Activation Of JNK Signaling Pathway Research Articles

MiR-92a/DUSP10/JNK signalling axis promotes human pancreatic cancer cells proliferation

Pancreatic cancer is one of the most common types of cancers in the whole world with a poor prognosis. Finding out how the cancer form and develop is the most important way to cure this cancer. miRNAs, 21–22 nucleotides regulatory small non-coding RNAs, have been found to be critical involved in the growth of pancreatic cancer. In this study, we found that miR-92a was up regulated in three kinds of human pancreatic cancer cell lines. There is a correlation between miR-92a and malignant degree of human pancreatic cancer cell lines. Then we found that miR-92a was essential for promoting cell proliferation in human pancreatic cancer. Inhibition of the function of miR-92a repressed the proliferation of pancreatic cancer cells. Further, we found that miR-92a enhanced the activation of JNK signalling pathway by directly targeting the JNK signalling inhibitor DUSP10. DUSP10 is responsible for miR-92a induced JNK signalling and cell proliferation. Altogether, our study showed a miR-92a/DUSP10/JNK signalling pathway that plays an important role in regulating the proliferation of pancreatic cancer cells.

JNK pathway may be involved in isoflurane-induced apoptosis in the hippocampi of neonatal rats

Previous studies have demonstrated that isoflurane, a commonly used volatile anesthetic, can induce widespread apoptosis in the neonatal animal brains and result in persistent cognitive impairment. Isoflurane-induced cytosolic Ca2+ overload and activation of mitochondrial pathway of apoptosis may be involved in this neurodegeneration. The c-Jun N-terminal kinase (JNK) signaling can regulate the expression of the Bcl-2 family members that modulates mitochondrial membrane integrity. Therefore, we hypothesize that JNK signaling pathway activation contributes to isoflurane-induced apoptosis in the brain. In this study, Sprague-Dawley neonatal rats at postnatal day 7 were exposed to 1.1% isoflurane or air for 4h. The JNK inhibitor SP600125 at 5μg, 10μg, 20μg, 30μg or the vehicle was intraventricularly administered before the exposure. Neuronal apoptosis in the hippocampi of neonatal rats was detected by TUNEL 6h after isoflurane or air exposure. The protein expression of phospho-JNK, phospho-c-Jun, and caspase-3 as well as the antiapoptotic protein Bcl-xL and Akt/glycogen synthase kinase (GSK)-3β pathway was detected by Western blotting. Isoflurane significantly increased apoptotic cells in the hippocampal CA1, CA3, and DG regions. The JNK inhibitor SP600125 dose-dependently inhibited isoflurane-induced neuronal apoptosis and increase of caspase-3 and phospho-JNK. SP600125 also attenuated isoflurane-induced down-regulation of Bcl-xL and maintained the activated Akt level to increase the phosphorylation of GSK-3β at Ser9. Our results indicate that JNK activation contributes to isoflurane-induced neuroapoptosis in the developing brain. Maintaining Bcl-xL and Akt activation may be involved in the neuroprotective effects of SP600125.

Characterization of a microtubule‐associated protein, doublecortin (DCX), as a substrate of c‐Jun N‐terminal Kinases (JNKs)

We are interested how docking motifs contribute to substrate recognition by JNK, a Ser/Thr protein kinase and member of the Mitogen‐Activated Protein Kinase (MAPK) family. The activation of JNK signaling pathways and concomitant phosphorylation of a range of JNK substrates has been implicated with regulation of critical events, including cell growth and proliferation, cell division and cell death. Specifically, we are defining the structural and functional properties of the microtubule‐associated protein doublecortin (DCX) as a JNK substrate. DCX is unusual with its evolutionary conserved DC domains, rather than a typical linear basic/hydrophobic motif, reported to mediate its interaction with JNK. Using mass spectrometry, we have identified a novel JNK‐mediated phosphorylation site in the DCX protein, and confirmed the phosphorylation of this site by mutagenesis and detection by phosphospecific antibodies. Through our use of NMR approaches, we are gaining new structural insights into DCX with the ultimate aim to identify the features of the JNK‐DC domain interaction interface that support this non‐conventional mode of JNK‐substrate recognition.

