AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry wa...AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry was performed on tissue array slides containing 423 human GC specimens. Using HER2-positve GC cell lines SNU-216 and NCI-N87, HER2 expression was silenced by RNA interference, and the activations of JNK and AKT were suppressed by SP600125 and LY294002, respectively. Transwell assay, Western blot, semi-quantitative reverse transcriptionpolymerase chain reaction and immunofluorescence staining were used in cell culture experiments. RESULTS In GC specimens, HER2, JNK, and AKT activations were positively correlated with each other. In vitro analysis revealed a positive regulatory feedback loop between HER2 and JNK in GC cell lines and the role of JNK as a downstream effector of AKT in the HER2/AKT signaling pathway. JNK inhibition suppressed migratory capacity through reversing EMT and dual inhibition of JNK and AKT induced a more profound effect on cancer cell motility.CONCLUSION HER2, JNK and AKT in human GC specimens are positively associated with each other. JNK and AKT, downstream effectors of HER2, co-operatively contribute to the metastatic potential of HER2-positive GC cells. Thus, targeting of these two molecules in combination with HER2 downregulation may be a good approach to combat HER2-positive GC.展开更多
Background C-Jun N-terminal kinase （JNK） signaling pathway plays a critical role in cerebral ischemia. Although the mechanistic basis for this activation of JNK1/2 is uncertain, oxidative stress may play a role. The...Background C-Jun N-terminal kinase （JNK） signaling pathway plays a critical role in cerebral ischemia. Although the mechanistic basis for this activation of JNK1/2 is uncertain, oxidative stress may play a role. The purpose of this study was to investigate whether the activation of JNK1/2 is associated with the production of endogenous nitric oxide （NO）. Methods Ischemia and reperfusion （I/R） was induced by cerebral four-vessel occlusion. Sprague-Dawley （SD） rats were divided into 6 groups： sham group, I/R group, neuronal nitric oxide synthase （nNOS） inhibitor （7-nitroindazole, 7-NI） given group, inducible nitric oxide synthase （iNOS） inhibitor （2-amino-5,6-dihydro-methylthiazine, AMT） given group, sodium chloride control group, and 1% dimethyl sulfoxide （DMSO） control group. The levels of protein expression and phospho-JNK1/2 were detected by Western blotting and the survival hippocampus neurons in CA1 zone were observed by cresyl violet staining. Results The study illustrated two peaks of JNK1/2 activation occurred at 30 minutes and 3 days during reperfusion. 7-NI inhibited JNK1/2 activation during the early reperfusion, whereas AMT preferably attenuated JNK1/2 activation during the later reperfusion. Administration of 7-NI and AMT can decrease I/R-induced neuronal loss in hippocampal CA1 region. Conclusion JNK1/2 activation is associated with endogenous NO in response to ischemic insult.展开更多
OBJECTIVE N2L is a novel lipoic acid-niacin dimer regulating lipid metabolism with multifunction,including antioxidant effect.We investigated the protective effect of N2L and the underlying mechanisms under the ferrop...OBJECTIVE N2L is a novel lipoic acid-niacin dimer regulating lipid metabolism with multifunction,including antioxidant effect.We investigated the protective effect of N2L and the underlying mechanisms under the ferroptosis inducer RAS-selective lethality 3(RSL3)treat⁃ment in HT22 cells.METHODS HT22 cells were pretreated with N2L and then were treated with RSL3 to establish a ferroptosis cell model.MTT assay was used to detect the cell survival rate.Free radical probe(dihydroethidium,DHE)and ferrous probe FerroOrange were used to detect the contents of free radicals and ferrous ions in cells.The ultrastructure of mitochondria of treat⁃ed cells was observed by transmission electron microscope.The expression of ferroptosis-relat⁃ed proteins acyl-CoA synthetase long-chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),cyclooxygenase-2(COX-2),ferritin Heavy Chain 1(FTH1),nuclear factor E2-related factor 2/heme oxygenase-1,and phosphoryla⁃tion levels of the c-Jun N-terminal kinase(JNK)/extracellular regulated protein kinases(ERK)pathway were detected by Western blotting.RE⁃SULTS RSL3 decreased the cell viability and induced excessive accumulation of(reactive oxy⁃gen species)ROS in HT22 cells.N2L pretreat⁃ment effectively protected HT22 cells against lipid peroxidation.What′s more,N2L recovered GPX4 protein expression and blocked the increase of COX-2 and ACSL4 expressions.Moreover,N2L also significantly prevented FTH1 from downregulation and maintained iron homeo⁃stasis.Finally,N2L pretreatment could decrease JNK/ERK activation induced by RSL3.CON⁃CLUSION N2L is an excellent ferroptosis inhibi⁃tor,and its anti-ferroptosis mechanism may be related to the reduction of lipid peroxidation and the regulation of iron homeostasis.展开更多
