Apoptotic Protease Activating Factor 1 (Apaf-1)–Independent Cell Death Suppression by Bcl-2

Reportedly, antiapoptotic Bcl-2 family proteins suppress apoptosis by binding to and inhibiting members of the CED-4 family of caspase activators. To explore this question, we used embryonic stem (ES) cells in which one (−/+) or both (−/−) copies of the gene encoding apoptotic protease activating factor 1 (Apaf-1), a CED-4 homologue, were disrupted by homologous recombination. Stable clones of heterozygous (−/+) and homozygous (−/−) Apaf-1 knockout ES cells that overexpressed Bcl-2 were generated. Withdrawal of serum growth factors or stimulation of heterozygous ES cells with staurosporine (STS), ultraviolet (UV)B irradiation, etoposide (VP16), or cisplatin induced apoptosis followed by cell death (determined by failure to exclude propidium iodide dye). These cell death stimuli also induced activation of several types of caspases and loss of mitochondrial membrane potential (ΔΨ) in heterozygous (+/−) Apaf-1 knockout ES cells. In addition, overexpression of Bcl-2 protected against these events in Apaf-1–expressing ES cells. In contrast, STS, UVB, and VP16 induced little or no caspase activation and apoptosis in homozygous (−/−) Apaf-1 knockout ES cells. Nevertheless, Apaf-1–deficient ES cells subjected to these cell death stimuli or deprived of growth factors did eventually die through a nonapoptotic mechanism associated with loss of ΔΨ. Moreover, Bcl-2 overprotection preserved ΔΨ, reduced the percentage of Apaf-1−/− ES cells undergoing cell death, and increased clonigenic survival. The extent of Bcl-2–mediated cytoprotection was not significantly different for heterozygous (−/+) versus homozygous (−/−) Apaf-1 knockout cells. Furthermore, although Bcl-2 could be readily coimmunoprecipitated with Bax, associations with Apaf-1 were undetectable under conditions where Apaf-1 interactions with procaspase-9 were observed. We conclude that Bcl-2 has cytoprotective functions independent of Apaf-1, preserving mitochondrial function through a caspase-independent mechanism.

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Deficiency of the Stress Kinase P38α Results in Embryonic Lethality

Journal of Experimental Medicine ◽

10.1084/jem.191.5.859 ◽

2000 ◽

Vol 191 (5) ◽

pp. 859-870 ◽

Cited By ~ 207

Author(s):

Melanie Allen ◽

Linne Svensson ◽

Marsha Roach ◽

John Hambor ◽

John McNeish ◽

...

Keyword(s):

Map Kinase ◽

Es Cells ◽

Interleukin 1 ◽

Embryonic Stem ◽

Wild Type ◽

Antiinflammatory Drugs ◽

Gene Encoding ◽

Significant Difference ◽

Induced Apoptosis

The mitogen-activated protein (MAP) kinase p38 is a key component of stress response pathways and the target of cytokine-suppressing antiinflammatory drugs (CSAIDs). A genetic approach was employed to inactivate the gene encoding one p38 isoform, p38α. Mice null for the p38α allele die during embryonic development. p38α1/− embryonic stem (ES) cells grown in the presence of high neomycin concentrations demonstrated conversion of the wild-type allele to a targeted allele. p38α−/− ES cells lacked p38α protein and failed to activate MAP kinase–activated protein (MAPKAP) kinase 2 in response to chemical stress inducers. In contrast, p38α1/+ ES cells and primary embryonic fibroblasts responded to stress stimuli and phosphorylated p38α, and activated MAPKAP kinase 2. After in vitro differentiation, both wild-type and p38α−/− ES cells yielded cells that expressed the interleukin 1 receptor (IL-1R). p38α1/+ but not p38α−/− IL-1R–positive cells responded to IL-1 activation to produce IL-6. Comparison of chemical-induced apoptosis processes revealed no significant difference between the p38α1/+ and p38α−/− ES cells. Therefore, these studies demonstrate that p38α is a major upstream activator of MAPKAP kinase 2 and a key component of the IL-1 signaling pathway. However, p38α does not serve an indispensable role in apoptosis.

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Caspase-8 deficiency induces a switch from TLR3 induced apoptosis to lysosomal cell death in neuroblastoma

Scientific Reports ◽

10.1038/s41598-021-89793-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Marie-Anaïs Locquet ◽

Gabriel Ichim ◽

Joseph Bisaccia ◽

Aurelie Dutour ◽

Serge Lebecque ◽

...