Monocytes expression of IL-12 related and IL-10 genes in association with development of colorectal cancer

The main regulator of anti-tumor immune response is the activity of monocytes, suggesting that the produced cytokines may have a prognostic role. This study investigates gene expression of interleukin (IL)-12-related cytokine and IL-10 in stimulated monocytes from colorectal cancer (CRC) patients. Relative quantification of IL-12A, IL-12B, IL-23A and IL-10 mRNA transcripts was performed on the third hours after stimulation by real-time qPCR. We also explored an inhibitor of JNK signaling pathway activation for the observed cytokine gene expression. A strong downregulation of IL-12B mRNA expression in CRC monocytes compared to healthy donors was observed. The rate of transcription of IL-12B in stimulated monocytes was associated with the stage of CRC. The expression of IL-12A gene in stimulated monocytes from patients with advanced was lower than early cancer. Moreover, we observed stage dependent JNK inhibition mediated reduction in IL-12A expression. The hyporesponsiveness was strongly expressed in monocytes from advanced then early stages of CRC. Expression of IL-10 mRNA was almost equally in CRC monocytes from early stages and healthy donors. We demonstrated that altered gene expression profiles of IL-12A, IL-12B, IL-23A at mRNA level in CRC monocytes was associated with tumor development and can be attributed to anticancer immune response.

Exercise-Induced Il-6 Gene Transcriptional in Rats Muscle Stimulated by Low Glycogen was Likely not Through Activation of Jnk Signaling Pathway

Exercise-Induced Il-6 Gene Transcriptional in Rats Muscle Stimulated by Low Glycogen was Likely not Through Activation of Jnk Signaling Pathway

P161 Cell-autonomous effect of microglia on the induction of Alzheimer’s disease neuronal cell cycle events

Introduction Massive neuronal loss is the prominent pathological hallmark feature of Alzheimer’s disease (AD). One of the ways post-mitotic neurons die is via their ectopic entry into cell-cycle, which often occurs at very early stages of the disease [1] . We have previously demonstrated occurrence of neuronal cell cycle events (CCEs) in a transgenic mouse model of AD (R1.40) similar to those occur in human AD [1] . Furthermore, these events were dependent upon amyloidogenic processing of amyloid-precursor protein [2] , precede deposition of senile plaques and coincide with elevated neuroinflammation. While we have also demonstrated that Tool-Like Receptor-4 (TLR4) mediated induction of neuroinflammation accelerated neuronal CCEs, and blocking inflammation via NSAIDs completely blocked neuronal CCEs and reduced microglial activation [3] , the source of neuroinflammation and molecular mechanisms for the induction of neuronal CCEs is unclear. Methods We utilized following methods: immunohistochemistry, purification and adoptive transfer of microglia, primary neurons and microglial cultures and generation and analysis of R1.40- Tnf α −/− mice. Results Here we demonstrate that neuroinflammation, cell autonomous to microglia, is capable of inducing neuronal CCEs. First, CD45+ microglia is present at six months of age in the affected region of R1.40 mouse brain and increases with age. Second, the F4/80+ microglia spatially co-exist with cyclin D1+ neurons in the temporal cortex of human AD brain. Notably, these cyclin D1+ neurons in the human brain are also TUNEL positive, suggesting that a portion of them is apoptotic. Third, purification and adoptive transfer of CD11b+ microglia derived form R1.40, but not from non-transgenic mice, was sufficient to induce cyclin D1 expression within the neurons of non-transgenic recipient mouse brain. Fourth, conditioned media (CM) derived from amyloid beta activated primary microglia significantly induced dose-dependent increase in DNA replication within primary cortical neurons and strongly correlated with increased levels of tumor-necrosis factor-α(TNFα) in the CM. Notably, blocking or activating TNFα pathway either via anti-TNFα antibody or recombinant human TNFα, significantly induced cyclin D1 expression and/or DNA replication in in vitro as well as in adoptive transfer studies. Generation of R1.40 mice deficient for TNFα (R1.40- Tnfa −/− ) displayed decreased microglial activation and showed significantly lower levels of cyclin D1+ neurons at six months of age. Finally, analysis of downstream molecules of TNFα receptor pathway suggested that neuronal CCEs is mediated via activation of JNK signaling pathway. Conclusion Together, our results provide first direct evidence that the neuroinflammation, cell-autonomous to microglia, induces AD-related neuronal CCEs and neurodegeneration via alteration of TNFR-MAPK signaling pathway.