Mixed lineage kinase 3 （MLK3） is a mitogen-activated protein kinase kinase kinase that is activated by tumor necrosis factor-α （TNF-α） and specifically activates c-Jun N-terminal kinase （JNK） on TNF-a stimulat...Mixed lineage kinase 3 （MLK3） is a mitogen-activated protein kinase kinase kinase that is activated by tumor necrosis factor-α （TNF-α） and specifically activates c-Jun N-terminal kinase （JNK） on TNF-a stimulation. The mecha- nism by which TNF-α activates MLK3 is still not known. TNF receptor-associated factors （TRAFs） are adapter molecules that are recruited to cytoplasmic end of TNF receptor and mediate the downstream signaling, including activation of JNK. Here, we report that MLK3 associates with TRAF2, TRAF5 and TRAF6; however only TRAF2 can significantly induce the kinase activity of MLK3. The interaction domain of TRAF2 maps to the TRAF domain and for MLK3 to its C-terminal half （amino acids 511-847）. Endogenous TRAF2 and MLK3 associate with each other in response to TNF-α treatment in a time-dependent manner. The association between MLK3 and TRAF2 mediates MLK3 activation and competition with the TRAF2 deletion mutant that binds to MLK3 attenuates MLK3 kinase activity in a dose-dependent manner, on TNF-α treatment. Furthermore the downstream target of MLK3, JNK was activated by TNF-α in a TRAF2-dependent manner. Hence, our data show that the direct interaction between TRAF2 and MLK3 is required for TNF-α-induced activation of MLK3 and its downstream target, JNK.展开更多
Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods...Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods:Human nasopharyngeal carcinoma CNE multicellular spheroids(MCS) were constructed with three dimensional cell culture methods.Western blot was employed to analyze the activity of JNK signaling pathway in MCS after X-ray irradiation,and the expression of caspase-3 protein before and after using SP600125(a special inhibitor of JNK).X-ray induced cell apoptosis in MCS before and after treated with SP600125 were detected by TUNEL.Results:The level of JNK phosphorylation in MCS was a dynamic course after radiation,and there was a phosphorylation peaks at 2 h later,the apoptotic rate of MCS(P < 0.05) and the expression of caspase-3 protein(P < 0.05) were significantly increased after treated with SP600125.Conclusion:The transient activation of JNK played a important role in sensitivity to radiotherapy of CNE MCS via mediating survival signals,blocking this pathway accelerate cell apoptosis,which may be related to the increased expression of caspase-3.展开更多
Background： Najanalgesin, a toxin isolated from the venom ofNaja nqja atra, has been shown to exert significant analgesic effects in a neuropathic pain model in rats. However, the molecular mechanism underlying this ...Background： Najanalgesin, a toxin isolated from the venom ofNaja nqja atra, has been shown to exert significant analgesic effects in a neuropathic pain model in rats. However, the molecular mechanism underlying this protective effect ofnajanalgesin is poorly understood. The present study sought to evaluate the intracellular signaling pathways that are involved in the antinociceptive effect of najanalgesin on neuropathic pain. Methods： The antinociceptive properties of najanalgesin were tested in hind paw withdrawal thresholds in response to mechanical stimulation. We analyzed the participation of the mitogen-activated protein kinase p38, extracellular-regulated kinase （ERK）, and c-Jun N-terminal kinase （JNK） by western blot analysis. This inhibition of JNK was confirmed by immunohistochemistry. Results： The phosphorylation levels of INK （as well as its downstream molecule c-Jun）, p38, and ERK were significantly increased after injury. Najanalgesin only inhibited JNK and c-Jun phosphorylation but had no effect on either ERK or p38. This inhibition of JNK was confirmed by immunohistochemistry, which suggested that the antinociceptive effect of najanalgesin on spinal nerve ligation-induced neuropathic pain in rats is associated with JNK activation in the spinal cord. Conclusion： The antinociceptive effect of najanalgesin thnctions by inhibiting the JNK in a neuropathic pain model.展开更多
Extracellular aggregation of amyloid-beta(Aβ)and intracellular tau tangles are two major pathogenic hallmarks and critical factors of Alzheimer’s disease.A linear interaction between Aβand tau protein has been char...Extracellular aggregation of amyloid-beta(Aβ)and intracellular tau tangles are two major pathogenic hallmarks and critical factors of Alzheimer’s disease.A linear interaction between Aβand tau protein has been characterized in several models.Aβinduces tau hyperphosphorylation through a complex mechanism;however,the master regulators involved in this linear process are still unclear.In our study with Drosophila melanogaster,we found that Aβregulated tau hyperphosphorylation and toxicity by activating c-Jun N-terminal kinase.Importantly,Aβtoxicity was dependent on tau hyperphosphorylation,and flies with hypophosphorylated tau were insulated against Aβ-induced toxicity.Strikingly,tau accumulation reciprocally interfered with Aβdegradation and correlated with the reduction in mRNA expression of genes encoding Aβ-degrading enzymes,including dNep1,dNep3,dMmp2,dNep4,and dIDE.Our results indicate that Aβand tau protein work synergistically to further accelerate Alzheimer’s disease progression and may be considered as a combined target for future development of Alzheimer’s disease therapeutics.展开更多
