scholarly journals Caspase-Independent Pathway is Related to Nilotinib Cytotoxicity in Cultured Cardiomyocytes

2017 ◽  
Vol 42 (6) ◽  
pp. 2182-2193 ◽  
Author(s):  
Qinghui Yang ◽  
Chunhui Zhang ◽  
Hong Wei ◽  
Zenghui Meng ◽  
Guangnan Li ◽  
...  
Keyword(s):  
Cathepsin B ◽  
Nuclear Translocation ◽  
Morphological Changes ◽  
H9c2 Cells ◽  
Mitochondrial Signaling ◽  
Leukemia Treatment ◽  
Induced Apoptosis ◽  
Apoptosis Inducing Factor ◽  
Cultured Cardiomyocytes

Background/Aims: Cardiotoxicity is a predominant side-effect of nilotinib during chronic myeloid leukemia treatment. The underlying molecular mechanism remains unclear. The role of autophagy and mitochondrial signaling was investigated in nilotinib-treated cardiac H9C2 cells. Methods: Cytotoxicity was assessed using Cell Death Detection kit. Immunoblot and immunofluorescence staining was performed, and cathepsin B and caspase3 activity was assessed in nilotinib-treated H9C2 cells with or without distinct pathway inhibitor or specific siRNA. Results: Nilotinib time- and dose-dependently induced H9C2 apoptosis, which was not completely prevented by the pan caspase inhibitor z-VAD-fmk. Following nilotinib treatment, mitochondrial membrane potential decreased significantly accompanied with remarkable morphological changes. Nuclear translocation of mitochondrial apoptosis inducing factor (AIF) and increased p53 was detected in nilotinib-treated cells. AIF knockdown prevented nilotinib-induced increase of p53 and apoptosis. Additionally, increased cathepsin B activity was detected, and inhibition of cathepsin B by CA-074Me prevented nilotinib-induced apoptosis and nuclear translocation of AIF. Moreover, increased Atg5 and transition of LC3-I to LC3-II was revealed following nilotinib treatment. Increased cathepsin B activity and apoptosis by nilotinib was significantly prohibited by specific autophagy inhibitor bafilomycin A and Atg5 knockdown. Conclusion: Our findings demonstrate that nilotinib increases autophagy and cathepsin B activity, leading to mitochondrial AIF release and nuclear translocation, which is responsible for p53 and apoptosis induction in H9C2 cells.

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

2006 ◽  
Vol 290 (1) ◽  
pp. C208-C221 ◽  
Author(s):  
Yoshito Ihara ◽  
Yoshishige Urata ◽  
Shinji Goto ◽  
Takahito Kondo
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Molecular Chaperone ◽  
Vital Role ◽  
H9c2 Cells ◽  
Cardiac Physiology ◽  
Caspase 12 ◽  
A Cell ◽  
Induced Apoptosis

Calreticulin (CRT), a Ca2+-binding molecular chaperone in the endoplasmic reticulum, plays a vital role in cardiac physiology and pathology. Oxidative stress is a main cause of myocardiac apoptosis in the ischemic heart, but the function of CRT under oxidative stress is not fully understood. In the present study, the effect of overexpression of CRT on susceptibility to apoptosis under oxidative stress was examined using myocardiac H9c2 cells transfected with the CRT gene. Under oxidative stress due to H2O2, the CRT-overexpressing cells were highly susceptible to apoptosis compared with controls. In the overexpressing cells, the levels of cytoplasmic free Ca2+ ([Ca2+]i) were significantly increased by H2O2, whereas in controls, only a slight increase was observed. The H2O2-induced apoptosis was enhanced by the increase in [Ca2+]i caused by thapsigargin in control cells but was suppressed by BAPTA-AM, a cell-permeable Ca2+ chelator in the CRT-overexpressing cells, indicating the importance of the level of [Ca2+]i in the sensitivity to H2O2-induced apoptosis. Suppression of CRT by the introduction of the antisense cDNA of CRT enhanced cytoprotection against oxidative stress compared with controls. Furthermore, we found that the levels of activity of calpain and caspase-12 were elevated through the regulation of [Ca2+]i in the CRT-overexpressing cells treated with H2O2 compared with controls. Thus we conclude that the level of CRT regulates the sensitivity to apoptosis under oxidative stress due to H2O2 through a change in Ca2+ homeostasis and the regulation of the Ca2+-calpain-caspase-12 pathway in myocardiac cells.

scholarly journals Activation of nuclear factor-κB during doxorubicin-induced apoptosis in endothelial cells and myocytes is pro-apoptotic: the role of hydrogen peroxide