Cyclophilin A inhibits rotavirus replication by facilitating host IFN-I production

Rotavirus (RV) infection causes serious dehydrating diarrhoea in infants and newborn animals. Our previous study revealed that cyclophilin A (CYPA), a peptidyl-prolyl cis–trans isomerase (PPIase), could be temporarily upregulated in RV-infected MA104 cells in early stage of infection (unpublished data). To find out the possible roles of CYPA in RV infection, we overexpressed and silenced CYPA in various cell lines by gene transfection and shRNA. We found that transfection of CYPA significantly inhibited RV replication, while silencing the expression of CYPA significantly increased RV replication. Accordingly, overexpression of CYPA significantly increased IFN-β production; while silencing CYPA significantly reduced IFN-β production. This effect of CYPA on IFN-β production was independent of its PPIase activity. Moreover, IFN-β secreted by host cells in RV infection had a critical repressive effect on viral replication. Finally, we found that inhibiting JNK pathway by SP600125 and JNK siRNA abrogated the effect of CYPA on IFN-β transcription in RV-infected MA104 cells. Together, our data suggested that CYPA inhibited RV replication by facilitating host IFN-β production, which was independent on the PPIase activity of CYPA but dependent on the activation of JNK signaling pathway.

Activation of c-Jun NH(2)-terminal kinase is required for porcine reproductive and respiratory syndrome virus-induced apoptosis but not for virus replication

Apoptosis of host cells plays a critical role in pathogenesis of virus infection. MAPK kinases especially stress-activated protein kinases c-Jun NH(2)-terminal kinase (SAPK/JNK) and p38 are often involved in virus-mediated apoptosis. It has been shown that porcine reproductive and respiratory syndrome virus (PRRSV) infection resulted in apoptosis of the host cells both in vitro and in vivo. The current investigation was initiated to determine whether stress-activated protein kinases JNK and p38 play a role in apoptosis induction by PRRSV infection. We examined phosphorylation of JNK and p38, and found that JNK but not p38 was activated in response to PRRSV infection. We then examined effects of this kinase on apoptosis induction and virus replication by using specific inhibitor. We found that JNK inhibition by its inhibitor SP600125 led to the abolishment of PRRSV-mediated apoptosis, but did not suppress virus replication. Further studies demonstrated that ROS generation was involved in JNK activation, and Bcl-2 family anti-apoptotic proteins Mcl-1 and Bcl-xl were downstream targets of JNK to mediate apoptosis. We conclude that activation of JNK signaling pathway is essential for PRRSV-mediated apoptosis but not for virus replication.

FTH1 binds to Daxx and inhibits Daxx-mediated cell apoptosis

As a highly conserved nuclear protein, death domain-associated protein (Daxx) plays an important role in transcriptional control, carcinogenesis, and resistance to virus infection and so on. In order to further investigate the mechanism of Daxx, the yeast two-hybrid technique was used to screen the intra-cellular proteins interacting with Daxx. And 13 positive colonies and three proteins interacting with Daxx were obtained. One of the candidate proteins was identified as ferritin, heavy polypeptide 1(FTH1). The interaction between Daxx and FTH1 was further supported by GST pull-down and co-immunoprecipitation respectively. Then Daxx was determined to induce apoptosis and FTH1 can inhibit Daxx-mediated apoptosis. Besides, it is found that Daxx mediated apoptosis through the Fas-Daxx-ASK1-JNK1 signaling pathway, while FTH1 can inhibit the activation of JNK signaling pathway. We present evidence to demonstrate the FTH1 and Daxx are able to participate in apoptosis pathway through JNK signal molecule and FTH1 can inhibit this pathway.

