucalcin levels were significantly increased in the brain tissues or the mitochondria obtained from regucalcin transgenic (RC TG) rats. The mitochondrial Ca(2+)-ATPase activity was significantly increased in RC TG rats as compared with that of wild-type rats. This study demonstrates that regucalcin has a role in the regulation of Ca(2+)-ATPase activity in the brain mitochondria of rats. 相似文献
6.
Phenylglyoxal (PGO) was used as a reagent for chemical modification of the ATP-binding site of Ca2+-transporting ATPase of rabbit skeletal muscle sarcoplasmic reticulum (SR-ATPase). When 1 mM PGO was reacted with SR-ATPase at 30°C at pH 8.5, PGO was bound to the ATPase molecule in two-to-one stoichiometry with concomitant loss of activity of the ATPase to form the phosphorylated intermediate (E-P). ATP and ADP prevented the binding of PGO and thereby protected the enzyme from inactivation. The SR membranes were labeled with [14C]PGO and then digested with pepsin to identify the attachment site of PGO. A 14C-labeled peptide (402lle-Arg*-Ser-Gly-Gln406) was purified to homogeneity by C18-reversed phase HPLC (Arg* denotes the binding site of [14C]PGO). These results indicate that Arg403 is located in the ATP binding site of the SR-ATPase. 相似文献
7.
Jens Peter Andersen 《Bioscience reports》1995,15(5):243-261
The results of site-directed mutagenesis studies of the sarcoplasmic reticulum Ca2+-ATPase are reviewed. More than 250 different point mutants have been expressed in cell culture and analysed by a panel of functional assays. Thereby, 40–50 important amino acid residues have been pinpointed, and the mutants have been assigned to functional classes: the Ca2+-affinity mutants, the phosphorylation-negative mutants, the ATP-affinity mutants, the E1P mutants, the E2P mutants, and the uncoupled mutants. Moreover, regions important to the specific inhibition by thapsigargin have been identified by analysis of Ca2+-ATPase/Na+, K+-ATPase chimeric constructs. 相似文献
8.
Gilchrist James S.C. Palahniuk Chris Bose Ratna 《Molecular and cellular biochemistry》1997,172(1-2):159-170
In this report we describe the application of spectroscopic methods to the study of Ca2+ release by isolated native sarcoplasmic reticulum (SR) membranes from rabbit skeletal muscle. To date, dual-wavelength spectroscopy of arsenazo III and antipyrylazo III difference absorbance have been the most common spectroscopic methods for the assay of SR Ca2+ transport. The utility of these methods is the ability to manipulate intraluminal Ca2+ loading of SR vesicles. These methods have also been useful for studying the effect of both agonists and antagonists upon SR Ca2+ release and Ca2+ uptake. In this study, we have developed the application of Calcium Green-2, a long-wavelength excitable fluorescent indicator, for the study of SR Ca2+ uptake and release. With this method we demonstrate how ryanodine receptor Ca2+ channel opening and closing is regulated in a complex manner by the relative distribution of Ca2+ between extraluminal and intraluminal Ca2+ compartments. Intraluminal Ca2+ is shown to be a key regulator of Ca2+ channel opening. However, these methods also reveal that the intraluminal Ca2+ threshold for Ca2+-induced Ca2+ release varies as a function of extraluminal Ca2+ concentration. The ability to study how the relative distribution of a finite pool of Ca2+ across the SR membrane influences Ca2+ uptake and Ca2+ release may be useful for understanding how the ryanodine receptor is regulated, in vivo. 相似文献
9.
