Role of calreticulin in the sensitivity of myocardiac H9c2 cells to oxidative stress caused by hydrogen peroxide

Ihara, Yoshito; Urata, Yoshishige; Goto, Shinji; Kondo, Takahito

doi:10.1152/ajpcell.00075.2005

Cited by 46 publications

(42 citation statements)

References 62 publications

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“…Cultured cells were harvested and lysed in lysis buffer [20 mmol/L Tris-HCl (pH 7.5), 130 mmol/L NaCl, and 1% NP40 including protease inhibitors (20 Amol/L amidinophenyl methanesulfonyl fluoride, 50 Amol/L pepstatin, and 50 Amol/L leupeptin)]. Protein samples were subjected to SDS-PAGE and then transferred to a nitrocellulose membrane as described (21). The membrane was blocked and then incubated with the primary antibody in TBS containing 0.05% Tween 20.…”

Section: Methodsmentioning

confidence: 99%

“…CRT is a multifunctional protein involved in many biological processes that include the regulation of Ca 2+ homeostasis (16), intercellular or intracellular signaling, gene expression (17), glycoprotein folding (18), and nuclear transport (19). We found that overexpression of CRT enhanced apoptosis in myocardiac H9c2 cells under conditions inductive to differentiation with retinoic acid (20) or under oxidative stress (21). Moreover, CRT regulates p53 function to induce apoptosis by affecting the rate of degradation and nuclear localization of p53 (12).…”

Section: Introductionmentioning

confidence: 94%

“…The cytoplasmic free Ca 2+ concentration, [Ca 2+ ] i , was measured with a dual-excitation wavelength spectrofluorophotometer (RF-5500, Shimadzu, Kyoto, Japan) using the fluorescent Ca 2+ indicator Fura-2 tetra (acetoxymethyl) ester (Fura-2-AM) essentially as described previously (21).…”

Section: Camentioning

confidence: 99%

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Calreticulin, a Molecular Chaperone in the Endoplasmic Reticulum, Modulates Radiosensitivity of Human Glioblastoma U251MG Cells

Okunaga

Urata²,

Goto³

et al. 2006

Cancer Research

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Radiotherapy is the primary and most important adjuvant therapy for malignant gliomas. Although the mechanism of radiation resistance in gliomas has been studied for decades, it is still not clear how the resistance is related with functions of molecular chaperones in the endoplasmic reticulum. Calreticulin (CRT) is a Ca 2+ -binding molecular chaperone in the endoplasmic reticulum. Recently, it was reported that changes in intracellular Ca 2+ homeostasis play a role in the modulation of apoptosis. In the present study, we found that the level of CRT was higher in neuroglioma H4 cells than in glioblastoma cells (U251MG and T98G), and was well correlated with the sensitivity to ;-irradiation. To examine the role of CRT in the radiosensitivity of malignant gliomas, the CRT gene was introduced into U251MG cells, which express low levels of CRT, and the effect of overexpression of CRT on the radiosensitivity was examined. The cells transfected with the CRT gene exhibited enhanced radiationinduced apoptosis compared with untransfected control cells. In CRT-overexpressing cells, cell survival signaling via Akt was markedly suppressed. Furthermore, the gene expression of protein phosphatase 2AcA (PP2AcA), which is responsible for the dephosphorylation and inactivation of Akt, was up-regulated in CRT-overexpressing cells, and the regulation was dependent on Ca 2+. Thus, overexpression of CRT modulates radiation-induced apoptosis by suppressing Akt signaling through the up-regulation of PP2AcA expression via altered Ca 2+ homeostasis. These results show the novel mechanism by which CRT is involved in the regulation of radiosensitivity and radiation-induced apoptosis in malignant glioma cells.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 94%

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Calreticulin, a Molecular Chaperone in the Endoplasmic Reticulum, Modulates Radiosensitivity of Human Glioblastoma U251MG Cells

Okunaga

Urata²,

Goto³

et al. 2006

Cancer Research

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“…H9c2 cells were seeded in quadruplicate in 24-well plates and the experimental groups were: i) control: CTRL, absence of any additional treatment; ii) H 2 O 2 : H9c2 were exposed to 50µM H 2 O 2 for 2 hours [20]; iii) CST: cells were exposed to CST 75nM for 3 hours; iv) CST+H 2 O 2 : cells were exposed to CST for 1 hour, then CST plus H 2 O 2 were continued for other 2 hours.…”

Section: Cell Culture and Drug Treatmentmentioning

confidence: 99%

“…In brief, JC-1 (10µM) was added to cultured cells [46] in control and after the treatment with H 2 O 2 (50µM) [20] with or without CST (75 nM). The ratio between red and green JC-1 florescence was taken as an index of mitochondrial membrane potential [25].…”