Keyword(s):

Cell Death ◽

Cancer Cells ◽

Neuroblastoma Cell ◽

Death Receptor ◽

Neuroblastoma Cell Line ◽

Caspase 8 ◽

Caspase Activation ◽

Extrinsic Pathway ◽

Lysosomal Membrane Permeabilization ◽

Induced Apoptosis

AbstractIn cancer cells only, TLR3 acquires death receptor properties by efficiently triggering the extrinsic pathway of apoptosis with Caspase-8 as apical protease. Here, we demonstrate that in the absence of Caspase-8, activation of TLR3 can trigger a form of programmed cell death, which is distinct from classical apoptosis. When TLR3 was activated in the Caspase-8 negative neuroblastoma cell line SH-SY5Y, cell death was accompanied by lysosomal permeabilization. Despite caspases being activated, lysosomal permeabilization as well as cell death were not affected by blocking caspase-activity, positioning lysosomal membrane permeabilization (LMP) upstream of caspase activation. Taken together, our data suggest that LMP with its deadly consequences represents a “default” death mechanism in cancer cells, when Caspase-8 is absent and apoptosis cannot be induced.

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Reactive oxygen species and caspase activation mediate silica-induced apoptosis in alveolar macrophages

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.2001.280.1.l10 ◽

2001 ◽

Vol 280 (1) ◽

pp. L10-L17 ◽

Cited By ~ 32

Author(s):

Han-Ming Shen ◽

Zhuo Zhang ◽

Qi-Feng Zhang ◽

Choon-Nam Ong

Keyword(s):

Reactive Oxygen Species ◽

Cell Death ◽

Alveolar Macrophages ◽

Caspase 3 ◽

Reactive Oxygen ◽

Parp Cleavage ◽

Oxygen Species ◽

Caspase Activation ◽

Caspase 9 ◽

Induced Apoptosis

Alveolar macrophages (AMs) are the principal target cells of silica and occupy a key position in the pathogenesis of silica-related diseases. Silica has been found to induce apoptosis in AMs, whereas its underlying mechanisms involving the initiation and execution of apoptosis are largely unknown. The main objective of the present study was to examine the form of cell death caused by silica and the mechanisms involved. Silica-induced apoptosis in AMs was evaluated by terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling assay and cell cycle/DNA content analysis. The elevated level of reactive oxygen species (ROS), caspase-9 and caspase-3 activation, and poly(ADP-ribose) polymerase (PARP) cleavage in silica-treated AMs were also determined. The results showed that there was a temporal pattern of apoptotic events in silica-treated AMs, starting with ROS formation and followed by caspase-9 and caspase-3 activation, PARP cleavage, and DNA fragmentation. Silica-induced apoptosis was significantly attenuated by a caspase-3 inhibitor, N-acetyl-Asp-Glu-Val-Asp aldehyde, and ebselen, a potent antioxidant. These findings suggest that apoptosis is an important form of cell death caused by silica exposure in which the elevated ROS level that results from silica exposure may act as an initiator, leading to caspase activation and PARP cleavage to execute the apoptotic process.

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Generation and Characterization of Smac/DIABLO-Deficient Mice

Molecular and Cellular Biology ◽

10.1128/mcb.22.10.3509-3517.2002 ◽

2002 ◽

Vol 22 (10) ◽

pp. 3509-3517 ◽

Cited By ~ 120

Author(s):

Hitoshi Okada ◽

Woong-Kyung Suh ◽

Jianping Jin ◽

Minna Woo ◽

Chunying Du ◽

...

Keyword(s):

Physiological Function ◽

Es Cells ◽

Embryonic Stem ◽

Caspase Activation ◽

Proapoptotic Protein ◽

Apoptosis Proteins ◽

Deficient Mice

ABSTRACT The mitochondrial proapoptotic protein Smac/DIABLO has recently been shown to potentiate apoptosis by counteracting the antiapoptotic function of the inhibitor of apoptosis proteins (IAPs). In response to apoptotic stimuli, Smac is released into the cytosol and promotes caspase activation by binding to IAPs, thereby blocking their function. These observations have suggested that Smac is a new regulator of apoptosis. To better understand the physiological function of Smac in normal cells, we generated Smac-deficient (Smac−/− ) mice by using homologous recombination in embryonic stem (ES) cells. Smac−/− mice were viable, grew, and matured normally and did not show any histological abnormalities. Although the cleavage in vitro of procaspase-3 was inhibited in lysates of Smac−/− cells, all types of cultured Smac−/− cells tested responded normally to all apoptotic stimuli applied. There were also no detectable differences in Fas-mediated apoptosis in the liver in vivo. Our data strongly suggest the existence of a redundant molecule or molecules capable of compensating for a loss of Smac function.

