Activation of c-Jun N-terminal kinase (JNK) is associated with a wide range of disparate cellular responses to extracellular stimuli. In mammals, three JNK isoforms are known, and their differential regulation occurs in a stimulus- or a cell type-dependent manner. However, the underlying mechanism of this differential regulation has not been clearly elucidated. Here we demonstrated that JNK1 and JNK3 were activated in SH-SY5Y cells after treatment with H2O2 or UV. In SH-SY5Y cells overexpressing mJIP1, a splicing variant of a JNK scaffold protein JIP1, the H2O2-induced activities of both JNK1 and JNK3 were significantly suppressed. In the same cell line, however, UV-induced JNK1 activity was significantly suppressed, but JNK3 activity was not. During the RA-induced differentiation of SH-SY5Y cells, JNK1 was activated, whereas JNK3 was not, and this JNK1 activation was completely abolished in the cells overexpressing mJIP1. These results suggest that JIP1 plays a role in the regulation of the isoform-specific activation of JNKs in stimulus-dependent manner.
Cell Line ; Humans* ; JNK Mitogen-Activated Protein Kinases ; Mammals ; Neuroblastoma* ; Protein Isoforms

Middle cerebral artery occlusion (MCAO) results in cell death by activation of complex signal pathways for cell death and survival. Hypothermia is a robust neuroprotectant, and its effect has often been attributed to various mechanisms, but it is not yet clear. Upstream from the cell death promoters and executioners are several enzymes that may activate several transcription factors involved in cell death and survival. In this study, we immunohistochemically examined the phosphorylation of mitogen- activated protein kinase, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 kinase during early period of the ischemic injury, following 2 hours (h) of transient MCAO. Increased phosphorylation of ERK and p38 was observed in the vessels at 3 h, neuron-like cells at 6 and 12 h and glia-like cells at 12 h. Activation of JNK was not remarkable, and a few cells showed active JNK following ischemia. Phosphorylation of Elk-1, a transcription factor, was reduced by ischemic insult. Hypothermia attenuated the activation of ERK, p38 and JNK, and inhibited reduction of Elk-1. These data suggest that signals via different MAPK family members converge on the cell damage process and hypothermia protects the brain by interfering with these pathways.
Brain ; Cell Death ; Humans ; Hypothermia* ; Infarction, Middle Cerebral Artery ; Ischemia ; JNK Mitogen-Activated Protein Kinases ; Mitogen-Activated Protein Kinases* ; Phosphorylation ; Phosphotransferases ; Protein Kinases ; Signal Transduction ; Stroke* ; Transcription Factors

JNK signaling pathway plays an important role in regulation of cell growth, proliferation, differentiation, migration and apoptosis. Studies have shown that JNK signaling pathway is closely related to autophagy, but the underlying mechanisms are not clear. Clarification of the correlation between the JNK signaling pathway and autophagy may provide opportunities for identifying potential molecular targets for the treatment of diseases associated with autophagy.
Autophagy ; Humans ; JNK Mitogen-Activated Protein Kinases ; MAP Kinase Signaling System

BACKGROUNDMesangial hypercellularity is a critical early histopathological finding in human and experimental glomerular diseases. Hyperlipidemia and the glomerular deposition of lipoproteins are commonly associated with mesangial hypercellularity and play an important pathobiological role in the development of glomerular diseases. The activated cytoplasmic mitogen-activated protein kinase (MAPK), including mainly extracellular-signal regulated protein kinase (ERK), c-Jun amino-terminal kinase (JNK), and p38, has been thought to translocate into the nucleus and activate various transcription factors and protooncogenes associated with cell growth and proliferation. Lipoprotein (a) (Lp(a)) has been shown to stimulate proliferation of mesangial cells, but the events of Lp(a) signaling have not yet been characterized. The purpose of this study was to investigate the signal transduction pathways involved in Lp(a)-induced cell proliferation and provide an evidence for the participation of Lp(a) in intracellular signaling pathways for mesangial cell proliferation.

