Involvement of the extracellular signal-regulated protein kinase (ERK) pathway in the induction of apoptosis by cadmium chloride in CCRF-CEM cells

When CCRF-CEM cells were incubated with 5–40 μM CdCl_2, apoptosis was observed most clearly at 10 μM. Prior to the development of apoptosis, mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK, were activated with different sensitivity to CdCl₂ exposure. ERK and p38 MAPK were phosphorylated with incubation of 1 μM CdCl_2, but higher than 20 μM CdCl₂ was required for the clear phosphorylation of JNK. In the time–course study, ERK and p38 MAPK were phosphorylated earlier than JNK after CdCl₂ exposure. The in vitro activities of MAPKs also increased in response to CdCl₂ exposure. Pretreatment with an intracellular Ca²⁺ chelator, 1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetra(acetoxymethyl) ester (BAPTA/AM), suppressed almost completely CdCl₂-induced phosphorylation of JNK and p38 MAPK, but not ERK phosphorylation, indicating that the activation of JNK and p38 MAPK depends on the intracellular Ca²⁺ but that of ERK does not. On the other hand, treatment with a MAPK/ERK kinase (MEK) inhibitor, U0126 (1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio]butadiene), suppressed CdCl₂-induced ERK activation and the apoptosis as well. The inhibition of p38 MAPK activity with SB203580 (4-[4-fluorophenyl]-2-[4-methylsulfinylphenyl]-5-[4-pyridyl]1H-imidazole) did not protect cells from apoptosis. The present results showed that the activation of ERK, JNK, and p38 MAPK is differently regulated in CCRF-CEM cells exposed to CdCl_2, and that the ERK pathway seems to be responsible for the induction of apoptosis by CdCl₂ exposure in this human T cell line.     

Indomethacin induces apoptosis in 786-O renal cell carcinoma cells by activating mitogen-activated protein kinases and AKT

Studies on chemoprevention of cancer are generating increasing interest. The anti-neoplastic effect of nonsteroidal anti-inflammatory drugs (NSAIDs) involves cyclooxygenase (COX)-dependent and COX-independent mechanisms. Evidence suggests that mitogen-activated protein kinases (MAPKs) may mediate apoptotic signaling induced by anti-neoplastic agents. While many reports have revealed the existence of MAPK activation in apoptosis induced by various stimuli, the signaling transduction pathways used by NSAIDs to trigger apoptosis in human renal cell carcinoma (RCC) remain largely unknown. Treatment of RCC 786-O cells with indomethacin resulted in growth regression and apoptosis. Caspase-dependent apoptosis was evidenced by the detection of enzymatic activities of caspase-3, caspase-6, and caspase-9 and suppression of toxicity using a caspase inhibitor. Indomethacin treatment was associated with increased expression of glucose-regulated protein 78 (GRP78) and C/EBP homologus protein (CHOP) and activation of ATF-6, characteristics of endoplasmic reticulum stress. In addition, the concomitant induction of peroxisome proliferator-activated receptor (PPAR), especially PPAR-beta, was apparent in treated cells. Western blotting revealed the activation of extracellular signal-regulated kinase (ERK), p38 MAPK, and c-Jun N-terminal kinase (JNK) with indomethacin treatment. Selective inhibitors of ERK, p38 MAPK, and JNK suppressed the induction of GRP78, CHOP, and PPAR-beta, attenuated indomethacin-induced cytotoxicity and reduced increased caspase activity. LY294002, a phosphoinositide-3 kinase (PI3K)/AKT inhibitor, and Trolox, an antioxidant, suppressed indomethacin-induced cytotoxicity and caspase activation. Furthermore, Trolox attenuated indomethacin-induced increased phosphorylation in ERK, p38 MAPK, JNK, and AKT. In conclusion, our findings establish a mechanistic link between the oxidative stress, PI3K/AKT pathway, MAPK pathway and indomethacin-induced cellular alterations and apoptosis in 786-O cells.     

