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Sample records for activates mapk signaling

  1. Mitogen-activated protein kinase phosphatase 1 negatively regulates MAPK signaling in mouse hypothalamus.

    PubMed

    Adachi, Koichi; Goto, Motomitsu; Onoue, Takeshi; Tsunekawa, Taku; Shibata, Miyuki; Hagimoto, Shigeru; Ito, Yoshihiro; Banno, Ryoichi; Suga, Hidetaka; Sugimura, Yoshihisa; Oiso, Yutaka; Arima, Hiroshi

    2014-05-21

    Mitogen-activated protein kinase phosphatase 1 (MKP-1) is shown to negatively regulate MAPK signaling in various peripheral tissues as well as the central nervous system such as cortex, striatum and hippocampus. In this study, we examined whether MKP-1 regulates MAPK signaling in the mouse hypothalamus. Intraperitoneal injection of TNFα significantly increased MKP-1 mRNA expression in paraventricular and arcuate nuclei in the hypothalamus. TNFα treatment induced increases in MKP-1 expression at both mRNA and protein levels, accompanied by the inactivation of MAPK signaling in mouse hypothalamic explants. Inhibition of MKP-1 by its inhibitor or siRNA increased MAPK activity in the explants. Our data indicate that MKP-1 negatively regulates MAPK signaling in the mouse hypothalamus.

  2. Calcium signaling and the MAPK cascade are required for sperm activation in Caenorhabditis elegans.

    PubMed

    Liu, Zhiyu; Wang, Bin; He, Ruijun; Zhao, Yanmei; Miao, Long

    2014-02-01

    In nematode, sperm activation (or spermiogenesis), a process in which the symmetric and non-motile spermatids transform into polarized and crawling spermatozoa, is critical for sperm cells to acquire fertilizing competence. SPE-8 dependent and SPE-8 independent pathways function redundantly during sperm activation in both males and hermaphrodites of Caenorhabditis elegans. However, the downstream signaling for both pathways remains unclear. Here we show that calcium signaling and the MAPK cascade are required for both SPE-8 dependent and SPE-8 independent sperm activation, implying that both pathways share common downstream signaling components during sperm activation. We demonstrate that activation of the MAPK cascade is sufficient to activate spermatids derived from either wild-type or spe-8 group mutant males and that activation of the MAPK cascade bypasses the requirement of calcium signal to induce sperm activation, indicating that the MAPK cascade functions downstream of or parallel with the calcium signaling during sperm activation. Interestingly, the persistent activation of MAPK in activated spermatozoa inhibits Major Sperm Protein (MSP)-based cytoskeleton dynamics. We demonstrate that MAPK plays dual roles in promoting pseudopod extension during sperm activation but also blocking the MSP-based, amoeboid motility of the spermatozoa. Thus, though nematode sperm are crawling cells, morphologically distinct from flagellated sperm, and the molecular machinery for motility of amoeboid and flagellated sperm is different, both types of sperm might utilize conserved signaling pathways to modulate sperm maturation.

  3. Canonical and alternative MAPK signaling.

    PubMed

    Pimienta, Genaro; Pascual, Jaime

    2007-11-01

    The archetype of MAPK cascade activation is somewhat challenged by the most recent discovery of unexpected phosphorylation patterns, alternative activation mechanisms and sub-cellular localization, in various members of this protein kinase family. In particular, activation by autophosphorylation pathways has now been described for the three best understood MAPK subgroups: ERK1/2; JNK1/2 and p38 alpha/beta. Also, a form of dosage compensation between homologs has been shown to occur in the case of ERK1/2 and JNK1/2. In this paper we summarize the MAPK activation pathway, with an emphasis on non-canonical examples. We use this information to propose a model for MAPK signal transduction that considers a cross-talk between MAPKs with different activation loop sequence motifs and unique C-terminal extensions. We highlight the occurrence of non-canonical substrate specificity during MAPK auto-activation, in strong connection with MAPK homo- and hetero-dimerization events.

  4. MAPK Cascades in Guard Cell Signal Transduction

    PubMed Central

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M.

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  5. MAPK Cascades in Guard Cell Signal Transduction.

    PubMed

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions.

  6. MAPK Cascades in Guard Cell Signal Transduction.

    PubMed

    Lee, Yuree; Kim, Yun Ju; Kim, Myung-Hee; Kwak, June M

    2016-01-01

    Guard cells form stomata on the epidermis and continuously respond to endogenous and environmental stimuli to fine-tune the gas exchange and transpirational water loss, processes which involve mitogen-activated protein kinase (MAPK) cascades. MAPKs form three-tiered kinase cascades with MAPK kinases and MAPK kinase kinases, by which signals are transduced to the target proteins. MAPK cascade genes are highly conserved in all eukaryotes, and they play crucial roles in myriad developmental and physiological processes. MAPK cascades function during biotic and abiotic stress responses by linking extracellular signals received by receptors to cytosolic events and gene expression. In this review, we highlight recent findings and insights into MAPK-mediated guard cell signaling, including the specificity of MAPK cascades and the remaining questions. PMID:26904052

  7. Inhibitory and activating functions for MAPK Kss1 in the S. cerevisiae filamentous-growth signalling pathway.

    PubMed

    Cook, J G; Bardwell, L; Thorner, J

    1997-11-01

    Mitogen-activated protein kinase (MAPK) cascades are conserved signalling modules that regulate responses to diverse extracellular stimuli, developmental cues and environmental stresses. A MAPK is phosphorylated and activated by a MAPK kinase (MAPKK), which is activated by an upstream protein kinase, such as Raf, Mos or a MAPKK kinase. Ste7, a MAPKK in the yeast Saccharomyces cerevisiae, is required for two developmental pathways: mating and invasive (filamentous) growth. Kss1 and Fus3, the MAPK targets of Ste7, are required for mating, but their role in invasive growth has been unclear. Because no other S. cerevisiae MAPK has been shown to function in invasive growth, it was proposed that Ste7 may have non-MAPK targets. We show instead that Kss1 is the principal target of Ste7 in the invasive-growth response in both haploids and diploids. We demonstrate further that Kss1 in its inactive form is a potent negative regulator of invasive growth. Ste7 acts to relieve this negative regulation by switching Kss1 from an inhibitor to an activator. These results indicate that this MAPK has a physiologically important function in its unactivated state. Comparison of normal and MAPK-deficient cells indicates that nitrogen starvation and activated Ras stimulate filamentous growth through both MAPK-independent and MAPK-dependent means. PMID:9363895

  8. Hog1 mitogen-activated protein kinase (MAPK) interrupts signal transduction between the Kss1 MAPK and the Tec1 transcription factor to maintain pathway specificity.

    PubMed

    Shock, Teresa R; Thompson, James; Yates, John R; Madhani, Hiten D

    2009-04-01

    In Saccharomyces cerevisiae, the mating, filamentous growth (FG), and high-osmolarity glycerol (HOG) mitogen-activated protein kinase (MAPK) signaling pathways share components and yet mediate distinct responses to different extracellular signals. Cross talk is suppressed between the mating and FG pathways because mating signaling induces the destruction of the FG transcription factor Tec1. We show here that HOG pathway activation results in phosphorylation of the FG MAPK, Kss1, and the MAPKK, Ste7. However, FG transcription is not activated because HOG signaling prevents the activation of Tec1. In contrast to the mating pathway, we find that the mechanism involves the inhibition of DNA binding by Tec1 rather than its destruction. We also find that nuclear accumulation of Tec1 is not affected by HOG signaling. Inhibition by Hog1 is apparently indirect since it does not require any of the consensus S/TP MAPK phosphorylation sites on Tec1, its DNA-binding partner Ste12, or the associated regulators Dig1 or Dig2. It also does not require the consensus MAPK sites of the Ste11 activator Ste50, in contrast to a recent proposal for a role for negative feedback in specificity. Our results demonstrate that HOG signaling interrupts the FG pathway signal transduction between the phosphorylation of Kss1 and the activation of DNA binding by Tec1. PMID:19218425

  9. MAPKs in development: insights from Dictyostelium signaling pathways

    PubMed Central

    Hadwiger, Jeffrey A.; Nguyen, Hoai-Nghia

    2011-01-01

    Mitogen activated protein kinases (MAPKs) play important roles in the development of eukaryotic organisms through the regulation of signal transduction pathways stimulated by external signals. MAPK signaling pathways have been associated with the regulation of cell growth, differentiation, and chemotaxis, indicating MAPKs contribute to a diverse set of developmental processes. In most eukaryotes, the diversity of external signals is likely to far exceed the diversity of MAPKs, suggesting that multiple signaling pathways might share MAPKs. Do different signaling pathways converge before MAPK function or can MAPKs maintain signaling specificity through interactions with specific proteins? The genetic and biochemical analysis of MAPK pathways in simple eukaryotes such as Dictyostelium offers opportunities to investigate functional specificity of MAPKs in G protein-mediated signal transduction pathways. This review considers the regulation and specificity of MAPK function in pathways that control Dictyostelium growth and development. PMID:21666837

  10. Hydrogen peroxide signaling mediator in the activation of p38 MAPK in vascular endothelial cells.

    PubMed

    Bretón-Romero, Rosa; Lamas, Santiago

    2013-01-01

    Substantial evidence suggests that a transient increase of hydrogen peroxide (H2O2) behaves as an intracellular messenger able to trigger the activation of different signaling pathways. These include phosphatases, protein kinases, and transcription factors among others; however, most of the studies have been performed using supraphysiological levels of H2O2. Reactive oxygen species (ROS) generation occurs under physiological conditions and different extracellular stimuli including cytokines, growth factors, and shear stress are able to produce both low levels of superoxide anion and H2O2. Here, we explore the redox-dependent activation of key signaling pathways induced by shear stress. We demonstrate that laminar shear stress (LSS) rapidly promotes a transient generation of H2O2 that is necessary for the activation of the stress-activated protein kinase p38 MAPK. We describe p38 MAPK as an early redox sensor in LSS. Our studies show that it is essential for the activation of endothelial nitric oxide synthase, the subsequent nitric oxide generation, and the protection of endothelial function.

  11. IGF-1 protects tubular epithelial cells during injury via activation of ERK/MAPK signaling pathway

    PubMed Central

    Wu, Zengbin; Yu, Yang; Niu, Lei; Fei, Aihua; Pan, Shuming

    2016-01-01

    Injury of renal tubular epithelial cells can induce acute renal failure and obstructive nephropathy. Previous studies have shown that administration of insulin-like growth factor-1 (IGF-1) ameliorates the renal injury in a mouse unilateral ureteral obstruction (UUO) model, whereas the underlying mechanisms are not completely understood. Here, we addressed this question. We found that the administration of IGF-1 significantly reduced the severity of the renal fibrosis in UUO. By analyzing purified renal epithelial cells, we found that IGF-1 significantly reduced the apoptotic cell death of renal epithelial cells, seemingly through upregulation of anti-apoptotic protein Bcl-2, at protein but not mRNA level. Bioinformatics analyses and luciferase-reporter assay showed that miR-429 targeted the 3′-UTR of Bcl-2 mRNA to inhibit its protein translation in renal epithelial cells. Moreover, IGF-1 suppressed miR-429 to increase Bcl-2 in renal epithelial cells to improve survival after UUO. Furthermore, inhibition of ERK/MAPK signaling pathway in renal epithelial cells abolished the suppressive effects of IGF-1 on miR-429 activation, and then the enhanced effects on Bcl-2 in UUO. Thus, our data suggest that IGF-1 may protect renal tubular epithelial cells via activation of ERK/MAPK signaling pathway during renal injury. PMID:27301852

  12. Extracellular ATP activates MAPK and ROS signaling during injury response in the fungus Trichoderma atroviride

    PubMed Central

    Medina-Castellanos, Elizabeth; Esquivel-Naranjo, Edgardo U.; Heil, Martin; Herrera-Estrella, Alfredo

    2014-01-01

    The response to mechanical damage is crucial for the survival of multicellular organisms, enabling their adaptation to hostile environments. Trichoderma atroviride, a filamentous fungus of great importance in the biological control of plant diseases, responds to mechanical damage by activating regenerative processes and asexual reproduction (conidiation). During this response, reactive oxygen species (ROS) are produced by the NADPH oxidase complex. To understand the underlying early signaling events, we evaluated molecules such as extracellular ATP (eATP) and Ca2+ that are known to trigger wound-induced responses in plants and animals. Concretely, we investigated the activation of mitogen-activated protein kinase (MAPK) pathways by eATP, Ca2+, and ROS. Indeed, application of exogenous ATP and Ca2+ triggered conidiation. Furthermore, eATP promoted the Nox1-dependent production of ROS and activated a MAPK pathway. Mutants in the MAPK-encoding genes tmk1 and tmk3 were affected in wound-induced conidiation, and phosphorylation of both Tmk1 and Tmk3 was triggered by eATP. We conclude that in this fungus, eATP acts as a damage-associated molecular pattern (DAMP). Our data indicate the existence of an eATP receptor and suggest that in fungi, eATP triggers pathways that converge to regulate asexual reproduction genes that are required for injury-induced conidiation. By contrast, Ca2+ is more likely to act as a downstream second messenger. The early steps of mechanical damage response in T. atroviride share conserved elements with those known from plants and animals. PMID:25484887

  13. Voltage-gated Na+ Channel Activity Increases Colon Cancer Transcriptional Activity and Invasion Via Persistent MAPK Signaling

    PubMed Central

    House, Carrie D.; Wang, Bi-Dar; Ceniccola, Kristin; Williams, Russell; Simaan, May; Olender, Jacqueline; Patel, Vyomesh; Baptista-Hon, Daniel T.; Annunziata, Christina M.; Silvio Gutkind, J.; Hales, Tim G.; Lee, Norman H.

    2015-01-01

    Functional expression of voltage-gated Na+ channels (VGSCs) has been demonstrated in multiple cancer cell types where channel activity induces invasive activity. The signaling mechanisms by which VGSCs promote oncogenesis remain poorly understood. We explored the signal transduction process critical to VGSC-mediated invasion on the basis of reports linking channel activity to gene expression changes in excitable cells. Coincidentally, many genes transcriptionally regulated by the SCN5A isoform in colon cancer have an over-representation of cis-acting sites for transcription factors phosphorylated by ERK1/2 MAPK. We hypothesized that VGSC activity promotes MAPK activation to induce transcriptional changes in invasion-related genes. Using pharmacological inhibitors/activators and siRNA-mediated gene knockdowns, we correlated channel activity with Rap1-dependent persistent MAPK activation in the SW620 human colon cancer cell line. We further demonstrated that VGSC activity induces downstream changes in invasion-related gene expression via a PKA/ERK/c-JUN/ELK-1/ETS-1 transcriptional pathway. This is the first study illustrating a molecular mechanism linking functional activity of VGSCs to transcriptional activation of invasion-related genes. PMID:26096612

  14. Voltage-gated Na+ Channel Activity Increases Colon Cancer Transcriptional Activity and Invasion Via Persistent MAPK Signaling

    NASA Astrophysics Data System (ADS)

    House, Carrie D.; Wang, Bi-Dar; Ceniccola, Kristin; Williams, Russell; Simaan, May; Olender, Jacqueline; Patel, Vyomesh; Baptista-Hon, Daniel T.; Annunziata, Christina M.; Silvio Gutkind, J.; Hales, Tim G.; Lee, Norman H.

    2015-06-01

    Functional expression of voltage-gated Na+ channels (VGSCs) has been demonstrated in multiple cancer cell types where channel activity induces invasive activity. The signaling mechanisms by which VGSCs promote oncogenesis remain poorly understood. We explored the signal transduction process critical to VGSC-mediated invasion on the basis of reports linking channel activity to gene expression changes in excitable cells. Coincidentally, many genes transcriptionally regulated by the SCN5A isoform in colon cancer have an over-representation of cis-acting sites for transcription factors phosphorylated by ERK1/2 MAPK. We hypothesized that VGSC activity promotes MAPK activation to induce transcriptional changes in invasion-related genes. Using pharmacological inhibitors/activators and siRNA-mediated gene knockdowns, we correlated channel activity with Rap1-dependent persistent MAPK activation in the SW620 human colon cancer cell line. We further demonstrated that VGSC activity induces downstream changes in invasion-related gene expression via a PKA/ERK/c-JUN/ELK-1/ETS-1 transcriptional pathway. This is the first study illustrating a molecular mechanism linking functional activity of VGSCs to transcriptional activation of invasion-related genes.

  15. Neural cell adhesion molecule modulates mesenchymal stromal cell migration via activation of MAPK/ERK signaling.

    PubMed

    Shi, Yu; Xia, Yin-Yan; Wang, Lei; Liu, Rui; Khoo, King-Shung; Feng, Zhi-Wei

    2012-10-15

    Mesenchymal Stromal Cells (MSCs) represent promising tools for cellular therapy owing to their multipotentiality and ability to localize to injured, inflamed sites and tumor. Various approaches to manipulate expression of MSC surface markers, including adhesion molecules and chemokine receptors, have been explored to enhance homing of MSCs. Recently, Neural Cell Adhesion Molecule (NCAM) has been found to be expressed on MSCs yet its function remains largely elusive. Herein, we show that bone marrow-derived MSCs from NCAM deficient mice exhibit defective migratory ability and significantly impaired adipogenic and osteogenic differentiation potential. We further explore the mechanism governing NCAM mediated migration of MSCs by showing the interplay between NCAM and Fibroblast Growth Factor Receptor (FGFR) induces activation of MAPK/ERK signaling, thereby the migration of MSCs. In addition, re-expression of NCAM180, but not NCAM140, could restore the defective MAPK/ERK signaling thereby the migration of NCAM deficient MSCs. Finally, we demonstrate that NCAM180 expression level could be manipulated by pro-inflammatory cytokine Tumor Necrosis Factor (TNF)-α treatment. Overall, our data reveal the vital function of NCAM in MSCs migration and differentiation thus raising the possibility of manipulating NCAM expression to enhance homing and therapeutic potential of MSCs in cellular therapy.

  16. Deoxycholic acid mediates non-canonical EGFR-MAPK activation through the induction of calcium signaling in colon cancer cells.

    PubMed

    Centuori, Sara M; Gomes, Cecil J; Trujillo, Jesse; Borg, Jamie; Brownlee, Joshua; Putnam, Charles W; Martinez, Jesse D

    2016-07-01

    Obesity and a western diet have been linked to high levels of bile acids and the development of colon cancer. Specifically, increased levels of the bile acid deoxycholic acid (DCA), an established tumor promoter, has been shown to correlate with increased development of colorectal adenomas and progression to carcinoma. Herein we investigate the mechanism by which DCA leads to EGFR-MAPK activation, a candidate mechanism by which DCA may promote colorectal tumorigenesis. DCA treated colon cancer cells exhibited strong and prolonged activation of ERK1/2 when compared to EGF treatment alone. We also showed that DCA treatment prevents EGFR degradation as opposed to the canonical EGFR recycling observed with EGF treatment. Moreover, the combination of DCA and EGF treatment displayed synergistic activity, suggesting DCA activates MAPK signaling in a non-canonical manner. Further evaluation showed that DCA treatment increased intracellular calcium levels and CAMKII phosphorylation, and that blocking calcium with BAPTA-AM abrogated MAPK activation induced by DCA, but not by EGF. Finally we showed that DCA-induced CAMKII leads to MAPK activation through the recruitment of c-Src. Taken together, we demonstrated that DCA regulates MAPK activation through calcium signaling, an alternative mechanism not previously recognized in human colon cancer cells. Importantly, this mechanism allows for EGFR to escape degradation and thus achieve a constitutively active state, which may explain its tumor promoting effects.

  17. Deoxycholic acid mediates non-canonical EGFR-MAPK activation through the induction of calcium signaling in colon cancer cells.

    PubMed

    Centuori, Sara M; Gomes, Cecil J; Trujillo, Jesse; Borg, Jamie; Brownlee, Joshua; Putnam, Charles W; Martinez, Jesse D

    2016-07-01

    Obesity and a western diet have been linked to high levels of bile acids and the development of colon cancer. Specifically, increased levels of the bile acid deoxycholic acid (DCA), an established tumor promoter, has been shown to correlate with increased development of colorectal adenomas and progression to carcinoma. Herein we investigate the mechanism by which DCA leads to EGFR-MAPK activation, a candidate mechanism by which DCA may promote colorectal tumorigenesis. DCA treated colon cancer cells exhibited strong and prolonged activation of ERK1/2 when compared to EGF treatment alone. We also showed that DCA treatment prevents EGFR degradation as opposed to the canonical EGFR recycling observed with EGF treatment. Moreover, the combination of DCA and EGF treatment displayed synergistic activity, suggesting DCA activates MAPK signaling in a non-canonical manner. Further evaluation showed that DCA treatment increased intracellular calcium levels and CAMKII phosphorylation, and that blocking calcium with BAPTA-AM abrogated MAPK activation induced by DCA, but not by EGF. Finally we showed that DCA-induced CAMKII leads to MAPK activation through the recruitment of c-Src. Taken together, we demonstrated that DCA regulates MAPK activation through calcium signaling, an alternative mechanism not previously recognized in human colon cancer cells. Importantly, this mechanism allows for EGFR to escape degradation and thus achieve a constitutively active state, which may explain its tumor promoting effects. PMID:27086143

  18. Small Molecule APY606 Displays Extensive Antitumor Activity in Pancreatic Cancer via Impairing Ras-MAPK Signaling

    PubMed Central

    Guo, Na; Liu, Zuojia; Zhao, Wenjing; Wang, Erkang; Wang, Jin

    2016-01-01

    Pancreatic cancer has been found with abnormal expression or mutation in Ras proteins. Oncogenic Ras activation exploits their extensive signaling reach to affect multiple cellular processes, in which the mitogen-activated protein kinase (MAPK) signaling exerts important roles in tumorigenesis. Therapies targeted Ras are thus of major benefit for pancreatic cancer. Although small molecule APY606 has been successfully picked out by virtual drug screening based on Ras target receptor, its in-depth mechanism remains to be elucidated. We herein assessed the antitumor activity of APY606 against human pancreatic cancer Capan-1 and SW1990 cell lines and explored the effect of Ras-MAPK and apoptosis-related signaling pathway on the activity of APY606. APY606 treatment resulted in a dose- and time-dependent inhibition of cancer cell viability. Additionally, APY606 exhibited strong antitumor activity, as evidenced not only by reduction in tumor cell invasion, migration and mitochondrial membrane potential but also by alteration in several apoptotic indexes. Furthermore, APY606 treatment directly inhibited Ras-GTP and the downstream activation of MAPK, which resulted in the down-regulation of anti-apoptotic protein Bcl-2, leading to the up-regulation of mitochondrial apoptosis pathway-related proteins (Bax, cytosolic Cytochrome c and Caspase 3) and of cyclin-dependent kinase 2 and Cyclin A, E. These data suggest that impairing Ras-MAPK signaling is a novel mechanism of action for APY606 during therapeutic intervention in pancreatic cancer. PMID:27223122

  19. [Studies on cell signaling immunomodulated murine peritoneal suppressor macrophages: LPS and PMA mediate the activation of RAF-1, MAPK p44 and MAPK p42 and p38 MAPK].

    PubMed

    Chang, Z L; Lin, M Q; Wang, M Z; Yao, Z

    1997-03-01

    Monocytes-macrophages which serve as host immune cells to kill pathogens can often be "activated" after exposing to viruses, bacteria, cytokines as well as chemical substances, However, it is paradoxical that highly activated macrophages can be induced to become the suppressor ones by live microbes, microbial products, tumor, and autoimmune disease, although the mechanism remains unknown. Our previous experimental studies have shown that immuno-suppressor activities of suppressor macrophages on T, B and NK cells can be prevented by the treatment with LPS or supernatant in vitro from mitogen-stimulated lymphocytes, while, at the same time, the tumoricidal activities of those macrophages can be kept or even enhanced following the same treatment. This phenomenon was then termed as "immune modulation" For the understanding of its mechanism, we are now undertaking signal transduction in modulated macrophages. Since mitogen-activated protein kinase (MAPK) is an integration point of different signal transduction pathways, its cascade and regulation of activation are being investigated extensively by the assay of electrophoresis mobility shift. Recent results suggested that interaction of ligand-receptor triggers protein tyrosine kinase(PTK) activation leading to Ras-GTP binding with Raf-1 to phosphorylate MAPK kinase (MAPKK), the specific activator of MAPK. It is reported that PKC-alpha can directly phosphorylate or activate Raf-1 in NIH3 T3 cells. Raf-1 (74 KDa), with an intrinsic serine (Ser)-threonine (The) kinase activity, becomes hyperphosphorylated after activation which can be followed by gel mobility shift test. It has also been shown that a variety of extracellular factors stimulate a pair of MAPK p44 and MAPK p42 of MAPK family members. A significant property of activation of ERK 1 and ERK 2 is the requirement for the phosphorylation of both Thr-183 and Tyr-185 (at TEY motif) within in its protein kinase subdomain VIII. More recently, two other MAPK subtypes, p

  20. CpG- and LPS-activated MAPK signaling in in vitro cultured salmon (Salmo salar) mononuclear phagocytes.

    PubMed

    Iliev, Dimitar B; Hansen, Tom; Jørgensen, Sven Martin; Krasnov, Aleksei; Jørgensen, Jorunn B

    2013-10-01

    The Mitogen-activated protein kinases (MAPK) are involved in transmitting intracellular signals downstream of diverse cell surface receptors and mediate the response to ligands such as growth factors, hormones and cytokines. In addition, MAPK are critically involved in the innate immune response to pathogen-derived substances, commonly referred to as pathogen-associated molecular patterns (PAMPs), such as bacterial lipopolysaccharide (LPS) and bacterial DNA rich in CpG dinucleotides. Currently, a great deal of knowledge is available about the involvement of MAPK in the innate immune response to PAMPs in mammals; however, little is known about the role of the different MAPK classes in the immune response to PAMPs in lower vertebrates. In the current study, p38 phosphorylation was induced by CpG oligonucleotides (ODNs) and LPS in primary salmon mononuclear phagocytes. Pre-treatment of the cells with a p38 inhibitor (SB203580) blocked the PAMP-induced p38 activity and suppressed the upregulation of most of the CpG- and LPS-induced transcripts highlighting the role of this kinase in the salmon innate immune response to PAMPs. In contrast to p38, the phosphorylation of extracellular signal-regulated kinase (ERK), a MAPK involved primarily in response to mitogens, was high in resting cells and, surprisingly, incubation with both CpG and control ODNs downregulated the phospho-ERK levels independently of p38 activation. The basal phospho-ERK level and the CpG-inducible p38 phosphorylation were greatly influenced by the length of in vitro incubation. The basal phospho-ERK level increased gradually throughout a 5-day culture period and was PI3K-dependent as demonstrated by its sensitivity to Wortmannin suggesting it is influenced by growth factors. Overall these data indicate that both basal and PAMP-induced activity of MAPKs might be greatly influenced by the differentiation status of salmon mononuclear phagocytes.

  1. Fasting potentiates the anticancer activity of tyrosine kinase inhibitors by strengthening MAPK signaling inhibition.

    PubMed

    Caffa, Irene; D'Agostino, Vito; Damonte, Patrizia; Soncini, Debora; Cea, Michele; Monacelli, Fiammetta; Odetti, Patrizio; Ballestrero, Alberto; Provenzani, Alessandro; Longo, Valter D; Nencioni, Alessio

    2015-05-20

    Tyrosine kinase inhibitors (TKIs) are now the mainstay of treatment in many types of cancer. However, their benefit is frequently short-lived, mandating the search for safe potentiation strategies. Cycles of fasting enhance the activity of chemo-radiotherapy in preclinical cancer models and dietary approaches based on fasting are currently explored in clinical trials. Whether combining fasting with TKIs is going to be potentially beneficial remains unknown. Here we report that starvation conditions increase the ability of commonly administered TKIs, including erlotinib, gefitinib, lapatinib, crizotinib and regorafenib, to block cancer cell growth, to inhibit the mitogen-activated protein kinase (MAPK) signaling pathway and to strengthen E2F-dependent transcription inhibition. In cancer xenografts models, both TKIs and cycles of fasting slowed tumor growth, but, when combined, these interventions were significantly more effective than either type of treatment alone. In conclusion, cycles of fasting or of specifically designed fasting-mimicking diets should be evaluated in clinical studies as a means to potentiate the activity of TKIs in clinical use.

  2. N-methyl-D-aspartate (NMDA) and the regulation of mitogen-activated protein kinase (MAPK) signaling pathways: a revolving neurochemical axis for therapeutic intervention?

    PubMed

    Haddad, John J

    2005-11-01

    Excitatory synaptic transmission in the central nervous system (CNS) is mediated by the release of glutamate from presynaptic terminals onto postsynaptic channels gated by N-methyl-D-aspartate (NMDA) and non-NMDA (AMPA and KA) receptors. Extracellular signals control diverse neuronal functions and are responsible for mediating activity-dependent changes in synaptic strength and neuronal survival. Influx of extracellular calcium ([Ca(2+)](e)) through the NMDA receptor (NMDAR) is required for neuronal activity to change the strength of many synapses. At the molecular level, the NMDAR interacts with signaling modules, which, like the mitogen-activated protein kinase (MAPK) superfamily, transduce excitatory signals across neurons. Recent burgeoning evidence points to the fact that MAPKs play a crucial role in regulating the neurochemistry of NMDARs, their physiologic and biochemical/biophysical properties, and their potential role in pathophysiology. It is the purpose of this review to discuss: (i) the MAPKs and their role in a plethora of cellular functions; (ii) the role of MAPKs in regulating the biochemistry and physiology of NMDA receptors; (iii) the kinetics of MAPK-NMDA interactions and their biologic and neurochemical properties; (iv) how cellular signaling pathways, related cofactors and intracellular conditions affect NMDA-MAPK interactions and (v) the role of NMDA-MAPK pathways in pathophysiology and the evolution of disease conditions. Given the versatility of the NMDA-MAPK interactions, the NMDA-MAPK axis will likely form a neurochemical target for therapeutic interventions.

  3. YES oncogenic activity is specified by its SH4 domain and regulates RAS/MAPK signaling in colon carcinoma cells.

    PubMed

    Dubois, Fanny; Leroy, Cédric; Simon, Valérie; Benistant, Christine; Roche, Serge

    2015-01-01

    Members of the SRC family of tyrosine kinases (SFK) display important functions in human cancer, but their specific role in tumorigenesis remains unclear. We previously demonstrated that YES regulates a unique oncogenic signaling important for colorectal cancer (CRC) progression that is not shared with SRC. Here, we addressed the underlying mechanism involved in this process. We show that YES oncogenic signaling relies on palmitoylation of its SH4 domain that controls YES localization in cholesterol-enriched membrane micro-domains. Specifically, deletion of the palmitoylation site compromised YES transforming activity, while addition of a palmitoylation site in the SH4 domain of SRC was sufficient for SRC to restore the transforming properties of cells in which YES had been silenced. Subsequently, SILAC phosphoproteomic analysis revealed that micro-domain-associated cell adhesive components and receptor tyrosine kinases are major YES substrates. YES also phosphorylates upstream regulators of RAS/MAPK signaling, including EGFR, SHC and SHP2, which were not targeted by SRC due to the absence of palmitoylation. Accordingly, EGFR-induced MAPK activity was attenuated by YES down-regulation, while increased RAS activity significantly restored cell transformation that was lost upon YES silencing. Collectively, these results uncover a critical role for the SH4 domain in the specification of SFK oncogenic activity and a selective role for YES in the induction of RAS/MAPK signaling in CRC cells.

  4. Interferon-β counter-regulates its own pro-apoptotic action by activating p38 MAPK signalling in human SH-SY5Y neuroblastoma cells.

    PubMed

    Dedoni, Simona; Olianas, Maria C; Onali, Pierluigi

    2014-10-01

    Type I interferons (IFNs) induce apoptosis of neuroblastoma cells, but the molecular mechanisms regulating this event have not been completely elucidated. Here, we investigated the role of p38 mitogen activated protein kinase (MAPK) activity, a key regulator of apoptosis and a known modulator of IFN-induced responses in non-neuronal cells. We show that in SH-SY5Y human neuroblastoma cells IFN-β induced a delayed and sustained increase of p38 MAPK activity through a novel mechanism involving the sequential activation of Janus kinase-signal transducer and activator of transcription-1 signalling, enhanced expression of the NADPH oxidase catalytic subunit gp91(phox), increased reactive oxygen species production and stimulation of the MAPK kinase kinase transforming growth factor-β-activated kinase 1. Either blockade of p38 MAPK by the second generation inhibitors BIRB0796 and VX745 or siRNA knockdown of p38α MAPK enhanced IFN-β-induced apoptosis of neuroblastoma cells. Exposure to IFN-β increased the phosphorylation of the small heat shock protein HSP27 at Ser15, Ser78 and Ser82 with a time course similar to p38 MAPK activation and this response was suppressed by either p38α MAPK depletion or pharmacological inhibition of p38 MAPK and MAPK-activated protein kinase 2 (MK2). Either silencing of HSP27 expression by siRNA or MK2 inhibition potentiated IFN-β-induced apoptotic death. These results indicate that IFN-β-induced apoptosis of human SH-SY5Y neuroblastoma cells is associated with a long-lasting up-regulation of p38 MAPK activity, stimulation of MK2 and phosphorylation of the pro-survival protein HSP27. Moreover, the data show that inhibition of p38 MAPK signalling potentiates the anti-neuroblastoma activity of the cytokine, indicating that this pathway mediates a counter-regulatory response.

  5. Phosphoproteomic analysis of protein kinase C signaling in Saccharomyces cerevisiae reveals Slt2 mitogen-activated protein kinase (MAPK)-dependent phosphorylation of eisosome core components.

    PubMed

    Mascaraque, Victoria; Hernáez, María Luisa; Jiménez-Sánchez, María; Hansen, Rasmus; Gil, Concha; Martín, Humberto; Cid, Víctor J; Molina, María

    2013-03-01

    The cell wall integrity (CWI) pathway of the model organism Saccharomyces cerevisiae has been thoroughly studied as a paradigm of the mitogen-activated protein kinase (MAPK) pathway. It consists of a classic MAPK module comprising the Bck1 MAPK kinase kinase, two redundant MAPK kinases (Mkk1 and Mkk2), and the Slt2 MAPK. This module is activated under a variety of stimuli related to cell wall homeostasis by Pkc1, the only member of the protein kinase C family in budding yeast. Quantitative phosphoproteomics based on stable isotope labeling of amino acids in cell culture is a powerful tool for globally studying protein phosphorylation. Here we report an analysis of the yeast phosphoproteome upon overexpression of a PKC1 hyperactive allele that specifically activates CWI MAPK signaling in the absence of external stimuli. We found 82 phosphopeptides originating from 43 proteins that showed enhanced phosphorylation in these conditions. The MAPK S/T-P target motif was significantly overrepresented in these phosphopeptides. Hyperphosphorylated proteins provide putative novel targets of the Pkc1-cell wall integrity pathway involved in diverse functions such as the control of gene expression, protein synthesis, cytoskeleton maintenance, DNA repair, and metabolism. Remarkably, five components of the plasma-membrane-associated protein complex known as eisosomes were found among the up-regulated proteins. We show here that Pkc1-induced phosphorylation of the eisosome core components Pil1 and Lsp1 was not exerted directly by Pkc1, but involved signaling through the Slt2 MAPK module.

  6. Thrombosis recanalization by paeoniflorin through the upregulation of urokinase-type plasminogen activator via the MAPK signaling pathway

    PubMed Central

    YE, SONGSHAN; MAO, BINGYU; YANG, LEI; FU, WEIYUN; HOU, JUNRAN

    2016-01-01

    Paeoniflorin, the major component of Paeonia lactiflora pall, has previously been reported to prevent thrombosis. Plasminogen activator urokinase (uPA) is a serine protease that markedly facilitates normal thrombosis resolution. Paeoniflorin and uPA have been linked to the mitogen-activated protein kinase (MAPK) signaling pathway. In the current study, the influence of paeoniflorin on the expression of uPA was investigated and the underlying regulatory mechanism was preliminarily determined. The prothrombotic state of the model animals treated with paeoniflorin were assessed by enzyme-linked immunosorbent assay (ELISA). Additionally, the cytotoxicity of paeoniflorin on human umbilical vein endothelial cell (HUVEC) cultures was estimated using a methyl thiazolyl tetrazolium assay and the possible pathways involved in the interaction between paeoniflorin and uPA were evaluated using western blot analysis. The ELISA results demonstrated that the levels of 6-keto prostaglandin F1a, fibronectin and uPA were significantly upregulated by treatment with paeoniflorin compared with control (P<0.05). By contrast, the expression of fibrinogen, D-dimer and thromboxane B2 were inhibited. With an increase in the concentration of paeoniflorin the cell viability of HUVECs decreased gradually. The results of western blot analysis demonstrated that paeoniflorin increased the phosphorylation of MAPK 14 (p38) and MAPK 8 (JNK). The present study demonstrated that paeoniflorin has the potential to improve the prethrombotic state and recanalize thrombosis by increasing the expression of uPA, which may be mediated via regulation of the p38 and JNK MAPK signaling pathways. However, this treatment effect was dependent on the concentration of paeoniflorin used, an unsuitable concentration of the agent would result in a negative effect on the anti-thrombosis pathways. PMID:27082639

  7. Activation of p21ras/MAPK signal transduction molecules decreases with age in mitogen-stimulated T cells from rats.

    PubMed

    Pahlavani, M A; Harris, M D; Richardson, A

    1998-04-10

    Signal transduction is ubiquitously involved in the initiation of physiological signals that lead to growth and proliferation of cells. The signaling cascade mediated by the mitogen-activated protein kinase (MAPK) is considered essential for T cell growth and function. Therefore, it was of interest to determine the influence of age on the induction of MAPK in mitogen-activated T cells. T cells from young (4-6 months) and old (24-26 months) rats responded to concanavalin A (Con A) stimulation by increasing MAPK, c-jun amino terminal kinase (JNK), and p21ras activities. The time course of induction of MAPK/JNK and p21ras activities was similar in T cells isolated from young and old rats. The induction of JNK activity did not change significantly with age; however, the induction of MAPK and p21ras activities was significantly less (50 to 65%) in T cells from old rats than in T cells from young rats. Although the relative protein levels of p42 and p44 MAPK did not change with age, the proportion of the phosphorylated p44 MAPK decreased with age. In addition, it was found that the in vitro kinase activities of the T cell receptor-associated protein tyrosine kinase Lck (p56Lck) and ZAP-70 but not Fyn (p59Fyn) were lower in T cells from old rats than in T cells from young rats. The decline in activities of these signaling molecules with age was not associated with changes in their corresponding protein levels. Thus, our results demonstrate that aging alters the activation of the signal transduction cascade that leads to T cell activation.

  8. Abrogation of NF-κB signaling in human neutrophils induces neutrophil survival through sustained p38-MAPK activation.

    PubMed

    Langereis, Jeroen D; Raaijmakers, Hanneke A J A; Ulfman, Laurien H; Koenderman, Leo

    2010-10-01

    NF-κB, an important transcription factor in the regulation of cellular inflammation, is one of the prime targets for novel anti-inflammatory therapeutics. Nowadays, anti-inflammatory therapies rely mostly on steroids, which among other effects, inhibit NF-κB activity. However, steroids have only limited efficacy in the treatment on neutrophil-driven diseases, such as COPD. Human neutrophils play an important role in the pathogenesis of COPD, and clearance of these cells by apoptosis is an effective pathway for resolution of inflammation. In this study, we tested the hypothesis that modulation of the NF-κB pathway in human neutrophils affects survival. Importantly, the pharmacological NF-κB inhibitor Bay 11-7082 inhibited NF-κB signaling in human neutrophils as expected. However, we found that complete inhibition of NF-κB activity with 10 μM Bay 11-7082 prolonged neutrophil survival significantly, which was not observed with inhibitors for other signaling pathways. Bay 11-7082-induced neutrophil survival was dependent on p38-MAPK kinase activity, as the p38 kinase activity inhibitor SB203580 abrogated this response completely. Bay 11-7082 induced rapid and sustained p38 activation that correlated with inhibited NF-κB signaling and prolonged neutrophil survival. The precise role of NF-κB in regulation of p38-MAPK activation remains to be established. Under these conditions of survival, the stability of Bcl-xL but not Mcl-1 was enhanced. Although inhibition of NF-κB leads to down-regulation of inflammatory genes in many cell types, our results illustrate that interference with basal NF-κB signaling in neutrophils as a drug target should be used with caution.

  9. MAPK Signaling and ERK1/2 bistability in Asthma

    PubMed Central

    Alam, Rafeul; Gorska, Magdalena M.

    2010-01-01

    Mitogen-activated protein kinases (MAPK) integrate signals from numerous receptors and translate these signals into cell functions. MAPKs are critical for immune cell metabolism, migration, production of pro-inflammatory mediators, survival, and differentiation. We provide a concise review of the involvement of MAPK in important cells of the immune system. Certain cell functions e.g. production of pro-inflammatory mediators resolve quickly and may require a transient MAPK activation, other processes such as cell differentiation and long-term survival may require persistent MAPK signal. The persistent MAPK signal is frequently a consequence of positive feedback loops or double negative feedback loops which perpetuate the signal after removal of an external cell stimulus. This self-perpetuated activation of a signaling circuit is a manifestation of its bistability. Bistable systems can exist in “on” and “off” states and both states are stable. We have demonstrated the existence of self-perpetuated activation mechanism for ERK1/2 in bronchial epithelial cells. This sustained activation of ERK1/2 supports long-term survival of these cells and primes them for cytokine transcription. ERK1/2 bistability arises from repetitive stimulation of the cell. The repeated stimulation (e.g. repeated viral infection or repeated allergen exposure) seems to be a common theme in asthma and other chronic illnesses. We thus hypothesize that the self-perpetuated ERK1/2 signal plays an important role in the pathogenesis of asthma. PMID:21121982

  10. ROCK activity affects IL-1-induced signaling possibly through MKK4 and p38 MAPK in Caco-2 cells.

    PubMed

    Banerjee, Sayantan; McGee, Dennis W

    2016-09-01

    Elevated levels of interleukin-1 (IL-1) accompany inflammatory bowel disease. IL-1-stimulated intestinal epithelial cells can secrete potent chemokines like CXCL8 to exacerbate inflammation. Previously, we found that inhibiting the Rho-associated kinase (ROCK) could inhibit IL-1- or TNF-α-induced CXCL8 secretion by the Caco-2 colonic epithelial cell line. This ROCK inhibition did not affect IκBα phosphorylation and degradation, but suppressed the phosphorylation of c-Jun N-terminal kinase (JNK). Therefore, ROCK must play an important role in epithelial cell CXCL8 responses through an effect on the JNK signaling pathway. Here, we extend these studies by showing that inhibiting ROCK suppressed the IL-1-induced phosphorylation of MKK4, a known activator of JNK, but not MKK7. Yet, ROCK inhibition had no significant effect on the IL-1-induced phosphorylation of extracellular-signal-regulated kinase (ERK) 1/2. Inhibiting ROCK also suppressed the phosphorylation of p38 MAPK after IL-1 stimulation, but this inhibition had no significant effect on the stability of CXCL8 messenger RNA (mRNA) after IL-1 stimulation. These results suggest that ROCK may be important in IL-1-induced signaling through MKK4 to JNK and the activation of p38 MAPK. Finally, inhibiting ROCK in IL-1 and TNF-α co-stimulated Caco-2 cells also resulted in a significant suppression of CXCL8 secretion and mRNA levels suggesting that inhibiting ROCK may be a mechanism to inhibit the overall response of epithelial cells to both cytokines. These studies indicate a novel signaling event, which could provide a target for suppressing intestinal epithelial cells (IEC) chemokine responses involved in mucosal inflammation.

  11. ROCK activity affects IL-1-induced signaling possibly through MKK4 and p38 MAPK in Caco-2 cells.

    PubMed

    Banerjee, Sayantan; McGee, Dennis W

    2016-09-01

    Elevated levels of interleukin-1 (IL-1) accompany inflammatory bowel disease. IL-1-stimulated intestinal epithelial cells can secrete potent chemokines like CXCL8 to exacerbate inflammation. Previously, we found that inhibiting the Rho-associated kinase (ROCK) could inhibit IL-1- or TNF-α-induced CXCL8 secretion by the Caco-2 colonic epithelial cell line. This ROCK inhibition did not affect IκBα phosphorylation and degradation, but suppressed the phosphorylation of c-Jun N-terminal kinase (JNK). Therefore, ROCK must play an important role in epithelial cell CXCL8 responses through an effect on the JNK signaling pathway. Here, we extend these studies by showing that inhibiting ROCK suppressed the IL-1-induced phosphorylation of MKK4, a known activator of JNK, but not MKK7. Yet, ROCK inhibition had no significant effect on the IL-1-induced phosphorylation of extracellular-signal-regulated kinase (ERK) 1/2. Inhibiting ROCK also suppressed the phosphorylation of p38 MAPK after IL-1 stimulation, but this inhibition had no significant effect on the stability of CXCL8 messenger RNA (mRNA) after IL-1 stimulation. These results suggest that ROCK may be important in IL-1-induced signaling through MKK4 to JNK and the activation of p38 MAPK. Finally, inhibiting ROCK in IL-1 and TNF-α co-stimulated Caco-2 cells also resulted in a significant suppression of CXCL8 secretion and mRNA levels suggesting that inhibiting ROCK may be a mechanism to inhibit the overall response of epithelial cells to both cytokines. These studies indicate a novel signaling event, which could provide a target for suppressing intestinal epithelial cells (IEC) chemokine responses involved in mucosal inflammation. PMID:27173611

  12. Titanium dioxide nanoparticles stimulate sea urchin immune cell phagocytic activity involving TLR/p38 MAPK-mediated signalling pathway.

    PubMed

    Pinsino, Annalisa; Russo, Roberta; Bonaventura, Rosa; Brunelli, Andrea; Marcomini, Antonio; Matranga, Valeria

    2015-09-28

    Titanium dioxide nanoparticles (TiO2NPs) are one of the most widespread-engineered particles in use for drug delivery, cosmetics, and electronics. However, TiO2NP safety is still an open issue, even for ethical reasons. In this work, we investigated the sea urchin Paracentrotus lividus immune cell model as a proxy to humans, to elucidate a potential pathway that can be involved in the persistent TiO2NP-immune cell interaction in vivo. Morphology, phagocytic ability, changes in activation/inactivation of a few mitogen-activated protein kinases (p38 MAPK, ERK), variations of other key proteins triggering immune response (Toll-like receptor 4-like, Heat shock protein 70, Interleukin-6) and modifications in the expression of related immune response genes were investigated. Our findings indicate that TiO2NPs influence the signal transduction downstream targets of p38 MAPK without eliciting an inflammatory response or other harmful effects on biological functions. We strongly recommend sea urchin immune cells as a new powerful model for nano-safety/nano-toxicity investigations without the ethical normative issue.

  13. Titanium dioxide nanoparticles stimulate sea urchin immune cell phagocytic activity involving TLR/p38 MAPK-mediated signalling pathway

    PubMed Central

    Pinsino, Annalisa; Russo, Roberta; Bonaventura, Rosa; Brunelli, Andrea; Marcomini, Antonio; Matranga, Valeria

    2015-01-01

    Titanium dioxide nanoparticles (TiO2NPs) are one of the most widespread-engineered particles in use for drug delivery, cosmetics, and electronics. However, TiO2NP safety is still an open issue, even for ethical reasons. In this work, we investigated the sea urchin Paracentrotus lividus immune cell model as a proxy to humans, to elucidate a potential pathway that can be involved in the persistent TiO2NP-immune cell interaction in vivo. Morphology, phagocytic ability, changes in activation/inactivation of a few mitogen-activated protein kinases (p38 MAPK, ERK), variations of other key proteins triggering immune response (Toll-like receptor 4-like, Heat shock protein 70, Interleukin-6) and modifications in the expression of related immune response genes were investigated. Our findings indicate that TiO2NPs influence the signal transduction downstream targets of p38 MAPK without eliciting an inflammatory response or other harmful effects on biological functions. We strongly recommend sea urchin immune cells as a new powerful model for nano-safety/nano-toxicity investigations without the ethical normative issue. PMID:26412401

  14. Mobile phone electromagnetic radiation activates MAPK signaling and regulates viability in Drosophila.

    PubMed

    Lee, Kyu-Sun; Choi, Jong-Soon; Hong, Sae-Yong; Son, Tae-Ho; Yu, Kweon

    2008-07-01

    Mobile phones are widely used in the modern world. However, biological effects of electromagnetic radiation produced by mobile phones are largely unknown. In this report, we show biological effects of the mobile phone 835 MHz electromagnetic field (EMF) in the Drosophila model system. When flies were exposed to the specific absorption rate (SAR) 1.6 W/kg, which is the proposed exposure limit by the American National Standards Institute (ANSI), more than 90% of the flies were viable even after the 30 h exposure. However, in the SAR 4.0 W/kg strong EMF exposure, viability dropped from the 12 h exposure. These EMF exposures triggered stress response and increased the production of reactive oxygen species. The EMF exposures also activated extracellular signal regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling, but not p38 kinase signaling. Interestingly, SAR 1.6 W/kg activated mainly ERK signaling and expression of an anti-apoptotic gene, whereas SAR 4.0 W/kg strongly activated JNK signaling and expression of apoptotic genes. In addition, SAR 4.0 W/kg amplified the number of apoptotic cells in the fly brain. These findings demonstrate that the exposure limit on electromagnetic radiation proposed by ANSI triggered ERK-survival signaling but the strong electromagnetic radiation activated JNK-apoptotic signaling in Drosophila.

  15. Tumor suppressor gene ING3 induces cardiomyocyte hypertrophy via inhibition of AMPK and activation of p38 MAPK signaling.

    PubMed

    Wang, Jiaojiao; Liu, Zhiping; Feng, Xiaojun; Gao, Si; Xu, Suowen; Liu, Peiqing

    2014-11-15

    Cardiac hypertrophy, an adaptive growth process that occurs in response to various pathophysiological stimuli, constitutes an important risk factor for the development of heart failure. However, the molecular mechanisms that regulate this cardiac growth response are not completely understood. Here we revealed that ING3 (inhibitor of growth family, member 3), a type II tumor suppressor, plays a critical role in the regulation of cardiac hypertrophy. ING3 expression was present in relatively high abundance in the heart, and was prominently upregulated in hypertrophic agonists angiotensin II (Ang II), phenylephrine (PE), or isoproterenol (ISO)-stimulated cardiomyocytes and in hearts of rat undergoing abdominal aortic constriction (AAC) surgery. In cardiomyocytes, overexpression of ING3 caused an increase in ANP, BNP and β-MHC mRNA levels and cell surface area, while depletion of ING3 attenuated PE-induced cardiomyocyte hypertrophy. Mechanistically, we have demonstrated that overexpression of ING3 could inactivate the AMPK and activate the canonical p38 MAPK signaling. Remarkably, AMPK agonist AICAR or p38 MAPK inhibitor SB203580 abrogated ING3-induced hypertrophic response in cardiomyocytes. In summary, our data disclose a novel role of ING3 as an inducer of pathological cardiac hypertrophy, suggesting that silencing of ING3 may be explored as a potential therapeutic target in preventing cardiac hypertrophy.