Experimental study of the gastric mucosa damage of the rat in stress ulcer reduced by JNK inhibitor

Objective To investigate the possibilities of c-Jun N-terminal kinase (JNK) inhibitor to reduce gastric mucosal injury in rats with stress ulcer.Methods Twenty-five male SD rats were randomly divided into five groups:control group,1,6,12 hours injured group,inhibitor of JNK intervention group.Immunohistochemistry and Western blotting were used to detect the expression and distribution of gastric mucosa p-JNK after injuried.Simultaneously,in each group of rats,gastric PH value,gastric mucosal blood flow,gastric mucosal injury index (UI) were measured,and the histology of gastric mucosa was observed.Results Compared with the control group,p-JNK in the injured groups were significantly expressed.Injured after 12 hours,p-JNK of epithelial cells in stress ulcer rats was strongly positive reaction.The injury of inhibitor group ( 15.60 ±2.70) was lighter than the 12 hours injured group (24.20 ±2.39) ( P ＜ 0.01 ).Between the 12 hours injured group (2.12 ± 0.14,26.90 ± 2.60) and inhibitor group ( 1.99 ±0.08,26.66 ±0.73),gastric PH value and gastric mucosal blood flow were lower than the control group ( 3.44 ± 0.03,35.46 ± 4.48 ) obviously ( P ＜ 0.01 ),but there was no significant difference be tween the two groups ( t = 0.082 ,P ＞ 0.05 ; t = 0.26 ,P ＞ 0.05 ) .Conclusion In the stress ulcer rats induced by fluid-percussion model,the activation of JNK signaling pathway may initiate and its inhibitors can reduce the gastric mucosal injury. Key words: Brain injury; Fluid-percussion model; Rat; c-Jun N-terminal kinase; Stress ulcer

Rab11 regulates JNK and Raf/MAPK‐ERK signalling pathways during Drosophila wing development

Developmental signalling pathways are regulated by intracellular vesicle trafficking in multicellular organisms. In our earlier communication, we have shown that mutation in Rab11 (a subfamily of the Ypt/Rab gene family) results in the activation of JNK signalling pathways in Drosophila eye. Here, we report that Rab11 regulates JNK and Raf/MAPK-ERK signalling pathways during Drosophila wing development. Using immunofluorescence and immunohistochemical analyses, we show that overexpression of Rab11 in mutant wing imaginal disc cells triggers the induction of apoptosis and activation of JNK and ERK. Further, using a genetic approach it has been shown that Rab11 interacts with the components of these pathways during Drosophila wing development. In addition to this, in Rab11 mutant wing imaginal discs JNK activity was monitored using puc(E)⁶⁹, a P-lacZ enhancer-trap line inserted in puckered (puc). A strong induction of puc in Rab11 mutant wing imaginal disc cells provided a strong support that Rab11 regulates the JNK signalling pathway during Drosophila wing development.

Abstract 228: Cucurbitacin-I (JSI-124) induces apoptosis in chronic lymphocytic leukemia (CLL) cells through inhibiting STAT3 activation and XIAP expression

Abstract Abstract AACR April-2010 Cucurbitacin-I (JSI-124) induces apoptosis in Chronic Lymphocytic Leukemia (CLL) cells through inhibiting STAT3 activation and XIAP expression. Ganchimeg Ishdorj, James B Johnston and Spencer B Gibson Manitoba Institute of Cell Biology, University of Manitoba, Winnipeg, Canada Chronic lymphocytic leukemia (CLL) is the most common adult leukemia in North America. Current therapies control the disease but eventually the disease recurs and becomes drug resistant. Phosphorylation of signal transducer and activator of transcription 3 (STAT3) plays a pivotal role in cell survival and induces proliferation, whereas deletion of STAT3 results in cell death. Constitutive phosphorylation of STAT3 on tyrosine 705 residues has been found in a number of solid tumors and hematologic malignancies. Unlike other tumors, STAT3 is constitutively phosphorylated (p) on serine 727 but not on tyrosine residues in CLL. These observations lead to investigation of the effect of STAT3/JAK2 inhibitor Cucurbitacin-I (JSI-124) in CLL cells. We proposed that the targeting STAT3 phosphorylation may induce apoptosis in CLL cells. We studied B-cell lines (BJAB, I-83, NALM-6) and primary B-cells from patients with CLL using immunoprecipitation and found that JSI-124 suppressed serine pSTAT3 and induced apoptosis of these cells as determined by FACS analysis and membrane permeability assay. Using western blotting analysis, caspase-8 activation was observed in these cells after JSI-124 treatment. Since the activation of caspases are inhibited by members of the inhibitor of apoptosis (IAP) and Bcl-2 protein families and STAT3 has been shown to regulate their expression, we investigated anti-apoptotic members of these families. JSI-124 treatment led to downregulation of IAP protein XIAP, while Bcl-xL and Mcl-1 levels remained unchanged. In addition, JSI-124 selectively induced activation of JNK signaling pathways in CLL cells. Taken together, our results indicate that JSI-124 causes apoptosis through alterations in signal transduction pathways and could be a useful treatment in CLL. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 228.