Jing Hui Xue Hiroko Takahashi Masayoshi Yamaguchi 《Journal of cellular biochemistry》2001,80(2):285-292
The effect of regucalcin, which is a regulatory protein of Ca2+ signaling, on Ca2+‐ATPase activity in isolated rat renal cortex mitochondria was investigated. The presence of regucalcin (50, 100, and 250 nM) in the enzyme reaction mixture led to a significant increase in Ca2+‐ATPase activity. Regucalcin significantly stimulated ATP‐dependent 45Ca2+ uptake by the mitochondria. Ruthenium red (10−6 M) or lanthunum chloride (10−6 M), an inhibitor of mitochondrial Ca2+ uptake, markedly inhibited regucalcin (100 nM)‐increased mitochondrial Ca2+‐ATPase activity and 45Ca2+ uptake. The effect of regucalcin (100 nM) in elevating Ca2+‐ATPase activity was completely prevented by the presence of digitonin (10−2%), a solubilizing reagent of membranous lipids, vanadate, an inhibitor of phosphorylation of ATPase, or dithiothreitol (50 mM), a protecting reagent of the sulfhydryl (SH) group of the enzyme. The activating effect of regucalcin (100 nM) on Ca2+‐ATPase activity was not further enhanced by calmodulin (0.30 μM) or dibutyryl cyclic AMP (10−4 M), which could increase Ca2+‐ATPase activity. Trifluoperazine (TFP; 50 μM), an antagonist of calmodulin, significantly decreased Ca2+‐ATPase activity. The activating effect of regucalcin on the enzyme was also seen in the presence of TFP, indicating that regucalcin's effect is not involved in mitochondrial calmodulin. The present study demonstrates that regucalcin can stimulate Ca2+‐pump activity in rat renal cortex mitochondria, and that the protein may act on an active site (SH group) related to phosphorylation of mitochondrial Ca2+‐ATPase. J. Cell. Biochem. 80:285–292, 2000. © 2000 Wiley‐Liss, Inc. 相似文献
10.
Topology of sarcoplasmic reticulum Ca2+-ATPase: an infrared study of thermal denaturation and limited proteolysis.
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I. Echabe U. Dornberger A. Prado F. M. Goi J. L. Arrondo 《Protein science : a publication of the Protein Society》1998,7(5):1172-1179
Sarcoplasmic reticulum Ca2+-ATPase structure and organization in the membrane has been studied by infrared spectroscopy by decomposition of the amide I band. Besides the component bands assignable to secondary structure elements such as alpha-helix, beta-sheet, etc...., two unusual bands, one at 1,645 cm(-1) in H2O buffer and the other at 1,625 cm(-1) in D2O buffer are present. By perturbing the protein using temperature and limited proteolysis, the band at 1,645 cm(-1) is tentatively assigned to alpha-helical segments located in the cytoplasmic domain and coupled to beta-sheet structure, whereas the band at 1,625 cm(-1) arises probably from monomer-monomer contacts in the native oligomeric protein. The secondary structure obtained is 33% alpha-helical segments in the transmembrane plus stalk domain; 20% alpha-helix and 22% beta-sheet in the cytoplasmic domain plus 19% turns and 6% unordered structure. Thermal unfolding of Ca2+-ATPase is a complex process that cannot be described as a two-state denaturation. The results obtained are compatible with the idea that the protein is an oligomer at room temperature. The loss of the 1,625 cm(-1) band upon heating would be consistent with a disruption of the oligomers in a process that later gives rise to aggregates (appearance of the 1,618 cm(-1) band). This picture would also be compatible with early results suggesting that processes governing Ca2+ accumulation and ATPase activity are uncoupled at temperatures above 37 degrees C, so that while ATPase activity proceeds at high rates, Ca2+ accumulation is inhibited. 相似文献
11.
Martin D. King Peter J. Quinn Felix M. Munkonge Thomas D. Madden 《Journal of bioenergetics and biomembranes》1987,19(1):45-52
Experiments were performed in which two batches of sarcoplasmic reticulum were isolated from rabbit hind leg muscle, one in the presence of dithiothreitol, the other in the absence of reducing agent. A comparative study was made of some of the properties of the two preparations, in particular, the Arrhenius behavior of the Ca2+-ATPase. The Ca2+-ATPase isolated in the absence of dithiothreitol is thermally unstable with the result that a triphasic Arrhenius plot was obtained. This triphasic behavior is largely the consequence of an uncoupling of the hydrolytic machinery from the calcium pump. In contrast, the sarcoplasmic reticulum preparation obtained in the presence of dithiothreitol is thermally stable and yields a linear Arrhenius plot. The difference in the Arrhenius behavior shown by the two preparations was abolished when the measurements of Ca2+-ATPase activity were made in the presence of the calcium ionophore, A23187. 相似文献
12.