  16. Integration analysis of MKK and MAPK family members highlights potential MAPK signaling modules in cotton.

    PubMed

    Zhang, Xueying; Xu, Xiaoyang; Yu, Yujia; Chen, Chuan; Wang, Jing; Cai, Caiping; Guo, Wangzhen

    2016-01-01

    Mitogen-activated protein kinase (MAPK) cascades play a crucial role in plant growth and development, as well as their biotic and abiotic stress responses. As a nodal point of the MAPK cascade, the MKK gene family has not been systematically studied in cotton. Here, we identified 11 putative MKK genes in the Gossypium raimondii genome. Phylogenetic analysis showed that the MKKs were supported by architectures of conserved protein motifs. Expression patterns of MKKs under hormone treatments or abiotic stresses revealed their diverse functions in stress responses. Based on a yeast two hybrid, a total of 63 interactive pairs of MKKs and MAPKs were identified in cotton. Among these, 40 interactive pairs were newly identified compared to that reported previously in Arabidopsis. Integration analysis of the interaction network and expression patterns of MKK and MAPK family members revealed 13 potential MAPK signaling modules that are involved in the complicated cross-talk between hormones and abiotic stresses. Taken together, our data enhance the understanding of the evolution and function of MAPK cascades in cotton, and lay the foundation for the improvement of various defense responses that use MAPK signaling modules in the future. PMID:27417377

  17. Integration analysis of MKK and MAPK family members highlights potential MAPK signaling modules in cotton

    PubMed Central

    Zhang, Xueying; Xu, Xiaoyang; Yu, Yujia; Chen, Chuan; Wang, Jing; Cai, Caiping; Guo, Wangzhen

    2016-01-01

    Mitogen-activated protein kinase (MAPK) cascades play a crucial role in plant growth and development, as well as their biotic and abiotic stress responses. As a nodal point of the MAPK cascade, the MKK gene family has not been systematically studied in cotton. Here, we identified 11 putative MKK genes in the Gossypium raimondii genome. Phylogenetic analysis showed that the MKKs were supported by architectures of conserved protein motifs. Expression patterns of MKKs under hormone treatments or abiotic stresses revealed their diverse functions in stress responses. Based on a yeast two hybrid, a total of 63 interactive pairs of MKKs and MAPKs were identified in cotton. Among these, 40 interactive pairs were newly identified compared to that reported previously in Arabidopsis. Integration analysis of the interaction network and expression patterns of MKK and MAPK family members revealed 13 potential MAPK signaling modules that are involved in the complicated cross-talk between hormones and abiotic stresses. Taken together, our data enhance the understanding of the evolution and function of MAPK cascades in cotton, and lay the foundation for the improvement of various defense responses that use MAPK signaling modules in the future. PMID:27417377

  18. Specificity of MAP kinase signaling in yeast differentiation involves transient versus sustained MAPK activation.

    PubMed

    Sabbagh, W; Flatauer, L J; Bardwell, A J; Bardwell, L

    2001-09-01

    Signals transmitted by common components often elicit distinct (yet appropriate) outcomes. In yeast, two developmental options-mating and invasive growth-are both regulated by the same MAP kinase cascade. Specificity has been thought to result from specialized roles for the two MAP kinases, Kss1 and Fus3, and because Fus3 prevents Kss1 from gaining access to the mating pathway. Kss1 has been thought to participate in mating only when Fus3 is absent. Instead, we show that Kss1 is rapidly phosphorylated and potently activated by mating pheromone in wild-type cells, and that this is required for normal pheromone-induced gene expression. Signal identity is apparently maintained because active Fus3 limits the extent of Kss1 activation, thereby preventing inappropriate signal crossover. PMID:11583629

  19. Noncanonical Wnt-4 signaling enhances bone regeneration of mesenchymal stem cells in craniofacial defects through activation of p38 MAPK.

    PubMed

    Chang, Jia; Sonoyama, Wataru; Wang, Zhuo; Jin, Qiming; Zhang, Chengfei; Krebsbach, Paul H; Giannobile, William; Shi, Songtao; Wang, Cun-Yu

    2007-10-19

    Mesenchymal stem cells (MSCs) are multipotent cells that can be differentiated into osteoblasts and provide an excellent cell source for bone regeneration and repair. Recently, the canonical Wnt/beta-catenin signaling pathway has been found to play a critical role in skeletal development and osteogenesis, implying that Wnts can be utilized to improve de novo bone formation mediated by MSCs. However, it is unknown whether noncanonical Wnt signaling regulates osteogenic differentiation. Here, we find that Wnt-4 enhanced in vitro osteogenic differentiation of MSCs isolated from human adult craniofacial tissues and promoted bone formation in vivo. Whereas Wnt-4 did not stabilize beta-catenin, it activated p38 MAPK in a novel noncanonical signaling pathway. The activation of p38 was dependent on Axin and was required for the enhancement of MSC differentiation by Wnt-4. Moreover, using two different models of craniofacial bone injury, we found that MSCs genetically engineered to express Wnt-4 enhanced osteogenesis and improved the repair of craniofacial defects in vivo. Taken together, our results reveal that noncanonical Wnt signaling could also play a role in osteogenic differentiation. Wnt-4 may have a potential use in improving bone regeneration and repair of craniofacial defects.

  20. Thrombosis recanalization by paeoniflorin through the upregulation of urokinase‑type plasminogen activator via the MAPK signaling pathway.

    PubMed

    Ye, Songshan; Mao, Bingyu; Yang, Lei; Fu, Weiyun; Hou, Junran

    2016-06-01

    Paeoniflorin, the major component of Paeonia lactiflora pall, has previously been reported to prevent thrombosis. Plasminogen activator urokinase (uPA) is a serine protease that markedly facilitates normal thrombosis resolution. Paeoniflorin and uPA have been linked to the mitogen‑activated protein kinase (MAPK) signaling pathway. In the current study, the influence of paeoniflorin on the expression of uPA was investigated and the underlying regulatory mechanism was preliminarily determined. The prothrombotic state of the model animals treated with paeoniflorin were assessed by enzyme‑linked immunosorbent assay (ELISA). Additionally, the cytotoxicity of paeoniflorin on human umbilical vein endothelial cell (HUVEC) cultures was estimated using a methyl thiazolyl tetrazolium assay and the possible pathways involved in the interaction between paeoniflorin and uPA were evaluated using western blot analysis. The ELISA results demonstrated that the levels of 6‑keto prostaglandin F1a, fibronectin and uPA were significantly upregulated by treatment with paeoniflorin compared with control (P<0.05). By contrast, the expression of fibrinogen, D‑dimer and thromboxane B2 were inhibited. With an increase in the concentration of paeoniflorin the cell viability of HUVECs decreased gradually. The results of western blot analysis demonstrated that paeoniflorin increased the phosphorylation of MAPK 14 (p38) and MAPK 8 (JNK). The present study demonstrated that paeoniflorin has the potential to improve the prethrombotic state and recanalize thrombosis by increasing the expression of uPA, which may be mediated via regulation of the p38 and JNK MAPK signaling pathways. However, this treatment effect was dependent on the concentration of paeoniflorin used, an unsuitable concentration of the agent would result in a negative effect on the anti‑thrombosis pathways. PMID:27082639

  1. p38 MAPK Signaling in Pemphigus: Implications for Skin Autoimmunity

    PubMed Central

    Mavropoulos, Athanasios; Orfanidou, Timoklia; Liaskos, Christos; Smyk, Daniel S.; Spyrou, Vassiliki; Sakkas, Lazaros I.; Rigopoulou, Eirini I.; Bogdanos, Dimitrios P.

    2013-01-01

    p38 mitogen activated protein kinase (p38 MAPK) signaling plays a major role in the modulation of immune-mediated inflammatory responses and therefore has been linked with several autoimmune diseases. The extent of the involvement of p38 MAPK in the pathogenesis of autoimmune blistering diseases has started to emerge, but whether it pays a critical role is a matter of debate. The activity of p38 MAPK has been studied in great detail during the loss of keratinocyte cell-cell adhesions and the development of pemphigus vulgaris (PV) and pemphigus foliaceus (PF). These diseases are characterised by autoantibodies targeting desmogleins (Dsg). Whether autoantibody-antigen interactions can trigger signaling pathways (such as p38 MAPK) that are tightly linked to the secretion of inflammatory mediators which may perpetuate inflammation and tissue damage in pemphigus remains unclear. Yet, the ability of p38 MAPK inhibitors to block activation of the proapoptotic proteinase caspase-3 suggests that the induction of apoptosis may be a consequence of p38 MAPK activation during acantholysis in PV. This review discusses the current evidence for the role of p38 MAPK in the pathogenesis of pemphigus. We will also present data relating to the targeting of these cascades as a means of therapeutic intervention. PMID:23936634

  2. Lipopolysaccharide signals activation of tumor necrosis factor biosynthesis through the ras/raf-1/MEK/MAPK pathway.

    PubMed Central

    Geppert, T. D.; Whitehurst, C. E.; Thompson, P.; Beutler, B.

    1994-01-01

    BACKGROUND: Lipopolysaccharide (LPS) is known to activate macrophages, causing the release of toxic cytokines that may provoke inflammation and shock. One of the most important and best studied of these cytokines is tumor necrosis factor (TNF). Details of the signaling pathway leading to TNF biosynthesis remain unclear. The pathway is branched in the sense that TNF gene transcription and TNF mRNA translation are both strongly stimulated by LPS. Recent evidence has indicated that MAP kinase homologs become phosphorylated in LPS-stimulated cells, suggesting their possible involvement in signal transduction. We sought to test this hypothesis. MATERIALS AND METHODS: Measurements of LPS-induced MEK and ERK2 activity were undertaken in LPS-sensitive and LPS-insensitive cells. Transfection studies, in which dominant inhibitors of ras and raf-1 were used to block signaling to the level of MAP kinase, were carried out in order to judge whether the TNF gene transcription and TNF mRNA translation are modulated through this pathway. RESULTS: In RAW 264.7 mouse macrophages, both ERK2 and MEK1 activity are induced by LPS treatment. In the same cell line, dominant negative inhibitors of ras and raf-1 block LPS-induced activation of the TNF promoter, as well as derepression of the translational blockade normally imposed by the TNF 3'-untranslated region. A constitutively active form of raf-1 (raf-BXB) was found to augment, but not replace, the LPS signal. In LPS-insensitive cells (RAW 264.7 x NIH 3T3 fusion hybrid cells and primary macrophages derived from C3H/HeJ mice), ERK2 activity was found to be refractory to induction by LPS. CONCLUSIONS: The ras/raf-1/MEK/MAPK pathway is chiefly responsible for transduction of the LPS signal to the level of the TNF gene and mRNA. raf and raf-1 lie upstream from (or actually represent) the physical branchpoints of the transcriptional and translation activation signals generated by LPS. The lesions that prevent LPS signaling in macrophages

  3. PBX3 promotes migration and invasion of colorectal cancer cells via activation of MAPK/ERK signaling pathway

    PubMed Central

    Han, Hai-Bo; Gu, Jin; Ji, Deng-Bo; Li, Zhao-Wei; Zhang, Yuan; Zhao, Wei; Wang, Li-Min; Zhang, Zhi-Qian

    2014-01-01

    AIM: To investigate the role of pre-B-cell leukemia homeobox (PBX)3 in migration and invasion of colorectal cancer (CRC) cells. METHODS: We detected PBX3 expression in five cell lines and surgical specimens from 111 patients with CRC using real-time reverse transcription-polymerase chain reaction. We forced expression of PBX3 in low metastatic HT-29 and SW480 cells and knocked down expression of PBX3 in highly metastatic LOVO and HCT-8 cells. Wound healing and Boyden chamber assays were used to detect cell migration and invasion after altered expression of PBX3. Western blot was performed to detect the change of signaling molecule ERK1/2 following PBX3 overexpression. RESULTS: High level of PBX3 expression was correlated with the invasive potential of CRC cells, and significantly associated with lymph node invasion (P = 0.02), distant metastasis (P = 0.04), advanced TNM stage (P = 0.03) and poor overall survival of patients (P < 0.05). Ectopic expression of PBX3 in low metastatic cells was shown to promote migration and invasion, while inhibited PBX3 expression in highly metastatic cells suppressed migration and invasion. Furthermore, upregulation of phosphorylated extracellular signal-regulated kinase (ERK)1/2 was found to be one of the targeted molecules responsible for PBX3-induced CRC cell migration and invasion. CONCLUSION: PBX3 induces invasion and metastasis of CRC cells partially through activation of the MAPK/ERK signaling pathway. PMID:25561793

  4. Biphasic activation of PI3K/Akt and MAPK/Erk1/2 signaling pathways in bovine herpesvirus type 1 infection of MDBK cells.

    PubMed

    Zhu, Liqian; Ding, Xiuyan; Zhu, Xiaofang; Meng, Songshu; Wang, Jianye; Zhou, Hong; Duan, Qiangde; Tao, Jie; Schifferli, Dieter M; Zhu, Guoqiang

    2011-04-14

    Many viruses have been known to control key cellular signaling pathways to facilitate the virus infection. The possible involvement of signaling pathways in bovine herpesvirus type 1 (BoHV-1) infection is unknown. This study indicated that infection of MDBK cells with BoHV-1 induced an early-stage transient and a late-stage sustained activation of both phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen activated protein kinases/extracellular signal-regulated kinase 1/2 (MAPK/Erk1/2) signaling pathways. Analysis with the stimulation of UV-irradiated virus indicated that the virus binding and/or entry process was enough to trigger the early phase activations, while the late phase activations were viral protein expression dependent. Biphasic activation of both pathways was suppressed by the selective inhibitor, Ly294002 for PI3K and U0126 for MAPK kinase (MEK1/2), respectively. Furthermore, treatment of MDBK cells with Ly294002 caused a 1.5-log reduction in virus titer, while U0126 had little effect on the virus production. In addition, the inhibition effect of Ly294002 mainly occurred at the post-entry stage of the virus replication cycle. This revealed for the first time that BoHV-1 actively induced both PI3K/Akt and MAPK/Erk1/2 signaling pathways, and the activation of PI3K was important for fully efficient replication, especially for the post-entry stage.

  5. Abnormal activity of the MAPK- and cAMP-associated signaling pathways in frontal cortical areas in postmortem brain in schizophrenia.

    PubMed

    Funk, Adam J; McCullumsmith, Robert E; Haroutunian, Vahram; Meador-Woodruff, James H

    2012-03-01

    Recent evidence suggests that schizophrenia may result from alterations of integration of signaling mediated by multiple neurotransmitter systems. Abnormalities of associated intracellular signaling pathways may contribute to the pathophysiology of schizophrenia. Proteins and phospho-proteins comprising mitogen activated protein kinase (MAPK) and 3'-5'-cyclic adenosine monophosphate (cAMP)-associated signaling pathways may be abnormally expressed in the anterior cingulate (ACC) and dorsolateral prefrontal cortex (DLPFC) in schizophrenia. Using western blot analysis we examined proteins of the MAPK- and cAMP-associated pathways in these two brain regions. Postmortem samples were used from a well-characterized collection of elderly patients with schizophrenia (ACC=36, DLPFC=35) and a comparison (ACC=33, DLPFC=31) group. Near-infrared intensity of IR-dye labeled secondary antisera bound to targeted proteins of the MAPK- and cAMP-associated signaling pathways was measured using LiCor Odyssey imaging system. We found decreased expression of Rap2, JNK1, JNK2, PSD-95, and decreased phosphorylation of JNK1/2 at T183/Y185 and PSD-95 at S295 in the ACC in schizophrenia. In the DLPFC, we found increased expression of Rack1, Fyn, Cdk5, and increased phosphorylation of PSD-95 at S295 and NR2B at Y1336. MAPK- and cAMP-associated molecules constitute ubiquitous intracellular signaling pathways that integrate extracellular stimuli, modify receptor expression and function, and regulate cell survival and neuroplasticity. These data suggest abnormal activity of the MAPK- and cAMP-associated pathways in frontal cortical areas in schizophrenia. These alterations may underlie the hypothesized hypoglutamatergic function in this illness. Together with previous findings, these data suggest that abnormalities of intracellular signaling pathways may contribute to the pathophysiology of schizophrenia. PMID:22048463

  6. Glycosylation defects activate filamentous growth Kss1 MAPK and inhibit osmoregulatory Hog1 MAPK.

    PubMed

    Yang, Hui-Yu; Tatebayashi, Kazuo; Yamamoto, Katsuyoshi; Saito, Haruo

    2009-05-20

    The yeast filamentous growth (FG) MAP kinase (MAPK) pathway is activated under poor nutritional conditions. We found that the FG-specific Kss1 MAPK is activated by a combination of an O-glycosylation defect caused by disruption of the gene encoding the protein O-mannosyltransferase Pmt4, and an N-glycosylation defect induced by tunicamycin. The O-glycosylated membrane proteins Msb2 and Opy2 are both essential for activating the FG MAPK pathway, but only defective glycosylation of Msb2 activates the FG MAPK pathway. Although the osmoregulatory HOG (high osmolarity glycerol) MAPK pathway and the FG MAPK pathway share almost the entire upstream signalling machinery, osmostress activates only the HOG-specific Hog1 MAPK. Conversely, we now show that glycosylation defects activate only Kss1, while activated Kss1 and the Ptp2 tyrosine phosphatase inhibit Hog1. In the absence of Kss1 or Ptp2, however, glycosylation defects activate Hog1. When Hog1 is activated by glycosylation defects in ptp2 mutant, Kss1 activation is suppressed by Hog1. Thus, the reciprocal inhibitory loop between Kss1 and Hog1 allows only one or the other of these MAPKs to be stably activated under various stress conditions. PMID:19369942

  7. Signal inhibition by a dynamically regulated pool of monophosphorylated MAPK

    PubMed Central

    Nagiec, Michal J.; McCarter, Patrick C.; Kelley, Joshua B.; Dixit, Gauri; Elston, Timothy C.; Dohlman, Henrik G.

    2015-01-01

    Protein kinases regulate a broad array of cellular processes and do so through the phosphorylation of one or more sites within a given substrate. Many protein kinases are themselves regulated through multisite phosphorylation, and the addition or removal of phosphates can occur in a sequential (processive) or a stepwise (distributive) manner. Here we measured the relative abundance of the monophosphorylated and dual-phosphorylated forms of Fus3, a member of the mitogen-activated protein kinase (MAPK) family in yeast. We found that upon activation with pheromone, a substantial proportion of Fus3 accumulates in the monophosphorylated state. Introduction of an additional copy of Fus3 lacking either phosphorylation site leads to dampened signaling. Conversely, cells lacking the dual-specificity phosphatase (msg5Δ) or that are deficient in docking to the MAPK-scaffold (Ste5ND) accumulate a greater proportion of dual-phosphorylated Fus3. The double mutant exhibits a synergistic, or “synthetic,” supersensitivity to pheromone. Finally, we present a predictive computational model that combines MAPK scaffold and phosphatase activities and is sufficient to account for the observed MAPK profiles. These results indicate that the monophosphorylated and dual-phosphorylated forms of the MAPK act in opposition to one another. Moreover, they reveal a new mechanism by which the MAPK scaffold acts dynamically to regulate signaling. PMID:26179917

  8. Dynamic signaling in the Hog1 MAPK pathway relies on high basal signal transduction.

    PubMed

    Macia, Javier; Regot, Sergi; Peeters, Tom; Conde, Núria; Solé, Ricard; Posas, Francesc

    2009-01-01

    Appropriate regulation of the Hog1 mitogen-activated protein kinase (MAPK) pathway is essential for cells to survive osmotic stress. Here, we show that the two sensing mechanisms upstream of Hog1 display different signaling properties. The Sho1 branch is an inducible nonbasal system, whereas the Sln1 branch shows high basal signaling that is restricted by a MAPK-mediated feedback mechanism. A two-dimensional mathematical model of the Snl1 branch, including high basal signaling and a Hog1-regulated negative feedback, shows that a system with basal signaling exhibits higher efficiency, with faster response times and higher sensitivity to variations in external signals, than would systems without basal signaling. Analysis of two other yeast MAPK pathways, the Fus3 and Kss1 signaling pathways, indicates that high intrinsic basal signaling may be a general property of MAPK pathways allowing rapid and sensitive responses to environmental changes. PMID:19318625

  9. The Conserved MAPK Site in E(spl)-M8, an Effector of Drosophila Notch Signaling, Controls Repressor Activity during Eye Development

    PubMed Central

    Bandyopadhyay, Mohna; Bishop, Clifton P.

    2016-01-01

    The specification of patterned R8 photoreceptors at the onset of eye development depends on timely inhibition of Atonal (Ato) by the Enhancer of split (E(spl) repressors. Repression of Ato by E(spl)-M8 requires the kinase CK2 and is inhibited by the phosphatase PP2A. The region targeted by CK2 harbors additional conserved Ser residues, raising the prospect of regulation via multi-site phosphorylation. Here we investigate one such motif that meets the consensus for modification by MAPK, a well-known effector of Epidermal Growth Factor Receptor (EGFR) signaling. Our studies reveal an important role for the predicted MAPK site of M8 during R8 birth. Ala/Asp mutations reveal that the CK2 and MAPK sites ensure that M8 repression of Ato and the R8 fate occurs in a timely manner and at a specific stage (stage-2/3) of the morphogenetic furrow (MF). M8 repression of Ato is mitigated by halved EGFR dosage, and this effect requires an intact MAPK site. Accordingly, variants with a phosphomimetic Asp at the MAPK site exhibit earlier (inappropriate) activity against Ato even at stage-1 of the MF, where a positive feedback-loop is necessary to raise Ato levels to a threshold sufficient for the R8 fate. Analysis of deletion variants reveals that both kinase sites (CK2 and MAPK) contribute to ‘cis’-inhibition of M8. This key regulation by CK2 and MAPK is bypassed by the E(spl)D mutation encoding the truncated protein M8*, which potently inhibits Ato at stage-1 of R8 birth. We also provide evidence that PP2A likely targets the MAPK site. Thus multi-site phosphorylation controls timely onset of M8 repressor activity in the eye, a regulation that appears to be dispensable in the bristle. The high conservation of the CK2 and MAPK sites in the insect E(spl) proteins M7, M5 and Mγ, and their mammalian homologue HES6, suggest that this mode of regulation may enable E(spl)/HES proteins to orchestrate repression by distinct tissue-specific mechanisms, and is likely to have broader

  10. Targeting MAPK Signaling in Age-Related Macular Degeneration

    PubMed Central

    Kyosseva, Svetlana V.

    2016-01-01

    Age-related macular degeneration (AMD) is a major cause of irreversible blindness affecting elderly people in the world. AMD is a complex multifactorial disease associated with demographic, genetics, and environmental risk factors. It is well established that oxidative stress, inflammation, and apoptosis play critical roles in the pathogenesis of AMD. The mitogen-activated protein kinase (MAPK) signaling pathways are activated by diverse extracellular stimuli, including growth factors, mitogens, hormones, cytokines, and different cellular stressors such as oxidative stress. They regulate cell proliferation, differentiation, survival, and apoptosis. This review addresses the novel findings from human and animal studies on the relationship of MAPK signaling with AMD. The use of specific MAPK inhibitors may represent a potential therapeutic target for the treatment of this debilitating eye disease. PMID:27385915

  11. γ-Enolase C-terminal peptide promotes cell survival and neurite outgrowth by activation of the PI3K/Akt and MAPK/ERK signalling pathways.

    PubMed

    Hafner, Anja; Obermajer, Nataša; Kos, Janko

    2012-04-15

    γ-Enolase, a glycolytic enzyme, is expressed specifically in neurons. It exerts neurotrophic activity and has been suggested to regulate growth, differentiation, survival and regeneration of neurons. In the present study, we investigated the involvement of γ-enolase in PI3K (phosphoinositide 3-kinase)/Akt and MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) signalling, the two pathways triggered predominantly by neurotrophic factors. Whereas the PI3K/Akt pathway, rather than the MAPK/ERK pathway, is involved in γ-enolase-enhanced cell survival, γ-enolase-stimulated neurite outgrowth requires both pathways, i.e. the activation of both PI3K and ERK1/2, leading to subsequent expression of the growth-cone-specific protein GAP-43 (growth-associated protein of 43 kDa). MEK (MAPK/ERK kinase) and PI3K inhibition blocked or attenuated the neurite outgrowth associated with dynamic remodelling of the actin-based cytoskeleton. We show that γ-enolase-mediated PI3K activation regulates RhoA kinase, a key regulator of actin cytoskeleton organization. Moreover, the inhibition of RhoA downstream effector ROCK (Rho-associated kinase) results in enhanced γ-enolase-induced neurite outgrowth, accompanied by actin polymerization and its redistribution to growth cones. Our results show that γ-enolase controls neuronal survival, differentiation and neurite regeneration by activating the PI3K/Akt and MAPK/ERK signalling pathways, resulting in downstream regulation of the molecular and cellular processes of cytoskeleton reorganization and cell remodelling, activation of transcriptional factors and regulation of the cell cycle.

  12. Transforming growth factor-beta1-induced activation of the Raf-MEK-MAPK signaling pathway in rat lung fibroblasts via a PKC-dependent mechanism.

    PubMed

    Axmann, A; Seidel, D; Reimann, T; Hempel, U; Wenzel, K W

    1998-08-19

    In fibroblasts transforming growth factor-beta1 (TGF-beta1) regulates cell proliferation and turnover of macromolecular components of the extracellular matrix. Here, intracellular signaling events in growth-inhibited embryonic rat lung fibroblasts (RFL-6) upon stimulation with TGF-beta1 were investigated. TGF-beta1 rapidly induced the activation of c-Raf-1, MEK-1, and MAPK p42 and p44. The activation of this pathway by TGF-beta1 did not depend on autocrine platelet-derived growth factor (PDGF) or basic fibroblast growth factor (bFGF). Inhibition of the binding of growth factors to their tyrosine kinase receptors did not affect MAPK activation by TGF-beta1. Ras activation by TGF-beta1 was significantly lower compared to the activation by PDGF or bFGF. The intracellular transduction of the TGF-beta1 signal was completely suppressed by depletion or inhibition of protein kinase C (PKC). It is shown that calcium-dependent isoforms of PKC are required for MAPK activation by TGF-beta1. PMID:9712718

  13. Lipopolysaccharide enhances bradykinin-induced signal transduction via activation of Ras/Raf/MEK/MAPK in canine tracheal smooth muscle cells

    PubMed Central

    Luo, Shue-Fen; Wang, Chuan-Chwan; Chiu, Chi-Tso; Chien, Chin-Sung; Hsiao, Li-Der; Lin, Chien-Huang; Yang, Chuen-Mao

    2000-01-01

    Bacterial lipopolysaccharide (LPS) was found to induce inflammatory responses and to enhance bronchial hyperreactivity to several contractile agonists. However, the implication of LPS in the pathogenesis of bronchial hyperreactivity was not completely understood. Therefore, in this study, we investigated the effect of LPS on mitogen-activated protein kinase (MAPK) activation associated with potentiation of bradykinin (BK)-induced inositol phosphates (IPs) accumulation and Ca2+ mobilization in canine cultured tracheal smooth muscle cells (TSMCs).LPS stimulated phosphorylation of p42/p44 MAPK in a time- and concentration-dependent manner using a Western blot analysis against a specific phosphorylated form of MAPK antibody. Maximal stimulation of the p42 and p44 MAPK isoforms occurred after 7 min-incubation and the maximal effect was achieved with 100 μg ml−1 LPS.Pretreatment of TSMCs with LPS potentiated BK-induced IPs accumulation and Ca2+ mobilization. However, there was no effect on the IPs response induced by endothelin-1, 5-hydroxytryptamine, and carbachol. In addition, pretreatment with PDGF-BB enhanced BK-induced IPs response.These enhancements by LPS and PDGF-BB might be due to an increase in BK B2 receptor density (Bmax) in TSMCs, characterized by competitive inhibition of [3H]-BK binding using B1 and B2 receptor-selective reagents.The enhancing effects of LPS and PDGF-BB were attenuated by PD98059, an inhibitor of MAPK kinase (MEK), suggesting that the effect of LPS may share a common signalling pathway with PDGF-BB in TSMCs.Furthermore, overexpression of dominant negative mutants, H-Ras-15A and Raf-N4, significantly suppressed p42/p44 MAPK activation induced by LPS and PDGF-BB, indicating that Ras and Raf may be required for activation of these kinases.These results suggest that the augmentation of BK-induced responses produced by LPS might be, at least in part, mediated through activation of Ras/Raf/MEK/MAPK pathway in TSMCs. PMID:10952668

  14. Ginsenoside-Rp1 inhibits platelet activation and thrombus formation via impaired glycoprotein VI signalling pathway, tyrosine phosphorylation and MAPK activation

    PubMed Central

    Endale, M; Lee, WM; Kamruzzaman, SM; Kim, SD; Park, JY; Park, MH; Park, TY; Park, HJ; Cho, JY; Rhee, MH

    2012-01-01

    BACKGROUND AND PURPOSE Ginsenosides are the main constituents for the pharmacological effects of Panax ginseng. Such effects of ginsenosides including cardioprotective and anti-platelet activities have shown stability and bioavailability limitations. However, information on the anti-platelet activity of ginsenoside-Rp1 (G-Rp1), a stable derivative of ginsenoside-Rg3, is scarce. We examined the ability of G-Rp1 to modulate agonist-induced platelet activation. EXPERIMENTAL APPROACH G-Rp1 in vitro and ex vivo effects on agonist-induced platelet-aggregation, granule-secretion, [Ca2+]i mobilization, integrin-αIIbβ3 activation were examined. Vasodilator-stimulated phosphoprotein (VASP) and MAPK expressions and levels of tyrosine phosphorylation of the glycoprotein VI (GPVI) signalling pathway components were also studied. G-Rp1 effects on arteriovenous shunt thrombus formation in rats or tail bleeding time and ex vivo coagulation time in mice were determined. KEY RESULT G-Rp1 markedly inhibited platelet aggregation induced by collagen, thrombin or ADP. While G-Rp1 elevated cAMP levels, it dose-dependently suppressed collagen-induced ATP-release, thromboxane secretion, p-selectin expression, [Ca2+]i mobilization and αIIbβ3 activation and attenuated p38MAPK and ERK2 activation. Furthermore, G-Rp1 inhibited tyrosine phosphorylation of multiple components (Fyn, Lyn, Syk, LAT, PI3K and PLCγ2) of the GPVI signalling pathway. G-Rp1 inhibited in vivo thrombus formation and ex vivo platelet aggregation and ATP secretion without affecting tail bleeding time and coagulation time, respectively. CONCLUSION AND IMPLICATIONS G-Rp1 inhibits collagen-induced platelet activation and thrombus formation through modulation of early GPVI signalling events, and this effect involves VASP stimulation, and ERK2 and p38-MAPK inhibition. These data suggest that G-Rp1 may have therapeutic potential for the treatment of cardiovascular diseases involving aberrant platelet activation. PMID

  15. Dietary influence on MAPK-signaling pathways and risk of colon and rectal cancer

    PubMed Central

    Slattery, Martha L.; Lundgreen, Abbie; Wolff, Roger K.

    2014-01-01

    Mitogen-activated protein kinase (MAPK) pathways regulate cellular functions including cell proliferation, differentiation, migration, and apoptosis. Associations between genes in the DUSP, ERK1/2, JNK, and p38 MAPK-signaling pathways and dietary factors associated with growth factors, inflammation, and oxidative stress and risk of colon and rectal cancer were evaluated. Data include colon cases (n=1555) and controls (n=1956) and rectal cases (n=754) and controls (n=959). Statistically significant interactions were observed for the MAPK-signaling pathways after adjustment for multiple comparisons. DUSP genes interacted with carbohydrates, mutagen index, calories, calcium, vitamin D, lycopene, dietary fats, folic acid, and selenium. MAPK1, MAPK3, MAPK1 and RAF1 within the ERK1/2 MAPK-signaling pathway interacted with dietary fats and cruciferous vegetables. Within the JNK MAPK-signaling pathway, interactions between MAP3K7 and protein, vitamin C, iron, folic acid, carbohydrates, and cruciferous vegetables; MAP3K10 and folic acid; MAP3K9 and lutein/zeaxanthin; MAPK8 and calcium; MAP3K3 and calcium and lutein; MAP3K1 and cruciferous vegetables. Interaction within the p38-signaling pathway included: MAPK14 with calories, carbohydrates saturated fat, selenium, vitamin C; MAP3K2 and carbohydrates, and folic acid. These data suggest that dietary factors involved in inflammation and oxidative stress interact with MAPK-signaling genes to alter risk of colorectal cancer. PMID:23859041

  16. Anti-Inflammatory Effects of α-Galactosylceramide Analogs in Activated Microglia: Involvement of the p38 MAPK Signaling Pathway

    PubMed Central

    Chung, Young Sun; Park, Seung Bum; Kim, Hee-Sun

    2014-01-01

    Microglial activation plays a pivotal role in the development and progression of neurodegenerative diseases. Thus, anti-inflammatory agents that control microglial activation can serve as potential therapeutic agents for neurodegenerative diseases. Here, we designed and synthesized α-galactosylceramide (α-GalCer) analogs to exert anti-inflammatory effects in activated microglia. We performed biological evaluations of 25 α-GalCer analogs and observed an interesting preliminary structure-activity relationship in their inhibitory influence on NO release and TNF-α production in LPS-stimulated BV2 microglial cells. After identification of 4d and 4e as hit compounds, we further investigated the underlying mechanism of their anti-inflammatory effects using RT-PCR analysis. We confirmed that 4d and 4e regulate the expression of iNOS, COX-2, IL-1β, and IL-6 at the mRNA level and the expression of TNF-α at the post-transcriptional level. In addition, both 4d and 4e inhibited LPS-induced DNA binding activities of NF-κB and AP-1 and phosphorylation of p38 MAPK without affecting other MAP kinases. When we examined the anti-inflammatory effect of a p38 MAPK-specific inhibitor, SB203580, on microglial activation, we observed an identical inhibitory pattern as that of 4d and 4e, not only on NO and TNF-α production but also on the DNA binding activities of NF-κB and AP-1. Taken together, these results suggest that p38 MAPK plays an important role in the anti-inflammatory effects of 4d and 4e via the modulation of NF-κB and AP-1 activities. PMID:24523867

  17. DARPP-32 is required for MAPK/ERK signaling in thyroid cells.

    PubMed

    Chocarro-Calvo, Ana; Zaballos, Miguel A; Santisteban, Pilar; García-Jiménez, Custodia

    2012-03-01

    Modulation of MAPK signaling duration by cAMP defines its physiological output by driving cells toward proliferation or differentiation. Understanding how the kinetics of MAPK signaling are integrated with other cellular signals is a key issue in development and cancer. Here we show that dopamine and cAMP-regulated neuronal phosphoprotein, 32 kDa (DARPP-32), a protein required for thyroid cell differentiation, determines whether MAPK/ERK activation is sustained or transient. Serum, a stimulus that activates MAPK signaling and does not independently increase DARPP-32 levels results in transient activation of the MAPK pathway. By contrast, TSH + (IGF-I) activate MAPK signaling but also independently increase DARPP-32 levels. Our results are consistent with a model in which maintenance of DARPP-32 expression by TSH + IGF-I leads to sustained MAPK signaling. Moreover, the sensitivity of MAPK/ERK signaling in thyroid cells is lost when de novo DARPP-32 expression is blocked by small interfering RNA. Because both DARPP-32 levels and function as inhibitor of protein phosphatase 1, a key inhibitor of MAPK kinase activity, are governed by cAMP/protein kinase A, the results may explain why in thyroid cells cAMP signaling downstream from TSH controls the duration of MAPK pathway activity. Thus, fine-tuning of DARPP-32 levels leads to changes in the kinetics or sensitivity of MAPK/ERK signaling. Given the implications of MAPK signaling in thyroid cancer and the loss of DARPP-32 in tumor and transformed thyroid cells, DARPP-32 may represent a key therapeutic target. PMID:22301787

  18. Contribution of S6K1/MAPK Signaling Pathways in the Response to Oxidative Stress: Activation of RSK and MSK by Hydrogen Peroxide

    PubMed Central

    Schneider, Taiane; Bonan, Carla Denise; Bartrons, Ramon; Ventura, Francesc; Rodrigues de Oliveira, Jarbas; Rosa, Jose Luis

    2013-01-01

    Cells respond to different kind of stress through the coordinated activation of signaling pathways such as MAPK or p53. To find which molecular mechanisms are involved, we need to understand their cell adaptation. The ribosomal protein, S6 kinase 1 (S6K1), is a common downstream target of signaling by hormonal or nutritional stress. Here, we investigated the initial contribution of S6K1/MAPK signaling pathways in the cell response to oxidative stress produced by hydrogen peroxide (H2O2). To analyze S6K1 activation, we used the commercial anti-phospho-Thr389-S6K1 antibody most frequently mentioned in the bibliography. We found that this antibody detected an 80-90 kDa protein that was rapidly phosphorylated in response to H2O2 in several human cells. Unexpectedly, this phosphorylation was insensitive to both mTOR and PI3K inhibitors, and knock-down experiments showed that this protein was not S6K1. RSK and MSK proteins were candidate targets of this phosphorylation. We demonstrated that H2O2 stimulated phosphorylation of RSK and MSK kinases at residues that are homologous to Thr389 in S6K1. This phosphorylation required the activity of either p38 or ERK MAP kinases. Kinase assays showed activation of RSK and MSK by H2O2. Experiments with mouse embryonic fibroblasts from p38 animals’ knockout confirmed these observations. Altogether, these findings show that the S6K1 signaling pathway is not activated under these conditions, clarify previous observations probably misinterpreted by non-specific detection of proteins RSK and MSK by the anti-phospho-Thr389-S6K1 antibody, and demonstrate the specific activation of MAPK signaling pathways through ERK/p38/RSK/MSK by H2O2. PMID:24058693

  19. Activation of transient receptor potential vanilloid 4 induces apoptosis in hippocampus through downregulating PI3K/Akt and upregulating p38 MAPK signaling pathways

    PubMed Central

    Jie, P; Hong, Z; Tian, Y; Li, Y; Lin, L; Zhou, L; Du, Y; Chen, L; Chen, L

    2015-01-01

    Transient receptor potential vanilloid 4 (TRPV4) is a calcium-permeable cation channel that is sensitive to cell swelling, arachidonic acid and its metabolites, epoxyeicosatrienoic acids, which are associated with cerebral ischemia. The activation of TRPV4 induces cytotoxicity in many types of cells, accompanied by an increase in the intracellular free calcium concentration. TRPV4 activation modulates the mitogen-activated protein kinase (MAPK) and phosphatidyl inositol 3 kinase (PI3K)/ protein kinase B (Akt) signaling pathways that regulate cell death and survival. Herein, we examined TRPV4-induced neuronal apoptosis by intracerebroventricular (ICV) injection of a TRPV4 agonist (GSK1016790A) and assessed its involvement in cerebral ischemic injury. ICV injection of GSK1016790A dose-dependently induced apoptosis in the mouse hippocampi (GSK-injected mice). The protein level of phosphorylated p38 MAPK (p-p38 MAPK) was markedly increased and that of phosphorylated c-Jun N-terminal protein kinase (p-JNK) was virtually unchanged. TRPV4 activation also decreased Bcl-2/Bax protein ratio and increased the cleaved caspase-3 protein level, and these effects were blocked by a PI3K agonist and a p38 MAPK antagonist, but were unaffected by a JNK antagonist. ICV injection of the TRPV4 antagonist HC-067047 reduced brain infarction after reperfusion for 48 h in mice with middle cerebral artery occlusion (MCAO). In addition, HC-067047 treatment attenuated the decrease in the phosphorylated Akt protein level and the increase in p-p38 MAPK protein level at 48 h after MCAO, while the increase in p-JNK protein level remained unchanged. Finally, the decreased Bcl-2/Bax protein ratio and the increased cleaved caspase-3 protein level at 48 h after MCAO were markedly attenuated by HC-067047. We conclude that activation of TRPV4 induces apoptosis by downregulating PI3K/Akt and upregulating p38 MAPK signaling pathways, which is involved in cerebral ischemic injury. PMID:26043075

  20. MAPK-Activated Protein Kinases (MKs): Novel Insights and Challenges.

    PubMed

    Gaestel, Matthias

    2015-01-01

    Downstream of MAPKs, such as classical/atypical ERKs and p38 MAPKs, but not of JNKs, signaling is often mediated by protein kinases which are phosphorylated and activated by MAPKs and, therefore, designated MAPK-activated protein kinases (MAPKAPKs). Recently, novel insights into the specificity of the assembly of MAPK/MAPKAPK hetero-dimeric protein kinase signaling complexes have been gained. In addition, new functional aspects of MKs have been described and established functions have been challenged. This short review will summarize recent developments including the linear motif (LM) in MKs, the ERK-independent activation of RSK, the RSK-independent effects of some RSK-inhibitors and the challenged role of MK5/PRAK in tumor suppression. PMID:26779481

  1. Total flavonoids of Hedyotis diffusa Willd inhibit inflammatory responses in LPS-activated macrophages via suppression of the NF-κB and MAPK signaling pathways

    PubMed Central

    CHEN, YUNLONG; LIN, YANYAN; LI, YACHAN; LI, CANDONG

    2016-01-01

    Nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) signaling pathways play a central role in inflammatory responses. Total flavonoids of Hedyotis diffusa Willd (TFHDW) are active compounds derived from Hedyotis diffusa Willd, which has been long used in Chinese traditional medicine for the treatment of various inflammatory diseases, including ulcerative colitis and bronchitis; however, the precise mechanisms underlying the effects of TFHDW are largely unknown. In the present study, the anti-inflammatory effect of TFHDW was evaluated and the underlying molecular mechanisms were investigated in an in vitro inflammatory model comprising lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. The results indicated that TFHDW inhibited the inflammatory response as it significantly reduced the LPS-induced expression of pro-inflammatory nitric oxide, tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1β in a concentration-dependent manner, without causing cytotoxicity. In addition, the mRNA expression of inducible nitric oxide synthase, TNF-α, IL-6 and IL-1β was suppressed by treatment with TFHDW in LPS-stimulated RAW 264.7 cells. Moreover, TFHDW treatment significantly inhibited the LPS-induced activation of NF-κB via the suppression of inhibitor of κB (IκB) phosphorylation, and reduced the phosphorylation of MAPK signaling molecules (p38, c-Jun N-terminal protein kinase and extracellular signal-regulated kinase 1/2), which resulted in the inhibition of cytokine expression. These findings suggest that TFHDW exerted anti-inflammatory activity via suppression of the NF-κB and MAPK signaling pathways. PMID:26998046

  2. p38 MAPK Signaling in Osteoblast Differentiation

    PubMed Central

    Rodríguez-Carballo, Eddie; Gámez, Beatriz; Ventura, Francesc

    2016-01-01

    The skeleton is a highly dynamic tissue whose structure relies on the balance between bone deposition and resorption. This equilibrium, which depends on osteoblast and osteoclast functions, is controlled by multiple factors that can be modulated post-translationally. Some of the modulators are Mitogen-activated kinases (MAPKs), whose role has been studied in vivo and in vitro. p38-MAPK modifies the transactivation ability of some key transcription factors in chondrocytes, osteoblasts and osteoclasts, which affects their differentiation and function. Several commercially available inhibitors have helped to determine p38 action on these processes. Although it is frequently mentioned in the literature, this chemical approach is not always as accurate as it should be. Conditional knockouts are a useful genetic tool that could unravel the role of p38 in shaping the skeleton. In this review, we will summarize the state of the art on p38 activity during osteoblast differentiation and function, and emphasize the triggers of this MAPK. PMID:27200351

  3. RNase activity of sialic acid-binding lectin from bullfrog eggs drives antitumor effect via the activation of p38 MAPK to caspase-3/7 signaling pathway in human breast cancer cells

    PubMed Central

    Kariya, Yukiko; Tatsuta, Takeo; Sugawara, Shigeki; Kariya, Yoshinobu; Nitta, Kazuo; Hosono, Masahiro

    2016-01-01

    Sialic acid-binding lectin obtained from bullfrog eggs (SBL) induces cell death in cancer cells but not in normal cells. This antitumor effect is mediated through its ribo-nuclease (RNase) activity. However, the underlying molecular mechanisms remain unclear. We found that the p38 mitogen-activated protein kinase (MAPK) signaling pathway was activated when SBL induced cell death in three human breast cancer cell lines: SK-BR-3, MCF-7, and MDA-MB231. The suppression of p38 MAPK phosphorylation by a p38 MAPK inhibitor as well as short interference RNA knockdown of p38 MAPK expression significantly decreased cell death and increased the cell viability of SBL-treated MDA-MB231 cells. H103A, an SBL mutant lacking in RNase activity, showed decreased SBL-induced cell death compared with native SBL. However, the loss of RNase activity of SBL had no effect on its internalization into cells. The H103A mutant also displayed decreased phosphorylation of p38 MAPK. Moreover, SBL promoted caspase-3/7 activation followed by a cleavage of poly (ADP-ribose)-polymerase, whereas the SBL mutant, H103A, lost this ability. The SBL-induced caspase-3/7 activation was suppressed by the p38 MAPK inhibitor, SB203580, as well as pan-caspase inhibitor, zVAD-fmk. In the presence of zVAD-fmk, the SBL-induced cell death was decreased. In addition, the cell viability of SBL-treated MDA-MB231 cells recovered by zVAD-fmk treatment. Taken together, our results suggest that the RNase activity of SBL leads to breast cancer cell death through the activation of p38 MAPK followed by the activation of caspase-3/7. PMID:27513956

  4. Coriolus versicolor mushroom polysaccharides exert immunoregulatory effects on mouse B cells via membrane Ig and TLR-4 to activate the MAPK and NF-κB signaling pathways.

    PubMed

    Yang, Shu-fa; Zhuang, Tai-feng; Si, Yan-mei; Qi, Ke-yan; Zhao, Juan

    2015-03-01

    This study aimed to characterize the immunopotentiating effects and immune receptors for Coriolus versicolor mushroom polysaccharides (CVP), a Chinese medicinal fungus that exerts anti-tumor activities by enhancing host immunity. Proliferation assays were used to determine whether CVP could activate splenocytes. Flow cytometry analysis and IgM and IgG detection were used to characterize CVP-binding cells. Immune receptors were analyzed in immunoprecipitation and western blot assays. The downstream signaling pathways were identified by western blotting or immunostaining. CVP significantly stimulated the proliferation of mouse splenocytes. Fluorescence-labeled CVP (fl-CVP) selectively stained mouse B cells, but not T cells. CVP induced the production of IgM and IgG1 with or without exogenous IL-4. Membrane Ig (B cell antigen-receptor, BCR) was identified as a CVP-binding protein in immunoprecipitation and western blot experiments. CVP-induced B cell proliferation could be significantly inhibited by anti-mouse immunoglobulin (Ig) blocking antibody (Fab) or in cells from TLR4-mutant mice (C3H/HeJ). Phosphorylation of ERK-1/2 and p38 MAPK were clearly increased in a time-dependent manner, as was the nuclear translocation of the cytosolic NF-κB p65 subunit after CVP stimulation. Together, we demonstrate that CVP can bind and induce B cell activation using membrane Ig and TLR-4 as potential immune receptors. CVP activates mouse B cells through the MAPK and NF-κB signaling pathways.

  5. Calcium Signaling Involvement in Cadmium-Induced Astrocyte Cytotoxicity and Cell Death Through Activation of MAPK and PI3K/Akt Signaling Pathways.

    PubMed

    Jiang, Jiao Hua; Ge, Guo; Gao, Kai; Pang, Ying; Chai, Rui Chao; Jia, Xi Hua; Kong, Jin Ge; Yu, Albert Cheung-Hoi

    2015-09-01

    Cadmium (Cd), a highly ubiquitous toxic heavy metal, can contaminate the environment, including agricultural soil, water and air, via industrial runoff and other sources of pollution. Cd accumulated in the body via direct exposure or through the food chain results in neurodegeneration and many other diseases. Previous studies on its toxicity in the central nervous system (CNS) focused mainly on neurons. To obtain a more comprehensive understanding of Cd toxicity for the CNS, we investigated how astrocytes respond to acute and chronic Cd exposure and its toxic molecular mechanisms. When primary cultures of cerebral cortical astrocytes incubated with 1-300 μM CdCl2, morphological changes, LDH release and cell death were observed in a time and dose-dependent manner. Further studies demonstrated that acute and chronic Cd treatment phosphorylated JNK, p38 and Akt to different degrees, while ERK1/2 was only phosphorylated under low doses of Cd (10 μM) exposure. Inhibition of JNK and PI3K/Akt, but not of p38, could partially protect astrocyte from cytotoxicity in chronic and acute Cd exposure. Moreover, Cd also induced a strong calcium signal, while BAPTA, a specific intracellular calcium (Ca(2+)) chelator, prevented Cd-induced intracellular increase of calcium levels in astrocytes; inhibited the Cd-induced activation of ERK1/2, JNK, p38 and Akt; and also significantly reduced astrocyte cell death. All of these results suggested that the Cd-Ca(2+)-MAPK and PI3K/Akt signaling pathways were involved in Cd-induced toxicity in astrocytes. This toxicity involvement indicates that these pathways may be exploited as a target for the prevention of Cd-induced neurodegenerative diseases. PMID:26248512

  6. Saikosaponin-D reduces cisplatin-induced nephrotoxicity by repressing ROS-mediated activation of MAPK and NF-κB signalling pathways.

    PubMed

    Ma, Xiaobin; Dang, Chengxue; Kang, Huafeng; Dai, Zhijun; Lin, Shuai; Guan, Haitao; Liu, Xiaoxu; Wang, Xijing; Hui, Wentao

    2015-09-01

    The nephrotoxicity induced by cisplatin (DDP) severely limits the clinical efficacy of this widely used anticancer agent. The observed nephrotoxicity may be the result of DDP-induced inflammation and apoptosis. Saikosaponin-D (SSD), a triterpenoid saponin, has numerous pharmacological properties. The goal of the present study was to investigate whether and how SSD protected against DDP-induced nephrotoxicity. Non-cytotoxic levels of SSD significantly increased the viability rate, improved the nuclear morphology, and attenuated the caspase-3 activation and programmed apoptosis of DDP-treated HK-2 cells. In addition, SSD treatment markedly inhibited the release of tumour necrosis factor (TNF)-α, interleukin-1β (IL-1β), and interleukin-6 (IL-6), as well as the production of nitric oxide and the expression of inducible nitric oxide synthase (iNOS) by these cells. More importantly, SSD effectively blocked the DDP-induced activation of NF-κB, P38, JNK, and MAPKs. Furthermore, we found that U0126 (a specific inhibitor of MAPKs) strongly inhibited the IKK/IκB/NF-κB-dependent release of pro-inflammatory cytokines and iNOS gene expression. Finally, we demonstrated that SSD decreased the level of reactive oxygen species (ROS) accumulation and that the specific ROS scavenger N-acetylcysteine (NAC) markedly inhibited the DDP-induced activation of MAPK and phosphorylation of the downstream signal NF-κB, which in turn reduced the levels of pro-inflammatory cytokine release and iNOS gene expression. Our results suggest that the SSD-mediated alleviation of DDP-induced nephrotoxicity was due to uncoupling of the ROS, P38, and JNK/NF-κB signalling pathways. PMID:26118633

  7. Thymol has antifungal activity against Candida albicans during infection and maintains the innate immune response required for function of the p38 MAPK signaling pathway in Caenorhabditis elegans.