Human cytomegalovirus induces caspase-dependent apoptosis of megakaryocytic CHRF-288-11 cells by activating the JNK pathway

Human cytomegalovirus (HCMV) infection is usually implicated in thrombocytopenia occurring in newborns and immunocompromised patients. However, the underlying mechanisms remain elusive. This study was conducted to investigate the effects of HCMV infection on the viability of megakaryocytic CHRF-288-11 cells and the underlying mechanisms involved. RT-PCR for determining mRNA expression of HCMV immediate early gene 1 and Western blot for measuring protein expression of late HCMV gene pp65 showed that CHRF-288-11 cells were susceptible to HCMV infection. HCMV infection reduced the viability of CHRF-288-11 cells via apoptosis in a dose- and time-dependent manner. Both caspase 3 and c-Jun terminal kinase (JNK) signaling pathway were activated in the HCMV-treated CHRF-288-11 cells. z-DEVD-fmk (a caspase inhibitor) and SP600125 (a JNK inhibitor) significantly prevented the death of CHRF-288-11 cells induced by HCMV, respectively. Furthermore, inhibition of JNK activity could reduce the formation of active caspase 3 induced by HCMV. Interestingly, the co-application of antivirus drug ganciclovir and SP600125 synergistically prevented the death of CHRF-288-11 cells induced by HCMV. Collectively, these findings suggest that HCMV infection may induce the caspase-dependent apoptosis of megakaryocytic CHRF-288-11 cells by the activation of JNK signaling pathway.

JNK1/2 siRNA inhibits transforming-growth factor-β1-induced connective tissue growth factor expression and fibrotic function in THSFs

Transforming-growth factor-beta1 (TGF-beta1) plays an important role in fibrosis by inducing connective tissue growth factor (CTGF) expression. In cornea, however, signaling pathway in the process is unclear. In this paper, real-time RT-PCR and Western blot analysis were used to examine the expression and TGF-beta1-induced upregulation of CTGF in telomerase-immortalized human cornea stroma fibroblast cells (THSFs). A detailed molecular mechanism for TGF-beta1-mediated cell proliferation and collagen I production was then investigated by detecting the activation of JNK pathway in TGF-beta1-stimulated THSFs and blocking the JNK pathway using JNK1/2 siRNA. Both TGF-beta1 and CTGF clearly induced cell proliferation in THSFs which was inhibited by functional blocking CTGF by anti-CTGF antibody. Blocking JNK pathway by JNK1/2 siRNA significantly inhibited induction of CTGF by TGF-beta1. Taken together, the data indicate that CTGF is critical to the fibrosis process in wounded human cornea, acting via upregulation and activation of JNK signaling pathway.