Roles of mitogen-activated protein kinase pathways for mediator release from human cultured mast cells

Human cultured mast cells (HCMC) secrete histamine, sulfidoleukotrienes (LTs), and prostaglandin D(2) (PGD(2)), and produce a variety of cytokines after aggregation of high-affinity receptors for IgE (FcepsilonRI). With respect to the mitogen-activated protein kinase (MAPK) family, extracellular signal-regulated kinases (ERKs), c-Jun NH(2)-terminal kinases (JNKs), and p38 mitogen-activated protein kinase (p38 MAPK) are known. To investigate the roles of these kinase pathways for mediator release from human mast cells, we examined the participation of the activation of these kinases in mediator release, using 1,4-diamino-2, 3-dicyano-1,4-bis(2-aminophenylthio)butadiene (U0126), an ERK pathway inhibitor, and 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imid azo le (SB203580), a p38 MAPK pathway inhibitor. U0126 inhibited ERK activation, LT and PGD(2) release, and granulocyte macrophage-colony stimulating factor (GM-CSF) production after stimulation of HCMC. SB203580, on the other hand, potentiated JNK activation and GM-CSF production. The findings of the present study demonstrated that: (i) the release of arachidonic acid metabolites is mediated by the ERK pathway; (ii) GM-CSF production may be driven by both the ERK and JNK pathways; and (iii) the p38 MAPK pathway negatively regulates the JNK pathway. This suggests that MAPK pathways play important roles in mediator release from human mast cells.     

Src tyrosine kinases mediate crystalline silica-induced NF-kappaB activation through tyrosine phosphorylation of IkappaB-alpha and p65 NF-kappaB in RAW 264.7 macrophages.

Protein tyrosine kinases (PTKs) and mitogen-activated protein kinases (MAPKs) have been demonstrated to play a crucial role in the signaling pathways induced by silica. In the present study, we investigated whether Src family TKs play a role in crystalline silica-induced NF-kappaB activation and whether NF-kappaB activation requires Src TK-dependent MAPK activity in RAW 264.7 cells, a mouse peritoneal macrophage cell line. Selective Src TK inhibitors, damnacanthal or PP1, inhibited silica-induced NF-kappaB activation in a dose-dependent manner. Furthermore, these kinase inhibitors suppressed silica-induced tyrosine phosphorylation of IkappaB-alpha and p65 NF-kappaB. Within a similar time frame, c-Src and Lck were physically associated with IkappaB-alpha and with p65 NF-kappaB. Silica stimulated the phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2), but not p38 MAPK and c-Jun NH(2)-terminal kinase 1 and 2 (JNK1/2). Damnacanthal or PP1 substantially blocked the silica-induced activation of ERK1/2. Moreover, PD98059, an inhibitor of ERK1/2, or SB203580, an inhibitor of p38 MAPK, failed to inhibit silica-induced NF-kappaB activation. These results suggest that c-Src and Lck act for silica-induced NF-kappaB activation by mediating the tyrosine phosphorylations of IkappaB-alpha and p65 NF-kappaB. However, the Src TK-dependent activation of ERK1/2 may not be involved in the silica signaling pathway leading to NF-kappaB activation.     