    PubMed

    Shu, Chengjie; Sun, Lingmei; Zhang, Weiming

    2016-08-01

    The Caenorhabditis elegans model can be used to study Candida albicans virulence and host immunity, as well as to identify plant-derived natural products to use against C. albicans. Thymol is a hydrophobic phenol compound from the aromatic plant thyme. In this study, the in vitro data demonstrated concentration-dependent thymol inhibition of both C. albicans growth and biofilm formation during different developmental phases. With the aid of the C. elegans system, we performed in vivo assays, and our results further showed the ability of thymol to increase C. elegans life span during infection, inhibit C. albicans colony formation in the C. elegans intestine, and increase the expression levels of host antimicrobial genes. Moreover, among the genes that encode the p38 MAPK signaling pathway, mutation of the pmk-1 or sek-1 gene decreased the beneficial effects of thymol's antifungal activity against C. albicans and thymol's maintenance of the innate immune response in nematodes. Western blot data showed the level of phosphorylation of pmk-1 was dramatically decreased against C. albicans. In nematodes, treatment with thymol recovered the dysregulation of pmk-1 and sek-1 gene expressions, the phosphorylation level of PMK-1 caused by C. albicans infection. Therefore, thymol may act, at least in part, through the function of the p38 MAPK signaling pathway to protect against C. albicans infection and maintain the host innate immune response to C. albicans. Our results indicate that the p38 MAPK signaling pathway plays a crucial role in regulating the beneficial effects observed after nematodes infected with C. albicans were treated with thymol. PMID:26783030

  8. Plastid-nucleus communication involves calcium-modulated MAPK signalling

    PubMed Central

    Guo, Hailong; Feng, Peiqiang; Chi, Wei; Sun, Xuwu; Xu, Xiumei; Li, Yuan; Ren, Dongtao; Lu, Congming; David Rochaix, Jean; Leister, Dario; Zhang, Lixin

    2016-01-01

    Chloroplast retrograde signals play important roles in coordinating the plastid and nuclear gene expression and are critical for proper chloroplast biogenesis and for maintaining optimal chloroplast functions in response to environmental changes in plants. Until now, the signals and the mechanisms for retrograde signalling remain poorly understood. Here we identify factors that allow the nucleus to perceive stress conditions in the chloroplast and to respond accordingly by inducing or repressing specific nuclear genes encoding plastid proteins. We show that ABI4, which is known to repress the LHCB genes during retrograde signalling, is activated through phosphorylation by the MAP kinases MPK3/MPK6 and the activity of these kinases is regulated through 14-3-3ω-mediated Ca2+-dependent scaffolding depending on the chloroplast calcium sensor protein CAS. These findings uncover an additional mechanism in which chloroplast-modulated Ca2+ signalling controls the MAPK pathway for the activation of critical components of the retrograde signalling chain. PMID:27399341

  9. The CORM ALF-186 Mediates Anti-Apoptotic Signaling via an Activation of the p38 MAPK after Ischemia and Reperfusion Injury in Retinal Ganglion Cells

    PubMed Central

    Ulbrich, Felix; Kaufmann, Kai B.; Meske, Alexander; Lagrèze, Wolf A.; Augustynik, Michael; Buerkle, Hartmut; Ramao, Carlos C.; Biermann, Julia

    2016-01-01

    Purpose Ischemia and reperfusion injury may induce apoptosis and lead to sustained tissue damage and loss of function, especially in neuronal organs. While carbon monoxide is known to exert protective effects after various harmful events, the mechanism of carbon monoxide releasing molecules in neuronal tissue has not been investigated yet. We hypothesize that the carbon monoxide releasing molecule (CORM) ALF-186, administered after neuronal ischemia-reperfusion injury (IRI), counteracts retinal apoptosis and its involved signaling pathways and consecutively reduces neuronal tissue damage. Methods IRI was performed in rat´s retinae for 1 hour. The water-soluble CORM ALF-186 (10 mg/kg) was administered intravenously via a tail vein after reperfusion. After 24 and 48 hours, retinal tissue was harvested to analyze mRNA and protein expression of Bcl-2, Bax, Caspase-3, ERK1/2, p38 and JNK. Densities of fluorogold pre-labeled retinal ganglion cells (RGC) were analyzed 7 days after IRI. Immunohistochemistry was performed on retinal cross sections. Results ALF-186 significantly reduced IRI mediated loss of RGC. ALF-186 treatment differentially affected mitogen-activated protein kinases (MAPK) phosphorylation: ALF-186 activated p38 and suppressed ERK1/2 phosphorylation, while JNK remained unchanged. Furthermore, ALF-186 treatment affected mitochondrial apoptosis, decreasing pro-apoptotic Bax and Caspase-3-cleavage, but increasing anti-apoptotic Bcl-2. Inhibition of p38-MAPK using SB203580 reduced ALF-186 mediated anti-apoptotic effects. Conclusion In this study, ALF-186 mediated substantial neuroprotection, affecting intracellular apoptotic signaling, mainly via MAPK p38. CORMs may thus represent a promising therapeutic alternative treating neuronal IRI. PMID:27764224

  10. Peroxiredoxins in Regulation of MAPK Signalling Pathways; Sensors and Barriers to Signal Transduction

    PubMed Central

    Latimer, Heather R.; Veal, Elizabeth A.

    2016-01-01

    Peroxiredoxins are highly conserved and abundant peroxidases. Although the thioredoxin peroxidase activity of peroxiredoxin (Prx) is important to maintain low levels of endogenous hydrogen peroxide, Prx have also been shown to promote hydrogen peroxide-mediated signalling. Mitogen activated protein kinase (MAPK) signalling pathways mediate cellular responses to a variety of stimuli, including reactive oxygen species (ROS). Here we review the evidence that Prx can act as both sensors and barriers to the activation of MAPK and discuss the underlying mechanisms involved, focusing in particular on the relationship with thioredoxin. PMID:26813660

  11. p38 MAPK Signaling in Postnatal Tendon Growth and Remodeling

    PubMed Central

    Schwartz, Andrew J.; Sarver, Dylan C.; Sugg, Kristoffer B.; Dzierzawski, Justin T.; Gumucio, Jonathan P.; Mendias, Christopher L.

    2015-01-01

    Tendon is a dynamic tissue whose structure and function is influenced by mechanical loading, but little is known about the fundamental mechanisms that regulate tendon growth and remodeling in vivo. Data from cultured tendon fibroblasts indicated that the p38 MAPK pathway plays an important role in tendon fibroblast proliferation and collagen synthesis in vitro. To gain greater insight into the mechanisms of tendon growth, and explore the role of p38 MAPK signaling in this process, we tested the hypotheses that inducing plantaris tendon growth through the ablation of the synergist Achilles tendon would result in rapid expansion of a neotendon matrix surrounding the original tendon, and that treatment with the p38 MAPK inhibitor SB203580 would prevent this growth. Rats were treated with vehicle or SB203580, and subjected to synergist ablation by bilateral tenectomy of the Achilles tendon. Changes in histological and biochemical properties of plantaris tendons were analyzed 3, 7, or 28 days after overload, and comparisons were made to non-overloaded animals. By 28 days after overload, tendon mass had increased by 30% compared to non-overloaded samples, and cross-sectional area (CSA) increased by around 50%, with most of the change occurring in the neotendon. The expansion in CSA initially occurred through the synthesis of a hyaluronic acid rich matrix that was progressively replaced with mature collagen. Pericytes were present in areas of active tendon growth, but never in the original tendon ECM. Inhibition of p38 MAPK resulted in a profound decrease in IL6 expression, and had a modest effect on the expression of other ECM and cell proliferation genes, but had a negligible impact on overall tendon growth. The combined results from this study provided novel insights into tendon mechanobiology, and suggest that p38 MAPK signaling does not appear to be necessary for tendon growth in vivo. PMID:25768932

  12. Thymoquinone prevents RANKL-induced osteoclastogenesis activation and osteolysis in an in vivo model of inflammation by suppressing NF-KB and MAPK Signalling.

    PubMed

    Thummuri, Dinesh; Jeengar, Manish Kumar; Shrivastava, Shweta; Nemani, Harishankar; Ramavat, Ravindar Naik; Chaudhari, Pradip; Naidu, V G M

    2015-09-01

    Osteoclasts are multinuclear giant cells responsible for bone resorption in inflammatory bone diseases such as osteoporosis, rheumatoid arthritis and periodontitis. Because of deleterious side effects with currently available drugs the search continues for novel effective and safe therapies. Thymoquinone (TQ), the major bioactive component of Nigella sativa has been investigated for its anti-inflammatory, antioxidant and anticancer activities. However, its effects in osteoclastogenesis have not been reported. In the present study we show for the first time that TQ inhibits nuclear factor-KB ligand (RANKL) induced osteoclastogenesis in RAW 264.7 and primary bone marrow derived macrophages (BMMs) cells. RANKL induced osteoclastogenesis is associated with increased expression of multiple transcription factors via activation of NF-KB, MAPKs signalling and reactive oxygen species (ROS). Mechanistically TQ blocked the RANKL induced NF-KB activation by attenuating the phosphorylation of IkB kinase (IKKα/β). Interestingly, in RAW 264.7 cells TQ inhibited the RANKL induced phosphorylation of MAPKs and mRNA expression of osteoclastic specific genes such as TRAP, DC-STAMP, NFATc1 and c-Fos. In addition, TQ also decreased the RANKL stimulated ROS generation in macropahges (RAW 264.7) and H2O2 induced ROS generation in osteoblasts (MC-3T3-E1). Consistent with in vitro results, TQ inhibited lipopolysaccharide (LPS) induced bone resorption by suppressing the osteoclastogenesis. Indeed, micro-CT analysis showed that bone mineral density (BMD) and bone architecture parameters were positively modulated by TQ. Taken together our data demonstrate that TQ has antiosteoclastogenic effect by inhibiting inflammation induced activation of MAPKs, NF-KB and ROS generation followed by suppressing the gene expression of c-Fos and NFATc1 in osteoclast precursors.

  13. Cadmium at nanomolar concentrations activates Raf-MEK-ERK1/2 MAPKs signaling via EGFR in human cancer cell lines.

    PubMed

    Ali, Imran; Damdimopoulou, Pauliina; Stenius, Ulla; Halldin, Krister

    2015-04-25

    Cadmium (Cd) is an environmental contaminant classified as carcinogenic to humans by the International Agency for Research on Cancer, supported by data from occupational exposure. Environmentally relevant dietary exposure to Cd has recently been associated with osteoporosis and cancers of the prostate, endometrium, and breast in the general population. The low exposure effects have been proposed to result from endocrine modulative properties of Cd, which mimic the physiological actions of estrogen and androgen. However, the mechanism of action of Cd is an unanswered question. We have shown previously, using mouse models, that canonical estrogen receptor signaling is not involved in estrogen mimicry effects of Cd. Instead, low-level Cd exposure stimulated the mitogen-activated protein kinases (MAPKs) ERK1/2 in these mice. Here we investigate further the ERK1/2 MAPK signaling activation by Cd in vitro by using nanomolar concentrations of cadmium chloride (CdCl2) in three different human carcinoma cell lines: HepG2, MCF-7, and ECC-1. The findings also were confirmed in previously collected mouse tissue samples. We show that 10(-8)M levels of CdCl2 activate ERK1/2 (Tyr 204) and the p53 specific ubiquitin ligase Mdm2 (Ser 166) via Raf and MEK by acting through the epidermal growth factor receptor (EGFR). Furthermore, our results suggest that the CdCl2-induced activation of ERK1/2 and Mdm2 may interfere with the p53 response to genotoxic compounds in cancer cell lines. Our data collectively suggest that nanomolar levels of CdCl2 activate Raf-MEK-ERK1/2 via EGFR. We hypothesize that this signaling cascade may be involved in observed low exposure effects of Cd in certain human populations.

  14. 24-Week Exposure to Oxidized Tyrosine Induces Hepatic Fibrosis Involving Activation of the MAPK/TGF-β1 Signaling Pathway in Sprague-Dawley Rats Model

    PubMed Central

    Li, Zhuqing Leslie; Shi, Yonghui; Le, Guowei; Ding, Yinyi; Zhao, Qi

    2016-01-01

    Scope. Oxidized tyrosine (O-Tyr) has been widely detected in many consumer protein products. O-Tyr products such as dityrosine (Dityr) and 3-nitrotyrosine (3-NT) are universal biomarkers of protein oxidation and have been demonstrated to be associated with metabolic disorders in biological system. Evaluation of potential intracorporal effects of dietary O-Tyr is important since the mechanism of biological impacts induced by oral oxidized protein products (OPPs) is still limited although we have proved that some dietary OPPs would induce oxidative injury to liver and kidney. Methods and Results. The present study aimed to investigate the dose-dependent hepatic injury caused by oral O-Tyr in rats. 24-week feeding of O-Tyr enhanced aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities, increased total bilirubin (TBiL) content, and led to oxidative damage in rats liver. Besides, O-Tyr distinctly increased the phosphorylation of p38 and ERK2 MAPKs and enhanced fibrosis-related TGF-β1 and Smad2/3 levels. Higher extracellular matrix (ECM) indexes (ICTP, PIIINP) and histological examination (HE and Masson staining) also supported dose-dependent hepatic fibrosis caused by O-Tyr. Conclusion. These findings reveal that O-Tyr may induce oxidative damage and hepatic fibrosis via MAPK/TGF-β1 signaling pathway, in which ROS together with malondialdehyde (MDA) and OPPs act as the pivotal mediators. PMID:26788244

  15. Inhibition of Advanced Glycation Endproduct Formation by Lotus Seedpod Oligomeric Procyanidins through RAGE-MAPK Signaling and NF-κB Activation in High-Fat-Diet Rats.

    PubMed

    Wu, Qian; Li, Shuyi; Li, Xiaopeng; Sui, Yong; Yang, Yan; Dong, Lihong; Xie, Bijun; Sun, Zhida

    2015-08-12

    This study investigated the protective properties of lotus seedpod oligomeric procyanidins (LSOPC) against nonalcoholic fatty liver disease (NAFLD) and its underlying mechanism. Sprague-Dawley (SD) male rats were fed a basic diet, a high-fat diet (HFD), or HFD plus 0.2 or 0.5% (w/w) LSOPC for 12 weeks. Administration of LSOPC markedly reduced serum and hepatic biochemical parameters and protein expression of advanced glycation endproducts (AGEs). Additionally, 0.5% (w/w) LSOPC treatment remarkably reversed the increasing tendency of receptor of advanced glycation endproduct (RAGE) to normal level. Furthermore, dietary LSOPC significantly decreased the protein levels of mitogen-activated protein kinases (MAPK) and nuclear factor-kappa B (NF-κB) and down-regulated genes involved in pro-inflammatory cytokines and adhesion molecules. Taken together, these findings demonstrate that LSOPC may protect obese rats with long-term HFD-induced NAFLD against RAGE-MAPK-NF-κB signaling suppression.

  16. MAPKs and Signal Transduction in the Control of Gastrointestinal Epithelial Cell Proliferation and Differentiation

    PubMed Central

    Osaki, Luciana H.; Gama, Patrícia

    2013-01-01

    Mitogen-activated protein kinase (MAPK) pathways are activated by several stimuli and transduce the signal inside cells, generating diverse responses including cell proliferation, differentiation, migration and apoptosis. Each MAPK cascade comprises a series of molecules, and regulation takes place at different levels. They communicate with each other and with additional pathways, creating a signaling network that is important for cell fate determination. In this review, we focus on ERK, JNK, p38 and ERK5, the major MAPKs, and their interactions with PI3K-Akt, TGFβ/Smad and Wnt/β-catenin pathways. More importantly, we describe how MAPKs regulate cell proliferation and differentiation in the rapidly renewing epithelia that lines the gastrointestinal tract and, finally, we highlight the recent findings on nutritional aspects that affect MAPK transduction cascades. PMID:23670595

  17. Trojan horse strategy in Agrobacterium transformation: abusing MAPK defense signaling.

    PubMed

    Djamei, Armin; Pitzschke, Andrea; Nakagami, Hirofumi; Rajh, Iva; Hirt, Heribert

    2007-10-19

    Nuclear import of transfer DNA (T-DNA) is a central event in Agrobacterium transformation of plant cells and is thought to occur by the hijacking of certain host cell proteins. The T-DNA-associated virulence protein VirE2 mediates this process by binding to the nuclear import machinery via the host cell factor VIP1, whose role in plants has been so far unknown. Here we show that VIP1 is a transcription factor that is a direct target of the Agrobacterium-induced mitogen-activated protein kinase (MAPK) MPK3. Upon phosphorylation by MPK3, VIP1 relocalizes from the cytoplasm to the nucleus and regulates the expression of the PR1 pathogenesis-related gene. MAPK-dependent phosphorylation of VIP1 is necessary for VIP1-mediated Agrobacterium T-DNA transfer, indicating that Agrobacterium abuses the MAPK-targeted VIP1 defense signaling pathway for nuclear delivery of the T-DNA complex as a Trojan horse. PMID:17947581

  18. Silica nanoparticles induce oxidative stress, inflammation, and endothelial dysfunction in vitro via activation of the MAPK/Nrf2 pathway and nuclear factor-κB signaling

    PubMed Central

    Guo, Caixia; Xia, Yinye; Niu, Piye; Jiang, Lizhen; Duan, Junchao; Yu, Yang; Zhou, Xianqing; Li, Yanbo; Sun, Zhiwei

    2015-01-01

    Despite the widespread application of silica nanoparticles (SiNPs) in industrial, commercial, and biomedical fields, their response to human cells has not been fully elucidated. Overall, little is known about the toxicological effects of SiNPs on the cardiovascular system. In this study, SiNPs with a 58 nm diameter were used to study their interaction with human umbilical vein endothelial cells (HUVECs). Dose- and time-dependent decrease in cell viability and damage on cell plasma-membrane integrity showed the cytotoxic potential of the SiNPs. SiNPs were found to induce oxidative stress, as evidenced by the significant elevation of reactive oxygen species generation and malondialdehyde production and downregulated activity in glutathione peroxidase. SiNPs also stimulated release of cytoprotective nitric oxide (NO) and upregulated inducible nitric oxide synthase (NOS) messenger ribonucleic acid, while downregulating endothelial NOS and ET-1 messenger ribonucleic acid, suggesting that SiNPs disturbed the NO/NOS system. SiNP-induced oxidative stress and NO/NOS imbalance resulted in endothelial dysfunction. SiNPs induced inflammation characterized by the upregulation of key inflammatory mediators, including IL-1β, IL-6, IL-8, TNFα, ICAM-1, VCAM-1, and MCP-1. In addition, SiNPs triggered the activation of the Nrf2-mediated antioxidant system, as evidenced by the induction of nuclear factor-κB and MAPK pathway activation. Our findings demonstrated that SiNPs could induce oxidative stress, inflammation, and NO/NOS system imbalance, and eventually lead to endothelial dysfunction via activation of the MAPK/Nrf2 pathway and nuclear factor-κB signaling. This study indicated a potential deleterious effect of SiNPs on the vascular endothelium, which warrants more careful assessment of SiNPs before their application. PMID:25759575

  19. Fluoxetine signature on hippocampal MAPK signalling in sex-dependent manner.

    PubMed

    Mitic, Milos; Lukic, Iva; Bozovic, Natalija; Djordjevic, Jelena; Adzic, Miroslav

    2015-02-01

    A growing body of evidence indicates that mitogen-activated protein kinase (MAPK) participates in various stress-induced responses and is considered to be one of the pathophysiological mechanisms in depression. Surprisingly, the effect of antidepressants on MAPKs is almost unexplored, particularly from the perspective of sexes. The present study investigates the cytoplasm-nuclear distribution of MAPK family, c-Jun N-terminal kinases (JNKs) 1, 2 and 3; extracellular signal-regulated kinases (ERKs) 1 and 2; and p38 kinases, as well as their phosphoisoforms in the hippocampus of chronically stressed female and male rats and upon chronic fluoxetine treatment. Additionally, we analysed crosstalk between MAPK signalling and depressive-like behaviour which correlated with brain-derived neurotrophic factor (BDNF) expression. Our results emphasize a gender-specific and compartment-dependent response of MAPKs to stress and fluoxetine. In females, stress decreased pp38 and pJNK and induced cytosolic retention of pERKs which reduced all nuclear pMAPKs. These changes correlated with altered BDNF expression and behaviour. Similarly, in males, stress decreased pp38 but promoted nuclear translocation of pJNKs and pERKs. These stress alterations of pMAPKs in males were not associated with BDNF expression and depressive-like behaviour. Fluoxetine treatment in stressed females upregulated whole pMAPK signalling particularly those in nucleus which was followed with BDNF expression and normalization of behaviour. In stressed males, fluoxetine affected only cytosolic pJNKs, while nuclear pMAPK signalling and BDNF expression were unaffected even though fluoxetine normalized behaviour. Overall, our results suggest existence of gender-specific mechanism of fluoxetine on nuclear pMAPK/BDNF signalling and depressive-like behaviour and reinforce the antidepressant dogma that females and males respond differently to certain antidepressants. PMID:24841087

  20. Fluoxetine signature on hippocampal MAPK signalling in sex-dependent manner.

    PubMed

    Mitic, Milos; Lukic, Iva; Bozovic, Natalija; Djordjevic, Jelena; Adzic, Miroslav

    2015-02-01

    A growing body of evidence indicates that mitogen-activated protein kinase (MAPK) participates in various stress-induced responses and is considered to be one of the pathophysiological mechanisms in depression. Surprisingly, the effect of antidepressants on MAPKs is almost unexplored, particularly from the perspective of sexes. The present study investigates the cytoplasm-nuclear distribution of MAPK family, c-Jun N-terminal kinases (JNKs) 1, 2 and 3; extracellular signal-regulated kinases (ERKs) 1 and 2; and p38 kinases, as well as their phosphoisoforms in the hippocampus of chronically stressed female and male rats and upon chronic fluoxetine treatment. Additionally, we analysed crosstalk between MAPK signalling and depressive-like behaviour which correlated with brain-derived neurotrophic factor (BDNF) expression. Our results emphasize a gender-specific and compartment-dependent response of MAPKs to stress and fluoxetine. In females, stress decreased pp38 and pJNK and induced cytosolic retention of pERKs which reduced all nuclear pMAPKs. These changes correlated with altered BDNF expression and behaviour. Similarly, in males, stress decreased pp38 but promoted nuclear translocation of pJNKs and pERKs. These stress alterations of pMAPKs in males were not associated with BDNF expression and depressive-like behaviour. Fluoxetine treatment in stressed females upregulated whole pMAPK signalling particularly those in nucleus which was followed with BDNF expression and normalization of behaviour. In stressed males, fluoxetine affected only cytosolic pJNKs, while nuclear pMAPK signalling and BDNF expression were unaffected even though fluoxetine normalized behaviour. Overall, our results suggest existence of gender-specific mechanism of fluoxetine on nuclear pMAPK/BDNF signalling and depressive-like behaviour and reinforce the antidepressant dogma that females and males respond differently to certain antidepressants.

  1. Osmotic Stress, not Aldose Reductase Activity, Directly induces Growth Factors and MAPK Signaling changes during Sugar Cataract Formation

    PubMed Central

    Zhang, Peng; Xing, Kuiyi; Randazzo, James; Blessing, Karen; Lou, Marjorie F.; Kador, Peter F.

    2012-01-01

    In sugar cataract formation in rats, aldose reductase (AR) actitvity is not only linked to lenticular sorbitol (diabetic) or galactitol (galactosemic) formation but also to signal transduction changes, cytotoxic signals and activation of apoptosis. Using both in vitro and in vivo techniques, the interrelationship between AR activity, polyol (sorbitol and galactitol) formation, osmotic stress, growth factor induction, and cell signaling changes have been investigated. For in vitro studies, lenses from Sprague Dawley rats were cultured for up to 48 hrs in TC-199-bicarbonate media containing either 30 mM fructose (control), or 30 mM glucose or galctose with/without the aldose reductase inhibitors AL1576 or tolrestat, the sorbitol dehydrogenase inhibitor (SDI) CP-470,711, or 15 mM mannitol (osmotic-compensated media). For in vivo studies, lenses were obtained from streptozotocin-induced diabetic Sprague Dawley rats fed diet with/without the ARIs AL1576 or tolrestat for 10 weeks. As expected, lenses cultured in high glucose / galactose media or from untreated diabetic rats all showed a decrease in the GSH pool that was lessened by ARI treatment. Lenses either from diabetic rats or from glucose/galactose culture conditions showed increased expression of basic-FGF, TGF-β, and increased signaling through P-Akt, P-ERK1/2 and P-SAPK/JNK which were also normalized by ARIs to the expression levels observed in non-diabetic controls. Culturing rat lenses in osomotically compensated media containing 30 mM glucose or galactose did not lead to increased growth factor expression or altered signaling. These studies indicate that it is the biophysical response of the lens to osmotic stress that results in an increased intralenticular production of basic-FGF and TGF-β and the altered cytotoxic signaling that is observed during sugar cataract formation. PMID:22710095

  2. HBx-related long non-coding RNA DBH-AS1 promotes cell proliferation and survival by activating MAPK signaling in hepatocellular carcinoma.

    PubMed

    Huang, Jin-lan; Ren, Ting-yu; Cao, Shun-wang; Zheng, Shi-hao; Hu, Xiu-mei; Hu, Yan-wei; Lin, Li; Chen, Jing; Zheng, Lei; Wang, Qian

    2015-10-20

    Accumulating evidence supports an important role for the hepatitis B virus x protein (HBx) in the pathogenesis of hepatitis B virus (HBV)-induced hepatocellular carcinoma (HCC), but the underlying mechanisms are not entirely clear. Here, we identified a novel long noncoding RNA (lncRNA) DBH-AS1 involved in the HBx-mediated hepatocarcinogenesis. The levels of DBH-AS1 were positively correlated with hepatitis B surface antigen (HBsAg) and tumor size in HCC tissues. Functionally, transgenic expression of DBH-AS1 significantly enhanced cell proliferation and tumorigenesis, whereas short hairpin RNA knockdown of DBH-AS1 caused an inhibition of cell proliferation. Mechanistically, overexpression of DBH-AS1 induced cell cycle progression by accelerating G1/S and G2/M transition concomitantly with upregulation of CDK6, CCND1, CCNE1 and downregulation of p16, p21 and p27. We also found that enhanced DBH-AS1 expression inhibited serum starvation-induced apoptosis of HCC cells. In contrast, suppressed DBH-AS1 expression had opposite effects. Furthermore, DBH-AS1 was shown to activate MAPK pathway. We also provide evidence that DBH-AS1 could be significantly induced by HBx protein and markedly down-regulated by p53. Thus, we concluded that DBH-AS1 can be induced by HBx and inactivated by p53, and consequently promote cell proliferation and cell survival through activation of MAPK signaling in HCC. Our study suggests that DBH-AS1 acts as an oncogene for HCC. PMID:26393879

  3. Transmembrane/cytoplasmic, rather than catalytic, domains of Mmp14 signal to MAPK activation and mammary branching morphogenesis via binding to integrin β1

    PubMed Central

    Mori, Hidetoshi; Lo, Alvin T.; Inman, Jamie L.; Alcaraz, Jordi; Ghajar, Cyrus M.; Mott, Joni D.; Nelson, Celeste M.; Chen, Connie S.; Zhang, Hui; Bascom, Jamie L.; Seiki, Motoharu; Bissell, Mina J.

    2013-01-01

    Epithelial cell invasion through the extracellular matrix (ECM) is a crucial step in branching morphogenesis. The mechanisms by which the mammary epithelium integrates cues from the ECM with intracellular signaling in order to coordinate invasion through the stroma to make the mammary tree are poorly understood. Because the cell membrane-bound matrix metalloproteinase Mmp14 is known to play a key role in cancer cell invasion, we hypothesized that it could also be centrally involved in integrating signals for mammary epithelial cells (MECs) to navigate the collagen 1 (CL-1)-rich stroma of the mammary gland. Expression studies in nulliparous mice that carry a NLS-lacZ transgene downstream of the Mmp14 promoter revealed that Mmp14 is expressed in MECs at the tips of the branches. Using both mammary organoids and 3D organotypic cultures, we show that MMP activity is necessary for invasion through dense CL-1 (3 mg/ml) gels, but dispensable for MEC branching in sparse CL-1 (1 mg/ml) gels. Surprisingly, however, Mmp14 without its catalytic activity was still necessary for branching. Silencing Mmp14 prevented cell invasion through CL-1 and disrupted branching altogether; it also reduced integrin β1 (Itgb1) levels and attenuated MAPK signaling, disrupting Itgb1-dependent invasion/branching within CL-1 gels. FRET imaging revealed that Mmp14 associates directly with Itgb1. We identified a domain of Mmp14 that is required for modulating the levels of Itgb1, MEC signaling and the rate of invasion within CL-1. These results shed light on hitherto undescribed non-proteolytic activities of Mmp14 that are necessary for the Itgb1-dependent biochemical and mechanical signals that regulate branching in the mammary epithelium. PMID:23250208

  4. MAPK Signaling in Cardiovascular Health and Disease: Molecular Mechanisms and Therapeutic Targets

    PubMed Central

    Muslin, Anthony J.

    2009-01-01

    Intracellular mitogen-activated protein kinase (MAPK) signaling cascades likely play an important role in the pathogenesis of cardiac and vascular disease. A substantial amount of basic science research has defined many of the details of MAPK pathway organization and activation, but the role of individual signaling proteins in the pathogenesis of various cardiovascular diseases is still being elucidated. In this review, the role of the MAPKs extracellular signal-regulated kinase (ERK), C-jun N-terminal kinase (JNK) and p38 MAPK in cardiac hypertrophy, cardiac remodeling after myocardial infarction, atherosclerosis and vascular restenosis will be examined with attention paid to genetically-modified murine model systems and to the use of pharmacologic inhibitors of protein kinases. Despite the complexities of this field of research, attractive targets for pharmacological therapy are emerging. PMID:18752467

  5. Ferulic Acid Administered at Various Time Points Protects against Cerebral Infarction by Activating p38 MAPK/p90RSK/CREB/Bcl-2 Anti-Apoptotic Signaling in the Subacute Phase of Cerebral Ischemia-Reperfusion Injury in Rats

    PubMed Central

    Cheng, Chin-Yi; Tang, Nou-Ying; Kao, Shung-Te; Hsieh, Ching-Liang

    2016-01-01

    Objectives This study aimed to evaluate the effects of ferulic acid (FA) administered at various time points before or after 30 min of middle cerebral artery occlusion (MCAo) followed by 7 d of reperfusion and to examine the involvement of mitogen-activated protein kinase (MAPK) signaling pathways in the cortical penumbra. Methods FA was intravenously administered to rats at a dose of 100 mg/kg 24 h before ischemia (B-FA), 2 h before ischemia (P-FA), immediately after ischemic insult (I-FA), 2 h after reperfusion (R-FA), or 24 h after reperfusion (D-FA). Results Our study results indicated that P-FA, I-FA, and R-FA effectively reduced cerebral infarct areas and neurological deficits. P-FA, I-FA, and R-FA significantly downregulated glial fibrillary acidic protein (GFAP), mitochondrial Bax, cytochrome c, and cleaved caspase-3 expression, and effectively restored the phospho-p38 MAPK (p-p38 MAPK)/p38 MAPK ratio, phospho-90 kDa ribosomal S6 kinase (p-p90RSK) expression, phospho-Bad (p-Bad) expression, the phospho-cAMP response element-binding protein (p-CREB)/CREB ratio, the cytosolic and mitochondrial Bcl-2/Bax ratios, and the cytosolic Bcl-xL/Bax ratio in the cortical penumbra 7 d after reperfusion. SB203580, a specific inhibitor of p38 MAPK, administered 30 min prior to ischemia abrogated the downregulating effects of I-FA on cerebral infarction, and mitochondrial Bax and cleaved caspase-3 expression, and the upregulating effects of I-FA on the p-p38 MAPK/p38 MAPK ratio, p-p90RSK expression, p-Bad expression, and the p-CREB/CREB, and cytosolic and mitochondrial Bcl-2/Bax ratios. Conclusions Our study results thus indicate that P-FA, I-FA, and R-FA effectively suppress reactive astrocytosis and exert neuroprotective effects against cerebral infarction by activating p38 MAPK signaling. The regulating effects of P-FA, I-FA, and R-FA on Bax-induced apoptosis result from activation of the p38 MAPK/p90RSK/CREB/Bcl-2 signaling pathway, and eventually contribute to

  6. Catalase, but not MnSOD, inhibits glucose deprivation-activated ASK1-MEK-MAPK signal transduction pathway and prevents relocalization of Daxx: hydrogen peroxide as a major second messenger of metabolic oxidative stress.

    PubMed

    Song, Jae J; Lee, Yong J

    2003-10-01

    Overexpression of catalase, but not manganese superoxide dismutase (MnSOD), inhibited glucose deprivation-induced cytotoxicity and c-Jun N-terminal kinase 1 (JNK1) activation in human prostate adenocarcinoma DU-145 cells. Suppression of JNK1 activation by catalase overexpression resulted from inhibition of apoptosis signal-regulating kinase 1 (ASK1) activation by preventing dissociation of thioredoxin (TRX) from ASK1. Overexpression of catalase also inhibited relocalization of Daxx from the nucleus to the cytoplasm as well as association of Daxx with ASK1 during glucose deprivation. Taken together, hydrogen peroxide (H(2)O(2)) rather than superoxide anion (O(2) (*-)) acts as a second messenger of metabolic oxidative stress to activate the ASK1-MAPK/extracellular signal-regulated kinase (ERK) kinase (MEK)-mitogen-activated protein kinase (MAPK) signal transduction pathway.

  7. MAPK/JNK signalling: a potential autophagy regulation pathway

    PubMed Central

    Zhou, Yuan-Yuan; Li, Ying; Jiang, Wei-Qin; Zhou, Lin-Fu

    2015-01-01

    Autophagy refers to a lysosomal degradative pathway or a process of self-cannibalization. This pathway maintains nutrients levels for vital cellular functions during periods of starvation and it provides cells with survival advantages under various stress situations. However, the mechanisms responsible for the induction and regulation of autophagy are poorly understood. The c-Jun NH2-terminal kinase (JNK) signal transduction pathway functions to induce defence mechanisms that protect organisms against acute oxidative and xenobiotic insults. This pathway has also been repeatedly linked to the molecular events involved in autophagy regulation. The present review will focus on recent advances in understanding of the relationship between mitogen-activated protein kinase (MAPK)/JNK signalling and autophagic cell death. PMID:26182361

  8. Ozone (O3) elicits neurotoxicity in spinal cord neurons (SCNs) by inducing ER Ca(2+) release and activating the CaMKII/MAPK signaling pathway.

    PubMed

    Li, Yun; Lin, Xiaowen; Zhao, XueJun; Xie, Juntian; JunNan, Wang; Sun, Tao; Fu, Zhijian

    2014-11-01

    Ozone (O3) is widely used in the treatment of spinal cord related diseases. Excess or accumulation of this photochemical air can however be neurotoxic. In this study, in vitro cultured Wister rat spinal cord neurons (SCNs) were used to investigate the detrimental effects and underlying mechanisms of O3. Ozone in a dose-dependent manner inhibited cell viability at a range of 20 to 500 μg/ml, with the dose at 40 μg/ml resulting in a decrease of cell viability to 75%. The cell death after O3 exposure was related to endoplasmic reticulum (ER) calcium (Ca(2+)) release. Intracellular Ca(2+) chelator, ER stabilizer (inositol 1,4,5-trisphosphate receptor (IP3R) antagonist and ryanodine receptor (RyR) antagonist) and calcium/calmodulin-dependent protein kinase II (CaMKII) antagonist could effectively block Ca(2+) mobilization and inhibit cell death following 40 μg/ml O3 exposure. In addition, ER Ca(2+) release due to O3 exposure enhanced phospho-p38 and phospho-JNK levels and apoptosis of SCNs through activating CaMKII. Based on these results, we confirm that ozone elicits neurotoxicity in SCNs via inducing ER Ca(2+) release and activating CaMKII/MAPK signaling pathway. Therefore, physicians should get attention to the selection of treatment concentrations of oxygen/ozone. And, approaches, such as chelating intracellular Ca(2+) and stabilizing neuronal Ca(2+) homeostasis could effectively ameliorate the neurotoxicity of O3.

  9. Wogonin suppresses TNF-{alpha}-induced MMP-9 expression by blocking the NF-{kappa}B activation via MAPK signaling pathways in human aortic smooth muscle cells

    SciTech Connect

    Lee, Syng-Ook; Jeong, Yun-Jeong; Yu, Mi Hee; Lee, Ji-Won; Hwangbo, Mi Hyang; Kim, Cheorl-Ho; Lee, In-Seon . E-mail: inseon@kmu.ac.kr

    2006-12-08

    Matrix metalloproteinase-9 (MMP-9) plays a major role in the pathogenesis of atherosclerosis and restenosis by regulating both migration and proliferation of vascular smooth muscle cells (VSMC) after an arterial injury. In this study, we examined the inhibitory effect of three major flavonoids in Scutellariae Radix, baicalin, baicalein, and wogonin, on TNF-{alpha}-induced MMP-9 expression in human aortic smooth muscle cells (HASMC). Wogonin, but not baicalin and baicalein, significantly and selectively suppressed TNF-{alpha}-induced MMP-9 expression in HASMC. Reporter gene, electrophoretic mobility shift, and Western blotting assays showed that wogonin inhibits MMP-9 gene transcriptional activity by blocking the activation of NF-{kappa}B via MAPK signaling pathways. Moreover, the Matrigel migration assay showed that wogonin reduced TNF-{alpha}-induced HASMC migration. These results suggest that wogonin effectively suppresses TNF-{alpha}-induced HASMC migration through the selective inhibition of MMP-9 expression and represents a potential agent for the prevention of vascular disorders related to the migration of VSMC.

  10. Tiron Inhibits UVB-Induced AP-1 Binding Sites Transcriptional Activation on MMP-1 and MMP-3 Promoters by MAPK Signaling Pathway in Human Dermal Fibroblasts.

    PubMed

    Lu, Jing; Guo, Jia-Hui; Tu, Xue-Liang; Zhang, Chao; Zhao, Mei; Zhang, Quan-Wu; Gao, Feng-Hou

    2016-01-01

    Recent research found that Tiron was an effective antioxidant that could act as the intracellular reactive oxygen species (ROS) scavenger or alleviate the acute toxic metal overload in vivo. In this study, we investigated the inhibitory effect of Tiron on matrix metalloproteinase (MMP)-1 and MMP-3 expression in human dermal fibroblast cells. Western blot and ELISA analysis revealed that Tiron inhibited ultraviolet B (UVB)-induced protein expression of MMP-1 and MMP-3. Real-time quantitative PCR confirmed that Tiron could inhibit UVB-induced mRNA expression of MMP-1 and MMP-3. Furthermore, Tiron significantly blocked UVB-induced activation of the MAPK signaling pathway and activator protein (AP)-1 in the downstream of this transduction pathway in fibroblasts. Through the AP-1 binding site mutation, it was found that Tiron could inhibit AP-1-induced upregulation of MMP-1 and MMP-3 expression through blocking AP-1 binding to the AP-1 binding sites in the MMP-1 and MMP-3 promoter region. In conclusion, Tiron may be a novel antioxidant for preventing and treating skin photoaging UV-induced. PMID:27486852

  11. Tiron Inhibits UVB-Induced AP-1 Binding Sites Transcriptional Activation on MMP-1 and MMP-3 Promoters by MAPK Signaling Pathway in Human Dermal Fibroblasts

    PubMed Central

    Zhang, Chao; Zhao, Mei; Zhang, Quan-Wu; Gao, Feng-Hou

    2016-01-01

    Recent research found that Tiron was an effective antioxidant that could act as the intracellular reactive oxygen species (ROS) scavenger or alleviate the acute toxic metal overload in vivo. In this study, we investigated the inhibitory effect of Tiron on matrix metalloproteinase (MMP)-1 and MMP-3 expression in human dermal fibroblast cells. Western blot and ELISA analysis revealed that Tiron inhibited ultraviolet B (UVB)-induced protein expression of MMP-1 and MMP-3. Real-time quantitative PCR confirmed that Tiron could inhibit UVB-induced mRNA expression of MMP-1 and MMP-3. Furthermore, Tiron significantly blocked UVB-induced activation of the MAPK signaling pathway and activator protein (AP)-1 in the downstream of this transduction pathway in fibroblasts. Through the AP-1 binding site mutation, it was found that Tiron could inhibit AP-1-induced upregulation of MMP-1 and MMP-3 expression through blocking AP-1 binding to the AP-1 binding sites in the MMP-1 and MMP-3 promoter region. In conclusion, Tiron may be a novel antioxidant for preventing and treating skin photoaging UV-induced. PMID:27486852

  12. The Role of MAPK Modules and ABA during Abiotic Stress Signaling.

    PubMed

    de Zelicourt, Axel; Colcombet, Jean; Hirt, Heribert

    2016-08-01

    To respond to abiotic stresses, plants have developed specific mechanisms that allow them to rapidly perceive and respond to environmental changes. The phytohormone abscisic acid (ABA) was shown to be a pivotal regulator of abiotic stress responses in plants, triggering major changes in plant physiology. The ABA core signaling pathway largely relies on the activation of SnRK2 kinases to mediate several rapid responses, including gene regulation, stomatal closure, and plant growth modulation. Mitogen-activated protein kinases (MAPKs) have also been implicated in ABA signaling, but an entire ABA-activated MAPK module was uncovered only recently. In this review, we discuss the evidence for a role of MAPK modules in the context of different plant ABA signaling pathways. PMID:27143288

  13. Berberine attenuates high glucose-induced fibrosis by activating the G protein-coupled bile acid receptor TGR5 and repressing the S1P2/MAPK signaling pathway in glomerular mesangial cells.

    PubMed

    Yang, Zhiying; Li, Jie; Xiong, Fengxiao; Huang, Junying; Chen, Cheng; Liu, Peiqing; Huang, Heqing

    2016-08-15

    Berberine (BBR) exerts powerful renoprotective effects on diabetic nephropathy (DN), but the underlying mechanisms remain unclear. We previously demonstrated that activation of the G protein-coupled bile acid receptor TGR5 ameliorates diabetic nephropathy by inhibiting the activation of the sphingosine 1-phosphate (S1P)/sphingosine 1-phosphate receptor 2 (S1P2) signaling pathway. In this study, we explored the role of TGR5 in the BBR-induced downregulation of sphingosine 1-phosphate receptor 2 (S1P2)/mitogen-activated protein kinase (MAPK)-mediated fibrosis in glomerular mesangial cells (GMCs). Results showed that, BBR suppressed the expression of FN, ICAM-1, and TGF-β1 in high-glucose cultures of GMCs, and the phosphorylation level of c-Jun/c-Fos was downregulated. The high glucose lowered TGR5 expression in a time-dependent manner; this effect was reversed by BBR in a dose-dependent manner. The TGR5 agonist INT-777 decreased the high glucose-induced FN, ICAM-1, and TGF-β1 protein contents. In addition, TGR5 siRNA blocked S1P2 degradation by BBR. And MAPK signaling, which plays important regulatory roles in the pathological progression of DN, was activated by TGR5 siRNA. Apart from this, MAPK signaling as well as FN, ICAM-1, and TGF-β1 suppressed by BBR under high glucose conditions were limited by TGR5 depletion. Thus, BBR decreases FN, ICAM-1, and TGF-β1 levels under high glucose conditions in GMCs possibly by activating TGR5 and inhibiting S1P2/MAPK signaling. PMID:27292312

  14. Age-dependent changes of the antioxidant system in rat livers are accompanied by altered MAPK activation and a decline in motor signaling

    PubMed Central

    Yang, Wei; Burkhardt, Britta; Fischer, Luise; Beirow, Maja; Bork, Nadja; Wönne, Eva C.; Wagner, Cornelia; Husen, Bettina; Zeilinger, Katrin; Liu, Liegang; Nussler, Andreas K.

    2015-01-01

    Aging is characterized by a progressive decrease of cellular functions, because cells gradually lose their capacity to respond to injury. Increased oxidative stress is considered to be one of the major contributors to age-related changes in all organs including the liver. Our study has focused on elucidating whether important antioxidative enzymes, the mTOR pathway, and MAPKs exhibit age-dependent changes in the liver of rats during aging. We found an age-dependent increase of GSH in the cytosol and mitochondria. The aged liver showed an increased SOD enzyme activity, while the CAT enzyme activity decreased. HO-1 and NOS-2 gene expression was lower in adult rats, but up-regulated in aged rats. Western blot analysis revealed that SOD1, SOD2, GPx, GR, γ-GCL, and GSS were age-dependent up-regulated, while CAT remained constant. We also demonstrated that the phosphorylation of Akt, JNK, p38, and TSC2Ser1254 decreased while ERK1/2 and TSC2Thr1462 increased age-dependently. Furthermore, our data show that the mTOR pathway seems to be activated in livers of aged rats, and hence stimulating cell proliferation/regeneration, as confirmed by an age-dependent increase of PCNA and p-eIF4ESer209 protein expression. Our data may help to explain the fact that liver cells only proliferate in cases of necessity, like injury and damage. In summary, we have demonstrated that, age-dependent changes of the antioxidant system and stress-related signaling pathways occur in the livers of rats, which may help to better understand organ aging. PMID:27004051

  15. Evolutionary history of mitogen-activated protein kinase (MAPK) genes in Lotus, Medicago, and Phaseolus

    PubMed Central

    Neupane, Achal; Nepal, Madhav P; Benson, Benjamin V; MacArthur, Kenton J; Piya, Sarbottam

    2013-01-01

    Mitogen-Activated Protein Kinase (MAPK) genes encode proteins that mediate various signaling pathways associated with biotic and abiotic stress responses in eukaryotes. The MAPK genes form a 3-tier signal transduction cascade between cellular stimuli and physiological responses. Recent identification of soybean MAPKs and availability of genome sequences from other legume species allowed us to identify their MAPK genes. The main objectives of this study were to identify MAPKs in 3 legume species, Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, and to assess their phylogenetic relationships. We used approaches in comparative genomics for MAPK gene identification and named the newly identified genes following Arabidopsis MAPK nomenclature model. We identified 19, 18, and 15 MAPKs and 7, 4, and 9 MAPKKs in the genome of Lotus japonicus, Medicago truncatula, and Phaseolus vulgaris, respectively. Within clade placement of MAPKs and MAPKKs in the 3 legume species were consistent with those in soybean and Arabidopsis. Among 5 clades of MAPKs, 4 founder clades were consistent to MAPKs of other plant species and orthologs of MAPK genes in the fifth clade-"Clade E" were consistent with those in soybean. Our results also indicated that some gene duplication events might have occurred prior to eudicot-monocot divergence. Highly diversified MAPKs in soybean relative to those in 3 other legume species are attributable to the polyploidization events in soybean. The identification of the MAPK genes in the legume species is important for the legume crop improvement; and evolutionary relationships and functional divergence of these gene members provide insights into plant genome evolution. PMID:24317362

  16. Baicalin Attenuates Hypoxia-Induced Pulmonary Arterial Hypertension to Improve Hypoxic Cor Pulmonale by Reducing the Activity of the p38 MAPK Signaling Pathway and MMP-9.

    PubMed

    Yan, Shuangquan; Wang, Yiran; Liu, Panpan; Chen, Ali; Chen, Mayun; Yao, Dan; Xu, Xiaomei; Wang, Liangxing; Huang, Xiaoying

    2016-01-01

    Baicalin has a protective effect on hypoxia-induced pulmonary hypertension in rats, but the mechanism of this effect remains unclear. Thus, investigating the potential mechanism of this effect was the aim of the present study. Model rats that display hypoxic pulmonary hypertension and cor pulmonale under control conditions were successfully generated. We measured a series of indicators to observe the levels of pulmonary arterial hypertension, pulmonary arteriole remodeling, and right ventricular remodeling. We assessed the activation of p38 mitogen-activated protein kinase (MAPK) in the pulmonary arteriole walls and pulmonary tissue homogenates using immunohistochemistry and western blot analyses, respectively. The matrix metalloproteinase- (MMP-) 9 protein and mRNA levels in the pulmonary arteriole walls were measured using immunohistochemistry and in situ hybridization. Our results demonstrated that baicalin not only reduced p38 MAPK activation in both the pulmonary arteriole walls and tissue homogenates but also downregulated the protein and mRNA expression levels of MMP-9 in the pulmonary arteriole walls. This downregulation was accompanied by the attenuation of pulmonary hypertension, arteriole remodeling, and right ventricular remodeling. These results suggest that baicalin may attenuate pulmonary hypertension and cor pulmonale, which are induced by chronic hypoxia, by downregulating the p38 MAPK/MMP-9 pathway. PMID:27688788

  17. Baicalin Attenuates Hypoxia-Induced Pulmonary Arterial Hypertension to Improve Hypoxic Cor Pulmonale by Reducing the Activity of the p38 MAPK Signaling Pathway and MMP-9

    PubMed Central

    Wang, Yiran; Chen, Ali; Chen, Mayun; Yao, Dan; Xu, Xiaomei; Wang, Liangxing

    2016-01-01

    Baicalin has a protective effect on hypoxia-induced pulmonary hypertension in rats, but the mechanism of this effect remains unclear. Thus, investigating the potential mechanism of this effect was the aim of the present study. Model rats that display hypoxic pulmonary hypertension and cor pulmonale under control conditions were successfully generated. We measured a series of indicators to observe the levels of pulmonary arterial hypertension, pulmonary arteriole remodeling, and right ventricular remodeling. We assessed the activation of p38 mitogen-activated protein kinase (MAPK) in the pulmonary arteriole walls and pulmonary tissue homogenates using immunohistochemistry and western blot analyses, respectively. The matrix metalloproteinase- (MMP-) 9 protein and mRNA levels in the pulmonary arteriole walls were measured using immunohistochemistry and in situ hybridization. Our results demonstrated that baicalin not only reduced p38 MAPK activation in both the pulmonary arteriole walls and tissue homogenates but also downregulated the protein and mRNA expression levels of MMP-9 in the pulmonary arteriole walls. This downregulation was accompanied by the attenuation of pulmonary hypertension, arteriole remodeling, and right ventricular remodeling. These results suggest that baicalin may attenuate pulmonary hypertension and cor pulmonale, which are induced by chronic hypoxia, by downregulating the p38 MAPK/MMP-9 pathway. PMID:27688788

  18. Baicalin Attenuates Hypoxia-Induced Pulmonary Arterial Hypertension to Improve Hypoxic Cor Pulmonale by Reducing the Activity of the p38 MAPK Signaling Pathway and MMP-9

    PubMed Central

    Wang, Yiran; Chen, Ali; Chen, Mayun; Yao, Dan; Xu, Xiaomei; Wang, Liangxing

    2016-01-01

    Baicalin has a protective effect on hypoxia-induced pulmonary hypertension in rats, but the mechanism of this effect remains unclear. Thus, investigating the potential mechanism of this effect was the aim of the present study. Model rats that display hypoxic pulmonary hypertension and cor pulmonale under control conditions were successfully generated. We measured a series of indicators to observe the levels of pulmonary arterial hypertension, pulmonary arteriole remodeling, and right ventricular remodeling. We assessed the activation of p38 mitogen-activated protein kinase (MAPK) in the pulmonary arteriole walls and pulmonary tissue homogenates using immunohistochemistry and western blot analyses, respectively. The matrix metalloproteinase- (MMP-) 9 protein and mRNA levels in the pulmonary arteriole walls were measured using immunohistochemistry and in situ hybridization. Our results demonstrated that baicalin not only reduced p38 MAPK activation in both the pulmonary arteriole walls and tissue homogenates but also downregulated the protein and mRNA expression levels of MMP-9 in the pulmonary arteriole walls. This downregulation was accompanied by the attenuation of pulmonary hypertension, arteriole remodeling, and right ventricular remodeling. These results suggest that baicalin may attenuate pulmonary hypertension and cor pulmonale, which are induced by chronic hypoxia, by downregulating the p38 MAPK/MMP-9 pathway.

  19. Systemic Regulation of RAS/MAPK Signaling by the Serotonin Metabolite 5-HIAA

    PubMed Central

    Schmid, Tobias; Snoek, L. Basten; Fröhli, Erika; van der Bent, M. Leontien; Kammenga, Jan; Hajnal, Alex

    2015-01-01

    Human cancer is caused by the interplay of mutations in oncogenes and tumor suppressor genes and inherited variations in cancer susceptibility genes. While many of the tumor initiating mutations are well characterized, the effect of genetic background variation on disease onset and progression is less understood. We have used C. elegans genetics to identify genetic modifiers of the oncogenic RAS/MAPK signaling pathway. Quantitative trait locus analysis of two highly diverged C. elegans isolates combined with allele swapping experiments identified the polymorphic monoamine oxidase A (MAOA) gene amx-2 as a negative regulator of RAS/MAPK signaling. We further show that the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA), which is a product of MAOA catalysis, systemically inhibits RAS/MAPK signaling in different organs of C. elegans. Thus, MAOA activity sets a global threshold for MAPK activation by controlling 5-HIAA levels. To our knowledge, 5-HIAA is the first endogenous small molecule that acts as a systemic inhibitor of RAS/MAPK signaling. PMID:25978500

  20. Jolkinolide B inhibits RANKL-induced osteoclastogenesis by suppressing the activation NF-κB and MAPK signaling pathways.