We propose an overview of the mechanism of Ca2+ transport through the sarcoplasmic reticulum membrane via the Ca2+-ATPase. We describe cytoplasmic calcium binding, calcium occlusion in the membrane and lumenal calcium dissociation. A channel-like structure is discussed and related to structural data on the membranous domain of the Ca2+-ATPase.Abbreviations SR
Sarcoplasmic Reticulum
- AMPPNP
adenylyl-imidodiphosphate
- AMPPCP
adenylyl (,-methylene)-diphosphonate
- FITC
fluorescein 5-isothiocyanate
- NBD
4-nitrobenzo-2-oxa-1,3-diazole
- DCCD
dicyclohexylcarbodiimide 相似文献
13.
Summary The two high affinity calcium binding sites of the cardiac (Ca2+ + Mg2+)-ATPase have been identified with the use of Eu3+. Eu3+ competes for the two high affinity calcium sites on the enzyme. With the use of laser-pulsed fluorescent spectroscopy, the environment of the two sites appear to be heterogeneous and contain different numbers of H2O molecules coordinated to the ion. The ion appears to be occluded even further in the presence of ATP. Using non-radiative energy transfer studies, we were able to estimate the distance between the two Ca2+ sites to be between 9.4 to 10.2 A in the presence of ATP. Finally, from the assumption that the calcium site must contain four carboxylic side chains to provide the 6–8 ligands needed to coordinate calcium, and based on our recently published data, we predict the peptidic backbone of the two sites. 相似文献
14.
S G O'Neal D B Rhoads E Racker 《Biochemical and biophysical research communications》1979,89(3):845-850
Vanadate is a potent inhibitor of the Ca2+-ATPase activity of sarcoplasmic reticulum in the presence of A-23187. The purified enzyme is sensitive to vanadate even in the absence of the ionophore. Ca2+ and norepinephrine protect the enzyme against inhibition of vanadate. The nonspecificity of vanadate is emphasized by the finding of inhibition of several other ATPases including the Ca2+Mg2+-ATPases of the ascites and human red cell plasma membranes, Mg2+-ATPase of the ascites plasma membrane, and the K+-ATPases of and hog gastric mucosal cell membranes. The ascites plasma membrane Ca2+-ATPase (an ecto ATPase) and mitochondrial ATPase are not inhibited by vanadate. 相似文献
15.
J. Dossett-Mercer H. Green E. R. Chin F. Grange 《Molecular and cellular biochemistry》1995,146(1):23-33
To examine the effect of short term intense activity on sarcoplasmic reticulum (SR) Ca2+ sequestering function, the gastrocnemius (G) muscles of 11 anaesthetized male rats (weight, 411±8 g,X±SE) were activated using supramaximal, intermittent stimulation (one train of 0.2 msec impulses per sec of 100 msec at 100 Hz). Homogenates were obtained from stimulated white (WG-S) and red (RG-S) tissues, assayed for Ca2+ uptake and maximal Ca2+ ATPase activity and compared to contralateral controls (WG-C, RG-C). Calcium uptake (nmoles/mg protein/min) determined using Indo-l and at [Ca2+]f concentrations between 300–400 nM was unaffected (p>0.05) by activity in both WG (6.14+0.43 vs 5.37+0.43) and RG (3.21+0.18 vs 3.07+0.20). Similarly, no effect (p>0.05) of contractile activity was found for maximal Ca2+ ATPase activity (mole/mg protein/min) determined spectrophotometrically in RG (0.276+0.03 vs 0.278+0.02). In WG, Ca2+ ATPase activity was 15% higher in WG-S compared to WG-C (0.412+0.03 vs 0.385+0.04). Repetitive stimulation resulted in a reduction in tetanic tension of 74% (p<0.05) by 2 min in the G muscle. By the end of the stimulation period, ATP concentration was reduced (p<0.05) by 57% in the WG and by 47% in the RG. These results indicate that the repeated generation of maximal tetanic force, at least for short term periods, need not adversely affectin vitro homogenate determination of Ca2+ sequestering function in spite of severe alterations in energy potential and that some other mechanism must be involved to explain the depression in Ca2+ uptake and Ca2+ ATPase activity previously noted with short term intense exercise. 相似文献
16.