MAPKs signaling in 5-LOX pathway-associated inflammatory responses

5-Lipoxygenase(5-LOX) is a crucial enzyme catalyzing arachidonic acid to form cysteinyl leukotrienes(Cys LTs, including LTC4, LTD4, LTE4). Cys LTs are potent inflammatory mediators that mediate many pathophysiological responses by activating two distinct G-protein-coupled receptors(CysLT1R and CysLT2R). 5-LOX pathway is implicated in inflammatory pathological processes. Mitogenactivated protein kinases(MAPKs) are important signal transduction pathways of complex cellular processes.Classical MAPK families in mammalian cells include the P38 MAPK, extracellular signal-regulated kinases(ERK)and c-Jun amino N-terminal kinase(JNK). Increasing evidence has suggested that MAPKs signaling acts as key modulator in 5-LOX pathway-related pathological processes. It was reported that P38 MAPK and 5-LOX were dramatically activated to mediate neuronal injury in oxygen-glucose deprivation(OGD)-treated PC12 cells,which could be inhibited by P38 MAPK inhibitor SB203580 and nonselective 5-LOX inhibitor caffeic acid. In primary cultured rat astrocytes, OGD induced increased expression of CysLT2R and aquaporin 4, and ischemic astrocyte injury. Also, OGD increased phosphorylation of ERK and P38 MAPK. These astrocyte responses were attenuated by P38 MAPK inhibitor SB203580, ERK inhibitor U0126, CysLT2R antagonist Bay Cys LT2 and non-selective Cys LTR antagonist Bay u9773, but not by JNK inhibitor SP600125 and CysLT1R antagonist montelukast. These data demonstrate the roles of ERK and P38 MAPK signaling pathways in 5-LOX and CysLT2R-mediated ischemic-like injury in neuron-like PC12 cel s and rat astrocytes. Furthermore, CysLT1R and ERK signaling were found to be involved in regulation of LTD4-stimulated expression of glial fibrillary acidic protein and cell proliferation in astrocytes. ERK inhibitor PD98059 and CysLT1R antagonist montelukast, MK-571 abolished Cys LTs-induced astrocyte proliferation. Vascular endothelial cells play a pivotal role in maintaining brain homeostasis. Endothelial cells undergo a number of pathological changes that can be characterized as uncontrolled cell proliferation, migration and dysfunction in respond to brain insults. In human endothelial cell line EA.hy926, LTD4-induced migration of endothelial cells could be modulated by CysLT1R via phosphorylation of ERK. LTD4 was also reported to enhanced cell proliferation mediated by activating CysLT1R in human umbilical vein endothelial cells(HUVECs), and this is regulated by ERK pathway. ERK inhibitor(U0126, PD98059), CysLT1R antagonist(montelukast, MK571) and dual antagonist(Bay u9773)suppressed LTD4-induced endothelial responses in EA.hy926 cells and HUVECs. Consistent with these results,a study showed that the involvement of ERK/early growth response-1(Egr-1) pathway in CysLT2R-mediated cytokine IL-8 production. ERK inhibitor U0126 and CysLT2R antagonist HAMI3379 inhibited Egr-1 and IL-8 expression as well as IL-8 release in LTC4 and LTD4-stimulated hCys LT2-HEK293 cells. In addition, LTD4 and LTC4 induced monocyte chemoattractant protein-1(MCP-1) by enhanced CysLT1R via phosphorylations of ERK and JNK in human monocytes/macrophages. Pretreatment with the ERK inhibitor PD98059 and JNK inhibitor SP600125 reduced MCP-1 production. CysLT1R antagonist pranlukast inhibited Cys LTs-induced phosphorylation of ERK and JNK as well as production of MCP-1. A better understanding of the roles of MAPKs signaling in 5-LOX pathway-associated pathological processes may open up new avenues for the development of therapeutic strategies targeting inflammatory diseases.     

Naringin Inhibits ROS‐activated MAPK Pathway in High Glucose‐induced Injuries in H9c2 Cardiac Cells

Naringin, an active flavonoid isolated from citrus fruit extracts, exhibits biological and pharmacological properties, such as antioxidant activity and antidiabetic effect. Mitogen‐activated protein kinase (MAPK) signalling pathway has been shown to participate in hyperglycaemia‐induced injury. The present study tested the hypothesis that naringin protects against high glucose (HG)‐induced injuries by inhibiting MAPK pathway in H9c2 cardiac cells. To examine this, the cells were treated with 35 mM glucose (HG) for 24 hr to establish a HG‐induced cardiomyocyte injury model. The cells were pre‐treated with 80 μM naringin for 2 hr before exposure to HG. The findings of this study showed that exposure of H9c2 cells to HG for 24 hr markedly induced injuries, as evidenced by a decrease in cell viability, increases in apoptotic cells and reactive oxygen species (ROS) production, as well as dissipation of mitochondrial membrance potential (MMP). These injuries were significantly attenuated by the pre‐treatment of cells with either naringin or SB203580 (a selective inhibitor of p38 MAPK) or U0126 (a selective inhibitor of extracellular signal regulated kinase 1/2, ERK1/2) or SP600125 (a selective inhibitor of c‐jun N‐termanal kinase, JNK) before exposure to HG, respectively. Furthermore, exposure of cells to HG increased the phosphorylation of p38 MAPK, ERK1/2 and JNK. The increased activation of MAPK pathway was ameliorated by pre‐treatment with either naringin or N‐acetyl‐L‐cysteine (NAC), a ROS scavenger, which also reduced HG‐induced cytotoxicity and apoptosis, leading to increase in cell viability and decrease in apoptotic cells. In conclusion, our findings provide new evidence for the first time that naringin protects against HG‐induced injuries by inhibiting the activation of MAPK (p38 MAPK, ERK1/2 and JNK) and oxidative stress in H9c2 cells.     