    PubMed

    Ma, Xiaojun; Liu, Yupeng; Zhang, Yao; Yu, Xiaobing; Wang, Weiming; Zhao, Dewei

    2014-03-01

    Osteoclasts together with osteoblasts play pivotal roles in bone remodeling. The unique function and ability of osteoclasts to resorb bone makes them critical in both normal bone homeostasis and pathologic bone diseases such as osteoporosis and rheumatoid arthritis. Thus, new compounds that may inhibit osteoclastogenesis and osteoclast function may be of great value in the treatment of osteoclast-related diseases. In the present study, we examined the effect of jolkinolide B (JB), isolated from the root of Euphorbia fischeriana Steud on receptor activator of NF-κB ligand (RANKL)-induced osteoclast formation. We found that JB inhibited RANKL-induced osteoclast differentiation from bone marrow macrophages (BMMs) without cytotoxicity. Furthermore, the expression of osteoclastic marker genes, such as tartrate-resistant acid phosphatase (TRAP), cathepsin K (CtsK), and calcitonin receptor (CTR), was significantly inhibited. JB inhibited RANKL-induced activation of NF-κB by suppressing RANKL-mediated IκBα degradation. Moreover, JB inhibited RANKL-induced phosphorylation of mitogen-activated protein kinases (p38, JNK, and ERK). This study thus identifies JB as an inhibitor of osteoclast formation and provides evidence that JB might be an alternative medicine for preventing and treating osteolysis.

  1. ECM compliance regulates osteogenesis by influencing MAPK signaling downstream of RhoA and ROCK.

    PubMed

    Khatiwala, Chirag B; Kim, Peter D; Peyton, Shelly R; Putnam, Andrew J

    2009-05-01

    The compliance of the extracellular matrix (ECM) regulates osteogenic differentiation by modulating extracellular signal-regulated kinase (ERK) activity. However, the molecular mechanism linking ECM compliance to the ERK-mitogen-activated protein kinase (MAPK) pathway remains unclear. Furthermore, RhoA has been widely implicated in integrin-mediated signaling and mechanotransduction. We studied the relationship between RhoA and ERK-MAPK signaling to determine their roles in the regulation of osteogenesis by ECM compliance. Inhibition of RhoA and ROCK in MC3T3-E1 pre-osteoblasts cultured on substrates of varying compliance reduced ERK activity, whereas constitutively active RhoA enhanced it. The expression of RUNX2, a potent osteogenic transcription factor, was increased on stiffer matrices and correlated with elevated ERK activity. Inhibition of RhoA, ROCK, or the MAPK pathway diminished RUNX2 activity and delayed the onset of osteogenesis as shown by altered osteocalcin (OCN) and bone sialoprotein (BSP) gene expression, alkaline phosphatase (ALP) activity, and matrix mineralization. These data establish that one possible mechanism by which ECM rigidity regulates osteogenic differentiation involves MAPK activation downstream of the RhoA-ROCK signaling pathway.

  2. Effects of acrylonitrile on lymphocyte lipid rafts and RAS/RAF/MAPK/ERK signaling pathways.

    PubMed

    Li, X J; Li, B; Huang, J S; Shi, J M; Wang, P; Fan, W; Zhou, Y L

    2014-09-26

    Acrylonitrile (ACN) is a widely used chemical in the production of plastics, resins, nitriles, acrylic fibers, and synthetic rubber. Previous epidemiological investigations and animal studies have confirmed that ACN affects the lymphocytes and spleen. However, the immune toxicity mechanism is unknown. Lipid rafts are cell membrane structures that are rich in cholesterol and involved in cell signal transduction. The B cell lymophoma-10 (Bcl10) protein is a joint protein that is important in lymphocyte development and signal pathways. This study was conducted to examine the in vitro effects of ACN. We separated lipid rafts, and analyzed Bcl10 protein and caveolin. Western blotting was used to detect mitogen-activated protein kinase (MAPK) and phosphorylated MAPK levels. The results indicated that with increasing ACN concentration, the total amount of Bcl10 remained stable, but was concentrated mainly in part 4 to part 11 in electrophoretic band district which is high density in gradient centrifugation. Caveolin-1 was evaluated as a lipid raft marker protein; caveolin-1 content and position were relatively unchanged. Western blotting showed that in a certain range, MAPK protein was secreted at a higher level. At some ACN exposure levels, MAPK protein secretion was significantly decreased compared to the control group (P < 0.05). These results indicate that ACN can cause immune toxicity by damaging lipid raft structures, causing Bcl10 protein and lipid raft separation and restraining Ras-Raf-MAPK-extracellular signal-regulated kinase signaling pathways.

  3. Effects of acrylonitrile on lymphocyte lipid rafts and RAS/RAF/MAPK/ERK signaling pathways.

    PubMed

    Li, X J; Li, B; Huang, J S; Shi, J M; Wang, P; Fan, W; Zhou, Y L

    2014-01-01

    Acrylonitrile (ACN) is a widely used chemical in the production of plastics, resins, nitriles, acrylic fibers, and synthetic rubber. Previous epidemiological investigations and animal studies have confirmed that ACN affects the lymphocytes and spleen. However, the immune toxicity mechanism is unknown. Lipid rafts are cell membrane structures that are rich in cholesterol and involved in cell signal transduction. The B cell lymophoma-10 (Bcl10) protein is a joint protein that is important in lymphocyte development and signal pathways. This study was conducted to examine the in vitro effects of ACN. We separated lipid rafts, and analyzed Bcl10 protein and caveolin. Western blotting was used to detect mitogen-activated protein kinase (MAPK) and phosphorylated MAPK levels. The results indicated that with increasing ACN concentration, the total amount of Bcl10 remained stable, but was concentrated mainly in part 4 to part 11 in electrophoretic band district which is high density in gradient centrifugation. Caveolin-1 was evaluated as a lipid raft marker protein; caveolin-1 content and position were relatively unchanged. Western blotting showed that in a certain range, MAPK protein was secreted at a higher level. At some ACN exposure levels, MAPK protein secretion was significantly decreased compared to the control group (P < 0.05). These results indicate that ACN can cause immune toxicity by damaging lipid raft structures, causing Bcl10 protein and lipid raft separation and restraining Ras-Raf-MAPK-extracellular signal-regulated kinase signaling pathways. PMID:25299088

  4. Novel human neutrophil agonistic properties of arsenic trioxide: involvement of p38 mitogen-activated protein kinase and/or c-jun NH2-terminal MAPK but not extracellular signal-regulated kinases-1/2.

    PubMed

    Binet, François; Girard, Denis

    2008-12-01

    Arsenic trioxide (ATO) is known for treating acute promyelocytic leukemia and for inducing apoptosis and mitogen-activated protein kinases (MAPKs) in promyelocytes and cancer cells. We recently reported that ATO induces neutrophil apoptosis. The aim of this study was to establish whether or not ATO recruits MAPKs in neutrophils, as well as to further investigate its agonistic properties. We found that ATO activates p38 and that, unlike H2O2, this response was not inhibited by exogenous catalase. Also, we demonstrated that ATO-induced p38 activation occurs before H2O2 generation and without a calcium burst. We next established that ATO recruits c-jun NH2-terminal (JNK) but not extracellular signal-regulated kinase 1 and 2 (Erk-1/2). Using pharmacological inhibitors, we found that the proapoptotic activity of ATO occurs by a MAPK-independent mechanism. In contrast, the ability of ATO to enhance adhesion, migration, phagocytosis, release, and activity of gelatinase and degranulation of secretory, specific, and gelatinase, but not azurophilic granules, is dependent upon activation of p38 and/or JNK. This is the first study establishing that ATO possesses important agonistic properties in human neutrophils. Given the central role of neutrophils in various inflammatory disorders, we propose that ATO might have broader therapeutic implications in clinics, especially for regulating inflammation.

  5. Expression profile of mitrogen-activated protein kinase (MAPK) signaling genes in the skeletal muscle & liver of rat with type 2 diabetes: Role in disease pathology

    PubMed Central

    Tang, Xiaoli; Deng, Libin; Xiong, Huangui; Li, Guilin; Lin, Jiari; Liu, Shuangmei; Xie, Jinyan; Liu, Jun; Kong, Fanjun; Tu, Guihua; Peng, Haiying; Liang, Shangdong

    2014-01-01

    Background & objectives: Type 2 diabetes (T2D) is characterized as hyperglycaemia caused by defects in insulin secretion, and it affects target tissues, such as skeletal muscle, liver and adipose tissue. Therefore, analyzing the changes of gene expression profiles in these tissues is important to elucidate the pathogenesis of T2D. We, therefore, measured the gene transcript alterations in liver and skeletal muscle of rat with induced T2D, to detect differentially expressed genes in liver and skeletal muscle and perform gene-annotation enrichment analysis. Methods: In the present study, skeletal muscle and liver tissue from 10 streptozotocin-induced diabetic rats and 10 control rats were analyzed using gene expression microarrays. KEGG pathways enriched by differentially expressed genes (DEGs) were identified by WebGestalt Expander and GATHER software. DEGs were validated by the method of real-time PCR and western blot. Results: From the 9,929 expressed genes across the genome, 1,305 and 997 differentially expressed genes (DEGs, P<0.01) were identified in comparisons of skeletal muscle and liver, respectively. Large numbers of DEGs (200) were common in both comparisons, which was clearly more than the predicted number (131 genes, P<0.001). For further interpretation of the gene expression data, three over-representation analysis softwares (WebGestalt, Expander and GATHER) were used. All the tools detected one KEGG pathway (MAPK signaling) and two GO (gene ontology) biological processes (response to stress and cell death), with enrichment of DEGs in both tissues. In addition, PPI (protein-protein interaction) networks constructed using human homologues not only revealed the tendency of DEGs to form a highly connected module, but also suggested a “hub” role of p38-MAPK-related genes (such as MAPK14) in the pathogenesis of T2D. Interpretation & conclusions: Our results indicated the considerably aberrant MAPK signaling in both insulin-sensitive tissues of T2D rat

  6. The BASL Polarity Protein Controls a MAPK Signaling Feedback Loop in Asymmetric Cell Division

    PubMed Central

    Zhang, Ying; Wang, Pengcheng; Shao, Wanchen; Zhu, Jian-Kang; Dong, Juan

    2015-01-01

    SUMMARY Cell polarization is linked to fate determination during asymmetric division of plant stem cells, but the underlying molecular mechanisms remain unknown. In Arabidopsis, BREAKING OF ASYMMETRY IN THE STOMATAL LINEAGE (BASL) is polarized to control stomatal asymmetric division. A MITOGEN-ACTIVATED PROTEIN KINASE (MAPK) cascade determines terminal stomatal fate by promoting the degradation of the lineage determinant SPEECHLESS (SPCH). Here we demonstrate that a positive feedback loop between BASL and the MAPK pathway constitutes a polarity module at the cortex. Cortical localization of BASL requires phosphorylation mediated by MPK3/6. Phosphorylated BASL functions as a scaffold and recruits the MAPKKK YODA and MPK3/6 to spatially concentrate signaling at the cortex. Activated MPK3/6 reinforces the feedback loop by phosphorylating BASL, and inhibits stomatal fate by phosphorylating SPCH. Polarization of the BASL-MAPK signaling feedback module represents a mechanism connecting cell polarity to fate differentiation during asymmetric stem cell division in plants. PMID:25843888

  7. A review of crosstalk between MAPK and Wnt signals and its impact on cartilage regeneration

    PubMed Central

    Zhang, Ying; Pizzute, Tyler; Pei, Ming

    2014-01-01

    Chondrogenesis is a developmental process that is controlled and coordinated by many growth and differentiation factors as well as environmental factors that initiate or suppress cellular signaling pathways and transcription of specific genes in a temporal-spatial manner. As key signaling molecules in regulating cell proliferation, homeostasis, and development, both mitogen-activated protein kinases (MAPK) and the Wnt family participate in morphogenesis and tissue patterning and play important roles in skeletal development, especially chondrogenesis. Recent findings suggest that both signals are also actively involved in arthritis and related diseases. Despite the fact that the crosstalk between MAPK and Wnt signaling has been implicated to play a significant role in cancer, few studies have summarized this interaction and crosstalk in regulating chondrogenesis. In this review, we focus on MAPK and Wnt signaling in reference to their relationships in different types of cells and particularly how this crosstalk might influence chondrogenesis and cartilage development. We also discuss how the interactions between MAPK and Wnt signaling might relate to cartilage related diseases such as osteoarthritis and explore the potential therapeutic targets for disease treatments. PMID:25312291

  8. The Dictyostelium MAPK ERK1 is phosphorylated in a secondary response to early developmental signaling

    PubMed Central

    Schwebs, David J.; Hadwiger, Jeffrey A.

    2014-01-01

    Previous reports have suggested that the two mitogen-activated protein kinases (MAPKs) in Dictyostelium discoideum, ERK1 and ERK2, can be directly activated in response to external cAMP even though these MAPKs play different roles in the developmental life cycle. To better characterize MAPK regulation, the levels of phosphorylated MAPKs were analyzed in response to external signals. Only ERK2 was rapidly phosphorylated in response to the chemoattractants, cAMP and folate. In contrast, the phosphorylation of ERK1 occurred as a secondary or indirect response to these stimuli and this phosphorylation was enhanced by cell-cell interactions, suggesting that other external signals can activate ERK1. The phosphorylation of ERK1 or ERK2 did not require the function of the other MAPK in these responses. Folate stimulation of a chimeric population of erk1− and gα4− cells revealed that the phosphorylation of ERK1 could be mediated through an intercellular signal other than folate. Loss of ERK1 function suppressed the developmental delay and the deficiency in anterior cell localization associated with gα5− mutants suggesting that ERK1 function can be down regulated through Gα5 subunit-mediated signaling. However, no major changes in the phosphorylation of ERK1 were observed in gα5− cells suggesting that the Gα5 subunit signaling pathway does not regulate the phosphorylation of ERK1. These findings suggest that the activation of ERK1 occurs as a secondary response to chemoattractants and that other cell-cell signaling mechanisms contribute to this activation. Gα5 subunit signaling can down regulate ERK1 function to promote prestalk cell development but not through major changes to the level of phosphorylated ERK1. PMID:25451080

  9. Crotoxin induces apoptosis and autophagy in human lung carcinoma cells in vitro via activation of the p38MAPK signaling pathway

    PubMed Central

    Han, Rong; Liang, Hui; Qin, Zheng-hong; Liu, Chun-yu

    2014-01-01

    Aim: Crotoxin (CrTX) is the primary toxin in South American rattlesnake (Crotalus durissus terrificus) venom, and exhibits antitumor and other pharmacological actions in vivo and in vitro. Here, we investigated the molecular mechanisms of the antitumor action of CrTX in human lung carcinoma cells in vitro. Methods: Human lung squamous carcinoma SK-MES-1 cells were tested. The cytotoxicity of CrTX was evaluated in both MTT and colony formation assays. Cell cycle was investigated with flow cytometry. Cell apoptosis was studied with Hoechst 33258 and Annexin V-FITC staining. The levels of relevant proteins were analyzed using Western blot assays. Results: CrTX (25, 50, 100 μmol/L) inhibited the growth and colony formation of SK-MES-1 cells in dose- and time-dependent manners. CrTX increased the proportion of S phase cells and dose-dependently induced cell apoptosis, accompanied by down-regulating the expression of proliferating cell nuclear antigen (PCNA), and increasing the level of cleaved caspase-3. Furthermore, CrTX dose-dependently increased the expression of autophagy-related proteins LC3-II and beclin 1, and decreased the level of p62 in the cells. Moreover, CrTX (50 μmol/L) significantly increased p38MAPK phosphorylation in the cells. Pretreatment of the cells with SB203580, a specific inhibitor of p38MAPK, blocked the inhibition of CrTX on cell proliferation, as well as CrTX-induced apoptosis and cleaved caspase-3 expression. Conclusion: The p38MAPK signaling pathway mediates CrTX-induced apoptosis and autophagy of human lung carcinoma SK-MES-1 cells in vitro. PMID:25132339

  10. PKG-Mediated MAPK Signaling Is Necessary for Long-Term Operant Memory in "Aplysia"

    ERIC Educational Resources Information Center

    Michel, Maximilian; Green, Charity L.; Eskin, Arnold; Lyons, Lisa C.

    2011-01-01

    Signaling pathways necessary for memory formation, such as the mitogen-activated protein kinase (MAPK) pathway, appear highly conserved across species and paradigms. Learning that food is inedible (LFI) represents a robust form of associative, operant learning that induces short- (STM) and long-term memory (LTM) in "Aplysia." We investigated the…

  11. Fisetin Inhibits Migration and Invasion of Human Cervical Cancer Cells by Down-Regulating Urokinase Plasminogen Activator Expression through Suppressing the p38 MAPK-Dependent NF-κB Signaling Pathway

    PubMed Central

    Chou, Ruey-Hwang; Hsieh, Shu-Ching; Yu, Yung-Luen; Huang, Min-Hsien; Huang, Yi-Chang; Hsieh, Yi-Hsien

    2013-01-01

    Fisetin (3,3’,4’,7-tetrahydroxyflavone), a naturally occurring flavonoid, has been reported to inhibit proliferation and induce apoptosis in several cancer types. However, its effect on the anti-metastatic potential of cervical cancer cells remains unclear. In the present study, we found that fisetin inhibits the invasion and migration of cervical cancer cells. The expression and activity of urokinase plasminogen activator (uPA) was significantly suppressed by fisetin in a dose-dependent manner. We also demonstrated that fisetin reduces the phosphorylation of p38 MAPK, but not that of ERK1/2, JNK1/2, or AKT. Addition of a p38 MAPK inhibitor, SB203580, further enhanced the inhibitory effect of fisetin on the expression and activity of uPA and the invasion and motility in cervical cancer cells. Fisetin suppressed the TPA (tetradecanoylphorbol-13-acetate)-induced activation of p38 MAPK and uPA, and inhibited the TPA-enhanced migratory and invasive abilities. Furthermore, the promoter activity of the uPA gene was dramatically repressed by fisetin, which disrupted the nuclear translocation of NF-κB and its binding amount on the promoter of the uPA gene, and these suppressive effects could be further enhanced by SB203580. This study provides strong evidence for the molecular mechanism of fisetin in inhibiting the aggressive phenotypes by repression of uPA via interruption of p38 MAPK-dependent NF-κB signaling pathway in cervical cancer cells and thus contributes insight to the potential of using fisetin as a therapeutic strategy against cervical cancer by inhibiting migration and invasion. PMID:23940799

  12. Fisetin inhibits migration and invasion of human cervical cancer cells by down-regulating urokinase plasminogen activator expression through suppressing the p38 MAPK-dependent NF-κB signaling pathway.

    PubMed

    Chou, Ruey-Hwang; Hsieh, Shu-Ching; Yu, Yung-Luen; Huang, Min-Hsien; Huang, Yi-Chang; Hsieh, Yi-Hsien

    2013-01-01

    Fisetin (3,3',4',7-tetrahydroxyflavone), a naturally occurring flavonoid, has been reported to inhibit proliferation and induce apoptosis in several cancer types. However, its effect on the anti-metastatic potential of cervical cancer cells remains unclear. In the present study, we found that fisetin inhibits the invasion and migration of cervical cancer cells. The expression and activity of urokinase plasminogen activator (uPA) was significantly suppressed by fisetin in a dose-dependent manner. We also demonstrated that fisetin reduces the phosphorylation of p38 MAPK, but not that of ERK1/2, JNK1/2, or AKT. Addition of a p38 MAPK inhibitor, SB203580, further enhanced the inhibitory effect of fisetin on the expression and activity of uPA and the invasion and motility in cervical cancer cells. Fisetin suppressed the TPA (tetradecanoylphorbol-13-acetate)-induced activation of p38 MAPK and uPA, and inhibited the TPA-enhanced migratory and invasive abilities. Furthermore, the promoter activity of the uPA gene was dramatically repressed by fisetin, which disrupted the nuclear translocation of NF-κB and its binding amount on the promoter of the uPA gene, and these suppressive effects could be further enhanced by SB203580. This study provides strong evidence for the molecular mechanism of fisetin in inhibiting the aggressive phenotypes by repression of uPA via interruption of p38 MAPK-dependent NF-κB signaling pathway in cervical cancer cells and thus contributes insight to the potential of using fisetin as a therapeutic strategy against cervical cancer by inhibiting migration and invasion.

  13. Cutaneous wound healing through paradoxical MAPK activation by BRAF inhibitors

    PubMed Central

    Escuin-Ordinas, Helena; Li, Shuoran; Xie, Michael W.; Sun, Lu; Hugo, Willy; Huang, Rong Rong; Jiao, Jing; de-Faria, Felipe Meira; Realegeno, Susan; Krystofinski, Paige; Azhdam, Ariel; Komenan, Sara Marie D.; Atefi, Mohammad; Comin-Anduix, Begoña; Pellegrini, Matteo; Cochran, Alistair J.; Modlin, Robert L.; Herschman, Harvey R.; Lo, Roger S.; McBride, William H.; Segura, Tatiana; Ribas, Antoni

    2016-01-01

    BRAF inhibitors are highly effective therapies for the treatment of BRAFV600-mutated melanoma, with the main toxicity being a variety of hyperproliferative skin conditions due to paradoxical activation of the mitogen-activated protein kinase (MAPK) pathway in BRAF wild-type cells. Most of these hyperproliferative skin changes improve when a MEK inhibitor is co-administered, as it blocks paradoxical MAPK activation. Here we show how the BRAF inhibitor vemurafenib accelerates skin wound healing by inducing the proliferation and migration of human keratinocytes through extracellular signal-regulated kinase (ERK) phosphorylation and cell cycle progression. Topical treatment with vemurafenib in two wound-healing mice models accelerates cutaneous wound healing through paradoxical MAPK activation; addition of a mitogen-activated protein kinase kinase (MEK) inhibitor reverses the benefit of vemurafenib-accelerated wound healing. The same dosing regimen of topical BRAF inhibitor does not increase the incidence of cutaneous squamous cell carcinomas in mice. Therefore, topical BRAF inhibitors may have clinical applications in accelerating the healing of skin wounds. PMID:27476449

  14. Cutaneous wound healing through paradoxical MAPK activation by BRAF inhibitors.

    PubMed

    Escuin-Ordinas, Helena; Li, Shuoran; Xie, Michael W; Sun, Lu; Hugo, Willy; Huang, Rong Rong; Jiao, Jing; de-Faria, Felipe Meira; Realegeno, Susan; Krystofinski, Paige; Azhdam, Ariel; Komenan, Sara Marie D; Atefi, Mohammad; Comin-Anduix, Begoña; Pellegrini, Matteo; Cochran, Alistair J; Modlin, Robert L; Herschman, Harvey R; Lo, Roger S; McBride, William H; Segura, Tatiana; Ribas, Antoni

    2016-08-01

    BRAF inhibitors are highly effective therapies for the treatment of BRAF(V600)-mutated melanoma, with the main toxicity being a variety of hyperproliferative skin conditions due to paradoxical activation of the mitogen-activated protein kinase (MAPK) pathway in BRAF wild-type cells. Most of these hyperproliferative skin changes improve when a MEK inhibitor is co-administered, as it blocks paradoxical MAPK activation. Here we show how the BRAF inhibitor vemurafenib accelerates skin wound healing by inducing the proliferation and migration of human keratinocytes through extracellular signal-regulated kinase (ERK) phosphorylation and cell cycle progression. Topical treatment with vemurafenib in two wound-healing mice models accelerates cutaneous wound healing through paradoxical MAPK activation; addition of a mitogen-activated protein kinase kinase (MEK) inhibitor reverses the benefit of vemurafenib-accelerated wound healing. The same dosing regimen of topical BRAF inhibitor does not increase the incidence of cutaneous squamous cell carcinomas in mice. Therefore, topical BRAF inhibitors may have clinical applications in accelerating the healing of skin wounds.

  15. Tumor Necrosis Factor-Like Weak Inducer of Apoptosis Accelerates the Progression of Renal Fibrosis in Lupus Nephritis by Activating SMAD and p38 MAPK in TGF-β1 Signaling Pathway

    PubMed Central

    Liu, Zhiqin; Xue, Leixi; Liu, Zhichun; Huang, Jun; Wen, Jian; Hu, Ji; Bo, Lin; Yang, Ru

    2016-01-01

    This study aim was to explore the effects of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) in lupus nephritis and its potential underlying mechanisms. MRL/lpr mice were used for in vivo experiments and human proximal tubular cells (HK2 cells) were used for in vitro experiments. Results showed that MRL/lpr mice treated with vehicle solution or LV-Control shRNA displayed significant proteinuria and severe renal histopathological changes. LV-TWEAK-shRNA treatment reversed these changes and decreased renal expressions of TWEAK, TGF-β1, p-p38 MAPK, p-Smad2, COL-1, and α-SMA proteins. In vitro, hTWEAK treatment upregulated the expressions of TGF-β1, p-p38 MAPK, p-SMAD2, α-SMA, and COL-1 proteins in HK2 cells and downregulated the expressions of E-cadherin protein, which were reversed by cotreatment with anti-TWEAK mAb or SB431542 treatment. These findings suggest that TWEAK may contribute to chronic renal changes and renal fibrosis by activating TGF-β1 signaling pathway, and phosphorylation of Smad2 and p38 MAPK proteins was also involved in this signaling pathway. PMID:27365897

  16. The Asian-American variant of human papillomavirus type 16 exhibits higher activation of MAPK and PI3K/AKT signaling pathways, transformation, migration and invasion of primary human keratinocytes.

    PubMed

    Hochmann, Jimena; Sobrinho, João S; Villa, Luisa L; Sichero, Laura

    2016-05-01

    Asian-American (AA) HPV-16 variants are associated with higher risk of cancer. Abnormal activation of intracellular signaling play a critical role in cancer development and progression. Our aim was to elucidate mechanisms underlying the higher oncogenic potential attributed to AA variant. We evaluated activation of MAPK and PI3K/AKT pathways in primary human keratinocytes (PHKs) transduced with E6/E7 of three HPV-16 variants: E-P, AA, E-350G. Phenotypes examined included migration, anchorage independent growth and invasion. AA PHKs presented the highest levels of active proteins involved in all cascades analyzed: MAPK-ERK, MAPK-p38 and PI3K-AKT. AA PHKs were more efficient in promoting anchorage independent growth, and in stimulating cell migration and invasion. MEK1 inhibition decreased migration. The mesenchymal phenotype marker vimentin was increased in AA PHKs. Our results suggest that MEK1, ERK2, AKT2 hyperactivation influence cellular behavior by means of GSK-3b inactivation and EMT induction prompting AA immortalized PHKs to more efficiently surpass carcinogenesis steps.

  17. p38γ and p38δ Mitogen Activated Protein Kinases (MAPKs), New Stars in the MAPK Galaxy.

    PubMed

    Escós, Alejandra; Risco, Ana; Alsina-Beauchamp, Dayanira; Cuenda, Ana

    2016-01-01

    The protein kinases p38γ and p38δ belong to the p38 mitogen-activated protein kinase (MAPK) family. p38MAPK signaling controls many cellular processes and is one of the most conserved mechanisms in eukaryotes for the cellular response to environmental stress and inflammation. Although p38γ and p38δ are widely expressed, it is likely that they perform specific functions in different tissues. Their involvement in human pathologies such as inflammation-related diseases or cancer is starting to be uncovered. In this article we give a general overview and highlight recent advances made in defining the functions of p38γ and p38δ, focusing in innate immunity and inflammation. We consider the potential of the pharmacological targeting of MAPK pathways to treat autoimmune and inflammatory diseases and cancer. PMID:27148533

  18. p38γ and p38δ Mitogen Activated Protein Kinases (MAPKs), New Stars in the MAPK Galaxy

    PubMed Central

    Escós, Alejandra; Risco, Ana; Alsina-Beauchamp, Dayanira; Cuenda, Ana

    2016-01-01

    The protein kinases p38γ and p38δ belong to the p38 mitogen-activated protein kinase (MAPK) family. p38MAPK signaling controls many cellular processes and is one of the most conserved mechanisms in eukaryotes for the cellular response to environmental stress and inflammation. Although p38γ and p38δ are widely expressed, it is likely that they perform specific functions in different tissues. Their involvement in human pathologies such as inflammation-related diseases or cancer is starting to be uncovered. In this article we give a general overview and highlight recent advances made in defining the functions of p38γ and p38δ, focusing in innate immunity and inflammation. We consider the potential of the pharmacological targeting of MAPK pathways to treat autoimmune and inflammatory diseases and cancer. PMID:27148533

  19. Kidney injury molecule-1 (KIM-1) mediates renal epithelial cell repair via ERK MAPK signaling pathway

    PubMed Central

    Zhang, Zhiwei; Cai, Cindy X

    2016-01-01

    The expression of kidney injury molecule-1 (KIM-1), a very promising sensitive and specific urinary biomarker for acute renal injury, is markedly upregulated in injured and regenerating renal proximal tubular epithelial cells following ischemic or toxic insults, suggesting a possible role for this molecule in renal repair process. In the present study we report that expression of KIM-1 facilitates renal tubular epithelial cell repair by promoting cell migration and proliferation. KIM-1 expression also enhances ERK MAPK activation, and the modulatory effect of KIM-1 on cellular repair process is likely mediated via ERK MAPK signaling pathway. PMID:27084535

  20. The role of MAPK signaling in patterning and establishing axial symmetry in the gastropod Haliotis asinina.

    PubMed

    Koop, Demian; Richards, Gemma S; Wanninger, Andreas; Gunter, Helen M; Degnan, Bernard M

    2007-11-01

    Gastropods are members of the Spiralia, a diverse group of invertebrates that share a common early developmental program, which includes spiral cleavage and a larval trochophore stage. The spiral cleavage program results in the division of the embryo into four quadrants. Specification of the dorsal (D) quadrant is intimately linked with body plan organization and in equally cleaving gastropods occurs when one of the vegetal macromeres makes contact with overlying micromeres and receives an inductive signal that activates a MAPK signaling cascade. Following the induction of the 3D macromere, the embryo begins to gastrulate and assumes a bilateral cleavage pattern. Here we inhibit MAPK activation in 3D with U0126 and examine its effect on the formation and patterning of the trochophore, using a suite of territory-specific markers. The head (pretrochal) region appears to maintain quadri-radial symmetry in U0126-treated embryos, supporting a role for MAPK signaling in 3D in establishing dorsoventral polarity in this region. Posterior (posttrochal) structures - larval musculature, shell and foot--fail to develop in MAPK inhibited trochophores. Inhibition of 3D specification by an alternative method--monensin treatment--yields similar abnormal trochophores. However, genes that are normally expressed in the ectodermal structures (shell and foot) are detected in U0126- and monensin-perturbed larvae in patterns that suggest that this region has latent dorsoventral polarity that is manifested even in the absence of D quadrant specification.

  1. A Functional Screen Reveals an Extensive Layer of Transcriptional and Splicing Control Underlying RAS/MAPK Signaling in Drosophila

    PubMed Central

    Ashton-Beaucage, Dariel; Udell, Christian M.; Gendron, Patrick; Sahmi, Malha; Lefrançois, Martin; Baril, Caroline; Guenier, Anne-Sophie; Duchaine, Jean; Lamarre, Daniel; Lemieux, Sébastien; Therrien, Marc

    2014-01-01

    The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway—including a new protein complex modulating RAF activation—we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing components can also

  2. Antagonistic control of cell fates by JNK and p38-MAPK signaling.

    PubMed

    Wada, T; Stepniak, E; Hui, L; Leibbrandt, A; Katada, T; Nishina, H; Wagner, E F; Penninger, J M

    2008-01-01

    During the development and organogenesis of all multicellular organisms, cell fate decisions determine whether cells undergo proliferation, differentiation, or aging. Two independent stress kinase signaling pathways, p38-MAPK, and JNKs, have evolved that relay developmental and environmental cues to determine cell responses. Although multiple stimuli can activate these two stress kinase pathways, the functional interactions and molecular cross-talks between these common second signaling cascades are poorly elucidated. Here we report that JNK and p38-MAPK pathways antagonistically control cellular senescence, oncogenic transformation, and proliferation in primary mouse embryonic fibroblasts (MEFs). Similarly, genetic inactivation of the JNK pathway results in impaired proliferation of fetal hepatoblasts in vitro and defective adult liver regeneration in vivo, which is rescued by inhibition of the p38-MAPK pathway. Thus, the balance between the two stress-signaling pathways, MKK7-JNK and MKK3/6-p38-MAPK, determines cell fate and links environmental and developmental stress to cell cycle arrest, senescence, oncogenic transformation, and adult tissue regeneration. PMID:17762881

  3. A novel lipopeptide from skin commensal activates TLR2/CD36-p38 MAPK signaling to increase antibacterial defense against bacterial infection.

    PubMed

    Li, Dongqing; Lei, Hu; Li, Zhiheng; Li, Hongquan; Wang, Yue; Lai, Yuping

    2013-01-01

    Staphylococcus epidermidis (S.epidermidis) plays important protective roles by directly producing or by stimulating hosts to produce antimicrobial peptides (AMPs) against pathogenic infections. Although several AMPs from S.epidermidis have been identified, molecules that stimulate hosts to produce AMPs remain largly unknown. Here we demonstrate that a new lipopeptide (named LP01) purified from S.epidermidis culture media has a unique structure with heneicosanoic acid (21 carbons) binding to lysine(11) of a peptide chain. In vitro LP01 increased the expression of β-defensin 2(hBD2) and hBD3 in neonatal human epidermal keratinocytes(NHEK), leading to increased capacity of cell lysates to inhibit the growth of S.aureus. In vivo LP01 induced the expression of mouse β-defensin 4(mBD4) to decrease the survival of local S.aureus in skin and systemic S.aureus survival in liver. The induction of beta-defensins by LP01 was dependent on TLR2 as Tlr2-deficient mice had decreased mBD4. Furthermore, knockdown of CD36 decreased the expression of hBD2 and hBD3, and p38 MAPK inhibitor significantly inhibited the expression of hBDs induced by LP01.Taken together, these findings demonstrate that lipopeptide LP01 from normal commensal S.epidermidis increases antimicrobial peptide hBD2 and hBD3 expression via the activation of TLR2/CD36-p38 MAPK, thus enhancing antimicrobial defense against pathogenic infections.

  4. In Vitro Manganese Exposure Disrupts MAPK Signaling Pathways in Striatal and Hippocampal Slices from Immature Rats

    PubMed Central

    Peres, Tanara Vieira; Pedro, Daniela Zótico; de Cordova, Fabiano Mendes; Lopes, Mark William; Gonçalves, Filipe Marques; Mendes-de-Aguiar, Cláudia Beatriz Nedel; Walz, Roger; Farina, Marcelo; Aschner, Michael; Leal, Rodrigo Bainy

    2013-01-01

    The molecular mechanisms mediating manganese (Mn)-induced neurotoxicity, particularly in the immature central nervous system, have yet to be completely understood. In this study, we investigated whether mitogen-activated protein kinases (MAPKs) and tyrosine hydroxylase (TH) could represent potential targets of Mn in striatal and hippocampal slices obtained from immature rats (14 days old). The aim of this study was to evaluate if the MAPK pathways are modulated after subtoxic Mn exposure, which do not significantly affect cell viability. The concentrations of manganese chloride (MnCl2; 10–1,000 μM) caused no change in cell viability in slices exposed for 3 or 6 hours. However, Mn exposure significantly increased extracellular signal-regulated kinase (ERK) 1/2, as well as c-Jun N-terminal kinase (JNK) 1/2/3 phosphorylation at both 3 and 6 hours incubations, in both brain structures. Furthermore, Mn exposure did not change the total content or phosphorylation of TH at the serine 40 site in striatal slices. Thus, Mn at concentrations that do not disrupt cell viability causes activation of MAPKs (ERK1/2 and JNK1/2/3) in immature hippocampal and striatal slices. These findings suggest that altered intracellular MAPKs signaling pathways may represent an early event concerning the effects of Mn in the immature brain. PMID:24324973

  5. Ssanghwa-tang, an oriental herbal cocktail, exerts anti-melanogenic activity by suppression of the p38 MAPK and PKA signaling pathways in B16F10 cells

    PubMed Central

    2013-01-01

    Background Ssanghwa-tang (SHT) is a widely used medication for the treatment of fatigue, pain, inflammation, hypothermia, erectile dysfunction, cancer, and osteoporosis in Asia, however, role of SHT on the melanin synthesis has not been checked previously. Thus, the present study was designed to determine the effect of SHT on α-melanocyte stimulating hormone (α-MSH)-induced melanogensis and its mechanisms of action in murine B16F10 melanoma cells. Method Cellular melanin content and tyrosinase activity in murine B16F10 melanoma cells were determined after α-MSH stimulation with or without pre-treatment of SHT at the concentration of 250 and 500 μg/ml. Expression level of tyrosinase, tyrosinase-related protein 1 (TRP-1), TRP-2, microphthalmia-associated transcription factor (MITF), and activation of c-AMP-dependent protein kinase (PKA), c-AMP-related element binding protein (CREB), and mitogen-activated protein kinases (MAPKs) were examined by Western blot analysis. Results SHT significantly inhibited α-MSH-induced melanin synthesis and tyrosinase activity, and also decreased α-MSH-induced expression of MITF, tyrosinase, and TRP-1. In addition, SHT remarkably suppressed tyrosinase, CRE, and MITF luciferase reporter activity in a resting state as well as in α-MSH-stimulating condition. Phosphorylation of p38 MAPK by α-MSH stimulation was efficiently blocked by SHT pre-treatment. Moreover, SHT as an herbal cocktail showed synergistic anti-melanogenic effect compared with that of each single constituent herb. Conclusion SHT efficiently inhibited c-AMP-induced melanin synthesis in B16F10 cells via suppression of PKA and p38 MAPK signaling pathways and subsequently decreased the level of CREB phosphorylation, MITF, and melanogenic enzymes. These results indicate that SHT may be useful as herbal medicine for treating hyperpigmentation and cosmetics as a skin-whitening agent. PMID:23981281

  6. Disorders of dysregulated signal traffic through the RAS-MAPK pathway: phenotypic spectrum and molecular mechanisms

    PubMed Central

    Tartaglia, Marco; Gelb, Bruce D.

    2010-01-01

    RAS GTPases control a major signaling network implicated in several cellular functions, including cell fate determination, proliferation, survival, differentiation, migration, and senescence. Within this network, signal flow through the RAF-MEK-ERK pathway, the first identified mitogen-associated protein kinase (MAPK) cascade, mediates early and late developmental processes controlling morphology determination, organogenesis, synaptic plasticity and growth. Signaling through the RAS-MAPK cascade is tightly controlled, and its enhanced activation represents a well-known event in oncogenesis. Unexpectedly, in the past few years, inherited dysregulation of this pathway has been recognized as the cause underlying a group of clinically related disorders sharing facial dysmorphism, cardiac defects, reduced postnatal growth, ectodermal anomalies, variable cognitive deficits and susceptibility to certain malignancies as major features. These disorders are caused by heterozygosity for mutations in genes encoding RAS proteins, regulators of RAS function, modulators of RAS interaction with effectors or downstream signal transducers. Here, we provide an overview of the phenotypic spectrum associated with germline mutations perturbing RAS-MAPK signaling, the unpredicted molecular mechanisms converging towards the dysregulation of this signaling cascade, and major genotype-phenotype correlations. PMID:20958325

  7. Tight interconnection and multi-level control of Arabidopsis MYB44 in MAPK cascade signalling.

    PubMed

    Persak, Helene; Pitzschke, Andrea

    2013-01-01

    Abiotic stress poses a huge, ever-increasing problem to plants and agriculture. The dissection of signalling pathways mediating stress tolerance is a prerequisite to develop more resistant plant species. Mitogen-activated protein kinase (MAPK) cascades are universal signalling modules. In Arabidopsis, the MAPK MPK3 and its upstream regulator MAPK kinase MKK4 initiate the adaptation response to numerous abiotic and biotic stresses. Yet, molecular steps directly linked with MKK4-MPK3 activation are largely unknown. Starting with a yeast-two-hybrid screen for interacting partners of MKK4, we identified a transcription factor, MYB44. MYB44 is controlled at multiple levels by and strongly inter-connected with MAPK signalling. As we had shown earlier, stress-induced expression of the MYB44 gene is regulated by a MPK3-targeted bZIP transcription factor VIP1. At the protein level, MYB44 interacts with MPK3 in vivo. MYB44 is phosphorylated by MPK3 in vitro at a single residue, Ser145. Although replacement of Ser145 by a non-phosphorylatable (S145A) or phosphomimetic (S145D) residue did not alter MYB44 subcellular localisation, dimerization behaviour nor DNA-binding characteristics, abiotic stress tolerance tests in stable transgenic Arabidopsis plants clearly related S145 phosphorylation to MYB44 function: Compared to Arabidopsis wild type plants, MYB44 overexpressing lines exhibit an enhanced tolerance to osmotic stress and are slightly more sensitive to abscisic acid. Interestingly, overexpression of the S145A variant revealed that impaired phosphorylation does not render the MYB44 protein non-functional. Instead, S145A lines are highly sensitive to abiotic stress, and thereby remarkably similar to mpk3-deficient plants. Its in vivo interaction with the nuclear sub-pools of both MPK3 and MKK4 renders MYB44 the first plant transcription factor to have a second function as putative MAPK cascade scaffolding protein.

  8. Cannabinoids inhibit the activation of ERK MAPK in PMA/Io-stimulated mouse splenocytes.

    PubMed

    Faubert Kaplan, Barbara L; Kaminski, Norbert E

    2003-10-01

    The mechanism of action of immune suppression by cannabinoids involves suppression of interleukin-2 (IL-2) production in phorbol ester plus calcium ionophore (PMA/Io)-stimulated lymphocytes. This decrease in IL-2 was due to inhibition of activator protein-1 (AP-1) and nuclear factor of activated T cells (NF-AT) transcription factors, both of which depend on proteins that are regulated by the extracellular signal-regulated kinase subgroup of the mitogen-activated protein kinases (ERK MAPK). Thus, the objective of the present study was to characterize the effects of cannabinoid compounds on ERK MAPK under conditions where IL-2 expression was suppressed. Using the MEK inhibitor PD098059 in order to assess the role of ERK MAPK in PMA/Io-stimulated splenocytes (SPLC), it was determined that IL-2 production and expression of c-fos and c-jun nuclear protein expression depended on activation of ERK MAPK. In response to PMA/Io, expression of nuclear phosphorylated ERK MAPK was rapidly induced, peaked at approximately 15 min, and was sustained for up to 240 min. Pretreatment with cannabinol (CBN) inhibited expression of phosphorylated ERK MAPK at several time points up to 240 min post cellular activation. Furthermore, WIN-55212-2, a synthetic cannabinoid, inhibited expression of phosphorylated ERK MAPK at 240 min post cellular activation. CBN did not induce activation of ERK MAPK in the absence of PMA/Io. Collectively, these studies suggest that cannabinoid-induced inhibition of IL-2 in PMA/Io-stimulated splenocytes might be due, in part, to inhibition of ERK MAPK activation.

  9. CARMA3 regulates the invasion, migration, and apoptosis of non-small cell lung cancer cells by activating NF-кB and suppressing the P38 MAPK signaling pathway.

    PubMed

    Xia, Z X; Li, Z X; Zhang, M; Sun, L M; Zhang, Q F; Qiu, X S

    2016-04-01

    In our previous study, CARMA3 overexpression in lung cancer cells promoted cell proliferation and invasion; however, the mechanism underlying the role of CARMA3 in cancer cell invasion remained unclear. In the present study, knockdown of CARMA3 in A549 and H1299 cells suppressed cell invasion and migration, and downregulated matrix metalloprotease 9 expression at the protein and mRNA levels, as shown by Western blotting and real-time PCR. CARMA3 knockdown increased cell apoptosis, as shown by flow cytometry, increased the mRNA and protein expression levels of Bax and Caspase3, and downregulated Bcl-2 in A549 and H1299 cells. Phosphorylated P38 levels increased and NF-кB activation decreased following knockdown of CARMA3. SB203580, a P38 MAPK inhibitor, activated NF-кB, increased cell migration, and inhibited cell apoptosis after knockdown of CARMA3 compared to knockdown of CARMA3 without SB203580. These findings indicate that CARMA3 may suppress the activation of the P38 MAPK signaling pathway to regulate invasion, migration and apoptosis of lung cancer cells by activating NF-кB (P65) in the nucleus.

  10. Ras-GTP dimers activate the mitogen-activated protein kinase (MAPK) pathway

    SciTech Connect

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li -Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-06-16

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referred to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors.

  11. Ras-GTP dimers activate the Mitogen-Activated Protein Kinase (MAPK) pathway

    PubMed Central

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li-Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-01-01

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referred to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors. PMID:26080442

  12. Ras-GTP dimers activate the mitogen-activated protein kinase (MAPK) pathway

    DOE PAGES

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li -Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-06-16

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referredmore » to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors.« less

  13. Reciprocal actions of NCAM and tPA via a Ras-dependent MAPK activation in rat hippocampal neurons.

    PubMed

    Son, Hyeon; Seuk Kim, Jin; Mogg Kim, Jung; Lee, Sang-Hun; Lee, Yong-Sung

    2002-10-25

    In an attempt to identify the functions of neural cell adhesion molecule (NCAM) and tissue plasminogen activator (tPA) in hippocampal synaptic plasticity, we investigated the relationship between the two molecules by focusing on mitogen-activated protein kinase (MAPK), an essential enzyme in this process. NCAM clustering in cultured hippocampal neurons transiently induced MAPK within 10min. Moreover, soluble NCAM also induced a Ras-dependent MAPK activation. Conversely, MAPK activation led to an increase in the expressions of all three isoforms of NCAM. Treatment of neurons with tPA and plasminogen induced a Ras-dependent MAPK activation and tPA-plasmin degradation of NCAM was mediated in a MAPK-dependent manner. Soluble NCAM transiently inhibited tPA mRNA expression levels in a MAPK-dependent manner, while stimulation of MAPK alone induced tPA reduction in cells. These results collectively indicate that NCAM and tPA reciprocally act as important regulators in the modulation of synaptic plasticity via a Ras-MAPK-involved signaling pathway. In turn, MAPK activation may cause tPA degradation or a decrease in expression to promote synaptic plasticity.

  14. Computational Modeling of PI3K/AKT and MAPK Signaling Pathways in Melanoma Cancer

    PubMed Central

    Pappalardo, Francesco; Russo, Giulia; Candido, Saverio; Pennisi, Marzio; Cavalieri, Salvatore; Motta, Santo; McCubrey, James A.; Nicoletti, Ferdinando; Libra, Massimo

    2016-01-01

    Background Malignant melanoma is an aggressive tumor of the skin and seems to be resistant to current therapeutic approaches. Melanocytic transformation is thought to occur by sequential accumulation of genetic and molecular alterations able to activate the Ras/Raf/MEK/ERK (MAPK) and/or the PI3K/AKT (AKT) signalling pathways. Specifically, mutations of B-RAF activate MAPK pathway resulting in cell cycle progression and apoptosis prevention. According to these findings, MAPK and AKT pathways may represent promising therapeutic targets for an otherwise devastating disease. Result Here we show a computational model able to simulate the main biochemical and metabolic interactions in the PI3K/AKT and MAPK pathways potentially involved in melanoma development. Overall, this computational approach may accelerate the drug discovery process and encourages the identification of novel pathway activators with consequent development of novel antioncogenic compounds to overcome tumor cell resistance to conventional therapeutic agents. The source code of the various versions of the model are available as S1 Archive. PMID:27015094

  15. Hyperhomocysteinemia inhibits satellite cell regenerative capacity through p38 alpha/beta MAPK signaling.

    PubMed

    Veeranki, Sudhakar; Lominadze, David; Tyagi, Suresh C

    2015-07-15

    Chronic failure in maintenance and regeneration of skeletal muscles leads to lower muscle mass (sarcopenia), muscle weakness, and poor response to injury. Evidence suggests that aberrant p38 MAPK signaling undermines the repair process after injury in aged mice. Previous studies have shown that hyperhomocysteinemia (HHcy) has been associated with muscle weakness and lower than normal body weights. However, whether or not HHcy condition also compromises skeletal muscle regenerative capabilities is not clear. In the current study, we show that CBS-/+ mice, a model for HHcy condition, exhibited compromised regenerative function and cell proliferation upon injury. However, there was no significant difference in Pax7 expression levels in the satellite cells from CBS-/+ mouse skeletal muscles. Interestingly, the satellite cells from CBS-/+ mice not only exhibited diminished in vitro proliferative capabilities, but also there was heightened oxidative stress. In addition, there was enhanced p38 MAPK activation as well as p16 and p21 expression in the CBS-/+ mouse satellite cells. Moreover, the C2C12 myoblasts also exhibited higher p38 MAPK activation and p16 expression upon treatment with homocysteine in addition to enhanced ROS presence. Tissue engraftment potential and regeneration after injury were restored to some extent upon treatment with the p38-MAPK inhibitor, SB203580, in the CBS-/+ mice. These results together suggest that HHcy-induced diminished satellite cell proliferation involves excessive oxidative stress and p38 MAPK signaling. Our study further proposes that HHcy is a potential risk factor for elderly frailty, and need to be considered as a therapeutic target while designing the alleviation interventions/postinjury rehabilitation measures for adults with HHcy.

  16. Trihydrophobin 1 Phosphorylation by c-Src Regulates MAPK/ERK Signaling and Cell Migration

    PubMed Central

    Wu, Weibin; Sun, Zhichao; Wu, Jingwen; Peng, Xiaomin; Gan, Huacheng; Zhang, Chunyi; Ji, Lingling; Xie, Jianhui; Zhu, Haiyan; Ren, Shifang

    2012-01-01

    c-Src activates Ras-MAPK/ERK signaling pathway and regulates cell migration, while trihydrophobin 1 (TH1) inhibits MAPK/ERK activation and cell migration through interaction with A-Raf and PAK1 and inhibiting their kinase activities. Here we show that c-Src interacts with TH1 by GST-pull down assay, coimmunoprecipitation and confocal microscopy assay. The interaction leads to phosphorylation of TH1 at Tyr-6 in vivo and in vitro. Phosphorylation of TH1 decreases its association with A-Raf and PAK1. Further study reveals that Tyr-6 phosphorylation of TH1 reduces its inhibition on MAPK/ERK signaling, enhances c-Src mediated cell migration. Moreover, induced tyrosine phosphorylation of TH1 has been found by EGF and estrogen treatments. Taken together, our findings demonstrate a novel mechanism for the comprehensive regulation of Ras/Raf/MEK/ERK signaling and cell migration involving tyrosine phosphorylation of TH1 by c-Src. PMID:22238675

  17. Anti-inflammatory activities of cardamonin from Alpinia katsumadai through heme oxygenase-1 induction and inhibition of NF-κB and MAPK signaling pathway in the carrageenan-induced paw edema.

    PubMed

    Li, Yuan-Yuan; Huang, Shyh-Shyun; Lee, Min-Min; Deng, Jeng-Shyan; Huang, Guan-Jhong

    2015-04-01

    Cardamonin is a chalcone isolated from Alpinia katsumadai. This study is aimed to evaluate treatment of cardamonin decreased the paw edema at the 5th hour after λ-carrageenan (Carr) administration and increased the activities of catalase (CAT) and superoxide dismutase (SOD) in the anti-inflammatory test. We also demonstrated that cardamonin significantly attenuated the malondialdehyde (MDA) level in the edema paw at the 5th hour after Carr injection. Cardamonin decreased the nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-6 levels on the serum level at the 5th hour after Carr injection. Western blotting revealed that cardamonin decreased Carr-induced inducible nitric oxide synthase (iNOS), cycloxyclase (COX-2), nuclear factor-κB (NF-κB) and MAPK [extracellular signal-regulated protein kinase (ERK), c-Jun NH(2)-terminal kinase (JNK), p38] expressions and increased heme oxygenase-1 (HO-1) expressions at the 5th hour in the edema paw. The anti-inflammatory mechanisms of cardamonin might be related to the decrease in the level of MDA, iNOS, COX-2, NF-κB, and MAPK and induction of the HO-1 expression in the edema paw via increasing the activities of CAT and SOD in the edema paw through the suppression of NO, TNF-α, IL-1β, and IL-6.