The Ca(2+)-ATPase activity of rat brain microsomes was studied in streptozotocin (STZ)-induced diabetes. Male rats, 200-250 g, were rendered diabetic by injection of STZ (45 mg kg(-1) body weight) via the teil vein. Brain tissues were collected at 1, 4 and 10 weeks after diabetes was induced for determination of Ca(2+)-ATPase activity, lipid peroxidation and tissue calcium levels. Diabetic rats had significantly elevated blood glucose levels compared to controls. Blood glucose levels were 92.92 +/- 1.22 mg dl(-1) (mean +/- SEM) for the control group, 362.50 +/- 9.61 mg dl(-1) at 1 week and >500 mg dl(-1) at 4, 8 and 10 weeks for the diabetics. Enzyme activities were significantly decreased at 1, 4, 8 and 10 weeks of diabetes relative to the control group (p < 0.001). Ca(2+)-ATPase activity was 0.084 +/- 0.008 U l(-1), 0.029 +/- 0.005 U l(-1), 0.029 +/- 0.006 U l(-1), 0.033 +/- 0.003 U l(-1) and 0.058 +/- 0.006 U l(-1) (mean +/- SEM) at control, 1, 4, 8 and 10 week of diabetes respectively. The change in calcium levels in diabetic rat brain at 8 and 10 weeks of diabetes was significantly higher than that of the control group (p < 0.05). On the other hand lipid peroxidation measured as TBARS (thiobarbituric acid reactive substances) was significantly higher at 8 and 10 weeks of diabetes (p < 0.05). The increase in lipid peroxidation observed in diabetic rat brain may be partly responsible for the decrease in calcium ATPase activity. 相似文献
17.
Changes in the fluoresence ofN-acetyl-N-(5-sulfo-1-naphthyl)ethylenediamine (EDANS), being attached to Cys-674 of sarcoplasmic reticulum Ca2+-ATPase without affecting the catalytic activity, as well as changes in the intrinsic tryptophan fluorescence were followed throughout the catalytic cycle by the steady-state measurements and the stopped-flow spectrofluorometry. EDANS-fluorescence changes reflect conformational changes near the ATP binding site in the cytoplasmic domain, while tryptophan-fluorescence changes most probably reflect conformational changes in or near the transmembrane domain in which the Ca2+ binding sites are located. Formation of the phosphoenzyme intermediates (EP) was also followed by the continuous flow-rapid quenching method. The kinetic analysis of EDANS-fluorescence changes andEP formation revealed that, when ATP is added to the calcium-activated enzyme, conformational changes in the ATP binding site occur in three successive reaction steps; conformational change in the calcium enzyme substrate complex, formation of ADP-sensitiveEP, and transition of ADP-sensitiveEP to ADP-insensitiveEP. In contrast, the ATP-induced tryptophan-fluorescence changes occur only in the latter two steps. Thus, we conclude that conformational changes in the ATP binding site in the cytoplasmic domain are transmitted to the Ca2+-binding sites in the transmembrane domain in these latter two steps.Abbreviations SR
sarcoplasmic reticulum
-
EP
phosphoenzyme
- EDANS
N-acetyl-N-(5-sulfo-1-naphthyl)ethylenediamine
- AMP-PCP
adenosine 5-(, -methylene)triphosphate
- NEM
N-ethylmaleimide 相似文献
18.