2002 ◽  
Vol 367 (3) ◽  
pp. 729-740 ◽  
Author(s):  
Suwei WANG ◽  
Srigiridhar KOTAMRAJU ◽  
Eugene KONOREV ◽  
Shasi KALIVENDI ◽  
Joy JOSEPH ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Nuclear Translocation ◽  
Nuclear Factor Κb ◽  
Nuclear Factor ◽  
Apoptotic Pathway ◽  
Toxic Side Effect ◽  
Rat Cardiomyocytes ◽  
A Cell ◽  
Induced Apoptosis

Doxorubicin (DOX) is a widely used anti-tumour drug. Cardiotoxicity is a major toxic side effect of DOX therapy. Although recent studies implicated an apoptotic pathway in DOX-induced cardiotoxicity, the mechanism of DOX-induced apoptosis remains unclear. In the present study, we investigated the role of reactive oxygen species and the nuclear transcription factor nuclear factor κB (NF-κB) during apoptosis induced by DOX in bovine aortic endothelial cells (BAECs) and adult rat cardiomyocytes. DOX-induced NF-κB activation is both dose- and time-dependent, as demonstrated using electrophoretic mobility-shift assay and luciferase and p65 (Rel A) nuclear-translocation assays. Addition of a cell-permeant iron metalloporphyrin significantly suppressed NF-κB activation and apoptosis induced by DOX. Overexpression of glutathione peroxidase, which detoxifies cellular H2O2, significantly decreased DOX-induced NF-κB activation and apoptosis. Inhibition of DOX-induced NF-κB activation by a cell-permeant peptide SN50 that blocks translocation of the NF-κB complex into the nucleus greatly diminished DOX-induced apoptosis. Apoptosis was inhibited when IκB mutant vector, another NF-κB inhibitor, was added to DOX-treated BAECs. These results suggest that NF-κB activation in DOX-treated endothelial cells and myocytes is pro-apoptotic, in contrast with DOX-treated cancer cells, where NF-κB activation is anti-apoptotic. Removal of intracellular H2O2 protects endothelial cells and myocytes from DOX-induced apoptosis, possibly by inhibiting NF-κB activation. These findings suggest a novel mechanism for enhancing the therapeutic efficacy of DOX.

scholarly journals Repression of BIRC5/Survivin by FOXO3/FKHRL1 Sensitizes Human Neuroblastoma Cells to DNA Damage-induced Apoptosis

2009 ◽  
Vol 20 (7) ◽  
pp. 2041-2048 ◽  
Author(s):  
Petra Obexer ◽  
Judith Hagenbuchner ◽  
Thomas Unterkircher ◽  
Nora Sachsenmaier ◽  
Christoph Seifarth ◽  
...  
Keyword(s):  
Nuclear Translocation ◽  
Chemotherapy Resistance ◽  
Neuroblastoma Cells ◽  
Phosphatidylinositol 3 Kinase ◽  
Human Neuroblastoma Cells ◽  
Hairpin Rna ◽  
Human Neuroblastoma ◽  
Dna Damaging Agents ◽  
Induced Apoptosis

The phosphatidylinositol 3-kinase (PI3K)–protein kinase B (PKB) pathway regulates survival and chemotherapy resistance of neuronal cells, and its deregulation in neuroblastoma (NB) tumors predicts an adverse clinical outcome. Here, we show that inhibition of PI3K-PKB signaling in human NB cells induces nuclear translocation of FOXO3/FKHRL1, represses the prosurvival protein BIRC5/Survivin, and sensitizes to DNA-damaging agents. To specifically address whether FKHRL1 contributes to Survivin regulation, we introduced a 4-hydroxy-tamoxifen-regulated FKHRL1(A3)ERtm allele into NB cells. Conditional FKHRL1 activation repressed Survivin transcription and protein expression. Transgenic Survivin exerted a significant antiapoptotic effect and prevented the accumulation of Bim and Bax at mitochondria, the loss of mitochondrial membrane potential as well as the release of cytochrome c during FKHRL1-induced apoptosis. In concordance, Survivin knockdown by retroviral short hairpin RNA technology accelerated FKHRL1-induced apoptosis. Low-dose activation of FKHRL1 sensitized to the DNA-damaging agents doxorubicin and etoposide, whereas the overexpression of Survivin diminished FKHRL1 sensitization to these drugs. These results suggest that repression of Survivin by FKHRL1 facilitates FKHRL1-induced apoptosis and sensitizes to cell death induced by DNA-damaging agents, which supports the central role of PI3K-PKB-FKHRL1 signaling in drug resistance of human NB.

scholarly journals p31 Deficiency Influences Endoplasmic Reticulum Tubular Morphology and Cell Survival

2009 ◽  
Vol 29 (7) ◽  
pp. 1869-1881 ◽  
Author(s):  
Takefumi Uemura ◽  
Takashi Sato ◽  
Takehiro Aoki ◽  
Akitsugu Yamamoto ◽  
Tetsuya Okada ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Er Stress ◽  
Nuclear Translocation ◽  
Marked Change ◽  
Conditional Knockout ◽  
Induced Apoptosis ◽  
Factor C ◽  
Er Membrane