  18. FSH-induced p38-MAPK-mediated dephosphorylation at serine 727 of the signal transducer and activator of transcription 1 decreases Cyp1b1 expression in mouse granulosa cells.

    PubMed

    Du, Xue-Hai; Zhou, Xiao-Long; Cao, Rui; Xiao, Peng; Teng, Yun; Ning, Cai-Bo; Liu, Hong-Lin

    2015-01-01

    Most mammalian follicles undergo atresia at various stages before ovulation, and granulosa cell apoptosis is a major cause of antral follicular atresia. Estradiol is an essential mitogen for granulosa cell proliferation in vivo and inhibition of apoptosis. The estradiol-producing capacity and metabolism levels are important for follicle health, and sufficient estradiol is necessary for follicle development and ovulation. Cyp1b1, a member of the cytochrome P450 1 subfamily, is responsible for the metabolism of a wide variety of halogenated and polycyclic aromatic hydrocarbons in diverse tissues. In mouse follicles, Cyp1b1 converts estradiol to 4-hydroxyestradiol. We investigated mouse granulosa cells (MGCs) in vivo and in vitro and found that Cyp1b1 played a crucial role in estradiol metabolism in dominant follicles. Follicle-stimulating hormone (FSH) decreased estrogen metabolism by reducing Cyp1b1 mRNA and protein levels in MGCs. Furthermore, FSH regulated signal transducer and activator of transcription 1 (STAT1), a significant transcription factor of Cyp1b1, by mediating the dephosphorylation of STAT1 on serine 727 (Ser(727)) in MGCs. p38 mitogen-activated protein kinase (MAPK) may be involved in the FSH-induced dephosphorylation of STAT1 on Ser(727) in MGCs. These results suggested that FSH functions via p38 MAPK-induced dephosphorylation at Ser(727) of STAT1 to downregulate Cyp1b1 expression and maintain the estradiol levels in mouse dominant follicles.

  19. EGFR/Ras/MAPK signaling mediates adult midgut epithelial homeostasis and regeneration in Drosophila

    PubMed Central

    Jiang, Huaqi; Grenley, Marc O.; Bravo, Maria-Jose; Blumhagen, Rachel Z.; Edgar, Bruce A.

    2010-01-01

    Many tissues in higher animals undergo dynamic homeostatic growth, wherein damaged or aged cells are replaced by the progeny of resident stem cells. To maintain homeostasis, stem cells must respond to tissue needs. Here we show that in response to damage or stress in the intestinal (midgut) epithelium of adult Drosophila, multiple EGFR ligands and rhomboids (intramembrane proteases that activate some EGFR ligands) are induced, leading to the activation of EGFR signaling in intestinal stem cells (ISCs). Activation of EGFR signaling promotes ISC division and midgut epithelium regeneration, thus maintaining tissue homeostasis. ISCs defective in EGFR signaling cannot grow or divide, are poorly maintained, and cannot support midgut epithelium regeneration following enteric infection by the bacterium, Pseudomonas entomophila. Furthermore, ISC proliferation induced by Jak/Stat signaling is dependent upon EGFR signaling. Thus the EGFR/Ras/MAPK signaling pathway plays central, essential roles in ISC maintenance and the feedback system that mediates intestinal homeostasis. PMID:21167805

  20. Hypoxia Drives Breast Tumor Malignancy through a TET-TNFα-p38-MAPK Signaling Axis.

    PubMed

    Wu, Min-Zu; Chen, Su-Feng; Nieh, Shin; Benner, Christopher; Ger, Luo-Ping; Jan, Chia-Ing; Ma, Li; Chen, Chien-Hung; Hishida, Tomoaki; Chang, Hong-Tai; Lin, Yaoh-Shiang; Montserrat, Nuria; Gascon, Pedro; Sancho-Martinez, Ignacio; Izpisua Belmonte, Juan Carlos

    2015-09-15

    Hypoxia is a hallmark of solid tumors that drives malignant progression by altering epigenetic controls. In breast tumors, aberrant DNA methylation is a prevalent epigenetic feature associated with increased risk of metastasis and poor prognosis. However, the mechanism by which hypoxia alters DNA methylation or other epigenetic controls that promote breast malignancy remains poorly understood. We discovered that hypoxia deregulates TET1 and TET3, the enzymes that catalyze conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), thereby leading to breast tumor-initiating cell (BTIC) properties. TET1/3 and 5hmC levels were closely associated with tumor hypoxia, tumor malignancy, and poor prognosis in breast cancer patients. Mechanistic investigations showed that hypoxia leads to genome-wide changes in DNA hydroxymethylation associated with upregulation of TNFα expression and activation of its downstream p38-MAPK effector pathway. Coordinate functions of TET1 and TET3 were also required to activate TNFα-p38-MAPK signaling as a response to hypoxia. Our results reveal how signal transduction through the TET-TNFα-p38-MAPK signaling axis is required for the acquisition of BTIC characteristics and tumorigenicity in vitro and in vivo, with potential implications for how to eradicate BTIC as a therapeutic strategy. PMID:26294212

  1. Signals from the AT2 (angiotensin type 2) receptor of angiotensin II inhibit p21ras and activate MAPK (mitogen-activated protein kinase) to induce morphological neuronal differentiation in NG108-15 cells.

    PubMed

    Gendron, L; Laflamme, L; Rivard, N; Asselin, C; Payet, M D; Gallo-Payet, N

    1999-09-01

    In a previous study, we had shown that activation of the AT2 (angiotensin type 2) receptor of angiotensin II (Ang II) induced morphological differentiation of the neuronal cell line NG108-15. In the present study, we investigated the nature of the possible intracellular mediators involved in the AT2 effect. We found that stimulation of AT2 receptors in NG108-15 cells resulted in time-dependent modulation of tyrosine phosphorylation of a number of cytoplasmic proteins. Stimulation of NG108-15 cells with Ang II induced a decrease in GTP-bound p21ras but a sustained increase in the activity of p42mapk and p44mapk as well as neurite outgrowth. Similarly, neurite elongation, increased polymerized tubulin levels, and increased mitogen-activated protein kinase (MAPK) activity were also observed in a stably transfected NG108-15 cell line expressing the dominant-negative mutant of p21ras, RasN17. These results support the observation that inhibition of p21ras did not impair the effect of Ang II on its ability to stimulate MAPK activity. While 10 microM of the MEK inhibitor, PD98059, only moderately affected elongation, 50 microM PD98059 completely blocked the Ang II- and the RasN17-mediated induction of neurite outgrowth. These results demonstrate that some of the events associated with the AT2 receptor-induced neuronal morphological differentiation of NG108-15 cells not only include inhibition of p21ras but an increase in MAPK activity as well, which is essential for neurite outgrowth.

  2. Nucleophosmin and Nucleolin Regulate K-Ras Plasma Membrane Interactions and MAPK Signal Transduction*

    PubMed Central

    Inder, Kerry L.; Lau, Chiyan; Loo, Dorothy; Chaudhary, Natasha; Goodall, Andrew; Martin, Sally; Jones, Alun; van der Hoeven, Dharini; Parton, Robert G.; Hill, Michelle M.; Hancock, John F.

    2009-01-01

    The spatial organization of Ras proteins into nanoclusters on the inner leaflet of the plasma membrane is essential for high fidelity signaling through the MAPK pathway. Here we identify two selective regulators of K-Ras nanoclustering from a proteomic screen for K-Ras interacting proteins. Nucleophosmin (NPM) and nucleolin are predominantly localized to the nucleolus but also have extranuclear functions. We show that a subset of NPM and nucleolin localizes to the inner leaflet of plasma membrane and forms specific complexes with K-Ras but not other Ras isoforms. Active GTP-loaded and inactive GDP-loaded K-Ras both interact with NPM, although NPM-K-Ras binding is increased by growth factor receptor activation. NPM and nucleolin both stabilize K-Ras levels on the plasma membrane, but NPM concurrently increases the clustered fraction of GTP-K-Ras. The increase in nanoclustered GTP-K-Ras in turn enhances signal gain in the MAPK pathway. In summary these results reveal novel extranucleolar functions for NPM and nucleolin as regulators of K-Ras nanocluster formation and activation of the MAPK pathway. The study also identifies a new class of K-Ras nanocluster regulator that operates independently of the structural scaffold galectin-3. PMID:19661056

  3. MAPK signaling cascades mediate distinct glucocorticoid resistance mechanisms in pediatric leukemia

    PubMed Central

    Jones, Courtney L.; Gearheart, Christy M.; Fosmire, Susan; Delgado-Martin, Cristina; Evensen, Nikki A.; Bride, Karen; Waanders, Angela J.; Pais, Faye; Wang, Jinhua; Bhatla, Teena; Bitterman, Danielle S.; de Rijk, Simone R.; Bourgeois, Wallace; Dandekar, Smita; Park, Eugene; Burleson, Tamara M.; Madhusoodhan, Pillai Pallavi; Teachey, David T.; Raetz, Elizabeth A.; Hermiston, Michelle L.; Müschen, Markus; Loh, Mignon L.; Hunger, Stephen P.; Zhang, Jinghui; Garabedian, Michael J.; Porter, Christopher C.

    2015-01-01

    The outcome for pediatric acute lymphoblastic leukemia (ALL) patients who relapse is dismal. A hallmark of relapsed disease is acquired resistance to multiple chemotherapeutic agents, particularly glucocorticoids. In this study, we performed a genome-scale short hairpin RNA screen to identify mediators of prednisolone sensitivity in ALL cell lines. The incorporation of these data with an integrated analysis of relapse-specific genetic and epigenetic changes allowed us to identify the mitogen-activated protein kinase (MAPK) pathway as a mediator of prednisolone resistance in pediatric ALL. We show that knockdown of the specific MAPK pathway members MEK2 and MEK4 increased sensitivity to prednisolone through distinct mechanisms. MEK4 knockdown increased sensitivity specifically to prednisolone by increasing the levels of the glucocorticoid receptor. MEK2 knockdown increased sensitivity to all chemotherapy agents tested by increasing the levels of p53. Furthermore, we demonstrate that inhibition of MEK1/2 with trametinib increased sensitivity of ALL cells and primary samples to chemotherapy in vitro and in vivo. To confirm a role for MAPK signaling in patients with relapsed ALL, we measured the activation of the MEK1/2 target ERK in matched diagnosis-relapse primary samples and observed increased phosphorylated ERK levels at relapse. Furthermore, relapse samples have an enhanced response to MEK inhibition compared to matched diagnosis samples in xenograft models. Together, our data indicate that inhibition of the MAPK pathway increases chemosensitivity to glucocorticoids and possibly other agents and that the MAPK pathway is an attractive target for prevention and/or treatment of relapsed disease. PMID:26324703

  4. MAPK signaling cascades mediate distinct glucocorticoid resistance mechanisms in pediatric leukemia.

    PubMed

    Jones, Courtney L; Gearheart, Christy M; Fosmire, Susan; Delgado-Martin, Cristina; Evensen, Nikki A; Bride, Karen; Waanders, Angela J; Pais, Faye; Wang, Jinhua; Bhatla, Teena; Bitterman, Danielle S; de Rijk, Simone R; Bourgeois, Wallace; Dandekar, Smita; Park, Eugene; Burleson, Tamara M; Madhusoodhan, Pillai Pallavi; Teachey, David T; Raetz, Elizabeth A; Hermiston, Michelle L; Müschen, Markus; Loh, Mignon L; Hunger, Stephen P; Zhang, Jinghui; Garabedian, Michael J; Porter, Christopher C; Carroll, William L

    2015-11-01

    The outcome for pediatric acute lymphoblastic leukemia (ALL) patients who relapse is dismal. A hallmark of relapsed disease is acquired resistance to multiple chemotherapeutic agents, particularly glucocorticoids. In this study, we performed a genome-scale short hairpin RNA screen to identify mediators of prednisolone sensitivity in ALL cell lines. The incorporation of these data with an integrated analysis of relapse-specific genetic and epigenetic changes allowed us to identify the mitogen-activated protein kinase (MAPK) pathway as a mediator of prednisolone resistance in pediatric ALL. We show that knockdown of the specific MAPK pathway members MEK2 and MEK4 increased sensitivity to prednisolone through distinct mechanisms. MEK4 knockdown increased sensitivity specifically to prednisolone by increasing the levels of the glucocorticoid receptor. MEK2 knockdown increased sensitivity to all chemotherapy agents tested by increasing the levels of p53. Furthermore, we demonstrate that inhibition of MEK1/2 with trametinib increased sensitivity of ALL cells and primary samples to chemotherapy in vitro and in vivo. To confirm a role for MAPK signaling in patients with relapsed ALL, we measured the activation of the MEK1/2 target ERK in matched diagnosis-relapse primary samples and observed increased phosphorylated ERK levels at relapse. Furthermore, relapse samples have an enhanced response to MEK inhibition compared to matched diagnosis samples in xenograft models. Together, our data indicate that inhibition of the MAPK pathway increases chemosensitivity to glucocorticoids and possibly other agents and that the MAPK pathway is an attractive target for prevention and/or treatment of relapsed disease.

  5. Thrombin Enhances NGF-Mediated Neurite Extension via Increased and Sustained Activation of p44/42 MAPK and p38 MAPK

    PubMed Central

    Mufti, Rania E.; Sarker, Krishna; Jin, Yan; Fu, Songbin; Rosales, Jesusa L.; Lee, Ki-Young

    2014-01-01

    Rapid neurite remodeling is fundamental to nervous system development and plasticity. It involves neurite extension that is regulated by NGF through PI3K/AKT, p44/42 MAPK and p38 MAPK. It also involves neurite retraction that is regulated by the serine protease, thrombin. However, the intracellular signaling pathway by which thrombin causes neurite retraction is unknown. Using the PC12 neuronal cell model, we demonstrate that thrombin utilizes the PI3K/AKT pathway for neurite retraction in NGF-differentiated cells. Interestingly, however, we found that thrombin enhances NGF-induced neurite extension in differentiating cells. This is achieved through increased and sustained activation of p44/42 MAPK and p38 MAPK. Thus, thrombin elicits opposing effects in differentiated and differentiating cells through activation of distinct signaling pathways: neurite retraction in differentiated cells via PI3K/AKT, and neurite extension in differentiating cells via p44/42 MAPK and p38 MAPK. These findings, which also point to a novel cooperative role between thrombin and NGF, have significant implications in the development of the nervous system and the disease processes that afflicts it as well as in the potential of combined thrombin and NGF therapy for impaired learning and memory, and spinal cord injury which all require neurite extension and remodeling. PMID:25061982

  6. Multi-Compartmentalisation in the MAPK Signalling Pathway Contributes to the Emergence of Oscillatory Behaviour and to Ultrasensitivity.

    PubMed

    Shuaib, Aban; Hartwell, Adam; Kiss-Toth, Endre; Holcombe, Mike

    2016-01-01

    Signal transduction through the Mitogen Activated Protein Kinase (MAPK) pathways is evolutionarily highly conserved. Many cells use these pathways to interpret changes to their environment and respond accordingly. The pathways are central to triggering diverse cellular responses such as survival, apoptosis, differentiation and proliferation. Though the interactions between the different MAPK pathways are complex, nevertheless, they maintain a high level of fidelity and specificity to the original signal. There are numerous theories explaining how fidelity and specificity arise within this complex context; spatio-temporal regulation of the pathways and feedback loops are thought to be very important. This paper presents an agent based computational model addressing multi-compartmentalisation and how this influences the dynamics of MAPK cascade activation. The model suggests that multi-compartmentalisation coupled with periodic MAPK kinase (MAPKK) activation may be critical factors for the emergence of oscillation and ultrasensitivity in the system. Finally, the model also establishes a link between the spatial arrangements of the cascade components and temporal activation mechanisms, and how both contribute to fidelity and specificity of MAPK mediated signalling. PMID:27243235

  7. Multi-Compartmentalisation in the MAPK Signalling Pathway Contributes to the Emergence of Oscillatory Behaviour and to Ultrasensitivity

    PubMed Central

    Shuaib, Aban; Hartwell, Adam; Kiss-Toth, Endre; Holcombe, Mike

    2016-01-01

    Signal transduction through the Mitogen Activated Protein Kinase (MAPK) pathways is evolutionarily highly conserved. Many cells use these pathways to interpret changes to their environment and respond accordingly. The pathways are central to triggering diverse cellular responses such as survival, apoptosis, differentiation and proliferation. Though the interactions between the different MAPK pathways are complex, nevertheless, they maintain a high level of fidelity and specificity to the original signal. There are numerous theories explaining how fidelity and specificity arise within this complex context; spatio-temporal regulation of the pathways and feedback loops are thought to be very important. This paper presents an agent based computational model addressing multi-compartmentalisation and how this influences the dynamics of MAPK cascade activation. The model suggests that multi-compartmentalisation coupled with periodic MAPK kinase (MAPKK) activation may be critical factors for the emergence of oscillation and ultrasensitivity in the system. Finally, the model also establishes a link between the spatial arrangements of the cascade components and temporal activation mechanisms, and how both contribute to fidelity and specificity of MAPK mediated signalling. PMID:27243235

  8. Mitophagy is primarily due to alternative autophagy and requires the MAPK1 and MAPK14 signaling pathways.

    PubMed

    Hirota, Yuko; Yamashita, Shun-ichi; Kurihara, Yusuke; Jin, Xiulian; Aihara, Masamune; Saigusa, Tetsu; Kang, Dongchon; Kanki, Tomotake

    2015-01-01

    In cultured cells, not many mitochondria are degraded by mitophagy induced by physiological cellular stress. We observed mitophagy in HeLa cells using a method that relies on the pH-sensitive fluorescent protein Keima. With this approach, we found that mitophagy was barely induced by carbonyl cyanide m-chlorophenyl hydrazone treatment, which is widely used as an inducer of PARK2/Parkin-related mitophagy, whereas a small but modest amount of mitochondria were degraded by mitophagy under conditions of starvation or hypoxia. Mitophagy induced by starvation or hypoxia was marginally suppressed by knockdown of ATG7 and ATG12, or MAP1LC3B, which are essential for conventional macroautophagy. In addition, mitophagy was efficiently induced in Atg5 knockout mouse embryonic fibroblasts. However, knockdown of RAB9A and RAB9B, which are essential for alternative autophagy, but not conventional macroautophagy, severely suppressed mitophagy. Finally, we found that the MAPKs MAPK1/ERK2 and MAPK14/p38 were required for mitophagy. Based on these findings, we conclude that mitophagy in mammalian cells predominantly occurs through an alternative autophagy pathway, requiring the MAPK1 and MAPK14 signaling pathways.

  9. 2-O Heparan Sulfate Sulfation by Hs2st Is Required for Erk/Mapk Signalling Activation at the Mid-Gestational Mouse Telencephalic Midline.

    PubMed

    Chan, Wai Kit; Howe, Katherine; Clegg, James M; Guimond, Scott E; Price, David J; Turnbull, Jeremy E; Pratt, Thomas

    2015-01-01

    Heparan sulfate (HS) is a linear carbohydrate composed of polymerized uronate-glucosamine disaccharide units that decorates cell surface and secreted glycoproteins in the extracellular matrix. In mammals HS is subjected to differential sulfation by fifteen different heparan sulfotransferase (HST) enzymes of which Hs2st uniquely catalyzes the sulfation of the 2-O position of the uronate in HS. HS sulfation is postulated to be important for regulation of signaling pathways by facilitating the interaction of HS with signaling proteins including those of the Fibroblast Growth Factor (Fgf) family which signal through phosphorylation of extracellular signal-regulated kinases Erk1/2. In the developing mouse telencephalon Fgf2 signaling regulates proliferation and neurogenesis. Loss of Hs2st function phenocopies the thinned cerebral cortex of mutant mice in which Fgf2 or Erk1/2 function are abrogated, suggesting the hypothesis that 2-O-sulfated HS structures play a specific role in Fgf2/Erk signaling pathway in this context in vivo. This study investigated the molecular role of 2-O sulfation in Fgf2/Erk signaling in the developing telencephalic midline midway through mouse embryogenesis at E12.5. We examined the expression of Hs2st, Fgf2, and Erk1/2 activity in wild-type and Hs2st-/- mice. We found that Hs2st is expressed at high levels at the midline correlating with high levels of Erk1/2 activation and Erk1/2 activation was drastically reduced in the Hs2st-/- mutant at the rostral telencephalic midline. We also found that 2-O sulfation is specifically required for the binding of Fgf2 protein to Fgfr1, its major cell-surface receptor at the rostral telencephalic midline. We conclude that 2-O sulfated HS structures generated by Hs2st are needed to form productive signaling complexes between HS, Fgf2 and Fgfr1 that activate Erk1/2 at the midline. Overall, our data suggest the interesting possibility that differential expression of Hs2st targets the rostral telencephalic

  10. LPS promote the odontoblastic differentiation of human dental pulp stem cells via MAPK signaling pathway.

    PubMed

    He, Wenxi; Wang, Zhihua; Luo, Zhirong; Yu, Qing; Jiang, Yong; Zhang, Yaqing; Zhou, Zeyuan; Smith, Anthony J; Cooper, Paul R

    2015-03-01

    Human dental pulp stem cells (hDPSCs) show significant potential for exploitation in novel regeneration strategies, although lack of understanding of their responses to bacterial challenge constrains their application. The present study aimed to investigate whether lipopolysaccharide (LPS), the major pathogenic factor of Gram-negative bacteria, regulates the differentiation of hDPSCs and which intracellular signaling pathways may be involved. LPS treatment significantly promoted the differentiation of hDPSCs demonstrable by increased mineralized nodule formation and mRNA expression of several odontoblastic markers in a dose-dependent manner. While inhibition of TLR4, p38, and ERK signaling markedly antagonized LPS-mediated differentiation of hDPSCs. The inhibition of JNK and NF-κB signaling had no detectable effect on LPS activation of hDPSCs. LPS stimulation resulted in phosphorylation of NF-κB p65, IκB-α, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK) in DPSCs in a time-dependent manner, which was markedly suppressed by their specific inhibitors, respectively. Data demonstrated that LPS promoted odontoblastic differentiation of hDPSCs via TLR4, ERK, and P38 MAPK signaling pathways, but not NF-κB signaling.

  11. Inhibition of MAPK by Prolactin Signaling through the Short Form of Its Receptor in the Ovary and Decidua

    PubMed Central

    Devi, Y. Sangeeta; Seibold, Anita M.; Shehu, Aurora; Maizels, Evelyn; Halperin, Julia; Le, Jamie; Binart, Nadine; Bao, Lei; Gibori, Geula

    2011-01-01

    Prolactin (PRL) is essential for normal reproduction and signals through two types of receptors, the short (PRL-RS) and long (PRL-RL) form. We have previously shown that transgenic mice expressing only PRL-RS (PRLR−/−RS) display abnormal follicular development and premature ovarian failure. Here, we report that MAPK, essential for normal follicular development, is critically inhibited by PRL in reproductive tissues of PRLR−/−RS mice. Consequently, the phosphorylation of MAPK downstream targets are also markedly inhibited by PRL without affecting immediate upstream kinases, suggesting involvement of MAPK specific phosphatase(s) in this inhibition. Similar results are obtained in a PRL-responsive ovary-derived cell line (GG-CL) that expresses only PRL-RS. However, we found the expression/activation of several known MAPK phosphatases not to be affected by PRL, suggesting a role of unidentified phosphatase(s). We detected a 27-kDa protein that binds to the intracellular domain of PRL-RS and identified it as dual specific phosphatase DUPD1. PRL does not induce expression of DUDP1 but represses its phosphorylation on Thr-155. We also show a physical association of this phosphatase with ERK1/2 and p38 MAPK. Using an in vitro phosphatase assay and overexpression studies, we established that DUPD1 is a MAPK phosphatase. Dual specific phosphatase inhibitors as well as siRNA to DUPD1, completely prevent PRL-mediated MAPK inhibition in ovarian cells. Our results strongly suggest that deactivation of MAPK by PRL/PRL-RS contributes to the severe ovarian defect in PRLR−/−RS mice and demonstrate the novel association of PRL-RS with DUPD1 and a role for this phosphatase in MAPK deactivation. PMID:21199871

  12. Hepatitis C virus E2 protein promotes human hepatoma cell proliferation through the MAPK/ERK signaling pathway via cellular receptors

    SciTech Connect

    Zhao Lanjuan; Wang Lu; Ren Hao; Cao Jie; Li Li; Ke Jinshan; Qi Zhongtian . E-mail: qizt53@hotmail.com

    2005-04-15

    Dysregulation of mitogen-activated protein kinase (MAPK) signaling pathways by various viruses has been shown to be responsible for viral pathogenicity. The molecular mechanism by which hepatitis C virus (HCV) infection caused human liver diseases has been investigated on the basis of abnormal intracellular signal events. Current data are very limited involved in transmembrane signal transduction triggered by HCV E2 protein. Here we explored regulation of the MAPK/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway by E2 expressed in Chinese hamster oval cells. In human hepatoma Huh-7 cells, E2 specifically activated the MAPK/ERK pathway including downstream transcription factor ATF-2 and greatly promoted cell proliferation. CD81 and low density lipoprotein receptor (LDLR) on the cell surface mediated binding of E2 to Huh-7 cells. The MAPK/ERK activation and cell proliferation driven by E2 were suppressed by blockage of CD81 as well as LDLR. Furthermore, pretreatment with an upstream kinase MEK1/2 inhibitor U0126 also impaired the MAPK/ERK activation and cell proliferation induced by E2. Our results suggest that the MAPK/ERK signaling pathway triggered by HCV E2 via its receptors maintains survival and growth of target cells.

  13. Xiang-Qi-Tang and its active components exhibit anti-inflammatory and anticoagulant properties by inhibiting MAPK and NF-κB signaling pathways in LPS-treated rat cardiac microvascular endothelial cells.

    PubMed

    He, Chang-Liang; Yi, Peng-Fei; Fan, Qiao-Jia; Shen, Hai-Qing; Jiang, Xiao-Lin; Qin, Qian-Qian; Song, Zhou; Zhang, Cui; Wu, Shuai-Cheng; Wei, Xu-Bin; Li, Ying-Lun; Fu, Ben-Dong

    2013-04-01

    Xiang-Qi-Tang (XQT) is a Chinese herbal formula containing Cyperus rotundus, Astragalus membranaceus and Andrographis paniculata. Alpha-Cyperone (CYP), astragaloside IV (AS-IV) and andrographolide (AND) are the three major active components in this formula. XQT may modulate the inflammatory or coagulant responses. We therefore assessed the effects of XQT on lipopolysaccharide (LPS)-induced inflammatory model of rat cardiac microvascular endothelial cells (RCMECs). XQT, CYP, AS-IV and AND inhibited the production of tumor necrosis factor alpha (TNF-α), intercellular cell adhesion molecule-1 (ICAM-1) and plasminogen activator inhibitor-1 (PAI-1), and up-regulated the mRNA expression of Kruppel-like factor 2 (KLF2). XQT and CYP inhibited the secretion of tissue factor (TF). To further explore the mechanism, we found that XQT, or its active components CYP, AS-IV and AND significantly inhibited extracellular signal-regulated kinase (ERK), c-jun NH2-terminal kinase (JNK) and p38 phosphorylation protein expression as well as decreased the phosphorylation levels of nuclear factor κB (NF-κB) p65 proteins in LPS-stimulated RCMECs. These results suggested that XQT and its active components inhibited the expression of inflammatory and coagulant mediators via mitogen-activated protein kinase (MAPKs) and NF-κB signaling pathways. These findings may contribute to future research on the action mechanisms of this formula, as well as therapy for inflammation- or coagulation-related diseases. PMID:23171279

  14. Inhibition of Raf/MAPK signaling in Xenopus oocyte extracts by Raf-1-specific peptides.

    PubMed

    Radziwill, G; Steinhusen, U; Aitken, A; Moelling, K

    1996-10-01

    Raf-1 is an upstream element of the mitogen-activated protein kinase (MAPK) pathway which leads to cell proliferation and differentiation. In this study Raf-1 derived peptides comprising the conserved amino acid residues Arg89 and Ser259, involved in binding of activated Ras and 14-3-3 proteins, respectively, were shown to interfere with MAPK activation in extracts from immature Xenopus oocytes. Lipids prepared from oocyte extracts can stimulate MAPK in a Ras- and protein kinase C-independent manner. This lipid-induced MAPK activation is blocked by a Raf-1 derived peptide comprising Ser259.

  15. Skb5, an SH3 adaptor protein, regulates Pmk1 MAPK signaling by controlling the intracellular localization of the MAPKKK Mkh1.

    PubMed

    Kanda, Yuki; Satoh, Ryosuke; Matsumoto, Saki; Ikeda, Chisato; Inutsuka, Natsumi; Hagihara, Kanako; Matzno, Sumio; Tsujimoto, Sho; Kita, Ayako; Sugiura, Reiko

    2016-08-15

    The mitogen-activated protein kinase (MAPK) cascade is a highly conserved signaling module composed of MAPK kinase kinases (MAPKKKs), MAPK kinases (MAPKK) and MAPKs. The MAPKKK Mkh1 is an initiating kinase in Pmk1 MAPK signaling, which regulates cell integrity in fission yeast (Schizosaccharomyces pombe). Our genetic screen for regulators of Pmk1 signaling identified Shk1 kinase binding protein 5 (Skb5), an SH3-domain-containing adaptor protein. Here, we show that Skb5 serves as an inhibitor of Pmk1 MAPK signaling activation by downregulating Mkh1 localization to cell tips through its interaction with the SH3 domain. Consistent with this, the Mkh1(3PA) mutant protein, with impaired Skb5 binding, remained in the cell tips, even when Skb5 was overproduced. Intriguingly, Skb5 needs Mkh1 to localize to the growing ends as Mkh1 deletion and disruption of Mkh1 binding impairs Skb5 localization. Deletion of Pck2, an upstream activator of Mkh1, impaired the cell tip localization of Mkh1 and Skb5 as well as the Mkh1-Skb5 interaction. Interestingly, both Pck2 and Mkh1 localized to the cell tips at the G1/S phase, which coincided with Pmk1 MAPK activation. Taken together, Mkh1 localization to cell tips is important for transmitting upstream signaling to Pmk1, and Skb5 spatially regulates this process. PMID:27451356

  16. Modulation of the Ras/MAPK signalling pathway by the redox function of selenoproteins in Drosophila melanogaster.

    PubMed

    Morey, M; Serras, F; Baguñà, J; Hafen, E; Corominas, M

    2001-10-01

    Modulation of reactive oxygen species (ROS) plays a key role in signal transduction pathways. Selenoproteins act controlling the redox balance of the cell. We have studied how the alteration of the redox balance caused by patufet (selD(ptuf)), a null mutation in the Drosophila melanogaster selenophosphate synthetase 1 (sps1) gene, which codes for the SelD enzyme of the selenoprotein biosynthesis, affects the Ras/MAPK signalling pathway. The selD(ptuf) mutation dominantly suppresses the phenotypes in the eye and the wing caused by hyperactivation of the Ras/MAPK cassette and the activated forms of the Drosophila EGF receptor (DER) and Sevenless (Sev) receptor tyrosine kinases (RTKs), which signal in the eye and wing, respectively. No dominant interaction is observed with sensitized conditions in the Wnt, Notch, Insulin-Pi3K, and DPP signalling pathways. Our current hypothesis is that selenoproteins selectively modulate the Ras/MAPK signalling pathway through their antioxidant function. This is further supported by the fact that a selenoprotein-independent increase in ROS caused by the catalase amorphic Cat(n1) allele also reduces Ras/MAPK signalling. Here, we present the first evidence for the role of intracellular redox environment in signalling pathways in Drosophila as a whole organism.

  17. Evaluating the Activity of the Filamentous Growth MAPK Pathway in Yeast

    PubMed Central

    Cullen, Paul J.

    2015-01-01

    Mitogen-activated protein kinase (MAPK) pathways are evolutionarily conserved signaling pathways that regulate diverse processes in eukaryotes. One such pathway regulates filamentous growth, a nutrient limitation response in budding yeast and other fungal species. This protocol describes three assays used to measure the activity of the filamentous growth pathway. First, western blotting for phosphorylated (activated) MAPKs (P~MAPKs; Slt2p, Kss1p, Fus3p, and Hog1p) provides a measure of MAPK activity in yeast and other fungal species. Second, the PGU1 gene is a transcriptional target of the filamentous growth pathway. Cells that undergo filamentous growth secrete Pgu1, an endopolygalacturonase that degrades the plant-specific polysaccharide pectin. We describe an assay that measures secreted pectinase activity, which reflects an active filamentous growth pathway. Finally, in yeast, two mucin-like glycoproteins, Msb2 and Flo11, regulate filamentous growth. Secretion of the processed and shed glycodomain of Msb2 is an indicator of MAPK activity. Flo11, the major adhesion molecule that controls filamentous growth and biofilm/mat formation, is also shed from cells. Detecting shed mucins with epitope-tagged versions of the proteins (secretion profiling) provides information about the regulation of filamentous growth across fungal species. PMID:25734070

  18. Intracellular signaling via ERK/MAPK completes the pathway for tubulogenic fibronectin in MDCK cells.

    PubMed

    Liu, Zhao; Greco, Andres J; Hellman, Nathan E; Spector, June; Robinson, Jonathan; Tang, Oliver T; Lipschutz, Joshua H

    2007-02-16

    A classic in vitro model of branching morphogenesis utilizes the Madin-Darby canine kidney (MDCK) cell line. MDCK Strain II cells form hollow monoclonal cysts in a three-dimensional collagen matrix over the course of 10 days and tubulate in response to hepatocyte growth factor (HGF). We and our colleagues previously showed that activation of the extracellular-signal regulated kinase (ERK, aka MAPK) pathway is necessary and sufficient to induce tubulogenesis in MDCK cells. We also showed in a microarray study that one of the genes upregulated by HGF was the known tubulogene fibronectin. Given that HGF activates a multitude of signaling pathways, including ERK/MAPK, to test the intracellular regulatory pathway, we used two distinct inhibitors of ERK activation (U0126 and PD098059). Following induction of MDCK Type II cells with HGF, tubulogenic fibronectin mRNA was upregulated fourfold by real-time PCR, and minimal or no change in fibronectin expression was seen when HGF was added with either U0126 or PD098059. We confirmed these results using an MDCK cell line inducible for Raf, which is upstream of ERK. Following activation of Raf, fibronectin mRNA and protein expression were increased to a similar degree as was seen following HGF induction. Furthermore, MDCK Strain I cells, which originate from collecting ducts and have constitutively active ERK, spontaneously initiate tubulogenesis. We show here that MDCK Strain I cells have high levels of fibronectin mRNA and protein compared to MDCK Strain II cells. When U0126 and PD098059 were added to MDCK Strain I cells, fibronectin mRNA, and protein levels were decreased to levels seen in MDCK Strain II cells. These data allow us to complete what we believe is the first description of a tubulogenic pathway from receptor/ligand (HGF/CMET), through an intracellular signaling pathway (ERK/MAPK), to transcription and, finally, secretion of a critical tubuloprotein (fibronectin).

  19. Induction of Macrophage Function in Human THP-1 Cells Is Associated with Rewiring of MAPK Signaling and Activation of MAP3K7 (TAK1) Protein Kinase

    PubMed Central

    Richter, Erik; Ventz, Katharina; Harms, Manuela; Mostertz, Jörg; Hochgräfe, Falko

    2016-01-01

    Macrophages represent the primary human host response to pathogen infection and link the immediate defense to the adaptive immune system. Mature tissue macrophages convert from circulating monocyte precursor cells by terminal differentiation in a process that is not fully understood. Here, we analyzed the protein kinases of the human monocytic cell line THP-1 before and after induction of macrophage differentiation by using kinomics and phosphoproteomics. When comparing the macrophage-like state with the monocytic precursor, 50% of the kinome was altered in expression and even 71% of covered kinase phosphorylation sites were affected. Kinome rearrangements are for example characterized by a shift of overrepresented cyclin-dependent kinases associated with cell cycle control in monocytes to calmodulin-dependent kinases and kinases involved in proinflammatory signaling. Eventually, we show that monocyte-to-macrophage differentiation is associated with major rewiring of mitogen-activated protein kinase signaling networks and demonstrate that protein kinase MAP3K7 (TAK1) acts as the key signaling hub in bacterial killing, chemokine production and differentiation. Our study proves the fundamental role of protein kinases and cellular signaling as major drivers of macrophage differentiation and function. The finding that MAP3K7 is central to macrophage function suggests MAP3K7 and its networking partners as promising targets in host-directed therapy for macrophage-associated disease. PMID:27066479

  20. Single-Cell Analysis Reveals that Insulation Maintains Signaling Specificity between Two Yeast MAPK Pathways with Common Components

    PubMed Central

    Patterson, Jesse C.; Klimenko, Evguenia S.; Thorner, Jeremy

    2014-01-01

    Eukaryotic cells use multiple mitogen-activated protein kinase (MAPK) cascades to evoke appropriate responses to external stimuli. In Saccharomyces cerevisiae, the MAPK Fus3 is activated by pheromone-binding G protein-coupled receptors to promote mating, whereas the MAPK Hog1 is activated by hyperosmotic stress to elicit the high osmolarity glycerol (HOG) response. Although these MAPK pathways share several upstream components, exposure to either pheromone or osmolyte alone triggers only the appropriate response. We used fluorescent localization- and transcription-specific reporters to assess activation of these pathways in individual cells on the minute and hour timescale, respectively. Dual activation of these two MAPK pathways occurred over a broad range of stimulant concentrations and temporal regimes in wild-type cells subjected to co-stimulation. Thus, signaling specificity is achieved through an “insulation” mechanism, not a “cross-inhibition” mechanism. Furthermore, we showed that there was a critical period during which Hog1 activity had to occur for proper insulation of the HOG pathway. PMID:20959523

  1. Genome-wide survey of yeast mutations leading to activation of the yeast cell integrity MAPK pathway: Novel insights into diverse MAPK outcomes

    PubMed Central

    2011-01-01

    Background The yeast cell wall integrity mitogen-activated protein kinase (CWI-MAPK) pathway is the main regulator of adaptation responses to cell wall stress in yeast. Here, we adopt a genomic approach to shed light on two aspects that are only partially understood, namely, the characterization of the gene functional catalog associated with CWI pathway activation and the extent to which MAPK activation correlates with transcriptional outcomes. Results A systematic yeast mutant deletion library was screened for constitutive transcriptional activation of the CWI-related reporter gene MLP1. Monitoring phospho-Slt2/Mpk1 levels in the identified mutants revealed sixty-four deletants with high levels of phosphorylation of this MAPK, including mainly genes related to cell wall construction and morphogenesis, signaling, and those with unknown function. Phenotypic analysis of the last group of mutants suggests their involvement in cell wall homeostasis. A good correlation between levels of Slt2 phosphorylation and the magnitude of the transcriptional response was found in most cases. However, the expression of CWI pathway-related genes was enhanced in some mutants in the absence of significant Slt2 phosphorylation, despite the fact that functional MAPK signaling through the pathway was required. CWI pathway activation was associated to increased deposition of chitin in the cell wall - a known survival compensatory mechanism - in about 30% of the mutants identified. Conclusion We provide new insights into yeast genes related to the CWI pathway and into how the state of activation of the Slt2 MAPK leads to different outcomes, discovering the versatility of this kind of signaling pathways. These findings potentially have broad implications for understanding the functioning of other eukaryotic MAPKs. PMID:21810245

  2. The Phosphotyrosine Phosphatase SHP-2 Participates in a Multimeric Signaling Complex and Regulates T Cell Receptor (TCR) coupling to the Ras/Mitogen-activated Protein Kinase (MAPK) Pathway in Jurkat T Cells

    PubMed Central

    Frearson, Julie A.; Alexander, Denis R.

    1998-01-01

    Src homology 2 (SH2) domain–containing phosphotyrosine phosphatases (SHPs) are increasingly being shown to play critical roles in protein tyrosine kinase–mediated signaling pathways. The role of SHP-1 as a negative regulator of T cell receptor (TCR) signaling has been established. To further explore the function of the other member of this family, SHP-2, in TCR-mediated events, a catalytically inactive mutant SHP-2 was expressed under an inducible promoter in Jurkat T cells. Expression of the mutant phosphatase significantly inhibited TCR-induced activation of the extracellular-regulated kinase (ERK)-2 member of the mitogen-activated protein kinase (MAPK) family, but had no effect on TCR-ζ chain tyrosine phosphorylation or TCR-elicited Ca2+ transients. Inactive SHP-2 was targeted to membranes resulting in the selective increase in tyrosine phosphorylation of three membrane-associated candidate SHP-2 substrates of 110 kD, 55-60 kD, and 36 kD, respectively. Analysis of immunoprecipitates containing inactive SHP-2 also indicated that the 110-kD and 36-kD Grb-2–associated proteins were putative substrates for SHP-2. TCR-stimulation of Jurkat T cells expressing wild-type SHP-2 resulted in the formation of a multimeric cytosolic complex composed of SHP-2, Grb-2, phosphatidylinositol (PI) 3′-kinase, and p110. A significant proportion of this complex was shown to be membrane associated, presumably as a result of translocation from the cytosol. Catalytically inactive SHP-2, rather than the wild-type PTPase, was preferentially localized in complex with Grb-2 and the p85 subunit of PI 3′-kinase, suggesting that the dephosphorylating actions of SHP-2 may regulate the association of these signaling molecules to the p110 complex. Our results show that SHP-2 plays a critical role in linking the TCR to the Ras/MAPK pathway in Jurkat T cells, and also provide some insight into the molecular interactions of SHP-2 that form the basis of this signal transduction process

  3. Cobalt oxide nanoparticles induced oxidative stress linked to activation of TNF-α/caspase-8/p38-MAPK signaling in human leukemia cells.

    PubMed

    Chattopadhyay, Sourav; Dash, Sandeep Kumar; Tripathy, Satyajit; Das, Balaram; Kar Mahapatra, Santanu; Pramanik, Panchanan; Roy, Somenath

    2015-06-01

    The purpose of this study was to determine the intracellular signaling transduction pathways involved in oxidative stress induced by nanoparticles in cancer cells. Activation of reactive oxygen species (ROS) has some therapeutic benefits in arresting the growth of cancer cells. Cobalt oxide nanoparticles (CoO NPs) are an interesting compound for oxidative cancer therapy. Our results showed that CoO NPs elicited a significant (P <0.05) amount of ROS in cancer cells. Co-treatment with N-aceyltine cystine (an inhibitor of ROS) had a protective role in cancer cell death induced by CoO NPs. In cultured cells, the elevated level of tumor necrosis factor-alpha (TNF-α) was noted after CoO NPs treatment. This TNF-α persuaded activation of caspase-8 followed by phosphorylation of p38 mitogen-activated protein kinase and induced cell death. This study showed that CoO NPs induced oxidative stress and activated the signaling pathway of TNF-α-Caspase-8-p38-Caspase-3 to cancer cells.

  4. Socs36E Controls Niche Competition by Repressing MAPK Signaling in the Drosophila Testis.

    PubMed

    Amoyel, Marc; Anderson, Jason; Suisse, Annabelle; Glasner, Johanna; Bach, Erika A

    2016-01-01

    The Drosophila testis is a well-established system for studying stem cell self-renewal and competition. In this tissue, the niche supports two stem cell populations, germ line stem cells (GSCs), which give rise to sperm, and somatic stem cells called cyst stem cells (CySCs), which support GSCs and their descendants. It has been established that CySCs compete with each other and with GSCs for niche access, and mutations have been identified that confer increased competitiveness to CySCs, resulting in the mutant stem cell and its descendants outcompeting wild type resident stem cells. Socs36E, which encodes a negative feedback inhibitor of the JAK/STAT pathway, was the first identified regulator of niche competition. The competitive behavior of Socs36E mutant CySCs was attributed to increased JAK/STAT signaling. Here we show that competitive behavior of Socs36E mutant CySCs is due in large part to unbridled Mitogen-Activated Protein Kinase (MAPK) signaling. In Socs36E mutant clones, MAPK activity is elevated. Furthermore, we find that clonal upregulation of MAPK in CySCs leads to their outcompetition of wild type CySCs and of GSCs, recapitulating the Socs36E mutant phenotype. Indeed, when MAPK activity is removed from Socs36E mutant clones, they lose their competitiveness but maintain self-renewal, presumably due to increased JAK/STAT signaling in these cells. Consistently, loss of JAK/STAT activity in Socs36E mutant clones severely impairs their self-renewal. Thus, our results enable the genetic separation of two essential processes that occur in stem cells. While some niche signals specify the intrinsic property of self-renewal, which is absolutely required in all stem cells for niche residence, additional signals control the ability of stem cells to compete with their neighbors. Socs36E is node through which these processes are linked, demonstrating that negative feedback inhibition integrates multiple aspects of stem cell behavior. PMID:26807580

  5. Socs36E Controls Niche Competition by Repressing MAPK Signaling in the Drosophila Testis

    PubMed Central

    Amoyel, Marc; Anderson, Jason; Suisse, Annabelle; Glasner, Johanna; Bach, Erika A.

    2016-01-01

    The Drosophila testis is a well-established system for studying stem cell self-renewal and competition. In this tissue, the niche supports two stem cell populations, germ line stem cells (GSCs), which give rise to sperm, and somatic stem cells called cyst stem cells (CySCs), which support GSCs and their descendants. It has been established that CySCs compete with each other and with GSCs for niche access, and mutations have been identified that confer increased competitiveness to CySCs, resulting in the mutant stem cell and its descendants outcompeting wild type resident stem cells. Socs36E, which encodes a negative feedback inhibitor of the JAK/STAT pathway, was the first identified regulator of niche competition. The competitive behavior of Socs36E mutant CySCs was attributed to increased JAK/STAT signaling. Here we show that competitive behavior of Socs36E mutant CySCs is due in large part to unbridled Mitogen-Activated Protein Kinase (MAPK) signaling. In Socs36E mutant clones, MAPK activity is elevated. Furthermore, we find that clonal upregulation of MAPK in CySCs leads to their outcompetition of wild type CySCs and of GSCs, recapitulating the Socs36E mutant phenotype. Indeed, when MAPK activity is removed from Socs36E mutant clones, they lose their competitiveness but maintain self-renewal, presumably due to increased JAK/STAT signaling in these cells. Consistently, loss of JAK/STAT activity in Socs36E mutant clones severely impairs their self-renewal. Thus, our results enable the genetic separation of two essential processes that occur in stem cells. While some niche signals specify the intrinsic property of self-renewal, which is absolutely required in all stem cells for niche residence, additional signals control the ability of stem cells to compete with their neighbors. Socs36E is node through which these processes are linked, demonstrating that negative feedback inhibition integrates multiple aspects of stem cell behavior. PMID:26807580

  6. Characterization of early events involved in human dendritic cell maturation induced by sensitizers: Cross talk between MAPK signalling pathways

    SciTech Connect

    Trompezinski, Sandra; Migdal, Camille; Tailhardat, Magalie; Le Varlet, Beatrice; Courtellemont, Pascal; Haftek, Marek; Serres, Mireille

    2008-08-01

    Dendritic cells (DCs), efficient-antigen presenting cells play an important role in initiating and regulating immune responses. DC maturation following exposure to nickel or DNCB induced an up-regulation of phenotypic markers and inflammatory cytokine secretion. Early intracellular mechanisms involved in DC maturation required to be precise. To address this purpose, DCs derived from human monocytes were treated with sensitizers (nickel, DNCB or thimerosal) in comparison with an irritant (SDS). Our data confirming the up-regulation of CD86, CD54 and cytokine secretion (IL-8 and TNF{alpha}) induced by sensitizers but not by SDS, signalling transduction involved in DC maturation was investigated using these chemicals. Kinase activity measurement was assessed using two new sensitive procedures (Face{sup TM} and CBA) requiring few cells. SDS did not induce changes in signalling pathways whereas NiSO{sub 4}, DNCB and thimerosal markedly activated p38 MAPK and JNK, in contrast Erk1/2 phosphorylation was completely inhibited by DNCB or thimerosal and only activated by nickel. A pre-treatment with p38 MAPK inhibitor (SB203580) suppressed Erk1/2 inhibition induced by DNCB or thimerosal demonstrating a direct interaction between p38 MAPK and Erk1/2. A pre-treatment with an antioxidant, N-acetyl-L-cysteine (NAC) markedly reduced Erk1/2 inhibition and p38 MAPK phosphorylation induced by DNCB and thimerosal, suggesting a direct activation of p38 MAPK via an oxidative stress and a regulation of MAPK signalling pathways depending on chemicals. Because of a high sensitivity of kinase activity measurements, these procedures will be suitable for weak or moderate sensitizer screening.

  7. Resokaempferol-mediated anti-inflammatory effects on activated macrophages via the inhibition of JAK2/STAT3, NF-κB and JNK/p38 MAPK signaling pathways.