METHODSLp(a) was isolated from a patient who was being treated with low density lipoprotein (LDL)-apheresis by density gradient ultracentrifugation and then chromatography. Human mesangial cells (HMCs) were isolated by the sequential sieving technique and stimulated with Lp(a) in different concentration and time course. The DNA synthesis of the cells was measured by [3H] thymidine incorporation for detecting the proliferation. The expression of all the three members of MAPK family, including ERK1/ERK2, JNK, and p38, and their phosphorylation were detected by Western blotting.

RESULTSLp(a) could induce a significant dose-dependent proliferation of HMCs. The 3H-TdR incorporation was 1.64+/-0.31, 1.69+/-0.48, 3.59+/-0.68 (P<0.01), 4.14+/-0.78 (P<0.01), and 4.05+/-0.55 (P<0.01) (10(3) cpm) at the Lp(a) concentration of 0, 5, 10, 25, and 50 microg/ml, respectively. Lp(a) induced an increase in ERK1/ERK2 phosphorylation between 5 and 60 minutes, and in JNK phosphorylation between 15 and 30 minutes after incubating with HMCs, whereas the level of p38 and its phosphorylation was not changed.

CONCLUSIONSLp(a) could stimulate the proliferation of HMCs by activiating the phosphorylation of ERK1/ERK2 and JNK MAPK signaling pathway, whereas p38 pathway had no effect on the Lp(a)-induced HMC proliferation, which indicated that three MAPKs seem to be distinctly involved in the effect. In particular, it also provides the evidence that Lp(a) may act as one of the major endogenous modulators for mitogenic signaling response and cell proliferation within the glomerulus.

Blotting, Western ; Cells, Cultured ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Lipoprotein(a) ; pharmacology ; Mesangial Cells ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Phosphorylation ; drug effects ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo detect protein expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2) in human hepatocellular carcinoma and adjacent non-neoplastic liver.

METHODSIn 16 surgically resected hepatocellular carcinoma and para-carcinoma tissues, Western blotting was used to detect expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2).

RESULTSIn all cases, ERK(1), ERK(2), p38 expression in hepatocellular carcinoma was significantly higher than that in para-carcinoma: integral optic density (IOD) of ERK(1) was 300 +/- 98 in carcinoma and 98 +/- 48 in para-carcinoma tissues (t = 2.519, P < 0.01); IOD of ERK(2) was 587 +/- 83 in carcinoma and 232 +/- 96 in para-carcinoma tissues (t = 2.745, P < 0.01); IOD of p38 was 270 +/- 85 in carcinoma and 107 +/- 88 in para-carcinoma tissues (t = 2.491, P < 0.01). JNK(1) expression in hepatocellular carcinoma was significantly lower than that in para-carcinoma; IOD of JNK(1) was 111 +/- 93 in carcinoma and 292 +/- 109 in para-carcinoma tissues (t = 2.473, P < 0.01). Protein levels of MEK(1) and MEK(2) in carcinoma were significantly higher than in para-carcinoma. IOD of MEK(1) was 1 418 +/- 244 in carcinoma and 806 +/- 90 in para-carcinoma tissues (t = 2.546, P < 0.01). IOD of MEK(2) was 1 041 +/- 122 in carcinoma and 468 +/- 40 in para-carcinoma tissues (t = 2.861, P < 0.01).

CONCLUSIONSERK(1), ERK(2), MEK(1) and MEK(2) in the signal transduction pathway for cell proliferation are significantly overexpressed and the expression of JNK(1) is lower in hepatocellular carcinoma. Their unbalance is one of the important reasons for the over growth and infinite proliferation of the hepatocellular carcinoma cell. The p38 and JNK(1) may be activated by different pathway.

Adult ; Aged ; Carcinoma, Hepatocellular ; enzymology ; Enzyme Activation ; Female ; Humans ; JNK Mitogen-Activated Protein Kinases ; Liver Neoplasms ; enzymology ; MAP Kinase Kinase 1 ; Male ; Middle Aged ; Mitogen-Activated Protein Kinase Kinases ; analysis ; Mitogen-Activated Protein Kinases ; metabolism ; Protein-Serine-Threonine Kinases ; analysis