The risk of hepatocellular carcinoma (HCC) development increases as hepatitis virus C (HCV)-related liver diseases progress, especially in patients with active inflammation. Insight into hepatic carcinogenesis have em...The risk of hepatocellular carcinoma (HCC) development increases as hepatitis virus C (HCV)-related liver diseases progress, especially in patients with active inflammation. Insight into hepatic carcinogenesis have emerged from recent detailed analyses of transforming growth factor-β and c-Jun-N-terminal kinase signaling processes directed by multiple phosphorylated (phospho)-isoforms of a Smad3 mediator. In the course of HCV-related chronic liver diseases, chronic inflammation and host genetic/epigenetic alterations additively shift the hepatocytic Smad3 phospho-isoform signaling from tumor suppression to carcinogenesis, increasing the risk of HCC. Chronic inflammation represents an early carcinogenic step that provides a nonmutagenic tumor-promoting stimulus. After undergoing successful antiviral therapy, patients with chronic hepatitis C could experience a lower risk of HCC as Smad3 phospho-isoform signaling reverses from potential carcinogenesis to tumor suppression. Even after HCV clearance, however, patients with cirrhosis could still develop HCC because of sustained, intense carcinogenic Smad3 phospho-isoform signaling that is possibly caused by genetic or epigenetic alterations. Smad3 phospho-isoforms should assist with evaluating the effectiveness of interventions aimed at reducing human HCC.展开更多
The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as oppos...The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as opposed to conventional GPCR signaling via G proteins. Several recent studies using HEK293 cells where all G proteins were genetically ablated or inactivated, or both non-visual arrestins were knocked out, demonstrated that ERK1/2 phosphorylation requires G protein activity, but does not necessarily require the presence of non-visual arrestins. This appears to contradict the prevailing paradigm. Here we discuss these results along with the recent data on gene edited cells and arrestinmediated signaling. We suggest that there is no real controversy. G proteins might be involved in the activation of the upstream-most MAP3Ks, although in vivo most MAP3K activation is independent of heterotrimeric G proteins, being initiated by receptor tyrosine kinases and/or integrins. As far as MAP kinases are concerned, the best-established role of arrestins is scaffolding of the three-tiered cascades(MAP3K-MAP2 K-MAPK). Thus, it seems likely that arrestins, GPCRbound and free, facilitate the propagation of signals in these cascades, whereas signal initiation via MAP3K activation may be independent of arrestins. Different MAP3Ks are activated by various inputs, some of which are mediated by G proteins, particularly in cell culture, where we artificially prevent signaling by receptor tyrosine kinases and integrins, thereby favoring GPCR-induced signaling. Thus, there is no reason to change the paradigm: Arrestins and G proteins play distinct non-overlapping roles in cell signaling.展开更多
CSN1 is a component of the COP9 signalosome(CSN),a conserved protein complex with pleiotropic functions in many organs and cell types.CSN regulates ubiquitinproteasome dependent protein degradation via the deneddylati...CSN1 is a component of the COP9 signalosome(CSN),a conserved protein complex with pleiotropic functions in many organs and cell types.CSN regulates ubiquitinproteasome dependent protein degradation via the deneddylation and the associated deubiquitination activities.In addition,CSN associates with protein kinases and modulates cell signaling,particularly the activator protein 1(AP-1)pathway.We have shown previously that CSN1 suppresses AP-1 transcription activity and inhibits ultraviolet(UV)and serum activation of c-fos expression.Here we show that CSN1 can inhibit phosphorylation of proto-oncogene c-Jun product and repress c-Jun dependent transcription.Further,CSN1 dramatically downregulates ectopic expression of c-Jun N-terminal kinase 1(JNK1)in cultured cells.The decline in JNK1 is not caused by excessive proteolysis or by 3′UTR-dependent mRNA instability,but by CSN1-dependent repression of one or multiple steps in transcriptional and posttranscriptional mechanisms.Thus,in contrast to CSN5/Jab1,which promotes AP-1 activity,CSN1 displays a negative effect on the AP-1 pathway.Finally,we discuss about the dynamic equilibrium of the CSN complexes in regulation of the AP-1 pathway.展开更多