    PubMed

    Yu, Qian; Zeng, KeWu; Ma, XiaoLi; Song, FangJiao; Jiang, Yong; Tu, PengFei; Wang, XueMei

    2016-09-01

    The excessive or prolonged production of inflammatory mediators can result in numerous chronic diseases, such as rheumatoid arthritis, atherosclerosis, diabetes, and cancer. Therefore, for many inflammatory-related diseases, pharmaceutical intervention is required to restrain the excessive release of such inflammatory mediators. Novel therapeutics and mechanistic insight are sought for the management of chronic inflammatory diseases. Resokaempferol (RES) is a type of flavonoid recently reported to demonstrate anti-cancer properties. However, the anti-inflammatory capacity of RES has not been studied to date. Therefore, this study investigated whether RES is capable of suppressing the inflammatory response to lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages and the mechanism by which this is achieved. We found that RES attenuated the LPS-induced production of nitric oxide (NO), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), monocyte chemotactic protein 1 (MCP-1) and IL-6. RES also inhibited the nuclear translocation of signal transducer and activator of transcription (STAT) 3 and reduced the LPS-mediated phosphorylation of Janus kinase (JAK) 2 and STAT3 at the sites of Ser727 and Tyr705. RES also inhibited the activation of NF-κB and JNK/p38 MAPK signaling pathways in LPS-induced RAW264.7 cells. Additionally, RES inhibited the activation of the JAK2/STAT3 pathway in exogenous IL-6-activated RAW264.7 macrophages. We conclude that RES inhibits the inflammatory response in activated macrophages by blocking the activation of the JAK2/STAT3 pathway by both LPS and IL-6 signaling. PMID:27261558

  8. Suppression of B-Raf(V600E) cancers by MAPK hyper-activation

    PubMed Central

    Eldad, Sophia; Smeir, Elia; Bar-Tana, Jacob

    2016-01-01

    B-Raf(V600E) activates MEK/MAPK signalling and acts as oncogenic driver of a variety of cancers, including melanoma, colorectal and papillary thyroid carcinoma. Specific B-Raf(V600E) kinase inhibitors (e.g., Vemurafenib) prove initial efficacy in melanoma followed shortly by acquired resistance, while failing in most other B-Raf(V600E) cancers due to primary resistance. Resistance is due to acquired mutations in the Ras/Raf/MEK/MAPK pathway and/or other oncogenic drivers that bypass B-Raf(V600E). Surprisingly, hyper-activation of MAPK by inhibiting its protein phosphatase 2A by a synthetic long-chain fatty acid analogue (MEDICA), results in oncogene-induced growth arrest and apoptosis of B-Raf(V600E) cancer cells. Growth arrest is accompanied by MAPK-mediated serine/threonine phosphorylation and suppression of a variety of oncogenic drivers that resist treatment by B-Raf(V600E) kinase inhibitors, including ErbB members, c-Met, IGFR, IRS, STAT3 and Akt. The combined activities of mutated B-Raf and MEDICA are required for generating hyper-activated MAPK, growth arrest and apoptosis, implying strict specificity for mutated B-Raf cancer cells. PMID:26959890

  9. Suppression of B-Raf(V600E) cancers by MAPK hyper-activation.

    PubMed

    Atiq, Rawan; Hertz, Rachel; Eldad, Sophia; Smeir, Elia; Bar-Tana, Jacob

    2016-04-01

    B-Raf(V600E) activates MEK/MAPK signalling and acts as oncogenic driver of a variety of cancers, including melanoma, colorectal and papillary thyroid carcinoma. Specific B-Raf(V600E) kinase inhibitors (e.g., Vemurafenib) prove initial efficacy in melanoma followed shortly by acquired resistance, while failing in most other B-Raf(V600E) cancers due to primary resistance. Resistance is due to acquired mutations in the Ras/Raf/MEK/MAPK pathway and/or other oncogenic drivers that bypass B-Raf(V600E). Surprisingly, hyper-activation of MAPK by inhibiting its protein phosphatase 2A by a synthetic long-chain fatty acid analogue (MEDICA), results in oncogene-induced growth arrest and apoptosis of B-Raf(V600E) cancer cells. Growth arrest is accompanied by MAPK-mediated serine/threonine phosphorylation and suppression of a variety of oncogenic drivers that resist treatment by B-Raf(V600E) kinase inhibitors, including ErbB members, c-Met, IGFR, IRS, STAT3 and Akt. The combined activities of mutated B-Raf and MEDICA are required for generating hyper-activated MAPK, growth arrest and apoptosis, implying strict specificity for mutated B-Raf cancer cells. PMID:26959890

  10. Hirudin inhibits cell growth via ERK/MAPK signaling in human glioma

    PubMed Central

    Zhao, Li

    2015-01-01

    Glioma is one of the most common intracranial tumors, and the prognosis is poor though more and more treatments are employed. ERK/MAPK signaling has been reported to be associated with glioma. In the present study, we aimed to investigate hirudin as an antineoplastic drug inhibits ERK/MAPK signaling in glioma growth in vitro. The cell proliferation and apoptosis rate were detected using MTS and Annexin V staining assay, cell cycle distribution was detected using flowcytometry assay. Furthermore, the relevant molecules of ERK/MAPK signaling were examined using Western blot analysis and immunofluorescence staining assay. We provide the first evidence that hirudin increase inactivation state of ERK1/2, down-regulate the expression of canonical ERK/MAPK signaling pathway and establish an important role for hirudin in the treatment of glioma cells. PMID:26885029

  11. Atorvastatin Prevents Glutamate Uptake Reduction Induced by Quinolinic Acid Via MAPKs Signaling.

    PubMed

    Vandresen-Filho, S; Martins, W C; Bertoldo, D B; Rieger, D K; Maestri, M; Leal, R B; Tasca, C I

    2016-08-01

    Statins have been shown to promote neuroprotection in a wide range of neurological disorders. However, the mechanisms involved in such effects of statins are not fully understood. Quinolinic acid (QA) is a neurotoxin that induces seizures when infused in vivo and promotes glutamatergic excitotoxicity in the central nervous system. The aim of this study was to evaluate the putative glutamatergic mechanisms and the intracellular signaling pathways involved in the atorvastatin neuroprotective effects against QA toxicity. Atorvastatin (10 mg/kg) treatment for 7 days prevented the QA-induced decrease in glutamate uptake, but had no effect on increased glutamate release induced by QA. Moreover, atorvastatin treatment increased the phosphorylation of ERK1 and prevented the decrease in Akt phosphorylation induced by QA. Neither atorvastatin treatment nor QA infusion altered glutamine synthetase activity or the levels of phosphorylation of p38(MAPK) or JNK1/2 during the evaluation. Inhibition of MEK/ERK signaling pathway, but not PI3K/Akt signaling, abolished the neuroprotective effect of atorvastatin against QA-induced decrease in glutamate uptake. Our data suggest that atorvastatin protective effects against QA toxicity are related to modulation of glutamate transporters via MAPK/ERK signaling pathway.

  12. FGFR2c-mediated ERK-MAPK activity regulates coronal suture development.

    PubMed

    Pfaff, Miles J; Xue, Ke; Li, Li; Horowitz, Mark C; Steinbacher, Derek M; Eswarakumar, Jacob V P

    2016-07-15

    Fibroblast growth factor receptor 2 (FGFR2) signaling is critical for proper craniofacial development. A gain-of-function mutation in the 2c splice variant of the receptor's gene is associated with Crouzon syndrome, which is characterized by craniosynostosis, the premature fusion of one or more of the cranial vault sutures, leading to craniofacial maldevelopment. Insight into the molecular mechanism of craniosynostosis has identified the ERK-MAPK signaling cascade as a critical regulator of suture patency. The aim of this study is to investigate the role of FGFR2c-induced ERK-MAPK activation in the regulation of coronal suture development. Loss-of-function and gain-of-function Fgfr2c mutant mice have overlapping phenotypes, including coronal synostosis and craniofacial dysmorphia. In vivo analysis of coronal sutures in loss-of-function and gain-of-function models demonstrated fundamentally different pathogenesis underlying coronal suture synostosis. Calvarial osteoblasts from gain-of-function mice demonstrated enhanced osteoblastic function and maturation with concomitant increase in ERK-MAPK activation. In vitro inhibition with the ERK protein inhibitor U0126 mitigated ERK protein activation levels with a concomitant reduction in alkaline phosphatase activity. This study identifies FGFR2c-mediated ERK-MAPK signaling as a key mediator of craniofacial growth and coronal suture development. Furthermore, our results solve the apparent paradox between loss-of-function and gain-of-function FGFR2c mutants with respect to coronal suture synostosis. PMID:27034231

  13. Ganoderma lucidum polysaccharide exerts anti-tumor activity via MAPK pathways in HL-60 acute leukemia cells.

    PubMed

    Yang, Guohua; Yang, Lei; Zhuang, Yun; Qian, Xifeng; Shen, Yunfeng

    2016-01-01

    In this study, we investigated the anti-tumor activity both in vitro and in vivo of a polysaccharide obtained from Ganoderma lucidum on HL-60 acute myeloid leukemia cells, and focused on its targeting effect on mitogen-activated protein kinase (MAPK) pathways. It was found by the methods such as western blot and flow cytometry (FCM), that G. lucidum polysaccharide (GLP) blocked the extracellular signal-regulated kinase/MAPK signaling pathway, simultaneously activated p38 and JNK MAPK pathways, and therefore regulated their downstream genes and proteins, including p53, c-myc, c-fos, c-jun, Bcl-2, Bax, cleaved caspase-3 and cyclin D1. As a result, cycle arrest and apoptosis of HL-60 cells were induced. Therefore, GLP exerted anti-tumor activity via MAPK pathways in HL-60 acute leukemia cells.

  14. The role of docking interactions in mediating signaling input, output, and discrimination in the yeast MAPK network.

    PubMed

    Reményi, Attila; Good, Matthew C; Bhattacharyya, Roby P; Lim, Wendell A

    2005-12-22

    Cells use a network of mitogen-activated protein kinases (MAPKs) to coordinate responses to diverse extracellular signals. Here, we examine the role of docking interactions in determining connectivity of the yeast MAPKs Fus3 and Kss1. These closely related kinases are activated by the common upstream MAPK kinase Ste7 yet generate distinct output responses, mating and filamentous growth, respectively. We find that docking interactions are necessary for communication with the kinases and that they can encode subtle differences in pathway-specific input and output. The cell cycle arrest mediator Far1, a mating-specific substrate, has a docking motif that selectively binds Fus3. In contrast, the shared partner Ste7 has a promiscuous motif that binds both Fus3 and Kss1. Structural analysis reveals that Fus3 interacts with specific and promiscuous peptides in conformationally distinct modes. Induced fit recognition may allow docking peptides to achieve discrimination by exploiting subtle differences in kinase flexibility. PMID:16364919

  15. Tetraspanin CD9 regulates osteoclastogenesis via regulation of p44/42 MAPK activity

    SciTech Connect

    Yi, TacGhee; Kim, Hye-Jin; Cho, Je-Yoel; Woo, Kyung Mi; Ryoo, Hyun-Mo; Kim, Gwan-Shik; Baek, Jeong-Hwa . E-mail: baekjh@snu.ac.kr

    2006-08-18

    Tetraspanin CD9 has been shown to regulate cell-cell fusion in sperm-egg fusion and myotube formation. However, the role of CD9 in osteoclast, another multinucleated cell type, is not still clear. Therefore, we investigated the role of CD9 in osteoclast differentiation. CD9 was expressed in osteoclast lineage cells and its expression level increased during the progression of RANKL-induced osteoclastogenesis. KMC8, a neutralizing antibody specific to CD9, significantly suppressed RANKL-induced multinucleated osteoclast formation and the mRNA expression of osteoclast differentiation marker genes. To define CD9-regulated osteoclastogenic signaling pathway, MAPK pathways were examined. KMC8 induced long-term phosphorylation of p44/42 MAPK, but not of p38 MAPK. Constitutive activation of p44/42 MAPK by overexpressing constitutive-active mutant of MEK1 almost completely blocked osteoclast differentiation. Taken together, these results suggest that CD9 expressed on osteoclast lineage cells might positively regulate osteoclastogenesis via the regulation of p44/42 MAPK activity.

  16. Dehydrocorydaline promotes myogenic differentiation via p38 MAPK activation

    PubMed Central

    Yoo, Miran; Lee, Sang-Jin; Kim, Yong Kee; Seo, Dong-Wan; Baek, Nam-In; Ryu, Jae-Ha; Kang, Jong-Sun; Bae, Gyu-Un

    2016-01-01

    Muscle regeneration is a coordinated process that involves proliferation and differentiation of muscle progenitor cells. Activation of MyoD is a key event in myogenic differentiation, which is regulated by p38 mitogen-activated protein kinases (MAPK). In a screen of natural compounds for the enhancement of MyoD activity, dehydrocorydaline (DHC) from the Corydalis tuber was identified. Treatment of C2C12 myoblasts with DHC increased the expression levels of muscle-specific proteins, including MyoD, myogenin and myosin heavy chain. In addition, C2C12 myoblasts exhibited enhanced multinucleated myotube formation without any cytotoxicity. Treatment with DHC elevated p38 MAPK activation and the interaction of MyoD with an E protein, which is likely to result in activation of MyoD and promotion of myoblast differentiation. Furthermore, defects in differentiation-induced p38 MAPK activation and myoblast differentiation induced by depletion of the promyogenic receptor protein Cdo in C2C12 myoblasts were restored by DHC treatment. In conclusion, these results indicated that DHC stimulates p38 MAPK activation, which can enhance heterodimerization of MyoD and E proteins, thus resulting in MyoD activation and myoblast differentiation. These findings suggested that DHC may be considered a potential therapeutic compound for the improvement of muscle stem cell regenerative capacity in injured muscle. PMID:27573543

  17. Antagonistic roles of the ERK and p38 MAPK signalling pathways in globin expression, haem biosynthesis and iron uptake.

    PubMed

    Mardini, Louay; Gasiorek, Jadwiga; Derjuga, Anna; Carrière, Lucie; Schranzhofer, Matthias; Paw, Barry H; Ponka, Prem; Blank, Volker

    2010-11-15

    Late-stage erythroid cells synthesize large quantities of haemoglobin, a process requiring the co-ordinated regulation of globin and haem synthesis as well as iron uptake. In the present study, we investigated the role of the ERK (extracellular-signal-regulated kinase) and p38 MAPK (mitogen-activated protein kinase) signalling pathways in MEL (mouse erythroleukaemia) cell differentiation. We found that treatment of HMBA (hexamethylene bisacetamide)-induced MEL cells with the ERK pathway inhibitor UO126 results in an increase in intracellular haem and haemoglobin levels. The transcript levels of the genes coding for β(major)-globin, the haem biosynthesis enzyme 5-aminolevulinate synthase 2 and the mitochondrial iron transporter mitoferrin 1 are up-regulated. We also showed enhanced expression of globin and transferrin receptor 1 proteins upon UO126 treatment. With respect to iron uptake, we found that ERK inhibitor treatment led to an increase in both haem-bound and total iron. In contrast, treatment of MEL cells with the p38 MAPK pathway inhibitor SB202190 had the opposite effect, resulting in decreased globin expression, haem synthesis and iron uptake. Reporter assays showed that globin promoter and HS2 enhancer-mediated transcription was under the control of MAPKs, as inhibition of the ERK and p38 MAPK pathways led to increased and decreased gene activity respectively. Our present results suggest that the ERK1/2 and p38α/β MAPKs play antagonistic roles in HMBA-induced globin gene expression and erythroid differentiation. These results provide a novel link between MAPK signalling and the regulation of haem biosynthesis and iron uptake in erythroid cells.

  18. Sorbitol induces apoptosis of human colorectal cancer cells via p38 MAPK signal transduction.

    PubMed

    Lu, Xue; Li, Chun; Wang, Yong-Kun; Jiang, Kun; Gai, Xiao-Dong

    2014-06-01

    Sorbitol has been reported to have anticancer effects in several tumor models, however its effects on colorectal cancer remain elusive. In the present study, the effects of sorbitol on growth inhibition and apoptosis in the colorectal cancer HCT116 cell line were evaluated and its mechanism of action was examined. An MTT assay was utilized to determine the effect of sorbitol on HCT116 cell proliferation at different time points and variable doses. Western blot analysis was used to examine the effect of sorbitol on apoptosis-related protein expression and the p38 MAPK signaling pathway. The results revealed that sorbitol may inhibit the growth of HCT116 cells in a time- and dose-dependent manner. Following treatment with sorbitol for 3 h, western blotting demonstrated cleavage of the caspase-3 zymogen protein and a cleavage product of poly (ADP-ribose) polymerase (PARP), a known substrate of caspase-3, was also evident. During sorbitol-induced apoptosis, the mitochondrial pathway was activated by a dose-dependent increase in Bax expression and cytochrome c release, while the expression of anti-apoptotic protein Bcl-2 was significantly decreased in a dose-dependent manner. The investigation for the downstream signal pathway revealed that sorbitol-induced apoptosis was mediated by an increase in phosphorylated p38 MAPK expression. Overall, the observations from the present study imply that sorbitol causes increased levels of Bax in response to p38 MAPK signaling, which results in the initiation of the mitochondrial death cascade. Therefore, sorbitol is a promising candidate as a potential chemotherapeutic agent for the treatment of colorectal cancer HCT116 cells.

  19. Estrogen receptor beta growth-inhibitory effects are repressed through activation of MAPK and PI3K signalling in mammary epithelial and breast cancer cells.

    PubMed

    Cotrim, C Z; Fabris, V; Doria, M L; Lindberg, K; Gustafsson, J-Å; Amado, F; Lanari, C; Helguero, L A

    2013-05-01

    Two thirds of breast cancers express estrogen receptors (ER). ER alpha (ERα) mediates breast cancer cell proliferation, and expression of ERα is the standard choice to indicate adjuvant endocrine therapy. ERbeta (ERβ) inhibits growth in vitro; its effects in vivo have been incompletely investigated and its role in breast cancer and potential as alternative target in endocrine therapy needs further study. In this work, mammary epithelial (EpH4 and HC11) and breast cancer (MC4-L2) cells with endogenous ERα and ERβ expression and T47-D human breast cancer cells with recombinant ERβ (T47-DERβ) were used to explore effects exerted in vitro and in vivo by the ERβ agonists 2,3-bis (4-hydroxy-phenyl)-propionitrile (DPN) and 7-bromo-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol (WAY). In vivo, ERβ agonists induced mammary gland hyperplasia and MC4-L2 tumour growth to a similar extent as the ERα agonist 4,4',4''-(4-propyl-(1H)-pyrazole-1,3,5-triyl) trisphenol (PPT) or 17β-estradiol (E2) and correlated with higher number of mitotic and lower number of apoptotic features. In vitro, in MC4-L2, EpH4 or HC11 cells incubated under basal conditions, ERβ agonists induced apoptosis measured as upregulation of p53 and apoptosis-inducible factor protein levels and increased caspase 3 activity, whereas PPT and E2 stimulated proliferation. However, when extracellular signal-regulated kinase 1 and 2 (ERK ½) were activated by co-incubation with basement membrane extract or epidermal growth factor, induction of apoptosis by ERβ agonists was repressed and DPN induced proliferation in a similar way as E2 or PPT. In a context of active ERK ½, phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/RAC-alpha serine/threonine-protein kinase (AKT) signalling was necessary to allow proliferation stimulated by ER agonists. Inhibition of MEK ½ with UO126 completely restored ERβ growth-inhibitory effects, whereas inhibition of PI3K by LY294002 inhibited ERβ-induced proliferation. These

  20. Regulation of Muscle Stem Cell Functions: A Focus on the p38 MAPK Signaling Pathway

    PubMed Central

    Segalés, Jessica; Perdiguero, Eusebio; Muñoz-Cánoves, Pura

    2016-01-01

    Formation of skeletal muscle fibers (myogenesis) during development and after tissue injury in the adult constitutes an excellent paradigm to investigate the mechanisms whereby environmental cues control gene expression programs in muscle stem cells (satellite cells) by acting on transcriptional and epigenetic effectors. Here we will review the molecular mechanisms implicated in the transition of satellite cells throughout the distinct myogenic stages (i.e., activation from quiescence, proliferation, differentiation, and self-renewal). We will also discuss recent findings on the causes underlying satellite cell functional decline with aging. In particular, our review will focus on the epigenetic changes underlying fate decisions and on how the p38 MAPK signaling pathway integrates the environmental signals at the chromatin to build up satellite cell adaptive responses during the process of muscle regeneration, and how these responses are altered in aging. A better comprehension of the signaling pathways connecting external and intrinsic factors will illuminate the path for improving muscle regeneration in the aged.

  1. Layer specific and general requirements for ERK/MAPK signaling in the developing neocortex

    PubMed Central

    Xing, Lei; Larsen, Rylan S; Bjorklund, George Reed; Li, Xiaoyan; Wu, Yaohong; Philpot, Benjamin D; Snider, William D; Newbern, Jason M

    2016-01-01

    Aberrant signaling through the Raf/MEK/ERK (ERK/MAPK) pathway causes pathology in a family of neurodevelopmental disorders known as 'RASopathies' and is implicated in autism pathogenesis. Here, we have determined the functions of ERK/MAPK signaling in developing neocortical excitatory neurons. Our data reveal a critical requirement for ERK/MAPK signaling in the morphological development and survival of large Ctip2+ neurons in layer 5. Loss of Map2k1/2 (Mek1/2) led to deficits in corticospinal tract formation and subsequent corticospinal neuron apoptosis. ERK/MAPK hyperactivation also led to reduced corticospinal axon elongation, but was associated with enhanced arborization. ERK/MAPK signaling was dispensable for axonal outgrowth of layer 2/3 callosal neurons. However, Map2k1/2 deletion led to reduced expression of Arc and enhanced intrinsic excitability in both layers 2/3 and 5, in addition to imbalanced synaptic excitation and inhibition. These data demonstrate selective requirements for ERK/MAPK signaling in layer 5 circuit development and general effects on cortical pyramidal neuron excitability. DOI: http://dx.doi.org/10.7554/eLife.11123.001 PMID:26848828

  2. Stepwise metamorphosis of the tubeworm Hydroides elegans is mediated by a bacterial inducer and MAPK signaling

    PubMed Central

    Shikuma, Nicholas J.; Antoshechkin, Igor; Medeiros, João M.; Pilhofer, Martin; Newman, Dianne K.

    2016-01-01

    Diverse animal taxa metamorphose between larval and juvenile phases in response to bacteria. Although bacteria-induced metamorphosis is widespread among metazoans, little is known about the molecular changes that occur in the animal upon stimulation by bacteria. Larvae of the tubeworm Hydroides elegans metamorphose in response to surface-bound Pseudoalteromonas luteoviolacea bacteria, producing ordered arrays of phage tail-like metamorphosis-associated contractile structures (MACs). Sequencing the Hydroides genome and transcripts during five developmental stages revealed that MACs induce the regulation of groups of genes important for tissue remodeling, innate immunity, and mitogen-activated protein kinase (MAPK) signaling. Using two MAC mutations that block P. luteoviolacea from inducing settlement or metamorphosis and three MAPK inhibitors, we established a sequence of bacteria-induced metamorphic events: MACs induce larval settlement; then, particular properties of MACs encoded by a specific locus in P. luteoviolacea initiate cilia loss and activate metamorphosis-associated transcription; finally, signaling through p38 and c-Jun N-terminal kinase (JNK) MAPK pathways alters gene expression and leads to morphological changes upon initiation of metamorphosis. Our results reveal that the intricate interaction between Hydroides and P. luteoviolacea can be dissected using genomic, genetic, and pharmacological tools. Hydroides' dependency on bacteria for metamorphosis highlights the importance of external stimuli to orchestrate animal development. The conservation of Hydroides genome content with distantly related deuterostomes (urchins, sea squirts, and humans) suggests that mechanisms of bacteria-induced metamorphosis in Hydroides may have conserved features in diverse animals. As a major biofouling agent, insight into the triggers of Hydroides metamorphosis might lead to practical strategies for fouling control. PMID:27551098

  3. Stepwise metamorphosis of the tubeworm Hydroides elegans is mediated by a bacterial inducer and MAPK signaling.

    PubMed

    Shikuma, Nicholas J; Antoshechkin, Igor; Medeiros, João M; Pilhofer, Martin; Newman, Dianne K

    2016-09-01

    Diverse animal taxa metamorphose between larval and juvenile phases in response to bacteria. Although bacteria-induced metamorphosis is widespread among metazoans, little is known about the molecular changes that occur in the animal upon stimulation by bacteria. Larvae of the tubeworm Hydroides elegans metamorphose in response to surface-bound Pseudoalteromonas luteoviolacea bacteria, producing ordered arrays of phage tail-like metamorphosis-associated contractile structures (MACs). Sequencing the Hydroides genome and transcripts during five developmental stages revealed that MACs induce the regulation of groups of genes important for tissue remodeling, innate immunity, and mitogen-activated protein kinase (MAPK) signaling. Using two MAC mutations that block P. luteoviolacea from inducing settlement or metamorphosis and three MAPK inhibitors, we established a sequence of bacteria-induced metamorphic events: MACs induce larval settlement; then, particular properties of MACs encoded by a specific locus in P. luteoviolacea initiate cilia loss and activate metamorphosis-associated transcription; finally, signaling through p38 and c-Jun N-terminal kinase (JNK) MAPK pathways alters gene expression and leads to morphological changes upon initiation of metamorphosis. Our results reveal that the intricate interaction between Hydroides and P. luteoviolacea can be dissected using genomic, genetic, and pharmacological tools. Hydroides' dependency on bacteria for metamorphosis highlights the importance of external stimuli to orchestrate animal development. The conservation of Hydroides genome content with distantly related deuterostomes (urchins, sea squirts, and humans) suggests that mechanisms of bacteria-induced metamorphosis in Hydroides may have conserved features in diverse animals. As a major biofouling agent, insight into the triggers of Hydroides metamorphosis might lead to practical strategies for fouling control.

  4. Stepwise metamorphosis of the tubeworm Hydroides elegans is mediated by a bacterial inducer and MAPK signaling.

    PubMed

    Shikuma, Nicholas J; Antoshechkin, Igor; Medeiros, João M; Pilhofer, Martin; Newman, Dianne K

    2016-09-01

    Diverse animal taxa metamorphose between larval and juvenile phases in response to bacteria. Although bacteria-induced metamorphosis is widespread among metazoans, little is known about the molecular changes that occur in the animal upon stimulation by bacteria. Larvae of the tubeworm Hydroides elegans metamorphose in response to surface-bound Pseudoalteromonas luteoviolacea bacteria, producing ordered arrays of phage tail-like metamorphosis-associated contractile structures (MACs). Sequencing the Hydroides genome and transcripts during five developmental stages revealed that MACs induce the regulation of groups of genes important for tissue remodeling, innate immunity, and mitogen-activated protein kinase (MAPK) signaling. Using two MAC mutations that block P. luteoviolacea from inducing settlement or metamorphosis and three MAPK inhibitors, we established a sequence of bacteria-induced metamorphic events: MACs induce larval settlement; then, particular properties of MACs encoded by a specific locus in P. luteoviolacea initiate cilia loss and activate metamorphosis-associated transcription; finally, signaling through p38 and c-Jun N-terminal kinase (JNK) MAPK pathways alters gene expression and leads to morphological changes upon initiation of metamorphosis. Our results reveal that the intricate interaction between Hydroides and P. luteoviolacea can be dissected using genomic, genetic, and pharmacological tools. Hydroides' dependency on bacteria for metamorphosis highlights the importance of external stimuli to orchestrate animal development. The conservation of Hydroides genome content with distantly related deuterostomes (urchins, sea squirts, and humans) suggests that mechanisms of bacteria-induced metamorphosis in Hydroides may have conserved features in diverse animals. As a major biofouling agent, insight into the triggers of Hydroides metamorphosis might lead to practical strategies for fouling control. PMID:27551098

  5. EPAS1 trans-activation during hypoxia requires p42/p44 MAPK.

    PubMed

    Conrad, P W; Freeman, T L; Beitner-Johnson, D; Millhorn, D E

    1999-11-19

    Hypoxia is a common environmental stress that regulates gene expression and cell function. A number of hypoxia-regulated transcription factors have been identified and have been shown to play critical roles in mediating cellular responses to hypoxia. One of these is the endothelial PAS-domain protein 1 (EPAS1/HIF2-alpha/HLF/HRF). This protein is 48% homologous to hypoxia-inducible factor 1-alpha (HIF1-alpha). To date, virtually nothing is known about the signaling pathways that lead to either EPAS1 or HIF1-alpha activation. Here we show that EPAS1 is phosphorylated when PC12 cells are exposed to hypoxia and that p42/p44 MAPK is a critical mediator of EPAS1 activation. Pretreatment of PC12 cells with the MEK inhibitor, PD98059, completely blocked hypoxia-induced trans-activation of a hypoxia response element (HRE) reporter gene by transfected EPAS1. Likewise, expression of a constitutively active MEK1 mimicked the effects of hypoxia on HRE reporter gene expression. However, pretreatment with PD98059 had no effect on EPAS1 phosphorylation during hypoxia, suggesting that MAPK targets other proteins that are critical for the trans-activation of EPAS1. We further show that hypoxia-induced trans-activation of EPAS1 is independent of Ras. Finally, pretreatment with calmodulin antagonists nearly completely blocked both the hypoxia-induced phosphorylation of MAPK and the EPAS1 trans-activation of HRE-Luc. These results demonstrate that the MAPK pathway is a critical mediator of EPAS1 activation and that activation of MAPK and EPAS1 occurs through a calmodulin-sensitive pathway and not through the GTPase, Ras. These results are the first to identify a specific signaling pathway involved in EPAS1 activation. PMID:10559262

  6. Reactive oxygen species mediate nitric oxide production through ERK/JNK MAPK signaling in HAPI microglia after PFOS exposure.

    PubMed

    Wang, Cheng; Nie, Xiaoke; Zhang, Yan; Li, Ting; Mao, Jiamin; Liu, Xinhang; Gu, Yiyang; Shi, Jiyun; Xiao, Jing; Wan, Chunhua; Wu, Qiyun

    2015-10-15

    Perfluorooctane sulfonate (PFOS), an emerging persistent contaminant that is commonly encountered during daily life, has been shown to exert toxic effects on the central nervous system (CNS). However, the molecular mechanisms underlying the neurotoxicity of PFOS remain largely unknown. It has been widely acknowledged that the inflammatory mediators released by hyper-activated microglia play vital roles in the pathogenesis of various neurological diseases. In the present study, we examined the impact of PFOS exposure on microglial activation and the release of proinflammatory mediators, including nitric oxide (NO) and reactive oxidative species (ROS). We found that PFOS exposure led to concentration-dependent NO and ROS production by rat HAPI microglia. We also discovered that there was rapid activation of the ERK/JNK MAPK signaling pathway in the HAPI microglia following PFOS treatment. Moreover, the PFOS-induced iNOS expression and NO production were attenuated after the inhibition of ERK or JNK MAPK by their corresponding inhibitors, PD98059 and SP600125. Interestingly, NAC, a ROS inhibitor, blocked iNOS expression, NO production, and activation of ERK and JNK MAPKs, which suggested that PFOS-mediated microglial NO production occurs via a ROS/ERK/JNK MAPK signaling pathway. Finally, by exposing SH-SY5Y cells to PFOS-treated microglia-conditioned medium, we demonstrated that NO was responsible for PFOS-mediated neuronal apoptosis.

  7. Cell-type specific crosstalk between p38 MAPK and Rho signaling in lung micro- and macrovascular barrier dysfunction induced by Staphylococcus aureus-derived pathogens

    PubMed Central

    Wu, Tinghuai; Xing, Junjie; Birukova, Anna A.

    2014-01-01

    Lung inflammation and alterations in endothelial cell (EC) micro- and macro-vascular permeability are key events to development of acute lung injury (ALI). Using ECs derived from human pulmonary artery (HPAECs) and lung microvasculature (HLMVECs), we investigated interplay between p38 stress MAPK and Rho GTPase signaling in the inflammatory and hyperpermeability response. Both cell types were treated with Staphylococcus aureus-derived peptidoglycan (PepG) and lipoteichoic acid (LTA) with or without pretreatment with p38 MAPK or Rho kinase inhibitors. LTA and PepG markedly increased permeability in both pulmonary macrovascular and microvascular EC. Agonist-induced hyper-permeability was accompanied by cytoskeletal remodeling, disruption of cell-cell contacts, formation of paracellular gaps, and activation of p38 MAPK, NFκB, and Rho/Rho kinase signaling. In macrovascular ECs, pharmacological inhibition of Rho kinase with Y27632 significantly suppressed p38 MAP kinase cascade activation, while inhibition of p38 MAPK with SB203580 had no effect on Rho activation. In contrast, inhibition of p38 MAPK in microvascular ECs suppressed LTA/PepG-induced activation of Rho, while Rho inhibitor suppressed activation of p38 MAPK. Inhibition of either p38 MAPK or Rho kinase substantially attenuated activation of NFκB signaling. These results demonstrate cell type-specific differences in signaling induced by Staphylococcus aureus-derived pathogens in pulmonary endothelium. Thus, although Gram-positive bacterial compounds caused barrier dysfunction in both EC types, it was induced by different pattern of crosstalk between Rho, p38 MAPK, and NFκB signaling. These observations may have important implications in defining microvasculature-specific therapeutic strategies aimed at the treatment of sepsis and acute lung injury induced by Gram-positive bacterial pathogens. PMID:23571093

  8. The Protective Effect of Beraprost Sodium on Diabetic Nephropathy by Inhibiting Inflammation and p38 MAPK Signaling Pathway in High-Fat Diet/Streptozotocin-Induced Diabetic Rats

    PubMed Central

    Peng, Li; Li, Jie; Xu, Yixing; Wang, Yangtian; Du, Hong; Shao, Jiaqing; Liu, Zhimin

    2016-01-01

    Background. p38 mitogen-activated protein kinase (MAPK) plays a crucial role in regulating signaling pathways implicated in inflammatory processes leading to diabetic nephropathy (DN). This study aimed to examine p38 MAPK activation in DN and determine whether beraprost sodium (BPS) ameliorates DN by inhibiting inflammation and p38 MAPK signaling pathway in diabetic rats. Methods. Forty male Sprague Dawley (SD) rats were randomly divided into the normal control group, type 2 diabetic group, and BPS treatment group. At the end of the 8-week experiment, we measured renal pathological changes and the activation of the p38 MAPK signaling pathway and inflammation. Result. After BPS treatment, renal function, 24-hour urine protein, lipid profiles, and blood glucose level were improved significantly; meanwhile, inflammation and the expression of p38 MAPK signaling pathway in the diabetic kidney were attenuated. Conclusions. BPS significantly prevented type 2 diabetes induced kidney injury characterized by renal dysfunction and pathological changes. The protective mechanisms are complicated but may be mainly attributed to the inhibition of the p38 MAPK signaling pathway and inflammation in the diabetic kidney. PMID:27212945

  9. Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK) Gene Family in Brassica rapa

    PubMed Central

    Yu, Hao; Qu, Cunmin; Tang, Zhanglin; Li, Jiana; Chai, Yourong; Liang, Ying

    2015-01-01

    Mitogen-activated protein kinase (MAPK) cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY) of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D). Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa. PMID:26173020

  10. Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK) Gene Family in Brassica rapa.

    PubMed

    Lu, Kun; Guo, Wenjin; Lu, Junxing; Yu, Hao; Qu, Cunmin; Tang, Zhanglin; Li, Jiana; Chai, Yourong; Liang, Ying

    2015-01-01

    Mitogen-activated protein kinase (MAPK) cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY) of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D). Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa.

  11. Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK) Gene Family in Brassica rapa.

    PubMed

    Lu, Kun; Guo, Wenjin; Lu, Junxing; Yu, Hao; Qu, Cunmin; Tang, Zhanglin; Li, Jiana; Chai, Yourong; Liang, Ying

    2015-01-01

    Mitogen-activated protein kinase (MAPK) cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY) of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D). Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa. PMID:26173020

  12. p38 MAPK is a likely component of the signal transduction pathway triggering rapid cold hardening in the flesh fly Sarcophaga crassipalpis.

    PubMed

    Fujiwara, Yoshihiro; Denlinger, David L

    2007-09-01

    Rapid cold hardening (RCH) is an adaptation enabling insects to quickly respond to low temperature, but little is known about the molecular events that trigger this response. In this study of the flesh fly Sarcophaga crassipalpis, we explore a possible role for mitogen-activated protein kinases (MAPKs) in the low temperature signaling that elicits RCH. We report that p38 MAPK from S. crassipalpis, which shows high cDNA sequence homology to p38 MAPKs from other insects and mammals, is rapidly activated at temperatures around 0 degrees C, temperatures that are most effective for inducing RCH. By contrast, low temperature does not activate either extracellular signal-regulated kinase (ERK) or Jun N-terminal kinase (JNK). An increase in phospho-p38 MAPK was observed within 10 min following exposure to 0 degrees C and reached its maximum level in 2 h. When flies were transferred from 0 to 25 degrees C, the level of phospho-p38 MAPK decreased immediately and reached trace levels by 3 h. Nondiapausing flies were much more responsive to p38 MAPK activation than cold-hardy diapausing pupae. Thus, p38 MAPK activation and RCH both show the same narrow ranges of temperature sensitivity, temporal profiles of activation and decay, and developmental specificity. These correlations suggest that p38 MAPK plays a potential role in regulating the induction of RCH. The p38 MAPK response was not dependent upon the brain, as evidenced by high activation in isolated abdomens exposed to low temperature. PMID:17766307

  13. TNF-α stimulates System A amino acid transport in primary human trophoblast cells mediated by p38 MAPK signaling.

    PubMed

    Aye, Irving L M H; Jansson, Thomas; Powell, Theresa L

    2015-10-01

    Maternal obesity and gestational diabetes mellitus (GDM) increase the risk of delivering infants that are large for gestational age with greater adiposity, who are prone to the development of metabolic disease in childhood and beyond. These maternal conditions are also associated with increased levels of the proinflammatory cytokine TNF-α in maternal tissues and the placenta. Recent evidence suggests that changes in placental amino acid transport contribute to altered fetal growth. TNF-α was previously shown to stimulate System A amino acid transport in primary human trophoblasts (PHTs), however the molecular mechanisms remain unknown. In this study, we tested the hypothesis that TNF-α regulates amino acid uptake in cultured PHTs by a mitogen-activated protein kinase (MAPK)-dependent mechanism. Treatment of PHTs with TNF-α significantly increased System A amino acid transport, as well as Erk and p38 MAPK signaling. Pharmacological antagonism of p38, but not Erk MAPK activity, inhibited TNF-α stimulated System A activity. Silencing of p38 MAPK using siRNA transfections prevented TNF-α stimulated System A transport in PHTs. TNF-α significantly increased the protein expression of System A transporters SNAT1 and SNAT2, but did not affect their mRNA expression. The effects of TNF-α on SNAT1 and SNAT2 protein expression were reversed by p38 MAPK siRNA silencing. In conclusion, TNF-α regulates System A activity through increased SNAT1 and SNAT2 transporter protein expression in PHTs. These findings suggest that p38 MAPK may represent a critical mechanistic link between elevated proinflammatory cytokines and increased placental amino acid transport in obese and GDM pregnancies associated with fetal overgrowth.

  14. TNF-α stimulates System A amino acid transport in primary human trophoblast cells mediated by p38 MAPK signaling.

    PubMed

    Aye, Irving L M H; Jansson, Thomas; Powell, Theresa L

    2015-10-01

    Maternal obesity and gestational diabetes mellitus (GDM) increase the risk of delivering infants that are large for gestational age with greater adiposity, who are prone to the development of metabolic disease in childhood and beyond. These maternal conditions are also associated with increased levels of the proinflammatory cytokine TNF-α in maternal tissues and the placenta. Recent evidence suggests that changes in placental amino acid transport contribute to altered fetal growth. TNF-α was previously shown to stimulate System A amino acid transport in primary human trophoblasts (PHTs), however the molecular mechanisms remain unknown. In this study, we tested the hypothesis that TNF-α regulates amino acid uptake in cultured PHTs by a mitogen-activated protein kinase (MAPK)-dependent mechanism. Treatment of PHTs with TNF-α significantly increased System A amino acid transport, as well as Erk and p38 MAPK signaling. Pharmacological antagonism of p38, but not Erk MAPK activity, inhibited TNF-α stimulated System A activity. Silencing of p38 MAPK using siRNA transfections prevented TNF-α stimulated System A transport in PHTs. TNF-α significantly increased the protein expression of System A transporters SNAT1 and SNAT2, but did not affect their mRNA expression. The effects of TNF-α on SNAT1 and SNAT2 protein expression were reversed by p38 MAPK siRNA silencing. In conclusion, TNF-α regulates System A activity through increased SNAT1 and SNAT2 transporter protein expression in PHTs. These findings suggest that p38 MAPK may represent a critical mechanistic link between elevated proinflammatory cytokines and increased placental amino acid transport in obese and GDM pregnancies associated with fetal overgrowth. PMID:26508738

  15. Pheromone-Induced Morphogenesis Improves Osmoadaptation Capacity by Activating the HOG MAPK Pathway**

    PubMed Central

    Baltanás, Rodrigo; Bush, Alan; Couto, Alicia; Durrieu, Lucía; Hohmann, Stefan; Colman-Lerner, Alejandro

    2013-01-01

    Environmental and internal conditions expose cells to a multiplicity of stimuli whose consequences are difficult to predict. Here, we investigate the response to mating pheromone of yeast cells adapted to high osmolarity. Events downstream of pheromone binding involve two mitogen-activated protein kinase (MAPK) cascades: the pheromone response (PR) and the cell-wall integrity response (CWI). Although these MAPK pathways share components with each and a third MAPK pathway, the high osmolarity response (HOG), they are normally only activated by distinct stimuli, a phenomenon called insulation. We found that in cells adapted to high osmolarity, PR activated the HOG pathway in a pheromone- and osmolarity- dependent manner. Activation of HOG by the PR was not due to loss of insulation, but rather a response to a reduction in internal osmolarity, which resulted from an increase in glycerol release caused by the PR. By analyzing single-cell time courses, we found that stimulation of HOG occurred in discrete bursts that coincided with the “shmooing” morphogenetic process. Activation required the polarisome, the cell wall integrity MAPK Slt2, and the aquaglyceroporin Fps1. HOG activation resulted in high glycerol turnover that improved adaptability to rapid changes in osmolarity. Our work shows how a differentiation signal can recruit a second, unrelated sensory pathway to enable responses to yeast to multiple stimuli. PMID:23612707

  16. Kappa Opioid Receptor-Induced Aversion Requires p38 MAPK Activation in VTA Dopamine Neurons

    PubMed Central

    Ehrich, Jonathan M.; Messinger, Daniel I.; Knakal, Cerise R.; Kuhar, Jamie R.; Schattauer, Selena S.; Bruchas, Michael R.; Zweifel, Larry S.; Kieffer, Brigitte L.; Phillips, Paul E.M.

    2015-01-01

    . Understanding how KOR activation produces dysphoria is key to the development of better analgesics and to defining how the endogenous dynorphin opioids produce their depression-like effects. Results in this study show that the aversive effects of κ receptor activation required arrestin-dependent p38α MAPK activation in dopamine neurons but did not require inhibition of dopamine release in the nucleus accumbens. Thus, contrary to the prevailing view, inhibition of mesolimbic dopamine release does not mediate the aversive effects of KOR activation and functionally selective κ opioids that do not activate arrestin signaling may be effective analgesics lacking dysphoric effects. PMID:26377476

  17. Morin suppresses inflammatory cytokine expression by downregulation of nuclear factor-κB and mitogen-activated protein kinase (MAPK) signaling pathways in lipopolysaccharide-stimulated primary bovine mammary epithelial cells.

    PubMed

    Wang, Jingjing; Guo, Changming; Wei, Zhengkai; He, Xuexiu; Kou, Jinhua; Zhou, Ershun; Yang, Zhengtao; Fu, Yunhe

    2016-04-01

    Morin, a flavonoid isolated from Chinese herbs of the Moraceae family, has been reported to possess antiinflammatory activity. However, the effects of morin on mastitis have not been investigated. The present study was conducted to elucidate the antiinflammatory properties of morin on lipopolysaccharide (LPS)-stimulated primary bovine mammary epithelial cells (bMEC). The viability of bMEC was analyzed by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium] assay. Subsequently, bMEC were stimulated with LPS in the presence or absence of morin. Gene expression of proinflammatory cytokines was determined by quantitative real-time PCR (qRT-PCR). Nuclear factor-κB (NF-κB), inhibitory kappa B (IκBα) protein, extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK) were detected by Western blotting. The results showed that cell viability was not affected by morin. Moreover, morin inhibited the gene expression of tumor necrosis factor-α (TNF-α), IL-6, and IL-1β in LPS-stimulated bMEC in a dose-dependent manner. Western blot analysis showed that morin suppressed the phosphorylation of IκBα, NF-κB unit p65, ERK, p38, and JNK in LPS-stimulated bMEC. In conclusion, the protective effects of morin on LPS-induced inflammatory response in bMEC may be due to its ability to suppress NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways. These findings suggest that morin may be used as antiinflammatory drug for mastitis.

  18. FGF/MAPK signaling sets the switching threshold of a bistable circuit controlling cell fate decisions in embryonic stem cells.

    PubMed

    Schröter, Christian; Rué, Pau; Mackenzie, Jonathan Peter; Martinez Arias, Alfonso

    2015-12-15

    Intracellular transcriptional regulators and extracellular signaling pathways together regulate the allocation of cell fates during development, but how their molecular activities are integrated to establish the correct proportions of cells with particular fates is not known. Here we study this question in the context of the decision between the epiblast (Epi) and the primitive endoderm (PrE) fate that occurs in the mammalian preimplantation embryo. Using an embryonic stem cell (ESC) model, we discover two successive functions of FGF/MAPK signaling in this decision. First, the pathway needs to be inhibited to make the PrE-like gene expression program accessible for activation by GATA transcription factors in ESCs. In a second step, MAPK signaling levels determine the threshold concentration of GATA factors required for PrE-like differentiation, and thereby control the proportion of cells differentiating along this lineage. Our findings can be explained by a simple mutual repression circuit modulated by FGF/MAPK signaling. This might be a general network architecture to integrate the activity of signal transduction pathways and transcriptional regulators, and serve to balance proportions of cell fates in several contexts.

  19. MAF mediates crosstalk between Ras-MAPK and mTOR signaling in NF1

    PubMed Central

    Brundage, Meghan E.; Tandon, Preeti; Eaves, David W.; Williams, Jon P.; Miller, Shyra J.; Hennigan, Robert H.; Jegga, Anil; Cripe, Timothy P.; Ratner, Nancy

    2014-01-01

    Mutations in the neurofibromatosis type 1 (NF1 tumor suppressor gene are common in cancer, and can cause resistance to therapy. Using transcriptome analysis we identified MAF as an NF1 regulated transcription factor, and verified MAF regulation through RAS/MAPK/AP-1 signaling in malignant peripheral nerve sheath tumor (MPNST) cell lines. MAF was also downregulated in human MPNST. Acute re-expression of MAF promoted expression of glial differentiation markers in MPNST cells in vitro, decreased self-renewal of embryonic precursors and transiently affected tumor cell phenotypes in vitro by increasing MPNST cell death and reducing metabolic activity and anchorage independent growth. Paradoxically, chronic MAF overexpression enhanced MPNST cell tumor growth in vivo, correlating with elevated pS6 in vitro and in vivo. RAD001 blocked MAF-mediated tumor growth, and MAF regulated the mTOR pathway through DEPTOR. MAPK inhibition with NF1 loss of function is predicted to show limited efficacy due to reactivation of mTOR signaling via MAF. PMID:24509877

  20. Weaning Induced Hepatic Oxidative Stress, Apoptosis, and Aminotransferases through MAPK Signaling Pathways in Piglets

    PubMed Central

    Luo, Zhen; Zhu, Wei; Guo, Qi; Luo, Wenli; Zhang, Jing; Xu, Weina

    2016-01-01

    This study investigated the effects of weaning on the hepatic redox status, apoptosis, function, and the mitogen-activated protein kinase (MAPK) signaling pathways during the first week after weaning in piglets. A total of 12 litters of piglets were weaned at d 21 and divided into the weaning group (WG) and the control group (CG). Six piglets from each group were slaughtered at d 0 (d 20, referred to weaning), d 1, d 4, and d 7 after weaning. Results showed that weaning significantly increased the concentrations of hepatic free radicals H2O2 and NO, malondialdehyde (MDA), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), while significantly decreasing the inhibitory hydroxyl ability (IHA) and glutathione peroxidase (GSH-Px), and altered the level of superoxide dismutase (SOD). The apoptosis results showed that weaning increased the concentrations of caspase-3, caspase-8, caspase-9 and the ratio of Bax/Bcl-2. In addition, aspartate aminotransferase transaminase (AST) and alanine aminotransferase (ALT) in liver homogenates increased after weaning. The phosphorylated JNK and ERK1/2 increased, while the activated p38 initially decreased and then increased. Our results suggested that weaning increased the hepatic oxidative stress and aminotransferases and initiated apoptosis, which may be related to the activated MAPK pathways in postweaning piglets. PMID:27807471

  1. Failure to Target RANKL Signaling Through p38-MAPK Results in Defective Osteoclastogenesis in the Microphthalmia Cloudy-Eyed Mutant.

    PubMed

    Carey, Heather A; Bronisz, Agnieszka; Cabrera, Jennifer; Hildreth, Blake E; Cuitiño, Maria; Fu, Qi; Ahmad, Asrar; Toribio, Ramiro E; Ostrowski, Michael C; Sharma, Sudarshana M

    2016-03-01

    The Microphthalmia-associated transcription factor (MITF) is a basic helix-loop-helix leucine zipper family factor that is essential for terminal osteoclast differentiation. Previous work demonstrates that phosphorylation of MITF by p38 MAPK downstream of Receptor Activator of NFkB Ligand (RANKL) signaling is necessary for MITF activation in osteoclasts. The spontaneous Mitf cloudy eyed (ce) allele results in production of a truncated MITF protein that lacks the leucine zipper and C-terminal end. Here we show that the Mitf(ce) allele leads to a dense bone phenotype in neonatal mice due to defective osteoclast differentiation. In response to RANKL stimulation, in vitro osteoclast differentiation was impaired in myeloid precursors derived from neonatal or adult Mitf(ce/ce) mice. The loss of the leucine zipper domain in Mitf(ce/ce) mice does not interfere with the recruitment of MITF/PU.1 complexes to target promoters. Further, we have mapped the p38 MAPK docking site within the region deleted in Mitf(ce). This interaction is necessary for the phosphorylation of MITF by p38 MAPK. Site-directed mutations in the docking site interfered with the interaction between MITF and its co-factors FUS and BRG1. MITF-ce fails to recruit FUS and BRG1 to target genes, resulting in decreased expression of target genes and impaired osteoclast function. These results highlight the crucial role of signaling dependent MITF/p38 MAPK interactions in osteoclast differentiation.

  2. Failure to Target RANKL Signaling Through p38-MAPK Results in Defective Osteoclastogenesis in the Microphthalmia Cloudy-Eyed Mutant.

    PubMed

    Carey, Heather A; Bronisz, Agnieszka; Cabrera, Jennifer; Hildreth, Blake E; Cuitiño, Maria; Fu, Qi; Ahmad, Asrar; Toribio, Ramiro E; Ostrowski, Michael C; Sharma, Sudarshana M

    2016-03-01

    The Microphthalmia-associated transcription factor (MITF) is a basic helix-loop-helix leucine zipper family factor that is essential for terminal osteoclast differentiation. Previous work demonstrates that phosphorylation of MITF by p38 MAPK downstream of Receptor Activator of NFkB Ligand (RANKL) signaling is necessary for MITF activation in osteoclasts. The spontaneous Mitf cloudy eyed (ce) allele results in production of a truncated MITF protein that lacks the leucine zipper and C-terminal end. Here we show that the Mitf(ce) allele leads to a dense bone phenotype in neonatal mice due to defective osteoclast differentiation. In response to RANKL stimulation, in vitro osteoclast differentiation was impaired in myeloid precursors derived from neonatal or adult Mitf(ce/ce) mice. The loss of the leucine zipper domain in Mitf(ce/ce) mice does not interfere with the recruitment of MITF/PU.1 complexes to target promoters. Further, we have mapped the p38 MAPK docking site within the region deleted in Mitf(ce). This interaction is necessary for the phosphorylation of MITF by p38 MAPK. Site-directed mutations in the docking site interfered with the interaction between MITF and its co-factors FUS and BRG1. MITF-ce fails to recruit FUS and BRG1 to target genes, resulting in decreased expression of target genes and impaired osteoclast function. These results highlight the crucial role of signaling dependent MITF/p38 MAPK interactions in osteoclast differentiation. PMID:26218069

  3. OsMAPK6, a mitogen-activated protein kinase, influences rice grain size and biomass production.

    PubMed

    Liu, Shuying; Hua, Lei; Dong, Sujun; Chen, Hongqi; Zhu, Xudong; Jiang, Jun'e; Zhang, Fang; Li, Yunhai; Fang, Xiaohua; Chen, Fan

    2015-11-01

    Grain size is an important agronomic trait in determining grain yield. However, the molecular mechanisms that determine the final grain size are not well understood. Here, we report the functional analysis of a rice (Oryza sativa L.) mutant, dwarf and small grain1 (dsg1), which displays pleiotropic phenotypes, including small grains, dwarfism and erect leaves. Cytological observations revealed that the small grain and dwarfism of dsg1 were mainly caused by the inhibition of cell proliferation. Map-based cloning revealed that DSG1 encoded a mitogen-activated protein kinase (MAPK), OsMAPK6. OsMAPK6 was mainly located in the nucleus and cytoplasm, and was ubiquitously distributed in various organs, predominately in spikelets and spikelet hulls, consistent with its role in grain size and biomass production. As a functional kinase, OsMAPK6 interacts strongly with OsMKK4, indicating that OsMKK4 is likely to be the upstream MAPK kinase of OsMAPK6 in rice. In addition, hormone sensitivity tests indicated that the dsg1 mutant was less sensitive to brassinosteroids (BRs). The endogenous BR levels were reduced in dsg1, and the expression of several BR signaling pathway genes and feedback-inhibited genes was altered in the dsg1 mutant, with or without exogenous BRs, indicating that OsMAPK6 may contribute to influence BR homeostasis and signaling. Thus, OsMAPK6, a MAPK, plays a pivotal role in grain size in rice, via cell proliferation, and BR signaling and homeostasis. PMID:26366992

  4. Overexpression of LRIG1 regulates PTEN via MAPK/MEK signaling pathway in esophageal squamous cell carcinoma

    PubMed Central

    Jiang, Xiaofang; Li, Huiwu

    2016-01-01

    The present study aimed to evaluate the role of leucine-rich repeats and immunoglobulin-like domain protein 1 (LRIG1) in the regulation of phosphatase and tensin homolog (PTEN) expression in esophageal carcinogenesis. LRIG1 was overexpressed in esophageal squamous cell carcinoma (ESCC) cell lines, and the effect of LRIG1 overexpression on the mRNA and protein expression levels of PTEN was evaluated by reverse transcription-quantitative polymerase chain reaction and western blotting. Furthermore, the effects of LRIG1 overexpression on the cell cycle distribution and apoptosis of ESCC cells were examined by flow cytometry. Various cell signaling pathway inhibitors were used to assess the effects of LRIG1 on downstream signaling in ESCC cell lines. In addition, the association between LRIG1 and PTEN expression was examined in 48 samples from patients with ESCC. LRIG1 overexpression was demonstrated to downregulate PTEN expression in ESCC cell lines, and promote their proliferation and inhibit apoptosis. In addition, LRIG1-mediated suppression of PTEN expression was inhibited by the U0126 inhibitor, which suggests that LRIG1 may inhibit the activation of PTEN signaling molecules by triggering the mitogen-activated protein kinase (MAPK)/MAPK kinase 1 (MEK) signaling pathway. In conclusion, the present study demonstrated that overexpression of LRIG1 significantly and adversely affected the survival of ESCC cells, and that the MAPK/MEK signaling pathway may be responsible for the repression of PTEN expression and function. PMID:27698691

  5. Overexpression of LRIG1 regulates PTEN via MAPK/MEK signaling pathway in esophageal squamous cell carcinoma

    PubMed Central

    Jiang, Xiaofang; Li, Huiwu

    2016-01-01

    The present study aimed to evaluate the role of leucine-rich repeats and immunoglobulin-like domain protein 1 (LRIG1) in the regulation of phosphatase and tensin homolog (PTEN) expression in esophageal carcinogenesis. LRIG1 was overexpressed in esophageal squamous cell carcinoma (ESCC) cell lines, and the effect of LRIG1 overexpression on the mRNA and protein expression levels of PTEN was evaluated by reverse transcription-quantitative polymerase chain reaction and western blotting. Furthermore, the effects of LRIG1 overexpression on the cell cycle distribution and apoptosis of ESCC cells were examined by flow cytometry. Various cell signaling pathway inhibitors were used to assess the effects of LRIG1 on downstream signaling in ESCC cell lines. In addition, the association between LRIG1 and PTEN expression was examined in 48 samples from patients with ESCC. LRIG1 overexpression was demonstrated to downregulate PTEN expression in ESCC cell lines, and promote their proliferation and inhibit apoptosis. In addition, LRIG1-mediated suppression of PTEN expression was inhibited by the U0126 inhibitor, which suggests that LRIG1 may inhibit the activation of PTEN signaling molecules by triggering the mitogen-activated protein kinase (MAPK)/MAPK kinase 1 (MEK) signaling pathway. In conclusion, the present study demonstrated that overexpression of LRIG1 significantly and adversely affected the survival of ESCC cells, and that the MAPK/MEK signaling pathway may be responsible for the repression of PTEN expression and function.