Sodium nitroprusside induces apoptosis of H9C2 cardiac muscle cells in a c-Jun N-terminal kinase-dependent manner

Sodium nitroprusside (SNP) induces apoptosis in H9C2 cardiac muscle cells. Treatment with an exogenous NO donor SNP (2 mM) to H9C2 cells resulted in apoptotic morphological changes; a bright blue-fluorescent condensed nuclei and chromatin fragmentation by fluorescence microscope of Hoechst 33258-staining. The activity of caspase-3 like protease was increased during SNP-induced cell death. However, the activity of caspase-1 like protease was not affected by SNP. Pretreatment with Z-VAD-FMK (a pan-caspase inhibitor) or Ac-DEVD-CHO (a specific caspase-3 inhibitor) abrogated the SNP-induced cell death. SNP markedly activated three MAP kinases (JNK/SAPK, ERK and p38 MAP kinase) in the cardiac muscle cells. In this study, selective inhibition of the ERK or p38 MAPK pathway (by PD98059 or SB203580, respectively) had no effect on the extent of SNP-induced apoptosis in cardiac muscle cells. In contrast, inhibition of the JNK pathway by transfection of a dominant negative mutant of JNK markedly reduced the extent of SNP-induced cell death. Taken together, we suggest that JNK/SAPK will be related to SNP-induced apoptosis of H9C2 cardiac muscle cells.     

Mycotoxin patulin activates the p38 kinase and JNK signaling pathways in human embryonic kidney cells.

Patulin (PAT), a mycotoxin mainly produced by Penicillium and Aspergillus, is frequently detected in moldy fruits and fruit products. Exposure of human embryonic kidney (HEK293) cells to PAT led to a dose- and time-dependent increase in the phosphorylation of two major mitogen-activated protein kinases (MAPKs), p38 kinase and c-Jun N-terminal kinase (JNK). The phosphorylated forms of MAPK kinase 4 (MKK4), c-Jun, and ATF-2 were also seen in PAT-treated cultures. The cell death caused by PAT was significantly reduced by the p38 kinase inhibitor, SB203580, but not by the JNK inhibitor, SP600125. Neither p38 kinase nor JNK played a role in the PAT-induced DNA damage. In PAT-treated cells, inactivation of double-stranded RNA-activated protein kinase R (PKR) by the inhibitor, adenine, markedly suppressed JNK and ERK phosphorylation. Treatment of HEK293 cells with PAT-cysteine adduct, a chemical derivative of PAT, showed no effect on MAPK signaling pathways, cell viability, or DNA integrity. These results indicate that PAT causes rapid activation of p38 kinase and JNK in HEK293 cells, but only the p38 kinase signaling pathway contributes to the PAT-induced cell death. PKR also plays a role in PAT-mediated MAPK activation.     

Activation of extracellular signal-regulated kinase during silibinin-protected, isoproterenol-induced apoptosis in rat cardiac myocytes is tyrosine kinase pathway-mediated and protein kinase C-dependent