This study aims to elucidate the antiproliferative mechanism of hydroxychavicol(HC).Its effects on cell cycle,apoptosis,and the expression of c-Jun N-terminal kinase(JNK)and P38 mitogen-activated protein kinase(MAPK)i...This study aims to elucidate the antiproliferative mechanism of hydroxychavicol(HC).Its effects on cell cycle,apoptosis,and the expression of c-Jun N-terminal kinase(JNK)and P38 mitogen-activated protein kinase(MAPK)in HT-29 colon cancer cells were investigated.HC was isolated from Piper betle leaf(PBL)and verified by high-performance liquid chromatography(HPLC),nuclear magnetic resonance(NMR),and gas chromatography-mass spectrometry(GC-MS).The cytotoxic effects of the standard drug 5-fluorouracil(5-FU),PBL water extract,and HC on HT-29 cells were measured after 24,48,and 72 h of treatment.Cell cycle and apoptosis modulation by 5-FU and HC treatments were investigated up to 30 h.Changes in phosphorylated JNK(pJNK)and P38(pP38)MAPK expression were observed up to 18 h.The half maximal inhibitory concentration(IC_(50))values of HC(30μg/mL)and PBL water extract(380μg/mL)were achieved at 24 h,whereas the IC_(50)of 5-FU(50μmol/L)was obtained at 72 h.Cell cycle arrest at the G0/G1 phase in HC-treated cells was observed from12 h onwards.Higher apoptotic cell death in HC-treated cells compared to 5-FU-treated cells(P<0.05)was observed.High expression of pJNK and pP38 MAPK was observed at 12 h in HC-treated cells,but not in 5-FU-treated HT-29 cells(P<0.05).It is concluded that HC induces cell cycle arrest and apoptosis of HT-29 cells,with these actions possibly mediated by JNK and P38 MAPK.展开更多
Objective： Salmonella enterica remains a major cause of food-borne disease in humans, and Salmonella Typhimurium （ST） contamination of poultry products is a worldwide problem. Since macrophages play an essential ro...Objective： Salmonella enterica remains a major cause of food-borne disease in humans, and Salmonella Typhimurium （ST） contamination of poultry products is a worldwide problem. Since macrophages play an essential role in controlling Salmonella infection, the aim of this study was to evaluate the effect of glycyrrhizic acid （GA） on immune function of chicken HD11 macrophages. Methods： Chicken HD11 macrophages were treated with GA （0, 12.5 25, 50, 100, 200, 400, or 800 pg/ml） and lipopolysaccharide （LPS, 500 ng/ml） for 3, 6, 12, 24, or 48 h. Evaluated responses included phagocytosis, bacteria-killing, gene expression of cell surface molecules （cluster of differentiation 40 （CD40）, CD80, CD83, and CD197） and antimicrobial effectors （inducible nitric oxide synthase （iNOS）, NADPH oxidase-1 （NOX-1）, interferon-γ （IFN-γ）, LPS-induced tumor necrosis factor （TNF）-α factor （LITAF）, interleukin-6 （IL-6） and IL-IO）, and production of nitric oxide （NO） and hydrogen peroxide （H202）. Results： GA increased the internalization of both fiuorescein isothiocyanate （FITC）-dextran and ST by HD11 cells and markedly decreased the intracellular survival of ST. We found that the messenger RNA （mRNA） expression of cell surface molecules （CD40, CDSO, CD83, and CD197） and cytokines （IFN-γ, IL-6, and IL-10） of HD11 cells was up-regulated following GA exposure. The expression of iNOS and NOX-1 was induced by GA and thereby the productions of NO and H202 in HD11 cells were enhanced. Notably, it was verified that nuclear factor-κB （NF-κB） and c-Jun N-terminal kinase （JNK） pathways were responsible for GA-induced synthesis of NO and IFN-γ gene expression. Conclusions： Taken together, these results suggested that GA exhibits a potent immune regulatory effect to activate chicken macrophages and enhances Salmonella-killing capacity.展开更多
Liver injury and acute liver failure caused by acetaminophen(APAP)overdose is the clinically most important drug toxicity in Western countries.Mechanistic investigations have revealed a central role of mitochondria in...Liver injury and acute liver failure caused by acetaminophen(APAP)overdose is the clinically most important drug toxicity in Western countries.Mechanistic investigations have revealed a central role of mitochondria in the pathophysiology.Excess formation of the reactive metabolite N-acetyl-p-benzoquinone imine(NAPQI)after an overdose leads to hepatic glutathione depletion,mitochondrial protein adducts formation and an initial oxidant stress,which triggers the activation of mitogen activated protein(MAP)kinase cascade ultimately leading to c-jun N-terminal kinase(JNK)phosphorylation.Phospho-JNK translocates to the mitochondria and amplifies the oxidative and nitrosative stress eventually causing the mitochondrial membrane permeability transition pore opening and cessation of adenosine triphosphate(ATP)synthesis.In addition,mitochondrial matrix swelling ruptures the outer membrane and releases endonucleases,which cause nuclear deoxyribonucleic acid(DNA)fragmentation.Together,the nuclear DNA damage and the extensive mitochondrial dysfunction result in necrotic cell death.However,the procell death signaling events are counteracted by adaptive responses such as autophagy and mitochondrial biogenesis.The improved mechanistic insight into the pathophysiology leads to better understanding of the mechanisms of action of the existing antidote N-acetylcysteine and justifies the clinical testing of novel therapeutics such as 4-methylpyrazole and calmangafodipir.展开更多