  6. Control of MAPK signaling specificity by a conserved residue in the MEK-binding domain of the yeast scaffold protein Ste5.

    PubMed

    Schwartz, Monica A; Madhani, Hiten D

    2006-06-01

    The yeast kinase scaffold Ste5 has been proposed to prevent unwanted cross-talk between the pheromone response pathway and other MAPK cascades. Protein fusion experiments have demonstrated that covalently tethering signaling components to each other or to Ste5 can determine the outcome of signaling. However, these do not fully test the role of scaffolds in signaling specificity, since fusing components precludes differential dissociation of subpopulations. We performed a targeted genetic screen on STE5 and repeatedly identified recessive mutations in a conserved residue, E756, in the Ste7/MEK-binding domain that caused erroneous activation of the filamentation MAPK pathway by pheromone signaling. Mutant cells exhibited a shift in the MAPK activation pattern such that the filamentation MAPK Kss1 was predominately activated in response to pheromone. Velocity sedimentation studies showed that the mutant scaffold was defective in binding to a phosphorylated subpopulation of Ste7. Our data suggest that increased dissociation of activated Ste7 kinase from the mutant scaffold may cause the observed shift in MAPK activation from Fus3 to Kss1 and the resulting loss of specificity. Cross-talk in ste5-E756G cells was due to both increased activation of Kss1 and reduced Fus3-dependent degradation of the filamentation pathway transcription factor Tec1. These studies demonstrate a role for an endogenous scaffold in signaling specificity. PMID:16463042

  7. The Mitogen-Activated Protein Kinase (MAPK) Pathway: Role in Immune Evasion by Trypanosomatids.

    PubMed

    Soares-Silva, Mercedes; Diniz, Flavia F; Gomes, Gabriela N; Bahia, Diana

    2016-01-01

    Leishmania spp. and Trypanosoma cruzi are the causative agents of leishmaniasis and Chagas disease, respectively, two neglected tropical diseases that affect about 25 million people worldwide. These parasites belong to the family Trypanosomatidae, and are both obligate intracellular parasites that manipulate host signaling pathways and the innate immune system to establish infection. Mitogen-activated protein kinases (MAPKs) are serine and threonine protein kinases that are highly conserved in eukaryotes, and are involved in signal transduction pathways that modulate physiological and pathophysiological cell responses. This mini-review highlights existing knowledge concerning the mechanisms that Leishmania spp. and T. cruzi have evolved to target the host's MAPK signaling pathways and highjack the immune response, and, in this manner, promote parasite maintenance in the host. PMID:26941717

  8. The Mitogen-Activated Protein Kinase (MAPK) Pathway: Role in Immune Evasion by Trypanosomatids

    PubMed Central

    Soares-Silva, Mercedes; Diniz, Flavia F.; Gomes, Gabriela N.; Bahia, Diana

    2016-01-01

    Leishmania spp. and Trypanosoma cruzi are the causative agents of leishmaniasis and Chagas disease, respectively, two neglected tropical diseases that affect about 25 million people worldwide. These parasites belong to the family Trypanosomatidae, and are both obligate intracellular parasites that manipulate host signaling pathways and the innate immune system to establish infection. Mitogen-activated protein kinases (MAPKs) are serine and threonine protein kinases that are highly conserved in eukaryotes, and are involved in signal transduction pathways that modulate physiological and pathophysiological cell responses. This mini-review highlights existing knowledge concerning the mechanisms that Leishmania spp. and T. cruzi have evolved to target the host’s MAPK signaling pathways and highjack the immune response, and, in this manner, promote parasite maintenance in the host. PMID:26941717

  9. p38 MAPK and PI3K/AKT Signalling Cascades inParkinson’s Disease

    PubMed Central

    Jha, Saurabh Kumar; Jha, Niraj Kumar; Kar, Rohan; Ambasta, Rashmi K; Kumar, Pravir

    2015-01-01

    Parkinson's disease (PD) is a chronic neurodegenerative condition which has the second largest incidence rate among all other neurodegenerative disorders barring Alzheimer's disease (AD). Currently there is no cure and researchers continue to probe the therapeutic prospect in cell cultures and animal models of PD. Out of the several factors contributing to PD prognosis, the role of p38 MAPK (Mitogen activated protein-kinase) and PI3K/AKT signalling module in PD brains is crucial because the impaired balance between the pro- apoptotic and anti-apoptotic pathways trigger unwanted phenotypes such as microglia activation, neuroinflammation, oxidative stress and apoptosis. These factors continue challenging the brain homeostasis in initial stages thereby essentially assisting the dopaminergic (DA) neurons towards progressive degeneration in PD. Neurotherapeutics against PD shall then be targeted against the misregulated accomplices of the p38 and PI3K/AKT cascades. In this review, we have outlined many such established mechanisms involving the p38 MAPK and PI3K/AKT pathways which can offer therapeutic windows for the rectification of aberrant DA neuronal dynamics in PD brains. PMID:26261796

  10. Down-regulation of the P-glycoprotein relevant for multidrug resistance by intracellular acidification through the crosstalk of MAPK signaling pathways.

    PubMed

    Jin, Weina; Lu, Ying; Li, Qinghua; Wang, Jian; Zhang, Hongju; Chang, Guoqiang; Lin, Yani; Pang, Tianxiang

    2014-09-01

    In our previous study, we have found that the tumor multidrug resistance mediated by P-glycoprotein could be reversed by sustained intracellular acidification through down-regulating the multidrug resistance gene 1 mRNA and P-glycoprotein expression. However, the molecular events linking the intracellular acidification and the regulation of P-glycoprotein remain unclear. In the present study, the molecular pathways involved in the regulation of P-glycoprotein expression by the intracellular acidification were investigated. We found that the P-glycoprotein expression was down-regulated by the intracellular acidification through inhibition of p38 mitogen-activated protein kinase (MAPK) and the activation of c-Jun N-terminal kinase (JNK) in the resisitant K562/DOX cells. In the sensitive K562 and HL60 cell lines, the changes of the p38 MAPK expression after the acidification are not as obvious as that of K562/DOX cells, but the activation of extracellular signal-regulated kinase (ERK) is also observed, which indicates that the down-regulation of p38 MAPK by the intracellular acidification might be the resistant cell line specific. Blockade of ERK and JNK signaling by the inhibitors or RNA interference increased p38MAPK activities suggesting that cross-talk within MAPKs is also important for this response. Our study provides the first direct evidence that the reversal of P-glycoprotein-mediated multidrug resistance by intracellular acidification is mediated by the crosstalk of MAPK signaling pathways.

  11. Morin Attenuates Ovalbumin-Induced Airway Inflammation by Modulating Oxidative Stress-Responsive MAPK Signaling

    PubMed Central

    Ma, Yuan; Ge, Ai; Zhu, Wen; Liu, Ya-Nan; Ji, Ning-Fei; Zha, Wang-Jian; Zhang, Jia-Xiang; Zeng, Xiao-Ning

    2016-01-01

    Asthma is one of the most common inflammatory diseases characterized by airway hyperresponsiveness, inflammation, and remodeling. Morin, an active ingredient obtained from Moraceae plants, has been demonstrated to have promising anti-inflammatory activities in a range of disorders. However, its impacts on pulmonary diseases, particularly on asthma, have not been clarified. This study was designed to investigate whether morin alleviates airway inflammation in chronic asthma with an emphasis on oxidative stress modulation. In vivo, ovalbumin- (OVA-) sensitized mice were administered with morin or dexamethasone before challenge. Bronchoalveolar lavage fluid (BALF) and lung tissues were obtained to perform cell counts, histological analysis, and enzyme-linked immunosorbent assay. In vitro, human bronchial epithelial cells (BECs) were challenged by tumor necrosis factor alpha (TNF-α). The supernatant was collected for the detection of the proinflammatory proteins, and the cells were collected for reactive oxygen species (ROS)/mitogen-activated protein kinase (MAPK) evaluations. Severe inflammatory responses and remodeling were observed in the airways of the OVA-sensitized mice. Treatment with morin dramatically attenuated the extensive trafficking of inflammatory cells into the BALF and inhibited their infiltration around the respiratory tracts and vessels. Morin administration also significantly suppressed goblet cell hyperplasia and collagen deposition/fibrosis and dose-dependently inhibited the OVA-induced increases in IgE, TNF-α, interleukin- (IL-) 4, IL-13, matrix metalloproteinase-9, and malondialdehyde. In human BECs challenged by TNF-α, the levels of proteins such as eotaxin-1, monocyte chemoattractant protein-1, IL-8 and intercellular adhesion molecule-1, were consistently significantly decreased by morin. Western blotting and the 2′,7′-dichlorofluorescein assay revealed that the increases in intracellular ROS and MAPK phosphorylation were abolished by

  12. Morin Attenuates Ovalbumin-Induced Airway Inflammation by Modulating Oxidative Stress-Responsive MAPK Signaling.

    PubMed

    Ma, Yuan; Ge, Ai; Zhu, Wen; Liu, Ya-Nan; Ji, Ning-Fei; Zha, Wang-Jian; Zhang, Jia-Xiang; Zeng, Xiao-Ning; Huang, Mao

    2016-01-01

    Asthma is one of the most common inflammatory diseases characterized by airway hyperresponsiveness, inflammation, and remodeling. Morin, an active ingredient obtained from Moraceae plants, has been demonstrated to have promising anti-inflammatory activities in a range of disorders. However, its impacts on pulmonary diseases, particularly on asthma, have not been clarified. This study was designed to investigate whether morin alleviates airway inflammation in chronic asthma with an emphasis on oxidative stress modulation. In vivo, ovalbumin- (OVA-) sensitized mice were administered with morin or dexamethasone before challenge. Bronchoalveolar lavage fluid (BALF) and lung tissues were obtained to perform cell counts, histological analysis, and enzyme-linked immunosorbent assay. In vitro, human bronchial epithelial cells (BECs) were challenged by tumor necrosis factor alpha (TNF-α). The supernatant was collected for the detection of the proinflammatory proteins, and the cells were collected for reactive oxygen species (ROS)/mitogen-activated protein kinase (MAPK) evaluations. Severe inflammatory responses and remodeling were observed in the airways of the OVA-sensitized mice. Treatment with morin dramatically attenuated the extensive trafficking of inflammatory cells into the BALF and inhibited their infiltration around the respiratory tracts and vessels. Morin administration also significantly suppressed goblet cell hyperplasia and collagen deposition/fibrosis and dose-dependently inhibited the OVA-induced increases in IgE, TNF-α, interleukin- (IL-) 4, IL-13, matrix metalloproteinase-9, and malondialdehyde. In human BECs challenged by TNF-α, the levels of proteins such as eotaxin-1, monocyte chemoattractant protein-1, IL-8 and intercellular adhesion molecule-1, were consistently significantly decreased by morin. Western blotting and the 2',7'-dichlorofluorescein assay revealed that the increases in intracellular ROS and MAPK phosphorylation were abolished by morin

  13. Eupatilin induces human renal cancer cell apoptosis via ROS-mediated MAPK and PI3K/AKT signaling pathways

    PubMed Central

    Zhong, Wei-Feng; Wang, Xiao-Hong; Pan, Bin; Li, Feng; Kuang, Lu; Su, Ze-Xuan

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K)/AKT and mitogen activated protein kinase (MAPK) signaling cascades have significant roles in cell proliferation, survival, angiogenesis and metastasis of tumor cells. Eupatilin, one of the major compounds present in Artemisia species, has been demonstrated to have antitumor properties. However, the effect of eupatilin in renal cell carcinoma (RCC) remains to be elucidated. Therefore, the present study investigated the biological effects and mechanisms of eupatilin in RCC cell apoptosis. The results of the present study demonstrated that eupatilin significantly induced cell apoptosis and enhanced the production of reactive oxygen species (ROS) in 786-O cells. In addition, eupatilin induced phosphorylation of p38α (Thr180/Tyr182), extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase 1/2 (Thr183/Tyr185), and decreased the phosphorylation of PI3K and AKT in 786-O cells in a concentration-dependent manner. Furthermore, the ROS inhibitor N-acetyl-L-cysteine was able to rescue the MAPK activation and PI3K/AKT inhibition induced by eupatilin. Taken together, the results of the present study provide evidence that inhibition of eupatilin induces apoptosis in human RCC via ROS-mediated activation of the MAPK signaling pathway and inhibition of the PI3K/AKT signaling pathway. Thus, eupatilin may serve as a potential therapeutic agent for the treatment of human RCC. PMID:27698876

  14. Eupatilin induces human renal cancer cell apoptosis via ROS-mediated MAPK and PI3K/AKT signaling pathways

    PubMed Central

    Zhong, Wei-Feng; Wang, Xiao-Hong; Pan, Bin; Li, Feng; Kuang, Lu; Su, Ze-Xuan

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K)/AKT and mitogen activated protein kinase (MAPK) signaling cascades have significant roles in cell proliferation, survival, angiogenesis and metastasis of tumor cells. Eupatilin, one of the major compounds present in Artemisia species, has been demonstrated to have antitumor properties. However, the effect of eupatilin in renal cell carcinoma (RCC) remains to be elucidated. Therefore, the present study investigated the biological effects and mechanisms of eupatilin in RCC cell apoptosis. The results of the present study demonstrated that eupatilin significantly induced cell apoptosis and enhanced the production of reactive oxygen species (ROS) in 786-O cells. In addition, eupatilin induced phosphorylation of p38α (Thr180/Tyr182), extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase 1/2 (Thr183/Tyr185), and decreased the phosphorylation of PI3K and AKT in 786-O cells in a concentration-dependent manner. Furthermore, the ROS inhibitor N-acetyl-L-cysteine was able to rescue the MAPK activation and PI3K/AKT inhibition induced by eupatilin. Taken together, the results of the present study provide evidence that inhibition of eupatilin induces apoptosis in human RCC via ROS-mediated activation of the MAPK signaling pathway and inhibition of the PI3K/AKT signaling pathway. Thus, eupatilin may serve as a potential therapeutic agent for the treatment of human RCC.

  15. The activation of p38MAPK and JNK pathways in bovine herpesvirus 1 infected MDBK cells.

    PubMed

    Zhu, Liqian; Yuan, Chen; Huang, Liyuan; Ding, Xiuyan; Wang, Jianye; Zhang, Dong; Zhu, Guoqiang

    2016-01-01

    We have shown previously that BHV-1 infection activates Erk1/2 signaling. Here, we show that BHV-1 provoked an early-stage transient and late-stage sustained activation of JNK, p38MAPK and c-Jun signaling in MDBK cells. C-Jun phosphorylation was dependent on JNK. These early events were partially due to the viral entry process. Unexpectedly, reactive oxygen species were not involved in the later activation phase. Interestingly, only activated JNK facilitated the viral multiplication identified through both chemical inhibitor and siRNA. Collectively, this study provides insight into our understanding of early stages of BHV-1 infection. PMID:27590675

  16. The SNA analysis of a minimal model for bistability in the MAPK signaling cascade model

    NASA Astrophysics Data System (ADS)

    Hadač, O.; Schreiber, I.

    2013-12-01

    Successive phosphorylation cascades mediated by mitogen-activated protein kinases (MAPKs) are known to act as switches initiating various cellular processes. In addition, models of the MAPK reaction network are displaying other nonlinear phenomena including bistability and periodic oscillations. Recently bistability has been explained as a consequence of interaction between single and double phosphorylation/dephosphorylation pathways in the Stage 2 subsystem of the Huang-Ferrell model of the MAPK and a core bistable model has been proposed. Here we focus on a detailed stability analysis of the steady states of this minimal model. The analysis uses methods of convex analysis and stoichiometric network theory.

  17. Atractylodin Inhibits Interleukin-6 by Blocking NPM-ALK Activation and MAPKs in HMC-1.

    PubMed

    Chae, Hee-Sung; Kim, Young-Mi; Chin, Young-Won

    2016-09-02

    Atractylodin is one of the major constituents of the rhizome of Atractylodes lancea, which is widely used in Korean traditional medicine as a remedy for the treatment of gastritis and gastric ulcers. Despite of a major constituent of widely used botanical to treat inflammatory responses little is known about anti-inflammatory effect of atractylodin in the human mast cell (HMC-1). Hence, we evaluated the effect of atractylodin on the release of IL-6, the involvement of nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) and mitogen-activated protein kinases (MAPKs) in phorbol-12-myristate-13-acetate and A23187-induced HMC-1. In addition, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), phospholipase C (PLC) gamma 1, and AKT phosphorylation relevant to NPM-ALK signal pathway were assessed. IL-6 levels in the HMC-1 stimulated by phorbol-12-myristate-13-acetate and A23187 were apparently decreased by the treatment of atractylodin. Concurrently, atractylodin not only inhibited the phosphorylation of NPM-ALK, but also suppressed the phosphorylation of JAK2, STAT3, PLC gamma 1, and AKT. Furthermore, the activated mitogen-activated protein kinases (MAPKs) by phorbol-12-myristate-13-acetate and A23187 were inhibited by atractylodin. These results suggested that atractylodin might have a potential regulatory effect on inflammatory mediator expression through blockade of both the phosphorylation of MAPKs and the NPM-ALK signaling pathway.

  18. Atractylodin Inhibits Interleukin-6 by Blocking NPM-ALK Activation and MAPKs in HMC-1.

    PubMed

    Chae, Hee-Sung; Kim, Young-Mi; Chin, Young-Won

    2016-01-01

    Atractylodin is one of the major constituents of the rhizome of Atractylodes lancea, which is widely used in Korean traditional medicine as a remedy for the treatment of gastritis and gastric ulcers. Despite of a major constituent of widely used botanical to treat inflammatory responses little is known about anti-inflammatory effect of atractylodin in the human mast cell (HMC-1). Hence, we evaluated the effect of atractylodin on the release of IL-6, the involvement of nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) and mitogen-activated protein kinases (MAPKs) in phorbol-12-myristate-13-acetate and A23187-induced HMC-1. In addition, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), phospholipase C (PLC) gamma 1, and AKT phosphorylation relevant to NPM-ALK signal pathway were assessed. IL-6 levels in the HMC-1 stimulated by phorbol-12-myristate-13-acetate and A23187 were apparently decreased by the treatment of atractylodin. Concurrently, atractylodin not only inhibited the phosphorylation of NPM-ALK, but also suppressed the phosphorylation of JAK2, STAT3, PLC gamma 1, and AKT. Furthermore, the activated mitogen-activated protein kinases (MAPKs) by phorbol-12-myristate-13-acetate and A23187 were inhibited by atractylodin. These results suggested that atractylodin might have a potential regulatory effect on inflammatory mediator expression through blockade of both the phosphorylation of MAPKs and the NPM-ALK signaling pathway. PMID:27598116

  19. Rewiring yeast osmostress signalling through the MAPK network reveals essential and non-essential roles of Hog1 in osmoadaptation.

    PubMed

    Babazadeh, Roja; Furukawa, Takako; Hohmann, Stefan; Furukawa, Kentaro

    2014-01-01

    Mitogen-activated protein kinases (MAPKs) have a number of targets which they regulate at transcriptional and post-translational levels to mediate specific responses. The yeast Hog1 MAPK is essential for cell survival under hyperosmotic conditions and it plays multiple roles in gene expression, metabolic regulation, signal fidelity and cell cycle regulation. Here we describe essential and non-essential roles of Hog1 using engineered yeast cells in which osmoadaptation was reconstituted in a Hog1-independent manner. We rewired Hog1-dependent osmotic stress-induced gene expression under the control of Fus3/Kss1 MAPKs, which are activated upon osmostress via crosstalk in hog1Δ cells. This approach revealed that osmotic up-regulation of only two Hog1-dependent glycerol biosynthesis genes, GPD1 and GPP2, is sufficient for successful osmoadaptation. Moreover, some of the previously described Hog1-dependent mechanisms appeared to be dispensable for osmoadaptation in the engineered cells. These results suggest that the number of essential MAPK functions may be significantly smaller than anticipated and that knockout approaches may lead to over-interpretation of phenotypic data. PMID:24732094

  20. Rewiring yeast osmostress signalling through the MAPK network reveals essential and non-essential roles of Hog1 in osmoadaptation

    PubMed Central

    Babazadeh, Roja; Furukawa, Takako; Hohmann, Stefan; Furukawa, Kentaro

    2014-01-01

    Mitogen-activated protein kinases (MAPKs) have a number of targets which they regulate at transcriptional and post-translational levels to mediate specific responses. The yeast Hog1 MAPK is essential for cell survival under hyperosmotic conditions and it plays multiple roles in gene expression, metabolic regulation, signal fidelity and cell cycle regulation. Here we describe essential and non-essential roles of Hog1 using engineered yeast cells in which osmoadaptation was reconstituted in a Hog1-independent manner. We rewired Hog1-dependent osmotic stress-induced gene expression under the control of Fus3/Kss1 MAPKs, which are activated upon osmostress via crosstalk in hog1Δ cells. This approach revealed that osmotic up-regulation of only two Hog1-dependent glycerol biosynthesis genes, GPD1 and GPP2, is sufficient for successful osmoadaptation. Moreover, some of the previously described Hog1-dependent mechanisms appeared to be dispensable for osmoadaptation in the engineered cells. These results suggest that the number of essential MAPK functions may be significantly smaller than anticipated and that knockout approaches may lead to over-interpretation of phenotypic data. PMID:24732094

  1. Regulation of MAPK/ERK Signaling and Photic Entrainment of the Suprachiasmatic Nucleus Circadian Clock by Raf Kinase Inhibitor Protein

    PubMed Central

    Antoun, Ghadi; Cannon, Pascale Bouchard; Cheng, Hai-Ying Mary

    2013-01-01

    Activation of the MAPK/ERK signaling cascade in the suprachiasmatic nucleus (SCN) is a key event that couples light to circadian clock entrainment. However, we do not fully understand the mechanisms that shape the properties of MAPK/ERK signaling in the SCN, and how these mechanisms may influence overt circadian rhythms. Here we show that Raf kinase inhibitor protein (RKIP) controls the kinetics of light-induced MAPK/ERK activity in the SCN and photic entrainment of behavioral rhythms. Light triggers robust phosphorylation of RKIP in the murine SCN and dissociation of RKIP and c-Raf. Overexpression of a nonphosphorylatable form of RKIP in the SCN of transgenic mice blocks light-induced ERK1/2 activation in the SCN and severely dampens light-induced phase delays in behavioral rhythms. Conversely, in RKIP knock-out (RKIP−/−) mice, light-induced ERK1/2 activity in the SCN is prolonged in the early and late subjective night, resulting in augmentation of the phase-delaying and -advancing effects of light. Reentrainment to an advancing light cycle was also accelerated in RKIP−/− mice. In relation to the molecular clockwork, genetic deletion of RKIP potentiated light-evoked PER1 and PER2 protein expression in the SCN in the early night. Additionally, RKIP−/− mice displayed enhanced transcriptional activation of mPeriod1 and the immediate early gene c-Fos in the SCN in response to a phase-delaying light pulse. Collectively, our data reveal an important role of RKIP in the regulation of MAPK/ERK signaling in the SCN and photic entrainment of the SCN clock. PMID:22492043

  2. MAPK specificity in the yeast pheromone response independent of transcriptional activation.

    PubMed

    Breitkreutz, A; Boucher, L; Tyers, M

    2001-08-21

    The mechanisms whereby different external cues stimulate the same mitogen-activated protein kinase (MAPK) cascade, yet trigger an appropriately distinct biological response, epitomize the conundrum of specificity in cell signaling. In yeast, shared upstream components of the mating pheromone and filamentous growth pathways activate two related MAPKs, Fus3 and Kss1, which in turn regulate programs of gene expression via the transcription factor Ste12. As fus3, but not kss1, strains are impaired for mating, Fus3 exhibits specificity for the pheromone response. To account for this specificity, it has been suggested that Fus3 physically occludes Kss1 from pheromone-activated signaling complexes, which are formed on the scaffold protein Ste5. However, we find that genome-wide expression profiles of pheromone-treated wild-type, fus3, and kss1 deletion strains are highly correlated for all induced genes and, further, that two catalytically inactive versions of Fus3 fail to abrogate the pheromone-induced transcriptional response. Consistently, Fus3 and Kss1 kinase activity is induced to an equivalent extent in pheromone-treated cells. In contrast, both in vivo and in an in vitro-reconstituted MAPK system, Fus3, but not Kss1, exhibits strong substrate selectivity toward Far1, a bifunctional protein required for polarization and G(1) arrest. This effect accounts for the failure to repress G(1)-S specific transcription in fus3 strains and, in part, explains the mating defect of such strains. MAPK specificity in the pheromone response evidently occurs primarily at the substrate level, as opposed to specific kinase activation by dedicated signaling complexes. PMID:11525741

  3. 1-Bromopropane induces macrophage activation via extracellular signal-regulated kinase 1/2 MAPK and NF-κB pathways.

    PubMed

    Han, Eun Hee; Hwang, Yong Pil; Lee, Kyung Jin; Jeong, Tae Cheon; Jeong, Hye Gwang

    2008-04-01

    1-Bromopropane (1-BP) has been used in the workplace as an alternative to ozone-depleting solvents. This study examined the effects of 1-BP on the production of nitric oxide (NO) and on proinflammatory cytokines, and analyzed the mechanisms involved in macrophages. 1-BP dose-dependently induced the production of NO and proinflammatory cytokines, such as IL-1β, IL-6, and TNF-α, and expression levels of these genes also increased in a dose-dependent manner. The NF-κB sites were identified in the promoter of the iNOS and proinflammatory cytokine genes. Transient transfection and electrophoretic mobility shift assays revealed that NF-κB-mediated the 1-BP-induced increase in the iNOS and proinflammatory cytokine expression levels. Pretreating the macrophages with the NF-κB inhibitor, BAY 11-7082, and the ERK inhibitor, PD98059, inhibited NO production and iNOS expression induced by 1-BP. This demonstrates that 1-BP stimulates macrophage activation via NF-κB transactivation and ERK1/2 MAP kinase phosphorylation. These results suggest that 1-BP has the potential to be inflammatory and that it has previously unrecognized immunomodulating activity.

  4. Gab1 and Mapk Signaling Are Essential in the Hair Cycle and Hair Follicle Stem Cell Quiescence.

    PubMed

    Akilli Öztürk, Özlem; Pakula, Hubert; Chmielowiec, Jolanta; Qi, Jingjing; Stein, Simone; Lan, Linxiang; Sasaki, Yoshiteru; Rajewsky, Klaus; Birchmeier, Walter

    2015-10-20

    Gab1 is a scaffold protein that acts downstream of receptor tyrosine kinases. Here, we produced conditional Gab1 mutant mice (by K14- and Krox20-cre) and show that Gab1 mediates crucial signals in the control of both the hair cycle and the self-renewal of hair follicle stem cells. Remarkably, mutant hair follicles do not enter catagen, the destructive phase of the hair cycle. Instead, hair follicle stem cells lose quiescence and become exhausted, and thus no stem cell niches are established in the bulges. Moreover, conditional sustained activation of Mapk signaling by expression of a gain-of-function Mek1(DD) allele (by Krox20-cre) rescues hair cycle deficits and restores quiescence of the stem cells. Our data thus demonstrate an essential role of Gab1 downstream of receptor tyrosine kinases and upstream of Shp2 and Mapk in the regulation of the hair cycle and the self-renewal of hair follicle stem cells.

  5. Antiallergic Activity of Ethanol Extracts of Arctium lappa L. Undried Roots and Its Active Compound, Oleamide, in Regulating FcεRI-Mediated and MAPK Signaling in RBL-2H3 Cells.

    PubMed

    Yang, Woong-Suk; Lee, Sung Ryul; Jeong, Yong Joon; Park, Dae Won; Cho, Young Mi; Joo, Hae Mi; Kim, Inhye; Seu, Young-Bae; Sohn, Eun-Hwa; Kang, Se Chan

    2016-05-11

    The antiallergic potential of Arctium lappa L. was investigated in Sprague-Dawley rats, ICR mice, and RBL-2H3 cells. Ethanol extract (90%) of A. lappa (ALE, 100 μg/mL) inhibited the degranulation rate by 52.9%, determined by the level of β-hexosaminidase. ALE suppressed passive cutaneous anaphylaxis (PCA) in rats and attenuated anaphylaxis and histamine release in mice. To identify the active compound of ALE, we subsequently fractionated and determined the level of β-hexosaminidase in all subfractions. Oleamide was identified as an active compound of ALE, which attenuated the secretion of histamine and the production of tumor necrosis factor (TNF)-α and interleukin-4 (IL-4) in cells treated with compound 48/80 or A23187/phorbol myristate acetate (PMA). Oleamide suppressed FcεRI-tyrosine kinase Lyn-mediated pathway, c-Jun N-terminal kinases (JNK/SAPK), and p38 mitogen-activated protein kinases (p38-MAPKs). These results showed that ALE and oleamide attenuated allergic reactions and should serve as a platform to search for compounds with antiallergic activity.

  6. Antiallergic Activity of Ethanol Extracts of Arctium lappa L. Undried Roots and Its Active Compound, Oleamide, in Regulating FcεRI-Mediated and MAPK Signaling in RBL-2H3 Cells.

    PubMed

    Yang, Woong-Suk; Lee, Sung Ryul; Jeong, Yong Joon; Park, Dae Won; Cho, Young Mi; Joo, Hae Mi; Kim, Inhye; Seu, Young-Bae; Sohn, Eun-Hwa; Kang, Se Chan

    2016-05-11

    The antiallergic potential of Arctium lappa L. was investigated in Sprague-Dawley rats, ICR mice, and RBL-2H3 cells. Ethanol extract (90%) of A. lappa (ALE, 100 μg/mL) inhibited the degranulation rate by 52.9%, determined by the level of β-hexosaminidase. ALE suppressed passive cutaneous anaphylaxis (PCA) in rats and attenuated anaphylaxis and histamine release in mice. To identify the active compound of ALE, we subsequently fractionated and determined the level of β-hexosaminidase in all subfractions. Oleamide was identified as an active compound of ALE, which attenuated the secretion of histamine and the production of tumor necrosis factor (TNF)-α and interleukin-4 (IL-4) in cells treated with compound 48/80 or A23187/phorbol myristate acetate (PMA). Oleamide suppressed FcεRI-tyrosine kinase Lyn-mediated pathway, c-Jun N-terminal kinases (JNK/SAPK), and p38 mitogen-activated protein kinases (p38-MAPKs). These results showed that ALE and oleamide attenuated allergic reactions and should serve as a platform to search for compounds with antiallergic activity. PMID:27087645

  7. TLN-4601 suppresses growth and induces apoptosis of pancreatic carcinoma cells through inhibition of Ras-ERK MAPK signaling

    PubMed Central

    2010-01-01

    Background TLN-4601 is a structurally novel farnesylated dibenzodiazepinone discovered using Thallion's proprietary DECIPHER® technology, a genomics and bioinformatics platform that predicts the chemical structures of secondary metabolites based on gene sequences obtained by scanning bacterial genomes. Our recent studies suggest that TLN-4601 inhibits the Ras-ERK MAPK pathway post Ras prenylation and prior to MEK activation. The Ras-ERK MAPK signaling pathway is a well-validated oncogenic cascade based on its central role in regulating the growth and survival of cells from a broad spectrum of human tumors. Furthermore, RAS isoforms are the most frequently mutated oncogenes, occurring in approximately 30% of all human cancers, and KRAS is the most commonly mutated RAS gene, with a greater than 90% incidence of mutation in pancreatic cancer. Results To evaluate whether TLN-4601 interferes with K-Ras signaling, we utilized human pancreatic epithelial cells and demonstrate that TLN-4601 treatment resulted in a dose- and time-dependent inhibition of Ras-ERK MAPK signaling. The compound also reduced Ras-GTP levels and induced apoptosis. Finally, treatment of MIA PaCa-2 tumor-bearing mice with TLN-4601 resulted in antitumor activity and decreased tumor Raf-1 protein levels. Conclusion These data, together with phase I/II clinical data showing tolerability of TLN-4601, support conducting a clinical trial in advanced pancreatic cancer patients. PMID:21044336

  8. Ribosome Synthesis and MAPK Activity Modulate Ionizing Radiation-Induced Germ Cell Apoptosis in Caenorhabditis elegans

    PubMed Central

    Eberhard, Ralf; Stergiou, Lilli; Hofmann, E. Randal; Hofmann, Jen; Haenni, Simon; Teo, Youjin; Furger, André; Hengartner, Michael O.

    2013-01-01

    Synthesis of ribosomal RNA by RNA polymerase I (RNA pol I) is an elemental biological process and is key for cellular homeostasis. In a forward genetic screen in C. elegans designed to identify DNA damage-response factors, we isolated a point mutation of RNA pol I, rpoa-2(op259), that leads to altered rRNA synthesis and a concomitant resistance to ionizing radiation (IR)-induced germ cell apoptosis. This weak apoptotic IR response could be phenocopied when interfering with other factors of ribosome synthesis. Surprisingly, despite their resistance to DNA damage, rpoa-2(op259) mutants present a normal CEP-1/p53 response to IR and increased basal CEP-1 activity under normal growth conditions. In parallel, rpoa-2(op259) leads to reduced Ras/MAPK pathway activity, which is required for germ cell progression and physiological germ cell death. Ras/MAPK gain-of-function conditions could rescue the IR response defect in rpoa-2(op259), pointing to a function for Ras/MAPK in modulating DNA damage-induced apoptosis downstream of CEP-1. Our data demonstrate that a single point mutation in an RNA pol I subunit can interfere with multiple key signalling pathways. Ribosome synthesis and growth-factor signalling are perturbed in many cancer cells; such an interplay between basic cellular processes and signalling might be critical for how tumours evolve or respond to treatment. PMID:24278030

  9. Involvement of p38 MAPK in the Anticancer Activity of Cultivated Cordyceps militaris.

    PubMed

    Chou, Shang-Min; Lai, Wan-Jung; Hong, Tzuwen; Tsai, Sheng-Hong; Chen, Yen-Hsun; Kao, Cheng-Hsiang; Chu, Richard; Shen, Tang-Long; Li, Tsai-Kun

    2015-01-01

    Cordyceps militaris is a traditional Chinese medicine frequently used for tonic and therapeutic purposes. Reports from our laboratory and others have demonstrated that extracts of the cultivated fruiting bodies of C. militaris (CM) exhibit a potent cytotoxic effect against many cancer cell lines, especially human leukemia cells. Here, we further investigated the underlying mechanism through which CM is cytotoxic to cancer cells. The CM-mediated induction of PARP cleavage and its related DNA damage signal (γH2AX) was diminished by caspase inhibitor I. In contrast, a ROS scavenger failed to prevent CM-mediated leukemia cell death. Moreover, two signaling molecules, AKT and p38 MAPK, were activated during the course of apoptosis induction. Employing MTT analysis, we found that a p38 MAPK inhibitor but not an AKT inhibitor could rescue cells from CM-mediated cell death, as well as inhibit the cleavage of PARP, formation of apoptotic bodies and up-regulation of the γH2AX signal. These results suggest that CM-mediated leukemia cell death occurs through the activation of the p38 MAPK pathway, indicating its potential therapeutic effects against human leukemia. PMID:26205966

  10. Neferine induces autophagy of human ovarian cancer cells via p38 MAPK/ JNK activation.

    PubMed

    Xu, Limei; Zhang, Xiyu; Li, Yinuo; Lu, Shuhua; Lu, Shan; Li, Jieyin; Wang, Yuqiong; Tian, Xiaoxue; Wei, Jian-Jun; Shao, Changshun; Liu, Zhaojian

    2016-07-01

    Ovarian cancer is the most lethal gynecological malignancy. Patients usually have poor prognosis because of late diagnosis, relapse, and chemoresistance. It is pressing to seek novel agent for the treatment of ovarian cancer. Neferine is a bisbenzylisoquinoline alkaloid isolated from the embryos of lotus (Nelumbo nucifera). In this study, we investigated the antitumor effect of neferine on ovarian cancer cells. We found that neferine exhibited growth-inhibitory effect on human ovarian cancer cells, whereas showing less cytotoxic to non-malignant fallopian tube epithelial cells. Furthermore, we demonstrated that neferine induced autophagy and inactivated the mTOR pathway. Finally, we found that both p38 MAPK and JNK signaling pathways were activated by neferine treatment and contributed to the induction of autophagy in ovarian cancer cells. In conclusion, our findings showed that neferine induced autophagy of human ovarian cancer cells via p38 MAPK/JNK activation. Neferine may be explored as a promising antitumoral agent in ovarian cancer. PMID:26738868

  11. BDNF-TrkB signaling through Erk1/2MAPK phosphorylation mediates the enhancement of fear memory induced by glucocorticoids

    PubMed Central

    Revest, J-M; Le Roux, A; Roullot-Lacarrière, V; Kaouane, N; Vallée, M; Kasanetz, F; Rougé-Pont, F; Tronche, F; Desmedt, A; Piazza, P V

    2014-01-01

    Activation of glucocorticoid receptors (GR) by glucocorticoid hormones (GC) enhances contextual fear memories through the activation of the Erk1/2MAPK signaling pathway. However, the molecular mechanism mediating this effect of GC remains unknown. Here we used complementary molecular and behavioral approaches in mice and rats and in genetically modified mice in which the GR was conditionally deleted (GRNesCre). We identified the tPA-BDNF-TrkB signaling pathway as the upstream molecular effectors of GR-mediated phosphorylation of Erk1/2MAPK responsible for the enhancement of contextual fear memory. These findings complete our knowledge of the molecular cascade through which GC enhance contextual fear memory and highlight the role of tPA-BDNF-TrkB-Erk1/2MAPK signaling pathways as one of the core effectors of stress-related effects of GC. PMID:24126929

  12. COMP-angiopoietin 1 increases proliferation, differentiation, and migration of stem-like cells through Tie-2-mediated activation of p38 MAPK and PI3K/Akt signal transduction pathways

    SciTech Connect

    Kook, Sung-Ho; Lim, Shin-Saeng; Cho, Eui-Sic; Lee, Young-Hoon; Han, Seong-Kyu; Lee, Kyung-Yeol; Kwon, Jungkee; Hwang, Jae-Won; Bae, Cheol-Hyeon; Seo, Young-Kwon; Lee, Jeong-Chae

    2014-12-12

    Highlights: • COMP-Ang1 induces Tie-2 activation in BMMSCs, but not in primary osteoblasts. • Tie-2 knockdown inhibits COMP-Ang1-stimulated proliferation and osteoblastogenesis. • Tie-2 knockdown prevents COMP-Ang1-induced activation of PI3K/Akt and p38 MAPK. • COMP-Ang1 induces migration of cells via activation of PI3K/Akt and CXCR4 pathways. • COMP-Ang1 stimulates in vivo migration of PDLSCs into a calvarial defect site of rats. - Abstract: Recombinant COMP-Ang1, a chimera of angiopoietin-1 (Ang1) and a short coiled-coil domain of cartilage oligomeric matrix protein (COMP), is under consideration as a therapeutic agent capable of inducing the homing of cells with increased angiogenesis. However, the potentials of COMP-Ang1 to stimulate migration of mesenchymal stem cells (MSCs) and the associated mechanisms are not completely understood. We examined the potential of COMP-Ang1 on bone marrow (BM)-MSCs, human periodontal ligament stem cells (PDLSCs), and calvarial osteoblasts. COMP-Ang1 augmented Tie-2 induction at protein and mRNA levels and increased proliferation and expression of runt-related transcription factor 2 (Runx2), osterix, and CXCR4 in BMMSCs, but not in osteoblasts. The COMP-Ang1-mediated increases were inhibited by Tie-2 knockdown and by treating inhibitors of phosphoinositide 3-kinase (PI3K), LY294002, or p38 mitogen-activated protein kinase (MAPK), SB203580. Phosphorylation of p38 MAPK and Akt was prevented by siRNA-mediated silencing of Tie-2. COMP-Ang1 also induced in vitro migration of BMMSCs and PDLSCs. The induced migration was suppressed by Tie-2 knockdown and by CXCR4-specific peptide antagonist or LY294002, but not by SB203580. Furthermore, COMP-Ang1 stimulated the migration of PDLSCs into calvarial defect site of rats. Collectively, our results demonstrate that COMP-Ang1-stimulated proliferation, differentiation, and migration of progenitor cells may involve the Tie-2-mediated activation of p38 MAPK and PI3K/Akt pathways.

  13. [RELATIONSHIP BETWEEN p53/p21/Rb AND MAPK SIGNALING PATHWAYS IN HUMAN ENDOMETRIUM-DERIVED STEM CELLS UNDER OXIDATIVE STRESS].

    PubMed

    Deryabin, P I; Borodkina, A V; Nikolsky, N N; Burova, E B

    2015-01-01

    Human endometrium-derived mesenchymal stem cells (hMESC) under the sublethal oxidative stress induced by H2O2 activate both p53/p21/Rb and p38MAPK/MAPKAPK-2 pathways that are responsible for the induction of hMESC premature senescence (Borodkina et al., 2014). However the mutual relations between p53/p21/Rb and MAPK signaling pathways, including ERK, p38 and JNK remain unexplored as yet. Here, we used the specific inhibitors--pifithrin-α (PFT), U0126, SB203580 and SP600125 to "switch off" one of the proteins in these cascades and to evaluate the functional status alterations of the rest proteins. Suppression each of the MAPK significantly increased the p53 phosphorylation levels, as well as p21 protein expression followed by Rb hypophosphorylation. On the other hand, PFT-induced p53 inhibition enhanced mostly the ERK1/2 activation compared with p38 and JNK. These results suppose the existence of the reciprocal negative regulation between p53- and MAPK-dependent signaling pathways. Analyzing the possible interactions among the members of the MAPK family, we showed that p38 and JNK can function as the ERK antagonists: JNK is capable to activate ERK, while p38 may block the ERK activation. Together, these results demonstrate complex links between different signaling cascades in stressed hMESC, implicating ERK, p38 and JNK in regulation of the premature senescence via p53/p21/Rb pathway.

  14. The role of p38 MAPK activation in auranofin-induced apoptosis of human promyelocytic leukaemia HL-60 cells

    PubMed Central

    Park, Seon-Joo; Kim, In-Sook

    2005-01-01

    In a previous study, we reported an antileukaemic activity of auranofin (AF), demonstrating its dual effects: on the induction of apoptotic cell death and its synergistic action with retinoic acid on cell differentiation. In this study, we investigated the downstream signalling events of AF-induced apoptosis to determine the molecular mechanisms of AF activity. Treatment of HL-60 cells with AF induced apoptosis in a concentration- and time-dependent manner. Western blot analysis showed that AF-induced apoptosis was accompanied by the activation of caspase-8, caspase-9, and caspase-3, and the release of cytochrome c from the mitochondria. The phosphorylation and kinase activities of p38 mitogen-activated protein kinase (p38 MAPK) increased gradually until 12 h after AF (2 μM) treatment, and p38 MAPK was also activated concentration-dependently. Pretreatment with SB203580, a specific inhibitor of p38 MAPK, significantly blocked DNA fragmentation and the cleavage of procaspase-8, procaspase-3, and poly-ADP-ribose polymerase (PARP), whereas SB203580 alone had no effect. Reactive oxygen species (ROS) were also detected within 1 h after AF treatment, and the antioxidant N-acetyl-L-cysteine (NAC) effectively protected the cells from apoptosis by inhibiting the phosphorylation of p38 MAPK and the activation of caspases. These results suggest that ROS generation and the subsequent activation of p38 MAPK are essential for the proapoptotic effects of AF in human promyelocytic leukaemia HL-60 cells. PMID:16086031

  15. Transforming growth factor-beta1 induces activation of Ras, Raf-1, MEK and MAPK in rat hepatic stellate cells.

    PubMed

    Reimann, T; Hempel, U; Krautwald, S; Axmann, A; Scheibe, R; Seidel, D; Wenzel, K W

    1997-02-10

    The transdifferentiation of hepatic stellate cells into myofibroblast-like cells and the proliferation of the transdifferentiated cells are controlled by TGF-beta1. Little is known about the intracellular signal transducers of TGF-beta1. In this paper we show that in cultured hepatic stellate cells TGF-beta1 induces activation of Ras, Raf-1, MEK and MAPK p42 and p44. The activation of MAPK depends on the activation of MEK. Our data exclude that the observed effects are mediated by a bFGF or PDGF autocrine loop. PMID:9038360

  16. Homology modeling studies of yeast Mitogen-Activated Protein Kinases (MAPKS): structural motifs as a basis for specificity.

    PubMed

    Smith, D L; Nilar, S H

    2010-06-01

    Mitogen-activated protein kinases (MAPKs) are key components of cellular signal transduction. It is the objective of this communication to demonstrate that insight into protein-protein interactions in the Common Docking motif of yeast mitogen-activated protein kinases can be obtained based on homology models. Homology models for four yeast MAPKs, FUS3, KSS1, HOG1 and MPK1 were built based on the X-ray structures of active and inactive rat ERK2. The structural motifs required for the basis of specificity were rationalized based on these structures. PMID:19995338

  17. RAS/MAPK activation drives resistance to Smo inhibition, metastasis and tumor evolution in Shh pathway-dependent tumors

    PubMed Central

    Zhao, Xuesong; Ponomaryov, Tatyana; Ornell, Kimberly J.; Zhou, Pengcheng; Dabral, Sukriti K.; Pak, Ekaterina; Li, Wei; Atwood, Scott X.; Whitson, Ramon J.; Chang, Anne Lynn S.; Li, Jiang; Oro, Anthony E.; Chan, Jennifer A.; Kelleher, Joseph F.; Segal, Rosalind A.

    2015-01-01

    Aberrant Shh signaling promotes tumor growth in diverse cancers. The importance of Shh signaling is particularly evident in medulloblastoma and basal cell carcinoma (BCC), where inhibitors targeting the Shh pathway component Smoothened (Smo) show great therapeutic promise. However, the emergence of drug resistance limits long-term efficacy and the mechanisms of resistance remain poorly understood. Using new medulloblastoma models, we identify two distinct paradigms of resistance to Smo inhibition. Sufu mutations lead to maintenance of the Shh pathway in the presence of Smo inhibitors. Alternatively activation of the RAS/MAPK pathway circumvents Shh pathway-dependency, drives tumor growth and enhances metastatic behavior. Strikingly, in BCC patients treated with Smo inhibitor, squamous cell cancers with RAS/MAPK activation emerged from the antecedent BCC tumors. Together these findings reveal a critical role of RAS/MAPK pathway in drug resistance and tumor evolution of Shh pathway-dependent tumors. PMID:26130651

  18. Ozone (O{sub 3}) elicits neurotoxicity in spinal cord neurons (SCNs) by inducing ER Ca{sup 2+} release and activating the CaMKII/MAPK signaling pathway

    SciTech Connect

    Li, Yun; Lin, Xiaowen; Zhao, XueJun; Xie, Juntian; JunNan, Wang; Sun, Tao; Fu, Zhijian

    2014-11-01

    Ozone (O{sub 3}) is widely used in the treatment of spinal cord related diseases. Excess or accumulation of this photochemical air can however be neurotoxic. In this study, in vitro cultured Wister rat spinal cord neurons (SCNs) were used to investigate the detrimental effects and underlying mechanisms of O{sub 3}. Ozone in a dose-dependent manner inhibited cell viability at a range of 20 to 500 μg/ml, with the dose at 40 μg/ml resulting in a decrease of cell viability to 75%. The cell death after O{sub 3} exposure was related to endoplasmic reticulum (ER) calcium (Ca{sup 2+}) release. Intracellular Ca{sup 2+} chelator, ER stabilizer (inositol 1,4,5-trisphosphate receptor (IP3R) antagonist and ryanodine receptor (RyR) antagonist) and calcium/calmodulin-dependent protein kinase II (CaMKII) antagonist could effectively block Ca{sup 2+} mobilization and inhibit cell death following 40 μg/ml O{sub 3} exposure. In addition, ER Ca{sup 2+} release due to O{sub 3} exposure enhanced phospho-p38 and phospho-JNK levels and apoptosis of SCNs through activating CaMKII. Based on these results, we confirm that ozone elicits neurotoxicity in SCNs via inducing ER Ca{sup 2+} release and activating CaMKII/MAPK signaling pathway. Therefore, physicians should get attention to the selection of treatment concentrations of oxygen/ozone. And, approaches, such as chelating intracellular Ca{sup 2+} and stabilizing neuronal Ca{sup 2+} homeostasis could effectively ameliorate the neurotoxicity of O{sub 3}. - Highlights: • Exposure to O{sub 3} can reduce the viability of SCNs and cause the cell death. • Exposure to O{sub 3} can trigger RyR and IP3R dependent intracellular Ca{sup 2+} release. • Exposure to O{sub 3} can enhance the phospho-CaMKII, phospho-JNK and phospho-p38 levels.