Chengjing Cao Timothy Lockwich Terrence L. Scott Robert Blumenthal Adil E. Shamoo 《Molecular and cellular biochemistry》1991,103(2):97-111
Summary In reconstituted rabbit skeletal muscle (Ca2+ + Mg2+)-ATPase proteoliposomes, Ca2+-uptake is decreased by more than 90% with T2 cleavage (Arg-198). However, no difference in the ATP dependence of hydrolysis activity is seen between SR and trypsin-treated SR. A large decrease in E-P formation and hydrolysis activity of the enzyme appear only at T3 cleavage, which represents the cleavage of A1 fragment to A1a + A1b forms. The disappearance of hydrolysis activity due to digestion is prior to the disappearance of E-P formation. No significant difference is found in the passive Ca2+ efflux between control SR and tryptically digested SR in the absence of Mg+ ruthenium red or in the presence of ATP. However, the passive Ca2+ efflux rate for tryptically digested SR is much larger than control SR in the presence of Mg2+ + ruthenium red. These results show that the Ca2+ channel cannot be closed after trypsin digestion of SR membranes by the presence of the Ca2+ channel inhibitors, Mg2+ and ruthenium red. In the reconstituted ATPase proteoliposomes, the Ca2+ efflux rates are the same regardless of digestion (T2); also, efflux is not affected by the presence or absence of Mg2+ + ruthenium red. These results indicate that T2 cleavage causes uncoupling of the Ca2+-pump from ATP hydrolytic activity.A theoretical model is developed in order to fit the extent of tryptic digestion of the A fragment of the (Ca2+ + Mg2+)-ATPase polypeptide with the loss of Ca2+-transport. Fits of the theoretical equations to the data are consistent with that Ca2+-transport system appears to require a dimer of the polypeptide (Ca2+ + Mg2+)-ATPase. 相似文献
19.
Yusuf Olgar Erkan Tuncay Sinan Degirmenci Deniz Billur Rimpy Dhingra Lorrie Kirshenbaum Belma Turan 《Journal of cellular and molecular medicine》2020,24(15):8567-8578
The prevalence of death from cardiovascular disease is significantly higher in elderly populations; the underlying factors that contribute to the age‐associated decline in cardiac performance are poorly understood. Herein, we identify the involvement of sodium/glucose co‐transporter gene (SGLT2) in disrupted cellular Ca2+‐homeostasis, and mitochondrial dysfunction in age‐associated cardiac dysfunction. In contrast to younger rats (6‐month of age), older rats (24‐month of age) exhibited severe cardiac ultrastructural defects, including deformed, fragmented mitochondria with high electron densities. Cardiomyocytes isolated from aged rats demonstrated increased reactive oxygen species (ROS), loss of mitochondrial membrane potential and altered mitochondrial dynamics, compared with younger controls. Moreover, mitochondrial defects were accompanied by mitochondrial and cytosolic Ca2+ ([Ca2+]i) overload, indicative of disrupted cellular Ca2+‐homeostasis. Interestingly, increased [Ca2+]i coincided with decreased phosphorylation of phospholamban (PLB) and contractility. Aged‐cardiomyocytes also displayed high Na+/Ca2+‐exchanger (NCX) activity and blood glucose levels compared with young‐controls. Interestingly, the protein level of SGLT2 was dramatically increased in the aged cardiomyocytes. Moreover, SGLT2 inhibition was sufficient to restore age‐associated defects in [Ca2+]i‐homeostasis, PLB phosphorylation, NCX activity and mitochondrial Ca2+‐loading. Hence, the present data suggest that deregulated SGLT2 during ageing disrupts mitochondrial function and cardiac contractility through a mechanism that impinges upon [Ca2+]i‐homeostasis. Our studies support the notion that interventions that modulate SGLT2‐activity can provide benefits in maintaining [Ca2+]i and cardiac function with advanced age. 相似文献
20.
The effects of tricyclohexyltin hydroxide (Plictran), an organotin acaricide, on 45Ca2+ uptake and Ca2+ ATPase were studied in vitro and in vivo in rat heart ventricular membrane vesicles, primarily sarcoplasmic reticulum. There was a concentration dependent inhibition of both 45Ca2+ uptake and Ca2+ ATPase in vivo as well as in vitro. Isoproterenol, a beta-adrenergic agonist, stimulated 45Ca2+ uptake and Ca2+ ATPase of sarcoplasmic reticulum and this was also inhibited by Plictran. Since cardiac relaxation is mediated by beta-adrenergic stimulation via Ca+ uptake by sarcoplasmic reticulum, the inhibition of calcium pump activity by Plictran may result in alterations in cardiac Ca2+ fluxes leading to cardiac dysfunction. 相似文献