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DMPK is a New Candidate Mediator of Tumor Suppressor p53-Dependent Cell Death

Molecules ◽

10.3390/molecules24173175 ◽

2019 ◽

Vol 24 (17) ◽

pp. 3175

Author(s):

Katsuhiko Itoh ◽

Takahiro Ebata ◽

Hiroaki Hirata ◽

Takeru Torii ◽

Wataru Sugimoto ◽

...

Keyword(s):

Dna Damage ◽

Cell Death ◽

Tumor Suppressor ◽

Dependent Manner ◽

Tumor Suppressor P53 ◽

P53 Family ◽

Gene Encoding ◽

Myotonic Dystrophy Protein Kinase ◽

Induced Apoptosis ◽

Actomyosin Contraction

Tumor suppressor p53 plays an integral role in DNA-damage induced apoptosis, a biological process that protects against tumor progression. Cell shape dramatically changes when cells undergo apoptosis, which is associated with actomyosin contraction; however, it remains entirely elusive how p53 regulates actomyosin contraction in response to DNA-damaging agents. To identify a novel p53 regulating gene encoding the modulator of myosin, we conducted DNA microarray analysis. We found that, in response to DNA-damaging agent doxorubicin, expression of myotonic dystrophy protein kinase (DMPK), which is known to upregulate actomyosin contraction, was increased in a p53-dependent manner. The promoter region of DMPK gene contained potential p53-binding sequences and its promoter activity was increased by overexpression of the p53 family protein p73, but, unexpectedly, not of p53. Furthermore, we found that doxorubicin treatment induced p73 expression, which was significantly attenuated by downregulation of p53. These data suggest that p53 induces expression of DMPK through upregulating p73 expression. Overexpression of DMPK promotes contraction of the actomyosin cortex, which leads to formation of membrane blebs, loss of cell adhesion, and concomitant caspase activation. Taken together, our results suggest the existence of p53-p73-DMPK axis which mediates DNA-damage induced actomyosin contraction at the cortex and concomitant cell death.

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The U95 Protein of Human Herpesvirus 6B Interacts with Human GRIM-19: Silencing of U95 Expression Reduces Viral Load and Abrogates Loss of Mitochondrial Membrane Potential

Journal of Virology ◽

10.1128/jvi.01156-07 ◽

2007 ◽

Vol 82 (2) ◽

pp. 1011-1020 ◽

Cited By ~ 23

Author(s):

W. M. Yeo ◽

Yuji Isegawa ◽

Vincent T. K. Chow

Keyword(s):

Cell Death ◽

Membrane Potential ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Human Herpesvirus ◽

Ultrastructural Changes ◽

Gene Encoding ◽

Interacting Protein ◽

Functional Aspects ◽

Infected Cells

ABSTRACT To better understand the pathogenesis of human herpesvirus 6 (HHV-6), it is important to elucidate the functional aspects of immediate-early (IE) genes at the initial phase of the infection. To study the functional role of the HHV-6B IE gene encoding U95, we generated a U95-Myc fusion protein and screened a pretransformed bone marrow cDNA library for U95-interacting proteins, using yeast-two hybrid analysis. The most frequently appearing U95-interacting protein identified was GRIM-19, which belongs to the family of genes associated with retinoid-interferon mortality and serves as an essential component of the oxidative phosphorylation system. This interaction was verified by both coimmunoprecipitation and confocal microscopic coimmunolocalization. Short-term HHV-6B infection of MT-4 T-lymphocytic cells induced syncytial formation, resulted in decreased mitochondrial membrane potential, and led to progressively pronounced ultrastructural changes, such as mitochondrial swelling, myelin-like figures, and a loss of cristae. Compared to controls, RNA interference against U95 effectively reduced the U95 mRNA copy number and abrogated the loss of mitochondrial membrane potential. Our results indicate that the high affinity between U95 early viral protein and GRIM-19 may be closely linked to the detrimental effect of HHV-6B infection on mitochondria. These findings may explain the alternative cell death mechanism of expiration, as opposed to apoptosis, observed in certain productively HHV-6B-infected cells. The interaction between U95 and GRIM-19 is thus functionally and metabolically significant in HHV-6B-infected cells and may be a means through which HHV-6B modulates cell death signals by interferon and retinoic acid.