基金Supported by SNUH Research Fund,Grant NO 04-2016-0220the Education and Research Encouragement Fund of Seoul National University Hospital(2015)
文摘AIM To investigated the relationships between HER2, c-Jun N-terminal kinase(JNK) and protein kinase B(AKT) with respect to metastatic potential of HER2-positive gastric cancer(GC) cells.METHODS Immunohistochemistry was performed on tissue array slides containing 423 human GC specimens. Using HER2-positve GC cell lines SNU-216 and NCI-N87, HER2 expression was silenced by RNA interference, and the activations of JNK and AKT were suppressed by SP600125 and LY294002, respectively. Transwell assay, Western blot, semi-quantitative reverse transcriptionpolymerase chain reaction and immunofluorescence staining were used in cell culture experiments. RESULTS In GC specimens, HER2, JNK, and AKT activations were positively correlated with each other. In vitro analysis revealed a positive regulatory feedback loop between HER2 and JNK in GC cell lines and the role of JNK as a downstream effector of AKT in the HER2/AKT signaling pathway. JNK inhibition suppressed migratory capacity through reversing EMT and dual inhibition of JNK and AKT induced a more profound effect on cancer cell motility.CONCLUSION HER2, JNK and AKT in human GC specimens are positively associated with each other. JNK and AKT, downstream effectors of HER2, co-operatively contribute to the metastatic potential of HER2-positive GC cells. Thus, targeting of these two molecules in combination with HER2 downregulation may be a good approach to combat HER2-positive GC.
基金This work was supported by a grant from the Project of China Postdoctoral Science Foundation （No. 20100480568）.
文摘Background C-Jun N-terminal kinase （JNK） signaling pathway plays a critical role in cerebral ischemia. Although the mechanistic basis for this activation of JNK1/2 is uncertain, oxidative stress may play a role. The purpose of this study was to investigate whether the activation of JNK1/2 is associated with the production of endogenous nitric oxide （NO）. Methods Ischemia and reperfusion （I/R） was induced by cerebral four-vessel occlusion. Sprague-Dawley （SD） rats were divided into 6 groups： sham group, I/R group, neuronal nitric oxide synthase （nNOS） inhibitor （7-nitroindazole, 7-NI） given group, inducible nitric oxide synthase （iNOS） inhibitor （2-amino-5,6-dihydro-methylthiazine, AMT） given group, sodium chloride control group, and 1% dimethyl sulfoxide （DMSO） control group. The levels of protein expression and phospho-JNK1/2 were detected by Western blotting and the survival hippocampus neurons in CA1 zone were observed by cresyl violet staining. Results The study illustrated two peaks of JNK1/2 activation occurred at 30 minutes and 3 days during reperfusion. 7-NI inhibited JNK1/2 activation during the early reperfusion, whereas AMT preferably attenuated JNK1/2 activation during the later reperfusion. Administration of 7-NI and AMT can decrease I/R-induced neuronal loss in hippocampal CA1 region. Conclusion JNK1/2 activation is associated with endogenous NO in response to ischemic insult.
文摘OBJECTIVE N2L is a novel lipoic acid-niacin dimer regulating lipid metabolism with multifunction,including antioxidant effect.We investigated the protective effect of N2L and the underlying mechanisms under the ferroptosis inducer RAS-selective lethality 3(RSL3)treat⁃ment in HT22 cells.METHODS HT22 cells were pretreated with N2L and then were treated with RSL3 to establish a ferroptosis cell model.MTT assay was used to detect the cell survival rate.Free radical probe(dihydroethidium,DHE)and ferrous probe FerroOrange were used to detect the contents of free radicals and ferrous ions in cells.The ultrastructure of mitochondria of treat⁃ed cells was observed by transmission electron microscope.The expression of ferroptosis-relat⁃ed proteins acyl-CoA synthetase long-chain family member 4(ACSL4),glutathione peroxidase 4(GPX4),cyclooxygenase-2(COX-2),ferritin Heavy Chain 1(FTH1),nuclear factor E2-related factor 2/heme oxygenase-1,and phosphoryla⁃tion levels of the c-Jun N-terminal kinase(JNK)/extracellular regulated protein kinases(ERK)pathway were detected by Western blotting.RE⁃SULTS RSL3 decreased the cell viability and induced excessive accumulation of(reactive oxy⁃gen species)ROS in HT22 cells.N2L pretreat⁃ment effectively protected HT22 cells against lipid peroxidation.What′s more,N2L recovered GPX4 protein expression and blocked the increase of COX-2 and ACSL4 expressions.Moreover,N2L also significantly prevented FTH1 from downregulation and maintained iron homeo⁃stasis.Finally,N2L pretreatment could decrease JNK/ERK activation induced by RSL3.CON⁃CLUSION N2L is an excellent ferroptosis inhibi⁃tor,and its anti-ferroptosis mechanism may be related to the reduction of lipid peroxidation and the regulation of iron homeostasis.