BACKGROUND: Previously, it has been suggested that lipopolysaccaride (LPS) stimulates the activation of the transcriptional factor activator protein (AP-1) which is in part regulated by activation of the c-Jun N-terminal kinase (JNK) / stress-activated protein kinase (SAPK) in the murine macrophage cell line RAW 264.7. METHODS: Consistent with this notion, we find that treatment of LPS on RAW 264.7 cells induces the generation of nitric oxide (NO) and results in the activation of JNK and treated with NO donors and NO inhibitors. RESULTS: NO donors including sodium nitroprusside (1 mM), GSNO (0.2 mM), or SNAP (0.5 mM) treatment of the macrophage cell line markedly induces the activation of JNK. However NGMMA (2 mM), a competitive inhibitor of NO, does not inhibit the activation of JNK induced by LPS. SIN-1, NO, and superoxide donor induce an activation of JNK that is slightly decreased by treatment with sodium dismutase whereas the activation of JNK is significantly augmented by adding sodium dismutase with catalase. C2 ceramide suppresses the generation of NO induced by LPS, but significantly increases the activity of JNK in vivo. LPS can induce the activation of JNK at 30 min after stimulation in RAW 264.7 cells. Exposure to SNP does not affect the enzymatic activity of JNK, immunoprecipitates, JNK, and c-Jun N-terminal proteins. CONCLUSIONS: These data suggest that even though NO is one of the major activators of JNK induced by LPS, there is, at least, an NO-independent JNK activation, signaling a pathway for LPS. Also, there may be an undefined NO-sensitive JNK-regulator (s) in vivo.
Catalase ; Cell Line ; Humans ; JNK Mitogen-Activated Protein Kinases ; Macrophages ; Nitric Oxide* ; Nitroprusside ; Protein Kinases* ; Sodium ; Superoxides ; Tissue Donors

Activation of c-Jun N-terminal kinase (JNK), a member of the mitogen-activated protein kinase family, is an important cellular response that modulates the outcome of the cells which are exposed to the tumor necrosis factor (TNF) or the genotoxic stress including DNA damaging agents. Although it is known that JNK is activated in response to genotoxic stress, neither the pathways to transduce signals to activate JNK nor the primary sensors of the cells that trigger the stress response have been identified. Here, we report that the receptor interacting protein (RIP), a key adaptor protein of TNF signaling, was required to activate JNK in the cells treated with certain DNA damaging agents such as adriamycin (Adr) and 1-beta-D-arabinofuranosylcytosine (Ara-C) that cause slow and sustained activation, but it was not required when treated with N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and short wavelength UV, which causes quick and transient activation. Our findings revealed that this sustained JNK activation was not mediated by the TNF (tumor necrosis factor) receptor signaling, but it required a functional ATM (ataxia telangiectasia) activity. In addition, JNK inhibitor SP-600125 significantly blocked the Adr-induced cell death, but it did not affect the cell death induced by MNNG. These findings suggest that the sustained activation of JNK mediated by RIP plays an important role in the DNA damage-induced cell death, and that the duration of JNK activation relays a different stress response to determine the cell fate.
Cell Death ; DNA ; DNA Damage ; Doxorubicin ; Humans ; JNK Mitogen-Activated Protein Kinases ; Methylnitronitrosoguanidine ; Necrosis ; Protein Kinases ; Tumor Necrosis Factor-alpha

OBJECTIVETo investigate the mechanism by which carbon monoxide inhibits the expression of adhesion molecules on human gingival fibroblasts (HGF) stimulated with inflammatory cytokines.

METHODSHGF were cultured in vitro, and stimulated with 50 ng x mL tumor necrosis factor-alpha (TNF-alpha) and 10 ng x mL(-1) interleukin-1beta (IL-1beta) concurrently in the presence or absence of carbon monoxide releasing molecule-3 (CORM-3) at 500 micromol x L-1. Expression of phosphorylated extracellular regulated protein kinase (ERK), phosphorylated c-Jun N-terminal kinase (NK) and phosphorylated p38 in mitogen-activated protein kinase(MAPK) pathway was studied by Western blot at 10 min and 20 min, respectively. Nuclear expression of nuclear factor-kappaB (NF-kappaB) was checked by Western blot after 4 h stimulation. In some experiments, cells were prestimulated by 1H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ) for 8 h before cytokine stimulation and the expression of intercellular adhesion molecule-1 (ICAM-1) was checked by Western blot after 24 h.