Aim： To investigate the mechanism of silibinin-protected isoproterenol-induced apoptosis in rat cardiac myocytes. Methods： The viability of rat cardiac myocytes was measured by MTT method. The apoptotic ratio was measured by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling. Protein kinase C （PKC） activity assay was carried out according to the instructions of the PepTag non-radioactive protein kinase C assay kit. Western blot analysis was used to evaluate the level of Ras, Raf-1 and mitogen-activated protein kinase （MAPK） expression. Results： The protective effects of silibinin were significantly sup- pressed by inhibitors, including genistein, manumycin A and GW5074 [inhibitors for protein tyrosine kinases （PTK）, Ras and Raf- 1, respectively]. The exposure of rat cardiac myocytes to isoproterenol alone caused decreased PKC activity, which was prevented by pretreatment with silibinin dose-dependently. Simultaneously, the increased expression of Ras and Raf- 1 activated by silibinin were blocked by the PKC inhibitor, stauroporine. In addition, the extracellularly responsive kinase （ERK） inhibitor, PD98059, suppressed silibinin-protected apoptosis, whereas the p38 MAPK inhibitor, SB203580, protected cardiac myocytes from isoproterenolinduced injury, and the c-Jun N-terminal kinase （JNK） inhibitor, SP600125 had no protective effects. Furthermore, Westem blot analysis showed that the expres- sion of phosphorylated ERK was increased by silibinin, the expression of phos- phorylated p38 MAPK was decreased and total ERK, p38, JNK and phosphorylated JNK MAPK did not change after treatment with both isoproterenol and silibinin. Furthermore, pretreatment of cardiac myocyte with PKC, Ras and Raf inhibitors significantly blocked ERK phosphorylation. Conclusion： Silibinin is suggested to protect isoproterenol-induced rat cardiac myocyte apoptosis by activating the tyrosine kinase pathway, PKC and MAPK pathways.     

p38 and JNK MAPK, but not ERK1/2 MAPK, play important role in colchicine-induced cortical neurons apoptosis

Colchicine is a microtubule interfering agent and is able to induce neural apoptosis. However, the intracellular pathway involved in its neurotoxicity is still unclear. In the present study, three of mitogen-activated protein kinases (MAPKs): p38, c-Jun N-terminal kinase (JNK) and extracellular-regulated kinase 1/2 (ERK1/2) were investigated in colchicine-induced apoptosis on cortical neurons for the first time. Our results showed that 1 microM colchicine administration in primarily cultured cortical neurons led to typical neuronal apoptosis, and the apoptosis was attenuated by taxol, a microtubule stabilizer. Moreover, activation of p38 MAPK was found for the first time, as well as that of JNK MAPK, but not of ERK1/2 MAPK, after colchicine exposure. Apoptosis was inhibited by p38 MAPK inhibitors, SB203580 (4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole), SB239063 (trans-1-(4-hydroxycyclohexyl)-4-(fluorophenyl)-5-(2-methoxypyrimidin-4-yl) imidazole), and JNK MAPK pathway inhibitors, CEP11004 (9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid, 2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-5,16-bis[[(1-methylethyl)thio]methyl]-1-oxo-, methyl ester, (9S,10R,12R)-), SP600125 (anthra[1,9-cd]pyrazol-6(2H)-one). However, PD98059 (2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one) and U0126 (1,4-diamino-2,3-dicyano-1, 4-bis[2-aminophenylthio]butadiene), ERK1/2 MAPK inhibitors, did not work. Furthermore, better neuronal protective effects were achieved by using JNK and the p38 MAPK inhibitors together as compared to that by using either alone. The results suggested that p38 MAPK, JNK MAPK, but not ERK1/2 MAPK may play pivotal role in colchicine's neurotoxicity in primarily cultured cortical neurons, and the protective effects of the inhibition of p38 or JNK MPAK on cortical neurons were synergistically.     

Signal transduction mediated by mitogen-activated protein kinases and apoptisis of tumor cells

Objective To make a review about MAPK and apoptosis of tumor cells.Methods We collected a large number of related experimental papers,and summarized key points.Results mitogen activated protein kinase(MAPK)is one of the four biggest signal transduction systems which contain four subtribes named p38,ERK5,ERK and JNK/SAPK respectively.MAPK pathways constitute numerous modular network that regulates a variety of physiological processes,such as cell growth,roliferation,differentiation,and apoptotic cell death.Specially,the function of induced apoptosis in tumor cells has gradually become main focus.Both in-vitro and ex-vivo findings demonstrated that in apoptotic tumor cells the level of phosphorylation of JNK kinase is significantly improved.That means JNK kinase is activated in these tumor cells.At the same time,contrary to JNK kinase,the activity of ERK kinase is usually weakened.So,for most apoptotic tumor cells,JNK is a positive factor,however,ERK kinases is a negative factor.As for the p38 kinase,which can be activated to outside stimulus,also has the promotion of apoptosis.Importantly,ERK kinase activity is suppressed by JNK/p38 kinase during apoptosis induction.Further study demonstrate that the regulatory mechanisms of MAPK in apoptotic tumor cells are:working on upstream of caspase;starting death receptor channel;activating pro-apoptotic Bcl-2 protein family;changing mitochondrial permeability;participating in Fas/Fasl-mediated apoptosis;enhancing the expression of TNF-a and so on.Conclusions The relationship between MAPK and apoptosis of tumor cells is intimate.It is expected to be a new target for tumor in clinical treatment.     