文摘Mixed lineage kinase 3 （MLK3） is a mitogen-activated protein kinase kinase kinase that is activated by tumor necrosis factor-α （TNF-α） and specifically activates c-Jun N-terminal kinase （JNK） on TNF-a stimulation. The mecha- nism by which TNF-α activates MLK3 is still not known. TNF receptor-associated factors （TRAFs） are adapter molecules that are recruited to cytoplasmic end of TNF receptor and mediate the downstream signaling, including activation of JNK. Here, we report that MLK3 associates with TRAF2, TRAF5 and TRAF6; however only TRAF2 can significantly induce the kinase activity of MLK3. The interaction domain of TRAF2 maps to the TRAF domain and for MLK3 to its C-terminal half （amino acids 511-847）. Endogenous TRAF2 and MLK3 associate with each other in response to TNF-α treatment in a time-dependent manner. The association between MLK3 and TRAF2 mediates MLK3 activation and competition with the TRAF2 deletion mutant that binds to MLK3 attenuates MLK3 kinase activity in a dose-dependent manner, on TNF-α treatment. Furthermore the downstream target of MLK3, JNK was activated by TNF-α in a TRAF2-dependent manner. Hence, our data show that the direct interaction between TRAF2 and MLK3 is required for TNF-α-induced activation of MLK3 and its downstream target, JNK.
文摘Objective:The aim of the study was to investigate the effect of c-Jun N-terminal protein kinase(JNK) signaling pathway on influencing the sensitivity to radiotherapy of human nasopharyngeal carcinoma CNE cells.Methods:Human nasopharyngeal carcinoma CNE multicellular spheroids(MCS) were constructed with three dimensional cell culture methods.Western blot was employed to analyze the activity of JNK signaling pathway in MCS after X-ray irradiation,and the expression of caspase-3 protein before and after using SP600125(a special inhibitor of JNK).X-ray induced cell apoptosis in MCS before and after treated with SP600125 were detected by TUNEL.Results:The level of JNK phosphorylation in MCS was a dynamic course after radiation,and there was a phosphorylation peaks at 2 h later,the apoptotic rate of MCS(P < 0.05) and the expression of caspase-3 protein(P < 0.05) were significantly increased after treated with SP600125.Conclusion:The transient activation of JNK played a important role in sensitivity to radiotherapy of CNE MCS via mediating survival signals,blocking this pathway accelerate cell apoptosis,which may be related to the increased expression of caspase-3.
基金This study was supported by the grants from the National Natural Science Foundation of China，the Shandong Province National Natural Science Foundation，the project from Weifang Science and Technology Bureau，the Weifang Medical Technology Innovation Project
文摘Background： Najanalgesin, a toxin isolated from the venom ofNaja nqja atra, has been shown to exert significant analgesic effects in a neuropathic pain model in rats. However, the molecular mechanism underlying this protective effect ofnajanalgesin is poorly understood. The present study sought to evaluate the intracellular signaling pathways that are involved in the antinociceptive effect of najanalgesin on neuropathic pain. Methods： The antinociceptive properties of najanalgesin were tested in hind paw withdrawal thresholds in response to mechanical stimulation. We analyzed the participation of the mitogen-activated protein kinase p38, extracellular-regulated kinase （ERK）, and c-Jun N-terminal kinase （JNK） by western blot analysis. This inhibition of JNK was confirmed by immunohistochemistry. Results： The phosphorylation levels of INK （as well as its downstream molecule c-Jun）, p38, and ERK were significantly increased after injury. Najanalgesin only inhibited JNK and c-Jun phosphorylation but had no effect on either ERK or p38. This inhibition of JNK was confirmed by immunohistochemistry, which suggested that the antinociceptive effect of najanalgesin on spinal nerve ligation-induced neuropathic pain in rats is associated with JNK activation in the spinal cord. Conclusion： The antinociceptive effect of najanalgesin thnctions by inhibiting the JNK in a neuropathic pain model.
基金supported by the National Basic Research Program of China,Nos.31700883(to YPH)and 91649118(to BZ)China Postdoctoral Science Foundation,No.2015M581072(to YPH)the Strategic Priority Research Program of the Chinese Academy of Sciences,No.XDB38000000(to JRW).
文摘Extracellular aggregation of amyloid-beta(Aβ)and intracellular tau tangles are two major pathogenic hallmarks and critical factors of Alzheimer’s disease.A linear interaction between Aβand tau protein has been characterized in several models.Aβinduces tau hyperphosphorylation through a complex mechanism;however,the master regulators involved in this linear process are still unclear.In our study with Drosophila melanogaster,we found that Aβregulated tau hyperphosphorylation and toxicity by activating c-Jun N-terminal kinase.Importantly,Aβtoxicity was dependent on tau hyperphosphorylation,and flies with hypophosphorylated tau were insulated against Aβ-induced toxicity.Strikingly,tau accumulation reciprocally interfered with Aβdegradation and correlated with the reduction in mRNA expression of genes encoding Aβ-degrading enzymes,including dNep1,dNep3,dMmp2,dNep4,and dIDE.Our results indicate that Aβand tau protein work synergistically to further accelerate Alzheimer’s disease progression and may be considered as a combined target for future development of Alzheimer’s disease therapeutics.