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CD27-Mediated Apoptosis Is Dependent on Siva-Induced Caspase Activation in Human Multiple Myeloma.

Blood ◽

10.1182/blood.v106.11.3398.3398 ◽

2005 ◽

Vol 106 (11) ◽

pp. 3398-3398 ◽

Cited By ~ 1

Author(s):

Yu-Tzu Tai ◽

Xian-Feng Li ◽

Rory Coffey ◽

Iris Breitkreutz ◽

Laurence Catley ◽

...

Keyword(s):

Multiple Myeloma ◽

Cell Death ◽

Cell Growth ◽

Growth Inhibition ◽

Plasma Cells ◽

Death Domain ◽

Caspase Activation ◽

Growth And Survival ◽

Induced Apoptosis ◽

Cell Growth And Survival

Abstract CD27, a member of tumor necrosis factor receptor superfamily that lacks a death domain in its cytoplasmic region, and its interaction with its ligand, CD70, is crucial for differentiation into plasma cells. In malignant B cells, aberrant expression and reverse signaling of CD70 might contribute to disease progression. Recent studies showed that CD27 is heterogeneously expressed on multiple myeloma (MM) plasma cells and the expression is reduced with the progression of MM. However, a possible role for the loss of CD27-CD70 interaction in myelomagenesis was never defined. In this study, we identify functional significance of CD27-CD70 interaction in 4 CD27-expressing MM lines and define mechanisms regulating CD27-mediated MM cell death. Using RT-PCR and flow cytometric analysis, we first found that all of MM lines highly express CD70 (n=10) and 4 MM lines 12BM, 12PE, 28BM, 28PE express CD27 on the cell surface. We next evaluated the effect of CD27 ligation, by CD70-transfected NIH3T3 cells (CD70 transfectant), on [3H] thymidine incorporation by CD27-expressing MM lines. CD27 ligation by CD70 transfectants inhibited DNA synthesis in these 4 CD27-expressing MM lines, but not the control transfectants. Conversely, a blocking anti-CD70 mAb blocked CD27-mediated growth inhibition in a dose-dependent manner, indicating induced growth inhibition specific triggered by CD27-CD70 interaction. Using MTT assay, CD27 ligation by CD70 transfectant also inhibited MM cell survival. IL-6 (20 ng/ml) could overcome the inhibitory effect triggered by CD27 ligation on MM cell growth and survival. In addition, CD27 ligation further enhanced Dex-induced MM cell death. Importantly, CD27-mediated MM cell death was also observed in 2 CD27-expressing patient MM cells. Since Siva is a death domain-containing proapoptotic protein identified as an intracellular ligand of CD27, we investigated its role in CD27-mediated apoptosis in MM cells. Overexpression of Siva by transducing adenovirus-expressing Siva (Ad-Siva-GFP) in 12BM MM line is sufficient to induce cell death whereas control adenovirus (Ad-GFP) transduction did not alter 12BM cell growth and survival. CD27 ligation by CD70 transfectants on Siva-overexpressing 12BM cells further enhanced Siva-induced apoptosis, as evidenced by increased subG0 fraction in cell cycle analysis. Thus, the apoptosis triggered by Siva overexpression was related to the CD27-mediated apoptotic pathway. We further determined caspase involvement in the Siva-induced apoptosis in the absence and presence of CD70 transfectants. Caspase 8 and caspase 9 activities were detected 24h following Ad-Siva-GFP transduction in 12BM cells, whereas caspas-3 activity was detected 48h after transduction. Coculture of Ad-Siva-GFP-transduced 12BM cells with CD70 transfectant further enhanced caspase activities. Therefore, overexpression of Siva is sufficient to induce apoptosis and CD27-mediated apoptosis is mediated by Siva-dependent caspase activation in MM. Furthermore, these results suggest that lack of CD27 may lead to evasion of apoptosis in human MM.

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Ultrastructural Evidences of Caspase-Dependent Platelet Generation in ES Cell-Differentiation System.

Blood ◽

10.1182/blood.v106.11.3621.3621 ◽

2005 ◽

Vol 106 (11) ◽

pp. 3621-3621

Author(s):

Yumiko Matsubara ◽

Mitsuru Murata ◽

Hidenori Suzuki ◽

Tamihiro Kamata ◽

Aya Shimizu ◽

...