Section: Measurement Of Mitochondrial Potentialmentioning

confidence: 99%

Catestatin reduces myocardial ischaemia/reperfusion injury: involvement of PI3K/Akt, PKCs, mitochondrial KATP channels and ROS signalling

Perrelli

Tullio

Angotti

et al. 2013

Pflugers Arch - Eur J Physiol

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Aims: Catestatin (CST) limits myocardial ischaemia/reperfusion (I/R) injury with unknown mechanisms. Clearly phosphoinositide-3-kinase (PI3K), protein-kinase-C (PKC) isoforms, including intra-mitochondrial PKCε, mitochondrial-K ATP (mitoK ATP ) channels and subsequent reactiveoxygen-species (ROS)-signalling play important roles in postconditioning cardioprotection, preventing mitochondrial permeability transition pore (mPTP) opening. Therefore, we studied the role of these extra-and intra-mitochondrial factors in CST-induced protection. Methods and Results: Isolated rat hearts and H9c2 cells underwent I/R and oxidative stress, respectively. In isolated hearts CST (75nM, CST-Post) given in early-reperfusion significantly reduced infarct-size, limited post-ischaemic contracture, and improved recovery of developed left ventricular pressure.PI3K inhibitor, LY-294002 (LY), large spectrum PKC inhibitor, Chelerythrine (CHE), specific PKCε inhibitor (εV1-2), mitoK ATP channel blocker, 5-Hydroxydecanoate (5HD) or ROS scavenger, 2-mercaptopropionylglycine (MPG) abolished the infarct-sparing effect of CST. Notably the CSTinduced contracture limitation was maintained during co-infusion of 5HD, MPG or εV1-2, but it was lost during co-infusion of LY or CHE. In H9c2 cells challenged with H 2 O 2 , mitochondrialdepolarization (an index of mPTP-opening studied with JC1-probe) was drastically limited by CST (75nM). Conclusions: Our results suggest that the protective signalling pathway activated by CST includes mitoK ATP channels, ROS-signalling and prevention of mPTP opening, with a central role for upstream PI3K/Akt and PKCs. In fact, all inhibitors completely abolished CST-infarct-sparing effect. Since CST-anti-contracture effect cannot be explained by intra-mitochondrial mechanisms (PKCε activation and mitoK ATP channel opening) or ROS-signalling, it is proposed that these downstream signals are part of a reverberant loop which re-activates upstream PKCs, which therefore play a pivotal role in CST-induced protection.

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Factors controlling the activity of the SERCA2a pump in the normal and failing heart

et al. 2009

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Heart failure is the leading cause of death in western countries and is often associated with impaired Ca(2+) handling in the cardiomyocyte. In fact, cardiomyocyte relaxation and contraction are tightly controlled by the activity of the cardiac sarco(endo)plasmic reticulum (ER/SR) Ca(2+) pump SERCA2a, pumping Ca(2+) from the cytosol into the lumen of the ER/SR. This review addresses three important facets that control the SERCA2 activity in the heart. First, we focus on the alternative splicing of the SERCA2 messenger, which is strictly regulated in the developing heart. This splicing controls the formation of three SERCA2 splice variants with different enzymatic properties. Second, we will discuss the role and regulation of SERCA2a activity in the normal and failing heart. The two well-studied Ca(2+) affinity modulators phospholamban and sarcolipin control the activity of SERCA2a within a narrow window. An aberrantly high or low Ca(2+) affinity is often observed in and may even trigger cardiac failure. Correcting SERCA2a activity might therefore constitute a therapeutic approach to improve the contractility of the failing heart. Finally, we address the controversies and unanswered questions of other putative regulators of the cardiac Ca(2+) pump, such as sarcalumenin, HRC, S100A1, Bcl-2, HAX-1, calreticulin, calnexin, ERp57, IRS-1, and -2.

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Role of calreticulin in the sensitivity of myocardiac H9c2 cells to oxidative stress caused by hydrogen peroxide

Abstract: Ihara, Yoshito, Yoshishige Urata, Shinji Goto, and Takahito Kondo. Role of calreticulin in the sensitivity of myocardiac H9c2 cells to oxidative stress caused by hydrogen peroxide.

Cited by 46 publications

References 62 publications

Calreticulin, a Molecular Chaperone in the Endoplasmic Reticulum, Modulates Radiosensitivity of Human Glioblastoma U251MG Cells

Calreticulin, a Molecular Chaperone in the Endoplasmic Reticulum, Modulates Radiosensitivity of Human Glioblastoma U251MG Cells

Catestatin reduces myocardial ischaemia/reperfusion injury: involvement of PI3K/Akt, PKCs, mitochondrial KATP channels and ROS signalling

Factors controlling the activity of the SERCA2a pump in the normal and failing heart

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