文摘The risk of hepatocellular carcinoma (HCC) development increases as hepatitis virus C (HCV)-related liver diseases progress, especially in patients with active inflammation. Insight into hepatic carcinogenesis have emerged from recent detailed analyses of transforming growth factor-β and c-Jun-N-terminal kinase signaling processes directed by multiple phosphorylated (phospho)-isoforms of a Smad3 mediator. In the course of HCV-related chronic liver diseases, chronic inflammation and host genetic/epigenetic alterations additively shift the hepatocytic Smad3 phospho-isoform signaling from tumor suppression to carcinogenesis, increasing the risk of HCC. Chronic inflammation represents an early carcinogenic step that provides a nonmutagenic tumor-promoting stimulus. After undergoing successful antiviral therapy, patients with chronic hepatitis C could experience a lower risk of HCC as Smad3 phospho-isoform signaling reverses from potential carcinogenesis to tumor suppression. Even after HCV clearance, however, patients with cirrhosis could still develop HCC because of sustained, intense carcinogenic Smad3 phospho-isoform signaling that is possibly caused by genetic or epigenetic alterations. Smad3 phospho-isoforms should assist with evaluating the effectiveness of interventions aimed at reducing human HCC.
基金Supported by National Institutes of Health RO1 grants,No.EY011500National Institutes of Health R35 grants,No.GM122491Cornelius Vanderbilt Endowed Chair(Vanderbilt University),No.NS065868(to Gurevich VV)and No.DA030103(to Gurevich EV)
文摘The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as opposed to conventional GPCR signaling via G proteins. Several recent studies using HEK293 cells where all G proteins were genetically ablated or inactivated, or both non-visual arrestins were knocked out, demonstrated that ERK1/2 phosphorylation requires G protein activity, but does not necessarily require the presence of non-visual arrestins. This appears to contradict the prevailing paradigm. Here we discuss these results along with the recent data on gene edited cells and arrestinmediated signaling. We suggest that there is no real controversy. G proteins might be involved in the activation of the upstream-most MAP3Ks, although in vivo most MAP3K activation is independent of heterotrimeric G proteins, being initiated by receptor tyrosine kinases and/or integrins. As far as MAP kinases are concerned, the best-established role of arrestins is scaffolding of the three-tiered cascades(MAP3K-MAP2 K-MAPK). Thus, it seems likely that arrestins, GPCRbound and free, facilitate the propagation of signals in these cascades, whereas signal initiation via MAP3K activation may be independent of arrestins. Different MAP3Ks are activated by various inputs, some of which are mediated by G proteins, particularly in cell culture, where we artificially prevent signaling by receptor tyrosine kinases and integrins, thereby favoring GPCR-induced signaling. Thus, there is no reason to change the paradigm: Arrestins and G proteins play distinct non-overlapping roles in cell signaling.
基金supported by research grants from the National Institutes of Health(GM61812)to NWthe Human Frontier Long Term Fellowship(LT0084/1998-M)to TTa collaborative grant from The Kyoto University Foundation(2007-2008)to NW,SM,and TT.
文摘CSN1 is a component of the COP9 signalosome(CSN),a conserved protein complex with pleiotropic functions in many organs and cell types.CSN regulates ubiquitinproteasome dependent protein degradation via the deneddylation and the associated deubiquitination activities.In addition,CSN associates with protein kinases and modulates cell signaling,particularly the activator protein 1(AP-1)pathway.We have shown previously that CSN1 suppresses AP-1 transcription activity and inhibits ultraviolet(UV)and serum activation of c-fos expression.Here we show that CSN1 can inhibit phosphorylation of proto-oncogene c-Jun product and repress c-Jun dependent transcription.Further,CSN1 dramatically downregulates ectopic expression of c-Jun N-terminal kinase 1(JNK1)in cultured cells.The decline in JNK1 is not caused by excessive proteolysis or by 3′UTR-dependent mRNA instability,but by CSN1-dependent repression of one or multiple steps in transcriptional and posttranscriptional mechanisms.Thus,in contrast to CSN5/Jab1,which promotes AP-1 activity,CSN1 displays a negative effect on the AP-1 pathway.Finally,we discuss about the dynamic equilibrium of the CSN complexes in regulation of the AP-1 pathway.