ML-7 amplifies the quinocetone-induced cell death through akt and MAPK-mediated apoptosis on HepG2 cell line

The study aims at evaluating the combination of the quinocetone and the ML-7 in preclinical hepatocellular carcinoma models. To this end, the effect of quinocetone and ML-7 on apoptosis induction and signaling pathways was analyzed on HepG2 cell lines. Here, we report that ML-7, in a nontoxic concentration, sensitized the HepG2 cells to quinocetone-induced cytotoxicity. Also, ML-7 profoundly enhances quinocetone-induced apoptosis in HepG2 cell line. Mechanistic investigations revealed that ML-7 and quinocetone act in concert to trigger the cleavage of caspase-8 as well as Bax/Bcl-2 ratio up-regulation and subsequent cleavage of Bid, capsases-9 and -3. Importantly, ML-7 weakened the quinocetone-induced Akt pathway activation, but strengthened the phosphorylation of p-38, ERK and JNK. Further treatment of Akt activator and p-38 inhibitor almost completely abolished the ML-7/quinocetone-induced apoptosis. In contrast, the ERK and JNK inhibitor aggravated the ML-7/quinocetone-induced apoptosis, indicating that the synergism critically depended on p-38 phosphorylation and HepG2 cells provoke Akt, ERK and JNK signaling pathways to against apoptosis. In conclusion, the rational combination of quinocetone and ML-7 presents a promising approach to trigger apoptosis in hepatocellular carcinoma, which warrants further investigation.     

Apoptosis contributes to the cytotoxicity induced by amodiaquine and its major metabolite N-desethylamodiaquine in hepatic cells

Amodiaquine (ADQ), an antimalarial drug used in endemic areas, has been reported to be associated with liver toxicity; however, the mechanism underlying its hepatoxicity remains unclear. In this study, we examined the cytotoxicity of ADQ and its major metabolite N-desethylamodiaquine (NADQ) and the effect of cytochrome P450 (CYP)-mediated metabolism on ADQ-induced cytotoxicity. After a 48-h treatment, ADQ and NADQ caused cytotoxicity and induced apoptosis in HepG2 cells; NADQ was slightly more toxic than ADQ. ADQ treatment decreased the levels of anti-apoptotic Bcl-2 family proteins, which was accompanied by an increase in the levels of pro-apoptotic Bcl-2 family proteins, indicating that ADQ-induced apoptosis was mediated by the Bcl-2 family. NADQ treatment markedly increased the phosphorylation of JNK, extracellular signal-regulated kinase (ERK1/2), and p38, indicating that NADQ-induced apoptosis was mediated by MAPK signaling pathways. Metabolic studies using microsomes obtained from HepG2 cell lines overexpressing human CYPs demonstrated that CYP1A1, 2C8, and 3A4 were the major enzymes that metabolized ADQ to NADQ and that CYP1A2, 1B1, 2C19, and 3A5 also metabolized ADQ, but to a lesser extent. The cytotoxicity of ADQ was increased in CYP2C8 and 3A4 overexpressing HepG2 cells compared to HepG2/CYP vector cells, confirming that NADQ was more toxic than ADQ. Moreover, treatment of CYP2C8 and 3A4 overexpressing HepG2 cells with ADQ increased the phosphorylation of JNK, ERK1/2, and p38, but not the expression of Bcl-2 family proteins. Our findings indicate that ADQ and its major metabolite NADQ induce apoptosis, which is mediated by members of the Bcl-2 family and the activation of MAPK signaling pathways, respectively.     