Keyword(s):

Electron Microscopy ◽

Cell Death ◽

Morphological Changes ◽

Es Cells ◽

Chromatin Condensation ◽

Embryonic Stem ◽

Morphological Evidence ◽

Hematopoietic Stem ◽

Platelet Production ◽

Sequential Experiments

Abstract Platelets are essential for thrombosis and hemostasis, and the elucidating the unique mechanism for platelet production from megakaryocyte is of major importance. However, extensive data on platelet generation have yet to be fully documented because it has been extremely difficult to obtain the sufficient amounts of hematopoietic stem cells. In this study, we therefore used murine embryonic stem (ES) cells that can proliferate and differentiate to megakaryocyte in the presence of thrombopoietin in vitro. ES-derived megakaryocytes and platelets were studied in detail by morphological analyses, and we especially focused on the relationship between cell death of megakaryocytes and platelet production in sequential experiments. A coculture system with ES/OP9 cells, stromal cells derived from M-CSF deficient mice, in the presence of thrombopoietin was used to differentiate mesodermal-like hematopoietic progenitors, immature megakaryocytes, mature megakaryocytes, and functional platelets from ES cells on days 5, 8, 12, and 15 of the culture, respectively, and the peak of cell count was observed on the day 12 for megakaryocytes and the day 15 for platelets. These were confirmed by morphology, Wright-Giemsa staining, CD41expression or fibrinogen binding assays. Interestingly, electron microscopy and immuno-electron microscopy during platelet production showed morphological changes supported by both “proplatelet theory” and “explosive fragmentation theory”, which have been controversial issues related to platelet biogenesis. On the days 8 and 12, megakaryocyte with tube-like (proplatelet-like) extensions of the periphery showed the expression of CD41, CD42c, or von Willebrand factor, in contrast, megakaryocyte-like large cells with smooth periphery had no expression of CD41 and CD42c. On the days 12 and 15, global fragmentation of megakaryocyte cytoplasm into individual platelets was frequently observed. For cell death of megakaryocytes, cells exhibit clear morphological evidence of nuclear change: chromatin condensation, typical of early apoptosis, on the day 8 and extensive condensation and apoptotic nucleus surrounded by a shortrim of cytoplasm in continuity with a portion of granulated cytoplasm on the days 12 and 15. These proceedings of cell death were confirmed by the TUNEL assay in each stage. Also, low production of platelet was observed by adding Z-DEVD-fmk, an inhibitor of caspases −3 and −7, on the day 8. Next, we examined the caspase activation in the different stage of the platelet production by western blotting, and anti-CD41 antibody was used to test the purity of the meg-lineage cells in each stage. Peak expressions of activated caspases −12, −9, and −7 or caspase 10 levels were observed on the day 8. Peak expression of activated caspase 3 was observed on the day 12. On the contrary, no different levels of caspase 6 were shown between the days 8 and 15. Together, present studies in the sequential experiments for megakaryocyte differentiation and platelet production develop previous theories for platelet generation and also suggest that the process for platelet production is focally associated with caspases −12, −10, −9, −7, and −3 dependent cell death of mrgakaryocytes.

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BMP4 and TGFbeta Differentially Regulate CD34+ Progenitor Development in Human Embryonic Stem Cells through SMAD-Dependent Pathway

Blood ◽

10.1182/blood.v112.11.889.889 ◽

2008 ◽

Vol 112 (11) ◽

pp. 889-889

Author(s):

ZacK Z. Wang ◽

Hao Bai ◽

Melanie Arzigian ◽

Yong-Xing Gao ◽

Wen-Shu Wu

Keyword(s):