基金supported by the Taylor’s Research Grant Scheme(No.TRGS/MFS/2/2013/SBS/003),Malaysia。
文摘This study aims to elucidate the antiproliferative mechanism of hydroxychavicol(HC).Its effects on cell cycle,apoptosis,and the expression of c-Jun N-terminal kinase(JNK)and P38 mitogen-activated protein kinase(MAPK)in HT-29 colon cancer cells were investigated.HC was isolated from Piper betle leaf(PBL)and verified by high-performance liquid chromatography(HPLC),nuclear magnetic resonance(NMR),and gas chromatography-mass spectrometry(GC-MS).The cytotoxic effects of the standard drug 5-fluorouracil(5-FU),PBL water extract,and HC on HT-29 cells were measured after 24,48,and 72 h of treatment.Cell cycle and apoptosis modulation by 5-FU and HC treatments were investigated up to 30 h.Changes in phosphorylated JNK(pJNK)and P38(pP38)MAPK expression were observed up to 18 h.The half maximal inhibitory concentration(IC_(50))values of HC(30μg/mL)and PBL water extract(380μg/mL)were achieved at 24 h,whereas the IC_(50)of 5-FU(50μmol/L)was obtained at 72 h.Cell cycle arrest at the G0/G1 phase in HC-treated cells was observed from12 h onwards.Higher apoptotic cell death in HC-treated cells compared to 5-FU-treated cells(P<0.05)was observed.High expression of pJNK and pP38 MAPK was observed at 12 h in HC-treated cells,but not in 5-FU-treated HT-29 cells(P<0.05).It is concluded that HC induces cell cycle arrest and apoptosis of HT-29 cells,with these actions possibly mediated by JNK and P38 MAPK.
基金Project supported by the National Natural Science Foundation of China(No.31472128)
文摘Objective： Salmonella enterica remains a major cause of food-borne disease in humans, and Salmonella Typhimurium （ST） contamination of poultry products is a worldwide problem. Since macrophages play an essential role in controlling Salmonella infection, the aim of this study was to evaluate the effect of glycyrrhizic acid （GA） on immune function of chicken HD11 macrophages. Methods： Chicken HD11 macrophages were treated with GA （0, 12.5 25, 50, 100, 200, 400, or 800 pg/ml） and lipopolysaccharide （LPS, 500 ng/ml） for 3, 6, 12, 24, or 48 h. Evaluated responses included phagocytosis, bacteria-killing, gene expression of cell surface molecules （cluster of differentiation 40 （CD40）, CD80, CD83, and CD197） and antimicrobial effectors （inducible nitric oxide synthase （iNOS）, NADPH oxidase-1 （NOX-1）, interferon-γ （IFN-γ）, LPS-induced tumor necrosis factor （TNF）-α factor （LITAF）, interleukin-6 （IL-6） and IL-IO）, and production of nitric oxide （NO） and hydrogen peroxide （H202）. Results： GA increased the internalization of both fiuorescein isothiocyanate （FITC）-dextran and ST by HD11 cells and markedly decreased the intracellular survival of ST. We found that the messenger RNA （mRNA） expression of cell surface molecules （CD40, CDSO, CD83, and CD197） and cytokines （IFN-γ, IL-6, and IL-10） of HD11 cells was up-regulated following GA exposure. The expression of iNOS and NOX-1 was induced by GA and thereby the productions of NO and H202 in HD11 cells were enhanced. Notably, it was verified that nuclear factor-κB （NF-κB） and c-Jun N-terminal kinase （JNK） pathways were responsible for GA-induced synthesis of NO and IFN-γ gene expression. Conclusions： Taken together, these results suggested that GA exhibits a potent immune regulatory effect to activate chicken macrophages and enhances Salmonella-killing capacity.
基金supported by USA National Institute of Diabetes and Digestive and Kidney Diseases(NIDDK)R01 NIDDK102142 and R01 NIDDK 070195.
文摘Liver injury and acute liver failure caused by acetaminophen(APAP)overdose is the clinically most important drug toxicity in Western countries.Mechanistic investigations have revealed a central role of mitochondria in the pathophysiology.Excess formation of the reactive metabolite N-acetyl-p-benzoquinone imine(NAPQI)after an overdose leads to hepatic glutathione depletion,mitochondrial protein adducts formation and an initial oxidant stress,which triggers the activation of mitogen activated protein(MAP)kinase cascade ultimately leading to c-jun N-terminal kinase(JNK)phosphorylation.Phospho-JNK translocates to the mitochondria and amplifies the oxidative and nitrosative stress eventually causing the mitochondrial membrane permeability transition pore opening and cessation of adenosine triphosphate(ATP)synthesis.In addition,mitochondrial matrix swelling ruptures the outer membrane and releases endonucleases,which cause nuclear deoxyribonucleic acid(DNA)fragmentation.Together,the nuclear DNA damage and the extensive mitochondrial dysfunction result in necrotic cell death.However,the procell death signaling events are counteracted by adaptive responses such as autophagy and mitochondrial biogenesis.The improved mechanistic insight into the pathophysiology leads to better understanding of the mechanisms of action of the existing antidote N-acetylcysteine and justifies the clinical testing of novel therapeutics such as 4-methylpyrazole and calmangafodipir.