Lysophosphatidylcholine up‐regulates human endothelial nitric oxide synthase gene transactivity by c‐Jun N‐terminal kinase signalling pathway

Human endothelial nitric oxide synthase (eNOS) plays a pivotal role in maintaining blood pressure homeostasis and vascular integrity. It has recently been reported that mitogen-activated protein kinases (MAPKs) are intimately implicated in expression of eNOS. However detailed mechanism mediated by them remains to be clarified. In this study, eNOS gene transactivity in human umbilical vein endothelial cells was up-regulated by stimulation of lysophosphatidylcholine (LPC). The stimulation of LPC highly activated both extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK), with differences in the dynamic processes of activation between them. Unexpectedly, p38 MAPK could not be activated by the stimulation of LPC. The activation of JNK signalling pathway by overexpression of JNK or its upstream kinase active mutant up-regulated the transactivity of eNOS significantly, but the activation of p38 signalling pathway down-regulated it largely. The inhibition of either ERK1/2 or JNK signalling pathway by kinase-selective inhibitors could markedly block the induction of the transactivity by LPC. It was observed by electrophoretic mobility shift assay that LPC stimulated both SP1 and AP1 DNA binding activity to go up. Additionally using decoy oligonucleotides proved that SP1 was necessary for maintaining the basal or stimulated transactivity, whereas AP1 contributed mainly to the increase of the stimulated transactivity. These findings indicate that the up-regulation of the eNOS gene transactivity by LPC involves the enhancement of SP1 transcription factor by the activation of JNK and ERK1/2 signalling pathways and AP1 transcription factor by the activation of JNK signalling pathway.

Protective effect of a standardized Ginkgo extract (ginaton) on renal ischemia/reperfusion injury via suppressing the activation of JNK signal pathway

A new standardized Ginkgo extract (ginaton) destined for i.v. injection was investigated in rats for its protective effect on renal ischemia/reperfusion injury. We report on the elucidation of the downstream mechanism of action of JNK on the renal ischemia/reperfusion injury, which can be explained as the decrease in JNK phosphorylation at 20 min and c-Jun phosphorylation (Ser63/73) at 3h after renal ischemia. At the same time, ginaton attenuated the increased expression of FasL at 3h and caspase3 immunoreactivity at 6h after renal ischemia. Furthermore, ginaton significantly decreased renal epithelial tubular cell apoptosis induced by renal ischemia/reperfusion, alleviating renal ischemia/reperfusion injury. These results cumulatively indicate that ginaton could suppress the JNK-c-Jun-FasL-caspase3 signaling cascade, protecting renal tubular epithelial cells against ischemia/reperfusion-induced apoptosis, which implies that antioxidants may be a potential and effective agent for prevention of the ischemic/reperfusion injury through the suppression extrinsic apoptotic signal pathway induced by JNK signal pathway.

AIP1 Is Critical in Transducing IRE1-mediated Endoplasmic Reticulum Stress Response

We have previously shown that ASK1-interacting protein 1 (AIP1) transduces tumor necrosis factor-induced ASK1-JNK signaling. Because endoplasmic reticulum (ER) stress activates ASK1-JNK signaling cascade, we investigated the role of AIP1 in ER stress-induced signaling. We created AIP1-deficient mice (AIP1-KO) from which mouse embryonic fibroblasts and vascular endothelial cells were isolated. AIP1-KO cells show dramatic reductions in ER stress-induced, but not oxidative stress-induced, ASK1-JNK activation and cell apoptosis. The ER stress-induced IRE1-JNK/XBP-1 axis, but not the PERK-CHOP1 axis, is blunted in AIP1-KO cells. ER stress induced formation of an AIP1-IRE1 complex, and the PH domain of AIP1 is critical for the IRE1 interaction. Furthermore, reconstitution of AIP1-KO cells with AIP1 wild type, not an AIP1 mutant with a deletion of the PH domain (AIP1-DeltaPH), restores ER stress-induced IRE1-JNK/XBP-1 signaling. AIP1-IRE1 association facilitates IRE1 dimerization, a critical step for activation of IRE1 signaling. More importantly, AIP1-KO mice show impaired ER stress-induced IRE1-dependent signaling in vivo. We conclude that AIP1 is essential for transducing the IRE1-mediated ER stress response.