RESULTSCORM-3 significantly inhibited the phosphorylation of MAPK p38 after 10 min stimulation with cytokines, but had no signifi-cant effect on the phosphorylation of ERK and JNK. CORM-3 significantly inhibited the nuclear expression of NF-KB-p65 on HGF after 4 h stimulation by inflammatory cytokines. The inhibitory effect of CORM-3 on the expression of ICAM-1 was not influenced by guanylate cyclase inhibitor ODQ.

CONCLUSIONThe inhibitory effect of carbon monoxide on the expression of adhesion molecules might be exerted by its inhibitory effect on the NF-kappaB activity and MAPK p38 phosphorylation.

Carbon Monoxide ; Cytokines ; Fibroblasts ; Gingiva ; Humans ; Intercellular Adhesion Molecule-1 ; JNK Mitogen-Activated Protein Kinases ; Mitogen-Activated Protein Kinases ; NF-kappa B ; Phosphorylation ; Tumor Necrosis Factor-alpha

Resveratrol (RSV) may provide numerous protective eff ects against chronic inflammatory diseases. Due to local hypoxia and hypertonicity, the renal medulla is subject to extreme oxidative stress, and aldehyde products formed during lipid peroxidation, such as 4-hydroxy-2-hexenal (HHE), might be responsible for tubular injury. This study aimed at investigating the eff ects of RSV on renal and its signaling mechanisms. While HHE treatment resulted in decreased expression of Sirt1, AQP2, and nuclear factor erythroid 2-related factor 2 (Nrf2), mouse cortical collecting duct cells (M1) cells treated with HHE exhibited increased activation of p38 MAPK, extracellular signal regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and increased expression of NOX4, p47(phox), Kelch ECH associating protein 1 (Keap1) and COX2. HHE treatment also induced NF-κB activation by promoting IκB-α degradation. Meanwhile, the observed increases in nuclear NF-κB, NOX4, p47(phox), and COX2 expression were attenuated by treatment with Bay 117082, N-acetyl-l-cysteine (NAC), or RSV. Our findings indicate that RSV inhibits the expression of inflammatory proteins and the production of reactive oxygen species in M1 cells by inhibiting NF-κB activation.
Acetylcysteine ; Animals ; Anoxia ; Bays ; JNK Mitogen-Activated Protein Kinases ; Lipid Peroxidation ; Mice* ; Oxidative Stress* ; p38 Mitogen-Activated Protein Kinases ; Phosphotransferases ; Reactive Oxygen Species ; Sirtuin 1

: To assess the () recombinant gingivalis gingipain R2 (rRgpB)-induced Ca mobilization in human gingival fibroblast (HGF) mediated by protease-activated receptor (PAR) and its downstream signal transduction pathways. : Flow cytometry was used to detect the expression of PAR in HGF. The proliferation of HGF was measured by CCK-8. The dynamic changes of intracellular Ca concentration in HGF induced by rRgpB and the blocking effect of PAR-1 antagonist were observed by laser confocal microscopy. Western blot was performed to determine the phosphorylation levels of c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) 1/2, p38 mitogen-activated protein kinase (p38 MAPK) and p65 in HGF. : PAR-1 and PAR-3 were expressed in HGF, and the rRgpB could promote the proliferation of HGF. rRgpB caused a transient increase in [Ca], which could be completely suppressed by vorapaxar, a PAR-1 antagonist. The phosphorylation levels of JNK, ERK1/2 and p65 were significantly up-regulated after the induction of rRgpB for and (all <0.05), which was completely inhibited by vorapaxar. However, the phosphorylation level of p38 MAPK had no significant change after rRgpB stimulation. : rRgpB causes an increase in [Ca] in HGF mediated by PAR-1. JNK, ERK1/2 and nuclear factor-κB may be involved in intracellular signal transduction after PAR-1 activation.
Fibroblasts ; Humans ; JNK Mitogen-Activated Protein Kinases/metabolism* ; MAP Kinase Signaling System ; Phosphorylation ; Signal Transduction ; p38 Mitogen-Activated Protein Kinases/metabolism*