Stem Cells ◽

Smooth Muscle ◽

Growth Factors ◽

Progenitor Cells ◽

Embryoid Body ◽

Es Cells ◽

Embryonic Stem ◽

Ips Cells ◽

Bmp Signaling ◽

Cd34 Cells

Abstract Pluripotent stem cells derived from patients, including embryonic stem (ES) cells and “induced pluripotent stem” (iPS) cells, are a promising area of regenerative medical research. A major roadblock toward human clinical therapies using ES cells or iPS cells is to define the factors that direct ES cell differentiation into lineage specific cells. We previously established a simple and efficient human embryonic stem cell (hESC) differentiation system to generate CD34+/CD31+ progenitor cells that gave rise to hematopoietic and endothelial cells (Nat Biotech.25:317, 2007). To advance potential clinical application and to define the effects of growth factors on hematopoietic and vascular differentiation, we assessed hESC differentiation on human feeder cells in serum-free condition without intermediate embryoid body (EB) formation. We investigated the roles of BMPs, TGFbeta, VEGF, and FGF2 in directing hESC differentiation. Growth factors were added into culture at different time points to test their stage specific roles. Our study demonstrated that BMP proteins, including BMP2, BMP4, and BMP7, but not BMP9, had synergic effects to VEGF and FGF-2 on hESC differentiation to CD34+/CD31+ progenitor cells. BMP4 was essential to initial CD34+/CD31+ cell development, whereas VEGF and FGF2 promoted the differentiation in later stage, suggesting the sequential roles of BMP4, VEGF and FGF2 in directing hESC differentiation to CD34+/CD31+ progenitor cells. TGFbeta or activin promoted hESC differentiation into CD34+/CD31− cells that were unable to give rise to hematopoietic, endothelial, and smooth muscle cells. Furthermore, TGFbeta or activin activated Smad2/3 signaling, and suppressed BMP4-induced CD34+/CD31+ cells. Microarray analysis revealed that BMP4-induced CD34+ cells expressed hematopoietic, endothelial and smooth muscle genes, including GATA2, gamma globins, VE-Cad, KDR, CD31, Tie2, and aortic smooth muscle actin, whereas TGFbeta-induced CD34+ cells expressed pluripotent markers and endoderm markers, including Oct3/4, Sox2, and Nanog, HHEX, GATA6, and FoxA2. Both canonical BMP signaling (Smad1/5/8-dependent) and non-canonical BMP signaling (p38 MAPK and p42 ERK pathway) were activated by BMP4 in hESCs. Dorsomorphin specifically inhibited BMP4-mediated phosphorylation of Smad1/5/8, and blocked hESC differentiation into CD34+/CD31+ cells. In summary, BMPs and TGFbeta regulate distinct populations of CD34+ cells in hESCs. BMP-Smad1/5/8 pathway is critical for hematopoietic and vascular progenitor development.

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UPR-mediated TRIB3 expression correlates with reduced AKT phosphorylation and inability of interleukin 6 to overcome palmitate-induced apoptosis in RINm5F cells

Journal of Endocrinology ◽

10.1677/joe-09-0356 ◽

2010 ◽

Vol 206 (2) ◽

pp. 183-193 ◽

Cited By ~ 19

Author(s):

José Edgar Nicoletti-Carvalho ◽

Tatiane C Araújo Nogueira ◽

Renata Gorjão ◽

Carla Rodrigues Bromati ◽

Tatiana S Yamanaka ◽

...

Keyword(s):

Cell Death ◽

Inflammatory Cytokines ◽

Interleukin 6 ◽

Common Mechanism ◽

Rinm5f Cells ◽

Β Cell ◽

Induced Apoptosis ◽

Unconditional Stimulation ◽

Stimulation Of ◽

Pro Inflammatory Cytokines

Unfolded protein response (UPR)-mediated pancreatic β-cell death has been described as a common mechanism by which palmitate (PA) and pro-inflammatory cytokines contribute to the development of diabetes. There are evidences that interleukin 6 (IL6) has a protective action against β-cell death induced by pro-inflammatory cytokines; the effects of IL6 on PA-induced apoptosis have not been investigated yet. In the present study, we have demonstrated that PA selectively disrupts IL6-induced RAC-alpha serine/threonine-protein kinase (AKT) activation without interfering with signal transducer and activator of transcription 3 phosphorylation in RINm5F cells. The inability of IL6 to activate AKT in the presence of PA correlated with an inefficient protection against PA-induced apoptosis. In contrast to PA, IL6 efficiently reduced apoptosis induced by pro-inflammatory cytokines. In addition, we have demonstrated that IL6 is unable to overcome PA-stimulated UPR, as assessed by activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP) expression, X-box binding protein-1 gene mRNA splicing, and pancreatic eukaryotic initiation factor-2α kinase phosphorylation, whereas no significant induction of UPR by pro-inflammatory cytokines was detected. This unconditional stimulation of UPR and apoptosis by PA was accompanied by the stimulation of CHOP and tribble3 (TRIB3) expression, irrespective of the presence of IL6. These findings suggest that IL6 is unable to protect pancreatic β-cells from PA-induced apoptosis because it does not repress UPR activation. In this way, CHOP and ATF4 might mediate PA-induced TRIB3 expression and, by extension, the suppression of IL6 activation of pro-survival kinase AKT.

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