Activation of endothelial cells to pathological status by down-regulation of connexin43

We investigated the effects of connexin43 (Cx43) down-regulation on endothelial function. We used two different sequences of Cx43-specific small interference RNA (siRNA) to reduce de novo synthesis of Cx43 in human aortic endothelial cells and then examined the expression profiles, proliferation activity and viability, and angiogenic potential. The involvement of mitogen-activated protein kinase signalling pathways was analysed. In parallel, the effect of inhibition of gap-junctional communication by connexin-mimetic peptides was evaluated. During the down-regulation of Cx43 by the siRNA, the cells exhibited impaired gap-junctional communication, proliferation, viability, and angiogenic potential. In addition, plasminogen activator inhibitor-1 (PAI-1) and von Willebrand factor were up-regulated. Furthermore, c-jun N-terminal kinase (JNK) and its downstream target c-jun were activated, while caspase-3, p38, and extracellular signal-regulated kinase remained unchanged. Inhibition of JNK by SP600125 blocked the siRNA-induced increased expression of PAI-1 and partially recovered the impaired angiogenic potential. Short-term inhibition of Cx43 channels by connexin-mimetic peptides did not activate JNK. Down-regulation of Cx43 inhibits gap-junctional communication and activates endothelial cells to pathological status, as characterized by up-regulation of coagulatory molecules and impairment of proliferation, viability, and angiogenesis. The processes are associated with activation of JNK signalling pathways and rectified by inhibition of the activation. These results suggest that inadequate expression of Cx43 per se impairs endothelial function by the activation of stress-activated protein kinase.

Protective Effect of PEG 35 000 Da on Renal Cells: Paradoxical Activation of JNK Signaling Pathway During Cold Storage

Polyethylene glycol (PEG), a high-molecular weight colloid, is added to preservation solutions in order to decrease cold- and ischemia-induced injuries of the grafted organ. We evaluated on LLC-PK1, a porcine proximal tubular epithelial cell line (1) the efficiency of several commercial preservation solutions (University of Wisconsin, Euro-Collins, Celsior, SCOT, IGL-1), and (2) whether adding PEG (400-35,000 Da) in a simple extracellular-type buffer modified cell integrity and mitogen-activated protein kinase (MAPK) signaling pathways. SCOT was the most efficient commercial solution. Moreover, only PEG 35,000 Da totally preserved cell viability, induced a decrease on reactive oxygen species production and a decrease on p38-MAPK activation. Furthermore PEG 35,000 Da stimulated c-Jun N-terminal kinase (JNK). However, the inhibition of JNK pathway, with the specific SP600125 inhibitor, in the presence of PEG 35,000 Da did not affect cell survival. We also confirmed on whole pig kidney the protective effect of PEG 35,000 Da on cold-induced tubular injuries. This study confirms PEG antioxidative properties, but we demonstrate that its effect on JNK signaling pathway had also a paradoxical effect on cell death. This sheds a new light on PEG effects during cell preservation, independently from the classical immuno-camouflaging hypothesis.

Neuroprotection against ischemic brain injury by SP600125 via suppressing the extrinsic and intrinsic pathways of apoptosis

Our previous studies and the others have strongly suggested that JNK signaling pathway plays a critical role in ischemic brain injury. Here, we reported that SP600125, a potent, cell-permeable, selective, and reversible inhibitor of c-Jun N-terminal kinase (JNK), potently decrease neuronal apoptosis induced by global ischemia/reperfusion in the vulnerable hippocampal CA1 subregion. As a result, SP600125 diminished the increased phosphorylation of c-Jun and the increased expression of FasL induced by ischemia/reperfusion in the vulnerable hippocampal CA1 subregion. At the same time, through inhibiting phosphorylation of Bcl-2 and the release of Bax from Bcl-2/Bax dimers, SP600125 attenuated Bax translocation to mitochondria and the release of cytochrome c induced by ischemia/reperfusion (I/R). Furthermore, the activation of caspase-3 induced by ischemia/reperfusion was also significantly suppressed by preinfusion of SP600125. Importantly, the same neuropotective effect was showed by administration of SP600125 both before and after ischemia. Thus, our findings imply that SP600125 can inhibit the activation of JNK signaling pathway and induce neuroprotection against ischemia/reperfusion in rat hippocampal CA1 region via suppressing the extrinsic and intrinsic pathways of apoptosis. Taken together, these results indicate that targeting the JNK pathway provides a promising therapeutic approach for ischemic brain injury.