Involvement of ROS in Curcumin-induced Autophagic Cell Death

Many anticancer agents as well as ionizing radiation have been shown to induce autophagy which is originally described as a protein recycling process and recently reported to play a crucial role in various disorders. In HCT116 human colon cancer cells, we found that curcumin, a polyphenolic phytochemical extracted from the plant Curcuma longa, markedly induced the conversion of microtubule-associated protein 1 light chain 3 (LC3)-I to LC3-II and degradation of sequestome-1 (SQSTM1) which is a marker of autophagosome degradation. Moreover, we found that curcumin caused GFP-LC3 formation puncta, a marker of autophagosome, and decrease of GFP-LC3 and SQSTM1 protein level in GFP-LC3 expressing HCT116 cells. It was further confirmed that treatment of cells with hydrogen peroxide induced increase of LC3 conversion and decrease of GFP-LC3 and SQSTM1 levels, but these changes by curcumin were almost completely blocked in the presence of antioxidant, N-acetylcystein (NAC), indicating that curcumin leads to reactive oxygen species (ROS) production, which results in autophagosome development and autolysosomal degradation. In parallel with NAC, SQSTM1 degradation was also diminished by bafilomycin A, a potent inhibitor of autophagosome-lysosome fusion, and cell viability assay was further confirmed that cucurmin-induced cell death was partially blocked by bafilomycin A as well as NAC. We also observed that NAC abolished curcumin-induced activation of extracelluar signal-regulated kinases (ERK) 1/2 and p38 mitogen-activated protein kinases (MAPK), but not Jun N-terminal kinase (JNK). However, the activation of ERK1/2 and p38 MAPK seemed to have no effect on the curcumin-induced autophagy, since both the conversion of LC3 protein and SQSTM1 degradation by curcumin was not changed in the presence of NAC. Taken together, our data suggest that curcumin induced ROS production, which resulted in autophagic activation and concomitant cell death in HCT116 human colon cancer cell. However, ROS-dependent activation of ERK1/2 and p38 MAPK, but not JNK, might not be involved in the curcumin-induced autophagy.     

p38 MAP kinase and ERK play an important role in nitric oxide-induced apoptosis of the mouse embryonic stem cells

Previous study showed that nitric oxide (NO) induces apoptosis in mouse embryonic stem (mES) cells, but the precise mechanism governing NO-induced apoptosis in mES remains unclear. This study investigated the mechanism of NO-induced apoptosis of mES cells via MAP kinase signaling pathway. Sodium nitroprusside (SNP), a NO donor, induced apoptosis in mES cells with enhanced production of reactive oxygen species (ROS). In addition, treatment with SNP induced the activation of caspase-3, -8 and -9 as well as mitogen-activated protein (MAP) kinases (JNK, p38 MAP kinase and ERK). However, pretreatment with the p38 MAP kinase inhibitor SB203580 and ERK inhibitor U0126 attenuated NO-induced cell toxicity, ROS production, and caspase-3 activation. Moreover, SB203580 inhibited the translocation of Bax from the cytosol to the mitochondria. Taken together, these results suggest that NO-induced apoptosis in mES cells was mediated through p38 MAP kinase/ERK signaling pathway by triggering caspases activation and Bax translocation from the cytosol to the mitochondria.     

Impact of DUSP1 on the apoptotic potential of deoxynivalenol in the epithelial cell line HepG2

The trichothecene deoxynivalenol (DON) is the most common mycotoxin contaminant of cereal-based food products. Several studies revealed DON as a potent inducer of the three major mitogen-activated protein kinases (MAPKs). Until now, little is known about the role of negative regulators of MAPK pathway in the cellular response to DON. In this report we evaluated, for the first time, the impact of mitogen-activated protein kinase phosphatases (MKPs), particularly dual specific phosphatase 1 (DUSP1), on the toxic potential of DON in the epithelial cell line HepG2. Our results indicate that both low and high concentrations of DON trigger a strong and sustained DUSP1 mRNA and protein expression, mediated by the sustained activation of MEK/ERK pathway. Furthermore, the expression of DUSP1 protein correlates with the inactivation of JNK1/2, whereas a sustained activation of p38 and ERK1/2 was observed in the presence of DON. In contrast, treatment of DUSP1 knock-down cells with DON triggers a prolonged activation of JNK1/2, which leads to the induction of apoptosis. Taken together, we propose DUSP1 as a novel target gene of DON, which is essential for the prevention of DON induced apoptosis in the epithelial cell line HepG2.