ABSTRACT p31, the mammalian orthologue of yeast Use1p, is an endoplasmic reticulum (ER)-localized soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (SNARE) that forms a complex with other SNAREs, particularly syntaxin 18. However, the role of p31 in ER function remains unknown. To determine the role of p31 in vivo, we generated p31 conditional knockout mice. We found that homozygous deletion of the p31 gene led to early embryonic lethality before embryonic day 8.5. Conditional knockout of p31 in brains and mouse embryonic fibroblasts (MEFs) caused massive apoptosis accompanied by upregulation of ER stress-associated genes. Microscopic analysis showed vesiculation and subsequent enlargement of the ER membrane in p31-deficient cells. This type of drastic disorganization in the ER tubules has not been demonstrated to date. This marked change in ER structure preceded nuclear translocation of the ER stress-related transcription factor C/EBP homologous protein (CHOP), suggesting that ER stress-induced apoptosis resulted from disruption of the ER membrane structure. Taken together, these results suggest that p31 is an essential molecule involved in the maintenance of ER morphology and that its deficiency leads to ER stress-induced apoptosis.

scholarly journals Arp2/3 complex activity is necessary for mouse ESC differentiation, times formative pluripotency, and enables lineage specification

2021 ◽  
Author(s):  
Francesca M Aloisio ◽  
Diane L Barber
Keyword(s):  
Stem Cells ◽  
Embryonic Development ◽  
Nuclear Translocation ◽  
Morphological Changes ◽  
Embryonic Stem ◽  
Complex Activity ◽  
Lineage Specification ◽  
Epithelial Plasticity ◽  
Established Role

Mouse embryonic stem cells (mESCs), a model for differentiation into primed epiblast-like cells (EpiLCs), have revealed transcriptional and epigenetic control of early embryonic development. The control and significance of morphological changes, however, remain less defined. We show marked changes in morphology and actin architectures during differentiation that depend on Arp2/3 complex but not formin activity. Inhibiting Arp2/3 complex activity pharmacologically or genetically does not block exit from naive pluripotency but attenuates increases in EpiLC markers. We find that inhibiting Arp2/3 complex activity delays formative pluripotency and causes globally defective lineage specification as indicated by RNA-sequencing, with significant effects on TBX3-depedendent transcriptional programs. We also identify two previously unreported indicators of mESC differentiation; MRTF and FHL2, which have inverse Arp2/3 complex-dependent nuclear translocation. Our findings on Arp2/3 complex activity in differentiation and the established role of formins in EMT indicate that these two actin nucleators regulate distinct modes of epithelial plasticity.

scholarly journals The Role of Sirtuin 1 in Palmitic-acid-induced Endoplasmic-reticulum Stress in Cardiac Myoblasts

2021 ◽  
Author(s):  
Hsiang-Yu Yang ◽  
Jhao-Ying Chen ◽  
Yen-Nien Huo ◽  
Pei-Ling Yu ◽  
Pei-Zhen Lin ◽  
...  
Keyword(s):  
Er Stress ◽  
Knockout Mice ◽  
High Fat Diet ◽  
Nuclear Translocation ◽  
Sirtuin 1 ◽  
H9c2 Cells ◽  
High Fat ◽  
Chop Expression ◽  
Sirt1 Activator

Abstract Background: Lipotoxicity causes endoplasmic reticulum (ER) stress, leading to cell apoptosis. Sirtuin 1 (Sirt1) regulates gene transcription and cellular metabolism. In this study, we investigated the role of Sirt1 in palmitate-induced ER stress. Methods: H9c2 myoblasts and heart-specific Sirt1 knockout mice fed a palmitate-enriched high-fat diet were used. Results: The high-fat diet induced C/EBP homologous protein (CHOP) and activating transcription factor 4 (ATF4) expression in both Sirt1 knockout mice and controls. Sirt1 knockout mice showed higher CHOP and ATF4 expression compared to those in control. Palmitic acid (PA) induced ATF4 and CHOP expression in H9c2 cells. PA-treated H9c2 cells showed decreased cytosolic NAD+/NADH alongside reduced Sirt1’s activity. H9c2 cells showed increased ATF4 and CHOP expression when transfected with plasmid encoding dominant negative mutant Sirt1. Sirt1 activator SRT1720 did not affect CHOP and ATF4 expression. Although SRT1720 enhanced nuclear translocation of ATF4, the extent of the binding of ATF4 to the CHOP promoter did not increase further in PA treated-H9c2 cells.Conclusion: PA-induced ER stress is mediated through upregulation of ATF4 and CHOP. Cytosolic NAD+ concentration is diminished by PA-induced ER stress, leading to decreased Sirt1 activity. The Sirt1 activator SRT1720 promotes nuclear translocation of ATF4 in PA-treated H9c2 cells.

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