  19. BMP9-Induced Survival Effect in Liver Tumor Cells Requires p38MAPK Activation

    PubMed Central

    García-Álvaro, María; Addante, Annalisa; Roncero, Cesáreo; Fernández, Margarita; Fabregat, Isabel; Sánchez, Aránzazu; Herrera, Blanca

    2015-01-01

    The study of bone morphogenetic proteins (BMPs) role in tumorigenic processes, and specifically in the liver, has gathered importance in the last few years. Previous studies have shown that BMP9 is overexpressed in about 40% of hepatocellular carcinoma (HCC) patients. In vitro data have also shown evidence that BMP9 has a pro-tumorigenic action, not only by inducing epithelial to mesenchymal transition (EMT) and migration, but also by promoting proliferation and survival in liver cancer cells. However, the precise mechanisms driving these effects have not yet been established. In the present work, we deepened our studies into the intracellular mechanisms implicated in the BMP9 proliferative and pro-survival effect on liver tumor cells. In HepG2 cells, BMP9 induces both Smad and non-Smad signaling cascades, specifically PI3K/AKT and p38MAPK. However, only the p38MAPK pathway contributes to the BMP9 growth-promoting effect on these cells. Using genetic and pharmacological approaches, we demonstrate that p38MAPK activation, although dispensable for the BMP9 proliferative activity, is required for the BMP9 protective effect on serum withdrawal-induced apoptosis. These findings contribute to a better understanding of the signaling pathways involved in the BMP9 pro-tumorigenic role in liver tumor cells. PMID:26343646

  20. Arabinosylated Lipoarabinomannan Skews Th2 Phenotype towards Th1 during Leishmania Infection by Chromatin Modification: Involvement of MAPK Signaling

    PubMed Central

    Bhattacharya, Parna; Gupta, Gaurav; Majumder, Saikat; Adhikari, Anupam; Banerjee, Sayantan; Halder, Kuntal; Bhattacharya Majumdar, Suchandra; Ghosh, Moumita; Chaudhuri, Shubho; Roy, Syamal; Majumdar, Subrata

    2011-01-01

    The parasitic protozoan Leishmania donovani is the causative organism for visceral leishmaniasis (VL) which persists in the host macrophages by deactivating its signaling machinery resulting in a critical shift from proinflammatory (Th1) to an anti-inflammatory (Th2) response. The severity of this disease is mainly determined by the production of IL-12 and IL-10 which could be reversed by use of effective immunoprophylactics. In this study we have evaluated the potential of Arabinosylated Lipoarabinomannan (Ara-LAM), a cell wall glycolipid isolated from non pathogenic Mycobacterium smegmatis, in regulating the host effector response via effective regulation of mitogen-activated protein kinases (MAPK) signaling cascades in Leishmania donovani infected macrophages isolated from BALB/C mice. Ara-LAM, a Toll-like receptor 2 (TLR2) specific ligand, was found to activate p38 MAPK signaling along with subsequent abrogation of extracellular signal–regulated kinase (ERKs) signaling. The use of pharmacological inhibitors of p38MAPK and ERK signaling showed the importance of these signaling pathways in the regulation of IL-10 and IL-12 in Ara-LAM pretreated parasitized macrophages. Molecular characterization of this regulation of IL-10 and IL-12 was revealed by chromatin immunoprecipitation assay (CHIP) which showed that in Ara-LAM pretreated parasitized murine macrophages there was a significant induction of IL-12 by selective phosphorylation and acetylation of histone H3 residues at its promoter region. While, IL-10 production was attenuated by Ara-LAM pretreatment via abrogation of histone H3 phosphorylation and acetylation at its promoter region. This Ara-LAM mediated antagonistic regulations in the induction of IL-10 and IL-12 genes were further correlated to changes in the transcriptional regulators Signal transducer and activator of transcription 3 (STAT3) and Suppressor of cytokine signaling 3 (SOCS3). These results demonstrate the crucial role played by Ara-LAM in

  1. Exocyst Sec10 protects renal tubule cells from injury by EGFR/MAPK activation and effects on endocytosis.

    PubMed

    Fogelgren, Ben; Zuo, Xiaofeng; Buonato, Janine M; Vasilyev, Aleksandr; Baek, Jeong-In; Choi, Soo Young; Chacon-Heszele, Maria F; Palmyre, Aurélien; Polgar, Noemi; Drummond, Iain; Park, Kwon Moo; Lazzara, Matthew J; Lipschutz, Joshua H

    2014-12-15

    Acute kidney injury is common and has a high mortality rate, and no effective treatment exists other than supportive care. Using cell culture models, we previously demonstrated that exocyst Sec10 overexpression reduced damage to renal tubule cells and speeded recovery and that the protective effect was mediated by higher basal levels of mitogen-activated protein kinase (MAPK) signaling. The exocyst, a highly-conserved eight-protein complex, is known for regulating protein trafficking. Here we show that the exocyst biochemically interacts with the epidermal growth factor receptor (EGFR), which is upstream of MAPK, and Sec10-overexpressing cells express greater levels of phosphorylated (active) ERK, the final step in the MAPK pathway, in response to EGF stimulation. EGFR endocytosis, which has been linked to activation of the MAPK pathway, increases in Sec10-overexpressing cells, and gefitinib, a specific EGFR inhibitor, and Dynasore, a dynamin inhibitor, both reduce EGFR endocytosis. In turn, inhibition of the MAPK pathway reduces ligand-mediated EGFR endocytosis, suggesting a potential feedback of elevated ERK activity on EGFR endocytosis. Gefitinib also decreases MAPK signaling in Sec10-overexpressing cells to levels seen in control cells and, demonstrating a causal role for EGFR, reverses the protective effect of Sec10 overexpression following cell injury in vitro. Finally, using an in vivo zebrafish model of acute kidney injury, morpholino-induced knockdown of sec10 increases renal tubule cell susceptibility to injury. Taken together, these results suggest that the exocyst, acting through EGFR, endocytosis, and the MAPK pathway is a candidate therapeutic target for acute kidney injury.

  2. Exocyst Sec10 protects renal tubule cells from injury by EGFR/MAPK activation and effects on endocytosis

    PubMed Central

    Fogelgren, Ben; Zuo, Xiaofeng; Buonato, Janine M.; Vasilyev, Aleksandr; Baek, Jeong-In; Choi, Soo Young; Chacon-Heszele, Maria F.; Palmyre, Aurélien; Polgar, Noemi; Drummond, Iain; Park, Kwon Moo; Lazzara, Matthew J.

    2014-01-01

    Acute kidney injury is common and has a high mortality rate, and no effective treatment exists other than supportive care. Using cell culture models, we previously demonstrated that exocyst Sec10 overexpression reduced damage to renal tubule cells and speeded recovery and that the protective effect was mediated by higher basal levels of mitogen-activated protein kinase (MAPK) signaling. The exocyst, a highly-conserved eight-protein complex, is known for regulating protein trafficking. Here we show that the exocyst biochemically interacts with the epidermal growth factor receptor (EGFR), which is upstream of MAPK, and Sec10-overexpressing cells express greater levels of phosphorylated (active) ERK, the final step in the MAPK pathway, in response to EGF stimulation. EGFR endocytosis, which has been linked to activation of the MAPK pathway, increases in Sec10-overexpressing cells, and gefitinib, a specific EGFR inhibitor, and Dynasore, a dynamin inhibitor, both reduce EGFR endocytosis. In turn, inhibition of the MAPK pathway reduces ligand-mediated EGFR endocytosis, suggesting a potential feedback of elevated ERK activity on EGFR endocytosis. Gefitinib also decreases MAPK signaling in Sec10-overexpressing cells to levels seen in control cells and, demonstrating a causal role for EGFR, reverses the protective effect of Sec10 overexpression following cell injury in vitro. Finally, using an in vivo zebrafish model of acute kidney injury, morpholino-induced knockdown of sec10 increases renal tubule cell susceptibility to injury. Taken together, these results suggest that the exocyst, acting through EGFR, endocytosis, and the MAPK pathway is a candidate therapeutic target for acute kidney injury. PMID:25298525

  3. Differential effects of chronic overload-induced muscle hypertrophy on mTOR and MAPK signaling pathways in adult and aged rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We examined activation of the mammalian target of rapamycin (mTOR) and mitogen-activated protein kinase (MAPK) signaling pathways in adult (Y; 6 mo old; n = 16) and aged (O; 30 mo old; n = 16) male rats (Fischer 344 x Brown Norway) subjected to chronic overload-induced muscle hypertrophy of the plan...

  4. Parainfluenza Virus Type 1 Induces Epithelial IL-8 Production via p38-MAPK Signalling

    PubMed Central

    Galván Morales, Miguel Ángel; Cabello Gutiérrez, Carlos; Mejía Nepomuceno, Fidencio; Valle Peralta, Leticia; Valencia Maqueda, Elba; Manjarrez Zavala, María Eugenia

    2014-01-01

    Human parainfluenza virus type 1 (HPIV-1) is the most common cause of croup in infants. The aim of this study was to describe molecular mechanisms associated with IL-8 production during HPIV-1 infection and the role of viral replication in MAPK synthesis and activation. An in vitro model of HPIV-1 infection in the HEp-2 and A549 cell lines was used; a kinetic-based ELISA for IL-8 detection was also used, phosphorylation of the mitogen-activated protein kinases (MAPKs) was identified by Western blot analysis, and specific inhibitors for each kinase were used to identify which MAPK was involved. Inactivated viruses were used to assess whether viral replication is required for IL-8 production. Results revealed a gradual increase in IL-8 production at different selected times, when phosphorylation of MAPK was detected. The secretion of IL-8 in the two cell lines infected with the HPIV-1 is related to the phosphorylation of the MAPK as well as viral replication. Inhibition of p38 suppressed the secretion of IL-8 in the HEp-2 cells. No kinase activation was observed when viruses were inactivated. PMID:25013817

  5. Regulation of Muscle Stem Cell Functions: A Focus on the p38 MAPK Signaling Pathway

    PubMed Central

    Segalés, Jessica; Perdiguero, Eusebio; Muñoz-Cánoves, Pura

    2016-01-01

    Formation of skeletal muscle fibers (myogenesis) during development and after tissue injury in the adult constitutes an excellent paradigm to investigate the mechanisms whereby environmental cues control gene expression programs in muscle stem cells (satellite cells) by acting on transcriptional and epigenetic effectors. Here we will review the molecular mechanisms implicated in the transition of satellite cells throughout the distinct myogenic stages (i.e., activation from quiescence, proliferation, differentiation, and self-renewal). We will also discuss recent findings on the causes underlying satellite cell functional decline with aging. In particular, our review will focus on the epigenetic changes underlying fate decisions and on how the p38 MAPK signaling pathway integrates the environmental signals at the chromatin to build up satellite cell adaptive responses during the process of muscle regeneration, and how these responses are altered in aging. A better comprehension of the signaling pathways connecting external and intrinsic factors will illuminate the path for improving muscle regeneration in the aged. PMID:27626031

  6. Regulation of Muscle Stem Cell Functions: A Focus on the p38 MAPK Signaling Pathway.

    PubMed

    Segalés, Jessica; Perdiguero, Eusebio; Muñoz-Cánoves, Pura

    2016-01-01

    Formation of skeletal muscle fibers (myogenesis) during development and after tissue injury in the adult constitutes an excellent paradigm to investigate the mechanisms whereby environmental cues control gene expression programs in muscle stem cells (satellite cells) by acting on transcriptional and epigenetic effectors. Here we will review the molecular mechanisms implicated in the transition of satellite cells throughout the distinct myogenic stages (i.e., activation from quiescence, proliferation, differentiation, and self-renewal). We will also discuss recent findings on the causes underlying satellite cell functional decline with aging. In particular, our review will focus on the epigenetic changes underlying fate decisions and on how the p38 MAPK signaling pathway integrates the environmental signals at the chromatin to build up satellite cell adaptive responses during the process of muscle regeneration, and how these responses are altered in aging. A better comprehension of the signaling pathways connecting external and intrinsic factors will illuminate the path for improving muscle regeneration in the aged. PMID:27626031

  7. Fus3-regulated Tec1 degradation through SCFCdc4 determines MAPK signaling specificity during mating in yeast.

    PubMed

    Chou, Song; Huang, Lan; Liu, Haoping

    2004-12-29

    Signaling specificity is fundamental for parallel mitogen-activated protein kinase (MAPK) cascades that control growth and differentiation in response to different stimuli. In Saccharomyces cerevisiae, components of the pheromone-responsive MAPK cascade activate Fus3 and Kss1 MAPKs to induce mating and Kss1 to promote filamentation. Active Fus3 is required to prevent the activation of the filamentation program during pheromone response. How Fus3 prevents the crossactivation is not clear. Here we show that Tec1, a cofactor of Ste12 for the expression of filamentation genes, is rapidly degraded during pheromone response. Fus3 but not Kss1 induces Tec1 ubiquination and degradation through the SCFCdc4 ubiquitin ligase. T273 in a predicted high-affinity Cdc4 binding motif is phosphorylated by Fus3 both in vitro and in vivo. Tec1T273V blocks Tec1 ubiquitination and degradation and allows the induction of filamentation genes in response to pheromone. Thus, Fus3 inhibits filamentous growth during mating by degrading Tec1. PMID:15620356

  8. On the origin of bistability in the Stage 2 of the Huang-Ferrell model of the MAPK signaling

    NASA Astrophysics Data System (ADS)

    Hadač, O.; Schreiber, I.; Přibyl, M.

    2013-02-01

    Mitogen-activated protein kinases (MAPKs) are important signal transducing enzymes, unique to eukaryotes, that are involved in many pathways of cellular regulation. Successive phosphorylation cascades mediated by MAPKs serve as sensitive switches initiating various cellular processes. Apart from this basic feature, the underlying reaction network is capable of displaying other nonlinear phenomena including bistable steady states and hysteresis as well as periodic oscillations. We show that from the mechanistic point of view, bistability is a consequence of interaction between single and double phosphorylation/dephosphorylation pathways in a Stage 2 subsystem of the Huang-Ferrell model. Within this subsystem we uncover the core subnetwork obtained by systematic reduction relying on the methods of stoichiometric network analysis. For the core model we show that there is either one stable steady state or three steady states of which two are stable and point out the role of interplay between the single and double phosphorylation subnetworks in generating bistability.

  9. Data-Derived Modeling Characterizes Plasticity of MAPK Signaling in Melanoma

    PubMed Central

    Bernardo-Faura, Marti; Massen, Stefan; Falk, Christine S.; Brady, Nathan R.; Eils, Roland

    2014-01-01

    The majority of melanomas have been shown to harbor somatic mutations in the RAS-RAF-MEK-MAPK and PI3K-AKT pathways, which play a major role in regulation of proliferation and survival. The prevalence of these mutations makes these kinase signal transduction pathways an attractive target for cancer therapy. However, tumors have generally shown adaptive resistance to treatment. This adaptation is achieved in melanoma through its ability to undergo neovascularization, migration and rearrangement of signaling pathways. To understand the dynamic, nonlinear behavior of signaling pathways in cancer, several computational modeling approaches have been suggested. Most of those models require that the pathway topology remains constant over the entire observation period. However, changes in topology might underlie adaptive behavior to drug treatment. To study signaling rearrangements, here we present a new approach based on Fuzzy Logic (FL) that predicts changes in network architecture over time. This adaptive modeling approach was used to investigate pathway dynamics in a newly acquired experimental dataset describing total and phosphorylated protein signaling over four days in A375 melanoma cell line exposed to different kinase inhibitors. First, a generalized strategy was established to implement a parameter-reduced FL model encoding non-linear activity of a signaling network in response to perturbation. Next, a literature-based topology was generated and parameters of the FL model were derived from the full experimental dataset. Subsequently, the temporal evolution of model performance was evaluated by leaving time-defined data points out of training. Emerging discrepancies between model predictions and experimental data at specific time points allowed the characterization of potential network rearrangement. We demonstrate that this adaptive FL modeling approach helps to enhance our mechanistic understanding of the molecular plasticity of melanoma. PMID:25188314

  10. Stimulatory Effects of Coumestrol on Embryonic and Fetal Development Through AKT and ERK1/2 MAPK Signal Transduction.

    PubMed

    Lim, Whasun; Song, Gwonhwa

    2016-12-01

    Successful establishment of pregnancy is required for fetal-maternal interactions regulating implantation, embryonic development and placentation. A uterine environment with insufficient growth factors and nutrients increases the incidence of intrauterine growth restriction (IUGR) leading to an impaired uterine environment. In the present study, we demonstrated the effects of the phytoestrogen coumestrol on conceptus development in the pig that is regarded as an excellent biomedical animal model for research on IUGR. Results of this study indicated that coumestrol induced migration of porcine trophectoderm (pTr) cells in a concentration-dependent manner. In response to coumestrol, the phosphorylation of AKT, P70S6K, S6, ERK1/2 MAPK, and P90RSK proteins were activated in pTr cells and ERK1/2 MAPK and P90RSK phosphorylation was prolonged for a longer period than for the other proteins. To identify the signal transduction pathway induced by coumestrol, pharmacological inhibitors U0126 (an ERK1/2 inhibitor) and LY294002 (a PI3K inhibitor) were used to pretreat pTr cells. The results showed that coumestrol-induced phosphorylation of ERK1/2 MAPK and P90RSK was blocked by U0126. In addition, the increased phosphorylation in response to coumestrol was completely inhibited following pre-treatment incubation of pTr cells in the presence of LY294002 and U0126. Furthermore, these two inhibitors suppressed the ability of coumestrol to induce migration of pTr cells. Collectively, these findings suggest that coumestrol affects embryonic development through activation of the PI3K/AKT and ERK1/2 MAPK cell signal transduction pathways and improvement in the uterine environment through coumestrol supplementation may provide beneficial effects of enhancing embryonic and fetal survival and development. J. Cell. Physiol. 231: 2733-2740, 2016. © 2016 Wiley Periodicals, Inc.

  11. Two guard cell-preferential MAPKs, MPK9 and MPK12, regulate YEL signalling in Arabidopsis guard cells.

    PubMed

    Salam, M A; Jammes, F; Hossain, M A; Ye, W; Nakamura, Y; Mori, I C; Kwak, J M; Murata, Y

    2013-05-01

    We report that two mitogen-activated protein kinases (MAPKs), MPK9 and MPK12, positively regulate abscisic acid (ABA)-induced stomatal closure in Arabidopsis thaliana. Yeast elicitor (YEL) induced stomatal closure accompanied by intracellular reactive oxygen species (ROS) accumulation and cytosolic free calcium concentration ([Ca(2+) ]cyt ) oscillation. In this study, we examined whether these two MAP kinases are involved in YEL-induced stomatal closure using MAPKK inhibitors, PD98059 and U0126, and MAPK mutants, mpk9, mpk12 and mpk9 mpk12. Both PD98059 and U0126 inhibited YEL-induced stomatal closure. YEL induced stomatal closure in the mpk9 and mpk12 mutants but not in the mpk9 mpk12 mutant, suggesting that a MAPK cascade involving MPK9 and MPK12 functions in guard cell YEL signalling. However, YEL induced extracellular ROS production, intracellular ROS accumulation and cytosolic alkalisation in the mpk9, mpk12 and mpk9 mpk12 mutants. YEL induced [Ca(2+) ]cyt oscillations in both wild type and mpk9 mpk12 mutant. These results suggest that MPK9 and MPK12 function redundantly downstream of extracellular ROS production, intracellular ROS accumulation, cytosolic alkalisation and [Ca(2+) ]cyt oscillation in YEL-induced stomatal closure in Arabidopsis guard cells and are shared with ABA signalling.

  12. Hypoxia differentially regulates the mitogen- and stress-activated protein kinases. Role of Ca2+/CaM in the activation of MAPK and p38 gamma.

    PubMed

    Conrad, P W; Millhorn, D E; Beitner-Johnson, D

    2000-01-01

    Hypoxic/ischemic trauma is a primary factor in the pathology of various vascular, pulmonary, and cerebral disease states. Yet, the signaling mechanisms by which cells respond and adapt to changes in oxygen levels are not clearly established. The effects of hypoxia on the stress- and mitogen-activated protein kinase (SAPK and MAPK) signaling pathways were studied in PC12 cells. Exposure to moderate hypoxia (5% O2) was found to progressively stimulate phosphorylation and activation of p38 gamma in particular, and also p38 alpha, two isoforms of the p38 family of stress-activated protein kinases. In contrast, hypoxia had no effect on enzyme activity of p38 beta, p38 beta 2, p38 delta, or on JNK, another stress-activated protein kinase. Prolonged hypoxia also induced phosphorylation and activation of p42/p44 MAPK, although this activation was modest when compared to NGF and UV-induced activation. We further showed that activation of p38 gamma and MAPK during hypoxia requires calcium, as treatment with Ca(2+)-free media or the calmodulin antagonist, W13, blocked the activation of p38 gamma and MAPK, respectively. These studies demonstrate that an extremely typical physiological stress (hypoxia) causes selective activation of specific elements of the SAPKs and MAPKs, and identifies Ca+2/CaM as a critical upstream activator. PMID:10849670

  13. Klotho Protects Dopaminergic Neuron Oxidant-Induced Degeneration by Modulating ASK1 and p38 MAPK Signaling Pathways

    PubMed Central

    Brobey, Reynolds K.; German, Dwight; Sonsalla, Patricia K.; Gurnani, Prem; Pastor, Johanne; Hsieh, C-C; Papaconstantinou, John; Foster, Philip P.; Kuro-o, Makoto; Rosenblatt, Kevin P.

    2015-01-01

    Klotho transgenic mice exhibit resistance to oxidative stress as measured by their urinal levels of 8-hydroxy-2-deoxyguanosine, albeit this anti-oxidant defense mechanism has not been locally investigated in the brain. Here, we tested the hypothesis that the reactive oxygen species (ROS)-sensitive apoptosis signal-regulating kinase 1 (ASK1)/p38 MAPK pathway regulates stress levels in the brain of these mice and showed that: 1) the ratio of free ASK1 to thioredoxin (Trx)-bound ASK1 is relatively lower in the transgenic brain whereas the reverse is true for the Klotho knockout mice; 2) the reduced p38 activation level in the transgene corresponds to higher level of ASK1-bound Trx, while the KO mice showed elevated p38 activation and lower level of–bound Trx; and 3) that 14-3-3ζ is hyper phosphorylated (Ser-58) in the transgene which correlated with increased monomer forms. In addition, we evaluated the in vivo robustness of the protection by challenging the brains of Klotho transgenic mice with a neurotoxin, MPTP and analyzed for residual neuron numbers and integrity in the substantia nigra pars compacta. Our results show that Klotho overexpression significantly protects dopaminergic neurons against oxidative damage, partly by modulating p38 MAPK activation level. Our data highlight the importance of ASK1/p38 MAPK pathway in the brain and identify Klotho as a possible anti-oxidant effector. PMID:26452228

  14. Osteoprotegerin induces podosome disassembly in osteoclasts through calcium, ERK, and p38 MAPK signaling pathways.

    PubMed

    Zhao, Hongyan; Liu, Xuezhong; Zou, Hui; Dai, Nannan; Yao, Lulian; Gao, Qian; Liu, Wei; Gu, Jianhong; Yuan, Yan; Bian, Jianchun; Liu, Zongping

    2015-02-01

    Osteoclasts are critical for bone resorption and use podosomes to attach to bone matrix. Osteoprotegerin (OPG) is a negative regulator of osteoclast function that can affect the formation and function of podosomes. However, the signaling pathways that link OPG to podosome function have not been well characterized. Therefore, this study examined the roles of intracellular calcium and MAPKs in OPG-induced podosome disassembly in osteoclasts. We assessed the effects of the intracellular calcium chelator Bapta-AM, ERK inhibitor U0126, and p38 inhibitor SB202190 on OPG-treated osteoclast differentiation, adhesion structures, intracellular free Ca(2+) concentration and the phosphorylation state of podosome associated proteins (Pyk2 and Src). Mouse monocytic RAW 264.7 cells were differentiated to osteoclasts using RANKL (30ng/mL) and M-CSF (25ng/mL). The cells were pretreated with Bapta-AM (5μM), U0126 (5μM), or SB202190 (10μM) for 30min, followed by 40ng/mL OPG for 3h. Osteoclastogenesis, adhesion structure, viability and morphology, intracellular free Ca(2+) concentration and the phosphorylation state of Pyk2 and Src were measured by TRAP staining, scanning electron microscopy, real-time cell analyzer, flow cytometry and western blotting, respectively. OPG significantly inhibited osteoclastogenesis, the formation of adhesion structures, and reduced the amount of phosphorylated Pyk2 and Src-pY527, but increased phosphorylation of Src-pY416. Bapta-AM, U0126, and SB202190 partially restored osteoclast differentiation and adhesion structures. Both Bapta-AM and U0126, but not SB202190, restored the levels of intracellular free Ca(2+) concentration, phosphorylated Pyk2 and Src-pY527. All three inhibitors blocked OPG-induced phosphorylation at Src-pY416. These results suggest OPG disrupts the attachment structures of osteoclasts and activates Src as an adaptor protein that competes for the reduced amount of phosphorylated Pyk2 through calcium- and ERK-dependent signaling

  15. cAMP blocks MAPK activation and sclerotial development via Rap-1 in a PKA-independent manner in Sclerotinia sclerotiorum.

    PubMed

    Chen, Changbin; Dickman, Martin B

    2005-01-01

    Sclerotinia sclerotiorum is a filamentous ascomycete phytopathogen able to infect an extremely wide range of cultivated plants. Our previous studies have shown that increases in cAMP levels result in the impairment of the development of the sclerotium, a highly differentiated structure important in the disease cycle of this fungus. cAMP also inhibits the activation of a S. sclerotiorum mitogen-activated protein kinase (MAPK), which we have previously shown to be required for sclerotial maturation; thus cAMP-mediated sclerotial inhibition is modulated through MAPK. However, the mechanism(s) by which cAMP inhibits MAPK remains unclear. Here we demonstrate that a protein kinase A (PKA)-independent signalling pathway probably mediates MAPK inhibition by cAMP. Expression of a dominant negative form of Ras, an upstream activator of the MAPK pathway, also inhibited sclerotial development and MAPK activation, suggesting that a conserved Ras/MAPK pathway is required for sclerotial development. Evidence from bacterial toxins that specifically inhibit the activity of small GTPases, suggested that Rap-1 or Ras is involved in cAMP action. The Rap-1 inhibitor, GGTI-298, restored MAPK activation in the presence of cAMP, further suggesting that Rap-1 is responsible for cAMP-dependent MAPK inhibition. Importantly, inhibition of Rap-1 is able to restore sclerotial development blocked by cAMP. Our results suggest a novel mechanism involving the requirement of Ras/MAPK pathway for sclerotial development that is negatively regulated by a PKA-independent cAMP signalling pathway. Cross-talk between these two pathways is mediated by Rap-1. PMID:15612936

  16. The PDZ Protein Canoe/AF-6 Links Ras-MAPK, Notch and Wingless/Wnt Signaling Pathways by Directly Interacting with Ras, Notch and Dishevelled

    PubMed Central

    Carmena, Ana; Speicher, Stephan; Baylies, Mary

    2006-01-01

    Over the past few years, it has become increasingly apparent that signal transduction pathways are not merely linear cascades; they are organized into complex signaling networks that require high levels of regulation to generate precise and unique cell responses. However, the underlying regulatory mechanisms by which signaling pathways cross-communicate remain poorly understood. Here we show that the Ras-binding protein Canoe (Cno)/AF-6, a PDZ protein normally associated with cellular junctions, is a key modulator of Wingless (Wg)/Wnt, Ras-Mitogen Activated Protein Kinase (MAPK) and Notch (N) signaling pathways cross-communication. Our data show a repressive effect of Cno/AF-6 on these three signaling pathways through physical interactions with Ras, N and the cytoplasmic protein Dishevelled (Dsh), a key Wg effector. We propose a model in which Cno, through those interactions, actively coordinates, at the membrane level, Ras-MAPK, N and Wg signaling pathways during progenitor specification. PMID:17183697

  17. Differential Activation of Mitogen-Activated Protein Kinases, ERK 1/2, p38(MAPK) and JNK p54/p46 During Postnatal Development of Rat Hippocampus.

    PubMed

    Costa, Ana Paula; Lopes, Mark William; Rieger, Débora K; Barbosa, Sabrina Giovana Rocha; Gonçalves, Filipe Marques; Xikota, João Carlos; Walz, Roger; Leal, Rodrigo B

    2016-05-01

    Mitogen-activated protein kinases (MAPKs) are a group of serine-threonine kinases, including p38(MAPK), ERK 1/2 and JNK p54/p46, activated by phosphorylation in response to extracellular stimuli. The early postnatal period is characterized by significant changes in brain structure as well as intracellular signaling. In the hippocampus MAPKs have been involved in the modulation of development and neural plasticity. However, the temporal profile of MAPK activation throughout the early postnatal development is incomplete. An understanding of this profile is important since slight changes in the activity of these enzymes, in response to environmental stress in specific developmental windows, might alter the course of development. The present study was undertaken to investigate the hippocampal differential activation of MAPK during postnatal period. MAPK activation and total content were evaluated by Western blotting of hippocampal tissue obtained from male Wistar rats at postnatal days (P) 1, 4, 7, 10, 14, 21, 30 and 60. The total content and phosphorylation of each MAPK was expressed as mean ± SEM and then calculates as a percentile compared to P1 (set at 100 %). The results showed: (1) phosphorylation peaks of p38(MAPK) at PN4 (p = 0.036) and PN10 to PN60; (2) phosphorylation of ERK1 and ERK2 were increased with age (ERK1 p = 0.0000005 and ERK2 p = 0.003); (3) phosphorylation profile of JNK p54/p46 was not changed during the period analyzed (JNKp56 p = 0.716 and JNKp46 p = 0.192). Therefore, the activity profile of ERK 1/2 and p38(MAPK) during postnatal development of rat hippocampus are differentially regulated. Our results demonstrate that ERK 1/2 and p38(MAPK) are dynamically regulated during postnatal neurodevelopment, suggesting temporal correlation of MAPK activity with critical periods when programmed cell death and synaptogenesis are occurring. This suggests an important role for these MAPKs in postnatal development of rat hippocampus.

  18. TLR4-Activated MAPK-IL-6 Axis Regulates Vascular Smooth Muscle Cell Function

    PubMed Central

    Lee, Guan-Lin; Wu, Jing-Yiing; Tsai, Chien-Sung; Lin, Chih-Yuan; Tsai, Yi-Ting; Lin, Chin-Sheng; Wang, Yi-Fu; Yet, Shaw-Fang; Hsu, Yu-Juei; Kuo, Cheng-Chin

    2016-01-01

    Migration of vascular smooth muscle cells (VSMCs) into the intima is considered to be a vital event in the pathophysiology of atherosclerosis. Despite substantial evidence supporting the pathogenic role of Toll-like receptor 4 (TLR4) in the progression of atherogenesis, its function in the regulation of VSMC migration remains unclear. The goal of the present study was to elucidate the mechanism by which TLR4 regulates VSMC migration. Inhibitor experiments revealed that TLR4-induced IL-6 secretion and VSMC migration were mediated via the concerted actions of MyD88 and TRIF on the activation of p38 MAPK and ERK1/2 signaling. Neutralizing anti-IL-6 antibodies abrogated TLR4-driven VSMC migration and F-actin polymerization. Blockade of p38 MAPK or ERK1/2 signaling cascade inhibited TLR4 agonist-mediated activation of cAMP response element binding protein (CREB). Moreover, siRNA-mediated suppression of CREB production repressed TLR4-induced IL-6 production and VSMC migration. Rac-1 inhibitor suppressed TLR4-driven VSMC migration but not IL-6 production. Importantly, the serum level of IL-6 and TLR4 endogenous ligand HMGB1 was significantly higher in patients with coronary artery diseases (CAD) than in healthy subjects. Serum HMGB1 level was positively correlated with serum IL-6 level in CAD patients. The expression of both HMGB1 and IL-6 was clearly detected in the atherosclerotic tissue of the CAD patients. Additionally, there was a positive association between p-CREB and HMGB1 in mouse atherosclerotic tissue. Based on our findings, we concluded that, upon ligand binding, TLR4 activates p38 MAPK and ERK1/2 signaling through MyD88 and TRIF in VSMCs. These signaling pathways subsequently coordinate an additive augmentation of CREB-driven IL-6 production, which in turn triggers Rac-1-mediated actin cytoskeleton to promote VSMC migration. PMID:27563891

  19. TLR4-Activated MAPK-IL-6 Axis Regulates Vascular Smooth Muscle Cell Function.

    PubMed

    Lee, Guan-Lin; Wu, Jing-Yiing; Tsai, Chien-Sung; Lin, Chih-Yuan; Tsai, Yi-Ting; Lin, Chin-Sheng; Wang, Yi-Fu; Yet, Shaw-Fang; Hsu, Yu-Juei; Kuo, Cheng-Chin

    2016-01-01

    Migration of vascular smooth muscle cells (VSMCs) into the intima is considered to be a vital event in the pathophysiology of atherosclerosis. Despite substantial evidence supporting the pathogenic role of Toll-like receptor 4 (TLR4) in the progression of atherogenesis, its function in the regulation of VSMC migration remains unclear. The goal of the present study was to elucidate the mechanism by which TLR4 regulates VSMC migration. Inhibitor experiments revealed that TLR4-induced IL-6 secretion and VSMC migration were mediated via the concerted actions of MyD88 and TRIF on the activation of p38 MAPK and ERK1/2 signaling. Neutralizing anti-IL-6 antibodies abrogated TLR4-driven VSMC migration and F-actin polymerization. Blockade of p38 MAPK or ERK1/2 signaling cascade inhibited TLR4 agonist-mediated activation of cAMP response element binding protein (CREB). Moreover, siRNA-mediated suppression of CREB production repressed TLR4-induced IL-6 production and VSMC migration. Rac-1 inhibitor suppressed TLR4-driven VSMC migration but not IL-6 production. Importantly, the serum level of IL-6 and TLR4 endogenous ligand HMGB1 was significantly higher in patients with coronary artery diseases (CAD) than in healthy subjects. Serum HMGB1 level was positively correlated with serum IL-6 level in CAD patients. The expression of both HMGB1 and IL-6 was clearly detected in the atherosclerotic tissue of the CAD patients. Additionally, there was a positive association between p-CREB and HMGB1 in mouse atherosclerotic tissue. Based on our findings, we concluded that, upon ligand binding, TLR4 activates p38 MAPK and ERK1/2 signaling through MyD88 and TRIF in VSMCs. These signaling pathways subsequently coordinate an additive augmentation of CREB-driven IL-6 production, which in turn triggers Rac-1-mediated actin cytoskeleton to promote VSMC migration. PMID:27563891

  20. Fasudil inhibits LPS-induced migration of retinal microglial cells via regulating p38-MAPK signaling pathway

    PubMed Central

    Xu, Fan; Xu, Yue; Zhu, Liqiong; Rao, Pinhong; Wen, Jiamin; Sang, Yunyun; Shang, Fu

    2016-01-01

    Purpose To investigate the effect and possible molecular mechanisms of fasudil on retinal microglial (RMG) cell migration. Methods Primary cultured RMG cells were incubated with lipopolysaccharide (LPS), fasudil, and/or SB203580 (a p38 inhibitor). RMG cell motility was determined with the scratch wound assay and the Transwell migration assay. The phosphorylation of p38 and levels of matrix metalloproteinase 2 (MMP-2) and MMP-9 were measured with western blot. Results In the scratch-induced migration assay, as well as in the Transwell migration assay, the results indicated that LPS stimulated the migratory potential of RMG cells and fasudil significantly reduced LPS-stimulated RMG cell migration in a concentration-dependent manner. However, fasudil had no effect on RMG cell migration in the absence of LPS stimulation. Moreover, fasudil reduced the level of phosphor-p38 mitogen-activated protein kinase (p-p38-MAPK) in a concentration-dependent manner, without effects on the levels of phospho-p44/42 (p-ERK1/2) and phospho-c-Jun N-terminal kinase (p-JNK). Cotreatment with SB203580 (a p38 inhibitor) and fasudil resulted in the synergistic reduction of MMP-2, MMP-9, and p-p38-MAPK, as well as a reduction in the LPS-stimulated migration capabilities of the RMG cells, suggesting fasudil suppresses the LPS-stimulated migration of RMG cells via directly downregulating the p38-MAPK signaling pathway. Conclusions Our studies indicated that fasudil inhibited LPS-stimulated RMG cell migration via suppression of the p38-MAPK signaling pathway. PMID:27441000

  1. Mitogen-activated protein kinase (MAPK) regulates the expression of progelatinase B (MMP-9) in breast epithelial cells.

    PubMed

    Reddy, K B; Krueger, J S; Kondapaka, S B; Diglio, C A

    1999-07-19

    Mitogen-activated protein kinases (MAPKs) play a major role in the mitogenic signal transduction pathway and are essential components of both growth and differentiation. Constitutive activation of the MAPK cascade is associated with the carcinogenesis and metastasis of human breast and renal cell carcinomas. The gelatinases B (MMP-9) and A (MMP-2) are 2 members of the matrix metalloproteinase (MMPs) family which are expressed in human cancers and thought to play a critical role in tumor cell invasion and metastasis. In a previous study, we have shown that EGF and amphiregulin upregulate MMP-9 in metastatic SKBR-3 cells but have no effect on MMP-2 secretion. We now investigated specific step(s) in EGF-induced signalling associated with regulation of cell proliferation and MMP-9 induction. EGF-induced signalling in SKBR-3 cells was blocked by relatively specific inhibitors either on ras (FPT inhibitor-1) or P13 kinase (Wortmannin) or by reduction in EGF-induced tyrosine kinase activity (RG 13022). Blocking these signalling pathways significantly inhibited of EGF-induced cell proliferation but only partially reduced in EGF-induced MMP-9 secretion. In contrast, when SKBR-3 cells were exposed to MEK inhibitor (PD 98059) or MAPK inhibitors (Apigenin or MAPK antisense phosphorothioate oligodeoxynucleotides), EGF-induced cell proliferation, MMP-9 induction and invasion through reconstituted basement membrane were significantly reduced. Our results suggest that interfering with MAPK activity may provide a novel means of controlling growth and invasiveness of tumors in which the signalling cascade is activated.

  2. Ghrelin Protects against Dexamethasone-Induced INS-1 Cell Apoptosis via ERK and p38MAPK Signaling

    PubMed Central

    2016-01-01

    Glucocorticoid excess induces apoptosis of islet cells, which may result in diabetes. In this study, we investigated the protective effect of ghrelin on dexamethasone-induced INS-1 cell apoptosis. Our data showed that ghrelin (0.1 μM) inhibited dexamethasone-induced (0.1 μM) apoptosis of INS-1 cells and facilitated cell proliferation. Moreover, ghrelin upregulated Bcl-2 expression, downregulated Bax expression, and decreased caspase-3 activity. The protective effect of ghrelin against dexamethasone-induced INS-1 cell apoptosis was mediated via growth hormone secretagogue receptor 1a. Further studies revealed that ghrelin increased ERK activation and decreased p38MAPK expression after dexamethasone treatment. Ghrelin-mediated protection of dexamethasone-induced apoptosis of INS-1 cells was attenuated using the ERK inhibitor U0126 (10 μM), and cell viability increased using the p38MAPK inhibitor SB203580 (10 μM). In conclusion, ghrelin could protect against dexamethasone-induced INS-1 cell apoptosis, at least partially via GHS-R1a and the signaling pathway of ERK and p38MAPK. PMID:27190513

  3. Michael addition of dehydroalanine-containing MAPK peptides to catalytic lysine inhibits the activity of phosphothreonine lyase.

    PubMed

    Zhang, Yuan; Yang, Ru; Huang, Juan; Liang, Qiujin; Guo, Yanmin; Bian, Weixiang; Luo, Lingfei; Li, Hongtao

    2015-11-30

    The phosphothreonine lyases OspF and SpvC irreversibly inactivate host dual-phosphorylated mitogen-activated protein kinases (MAPKs) [pThr-X-pTyr motif] through β-elimination. We found that dual-phosphorylated (pSer-X-pTyr) MAPK substrate peptides and their resulting catalytic products cross-link to OspF and SpvC. Mass spectrometry results revealed that these linkages form between lysine, which acts as a general base, and dehydroalanine (Dha) on catalytic products. The nucleophilic addition efficiency is dependent on the K136 residue being in a deprotonated state. Peptide cross-linking inhibits the activity of SpvC and blocks the inactivation of MAPK signaling by SpvC. Small compounds mimicking these sequences may act as phosphothreonine lyase inhibitors. PMID:26519561

  4. Cadmium induces vascular permeability via activation of the p38 MAPK pathway

    SciTech Connect

    Dong, Fengyun; Guo, Fang; Li, Liqun; Guo, Ling; Hou, Yinglong; Hao, Enkui; Yan, Suhua; Allen, Thaddeus D.; Liu, Ju

    2014-07-18

    Highlights: • Low-dose cadmium (Cd) induces vascular hyper-permeability. • p38 MAPK mediates Cd-induced disruption of endothelial cell barrier function. • SB203850 inhibits Cd-induced membrane dissociation of VE-cadherin and β-catenin. • SB203850 reduces Cd-induced expression and secretion of TNF-α. - Abstract: The vasculature of various organs is a targeted by the environmental toxin, cadmium (Cd). However, mechanisms leading to pathological conditions are poorly understood. In the present study, we examined the effect of cadmium chloride (CdCl{sub 2}) on human umbilical vein endothelial cells (HUVECs). At 4 μM, CdCl{sub 2} induced a hyper-permeability defect in HUVECs, but not the inhibition of cell growth up to 24 h. This effect of CdCl{sub 2} was dependent on the activation of the p38 mitogen-activated protein kinase (MAPK) pathway. The p38 MAPK inhibitor SB203850 suppressed the CdCl{sub 2}-induced alteration in trans-endothelial electrical resistance in HUVEC monolayers, a model measurement of vascular endothelial barrier integrity. SB203850 also inhibited the Cd-induced membrane dissociation of vascular endothelial (VE) cadherin and β-catenin, the important components of the adherens junctional complex. In addition, SB203850 reduces the Cd-induced expression and secretion of tumor necrosis factor α (TNF-α). Taken together, our findings suggest that Cd induces vascular hyper-permeability and disruption of endothelial barrier integrity through stimulation of p38 MAPK signaling.

  5. Inflammatory MAPK and NF-κB signaling pathways differentiated hepatitis potential of two agglomerated titanium dioxide particles.

    PubMed

    Chen, Jin; Zhang, Jianying; Cao, Junmei; Xia, Zongping; Gan, Jay

    2016-03-01

    TiO2 nanoparticles (TiO2NPs) consumption and deposit in liver have possible implications for hepatitis risks. Specific signal dysregulation at early inflammatory responses needs to be characterized in TiO2NP-induced hepatopathy. MAPK and NF-κB signaling pathways are known to participate in inflammation and respond sensitively to chemical agents, making them preferable biomarkers for hepatitis. In the present study, dynamic activation of MAPK and NF-κB pathways were explored by immunoblotting and NF-κB luciferase reporter assay depending on characterization of TiO2NP agglomeration in human HepG2 cells. Inflammatory and cytotoxic potential of TiO2NPs were determined by qRT-PCR and WST-1 assay. AFM and TEM analyses uncovered ultrastructure damages underlying hepatotoxicity of TiO2NPs. Rod-like rutile agglomerated smaller and induced more pronounced cytotoxicity and immunogenicity than anatase. Correspondingly, though both rutile and anatase significantly activated p38, ERK1/2 and NF-κB pathways, rutile accelerated the maximum phosphorylation of ERK1/2 and elevated the potency of IκBα phosphorylation to its bell curve shape in comparison with a lower and sigmoid type of IκBα phosphorylation for anatase. Furthermore, cell elasticity indicated by Young's modulus and adhesion force increased accompanied with mitochondria damage, contributing to signal dysregulation and hepatotoxicity. The results suggested that differential activation of MAPK and NF-κB pathways could be early predictors for distinct hepatitis risks of two agglomerated TiO2NPs.

  6. Both mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 and phosphatidylinositide-3-OH kinase (PI3K)/Akt pathways regulate activation of E-twenty-six (ETS)-like transcription factor 1 (Elk-1) in U138 glioblastoma cells.

    PubMed

    Mut, Melike; Lule, Sevda; Demir, Ozlem; Kurnaz, Isil Aksan; Vural, Imran

    2012-02-01

    Epidermal growth factor (EGF) and its receptor (EGFR) have been shown to play a significant role in the pathogenesis of glioblastoma. In our study, the EGFR was stimulated with EGF in human U138 glioblastoma cells. We show that the activated mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 pathway phosphorylated the E twenty-six (ETS)-like transcription factor 1 (Elk-1) mainly at serine 383 residue. Mitogen-activated protein kinase kinase (MEK) 1/2 inhibitor, UO126 and ERK inhibitor II, FR180204 blocked the Elk-1 phosphorylation and activation. The phosphatidylinositide-3-OH kinase (PI3K)/Akt pathway was also involved in the Elk-1 activation. Activation of the Elk-1 led to an increased survival and a proliferative response with the EGF stimulation in the U138 glioblastoma cells. Knocking-down the Elk-1 using an RNA interference technique caused a decrease in survival of the unstimulated U138 glioblastoma cells and also decreased the proliferative response to the EGF stimulation. The Elk-1 transcription factor was important for the survival and proliferation of U138 glioblastoma cells upon the stimulation of EGFR with EGF. The MAPK/ERK1/2 and PI3K/Akt pathways regulated this response via activation of the Elk-1 transcription factor. The Elk-1 may be one of the convergence points for pathways located downstream of EGFR in glioblastoma cells. Utilization of the Elk-1 as a therapeutic target may lead to a novel strategy in treatment of glioblastoma.

  7. Inhibition of MAPK and NF-κB signaling pathways alleviate carbon tetrachloride (CCl4)-induced liver fibrosis in Toll-like receptor 5 (TLR5) deficiency mice.

    PubMed

    Shu, Ming; Huang, Dan-dan; Hung, Zuo-an; Hu, Xiao-rong; Zhang, Shun

    2016-02-26

    Current researches showed that TLR family plays an important role in liver fibrosis, yet the molecular mechanism by which this occurs is not fully explained. In this study, we investigated the role of TLR5 in carbon tetrachloride-induced liver fibrosis, and further examined wether TLR5 knockout attenuated tetrachloride-induced liver fibrosis by inhibiting hepatic stellate cells activation via modulating NF-κB and MAPK signaling pathways. Our results found that carbon tetrachloride induced liver function injury in WT mice with a inflammatory responses through the activation of NF-κB and MAPK signaling pathways, resulting in hepatic stellate cells activation. In contrast, TLR5 deficiency mice after carbon tetrachloride administration reduced NF-κB and MAPK signaling pathways activation, which down regulated hepatic stellate cells activation. In addition, alpha smooth muscle-actin as marker of hepatic stellate cells further indicated that TLR5 knockout mice have a lower collagen accumulation in liver tissue than WT mice after carbon tetrachloride administration, resulting in inhibition of NF-κB and MAPK signaling pathways activation. Moreover, in vitro experiment of hepatic stellate cells challenged with LPS or TGF-β, further indicated that NF-κB and MAPK were involved in liver fibrosis development, leading to α-SMA expression and inflammation infiltration. However, cells from TLR5(-)(/-) may weaken phosphorylation levels of signal pathways, finally suppress progress of collagen accumulation and inflammatory responses. These results suggest a new therapeutic approach or target to protect against fibrosis caused by chronic liver diseases.

  8. Effect of electroacupuncture stimulation at Zusanli acupoint (ST36) on gastric motility: possible through PKC and MAPK signal transduction pathways

    PubMed Central

    2014-01-01

    Background Electroacupuncture (EA) stimulation has been shown to have a great therapeutic potential for treating gastrointestinal motility disorders. However, no evidence has clarified the mechanisms contributing to the effects of EA stimulation at the Zusanli acupoint (ST.36). This study was designed to investigate the regulative effect of EA stimulation at the ST.36 on gastric motility and to explore its possible mechanisms. Methods Thirty Sprague-Dawley rats were randomly divided into three groups: the ST.36 group, the non-acupoint group, and the control group. EA stimulation was set at 2 Hz, continuous mode, and 1 V for 30 min. The frequency and average peak amplitude of gastric motility were measured by electrogastrography. The protein kinase C (PKC) and mitogen-activated protein kinase (MAPK) signaling pathways were assessed using real-time polymerase chain reactions. Caldesmon (CaD) and calponin (CaP) protein expression in the gastric antrum were detected on Western blots. A Computed Video Processing System was used to evaluate morphological changes in smooth muscle cells (SMCs) from the gastric antrum. Results EA stimulation at ST.36 had a dual effect on the frequency and average peak amplitude. Additionally, EA stimulation at ST.36 regulated the expression of some genes in the PKC and MAPK signaling pathways, and it regulated the expression of the CaD and CaP proteins. EA serum induced SMC contractility. Promotion of gastric motility may correlate with up-regulation of MAPK6 (ERK3), MAPK13, and Prostaglandin-endoperoxide synthase 2 (PTGS2) gene expression, and the down-regulation of the collagen, type I, alpha 1 (COL1A1) gene and CaD and CaP protein expression. Inhibition of gastric motility may correlate with down-regulation of the Interleukin-1 receptor type 2 (IL1R2) and Matrix metalloproteinase-9 (MMP9) genes, and up-regulation of CaD and CaP protein expression. Conclusions EA stimulation at ST.36 regulated gastric motility, and the effects were

  9. MAPK signaling determines anxiety in the juvenile mouse brain but depression-like behavior in adults.

    PubMed

    Wefers, Benedikt; Hitz, Christiane; Hölter, Sabine M; Trümbach, Dietrich; Hansen, Jens; Weber, Peter; Pütz, Benno; Deussing, Jan M; de Angelis, Martin Hrabé; Roenneberg, Till; Zheng, Fang; Alzheimer, Christian; Silva, Alcino; Wurst, Wolfgang; Kühn, Ralf

    2012-01-01

    MAP kinase signaling has been implicated in brain development, long-term memory, and the response to antidepressants. Inducible Braf knockout mice, which exhibit protein depletion in principle forebrain neurons, enabled us to unravel a new role of neuronal MAPK signaling for emotional behavior. Braf mice that were induced during adulthood showed normal anxiety but increased depression-like behavior, in accordance with pharmacological findings. In contrast, the inducible or constitutive inactivation of Braf in the juvenile brain leads to normal depression-like behavior but decreased anxiety in adults. In juvenile, constitutive mutants we found no alteration of GABAergic neurotransmission but reduced neuronal arborization in the dentate gyrus. Analysis of gene expression in the hippocampus revealed nine downregulated MAPK target genes that represent candidates to cause the mutant phenotype.Our results reveal the differential function of MAPK signaling in juvenile and adult life phases and emphasize the early postnatal period as critical for the determination of anxiety in adults. Moreover, these results validate inducible gene inactivation as a new valuable approach, allowing it to discriminate between gene function in the adult and the developing postnatal brain. PMID:22529971

  10. The role of MAPK signaling pathway in the Her-2-positive meningiomas

    PubMed Central

    Wang, Zhaoyin; Wang, Weijia; Xu, Shan; Wang, Shanshan; Tu, Yi; Xiong, Yifeng; Mei, Jinhong; Wang, Chunliang

    2016-01-01

    Meningiomas are common types of adult nerve system tumors. Although most cases are considered benign, due to its high rate of recurrence and easy malignant progression to anaplastic meningioma they present a puzzle for the current treatment. The HER-2 oncogene has important value for meningioma cells development and progression. So far, little is known about the effect on the exact underlying signal pathway and molecular mechanisms of HER-2-positive meningioma cells. The goal of the present study was to determine the effects of HER-2 gene and possible involvement of MAPK signal pathway in human malignant meningioma. We applied q-PCR analysis, immunofluorescence (IF) staining, western blot analysis, animal model, MAPK inhibition, MTT assay and cell invasion analysis for the investigation. The results demonstrated that the downregulation of the expression of HER-2 significantly inhibited cell motility and proliferation of human meningioma cells in vivo. Accordingly, in the HER-2-overexpression meningioma cells with the inhibition of ERK1/2, ERK5, JNK, in the cells with the ERK1/2, ERK5 inhibition, protein expression was markedly suppressed as well as the cell proliferation resistance. No difference was observed in the HER-2-overexpression meningioma cells with the inhibition of JNK. These findings suggest that HER-2 gene can affect the proliferation ability of human meningioma cells in vivo and MAPK signal pathway may contribute to the carcinogenesis and development of human meningiomas combinating with HER-2. PMID:27279438

  11. The role of MAPK signaling pathway in the Her-2-positive meningiomas.

    PubMed

    Wang, Zhaoyin; Wang, Weijia; Xu, Shan; Wang, Shanshan; Tu, Yi; Xiong, Yifeng; Mei, Jinhong; Wang, Chunliang

    2016-08-01

    Meningiomas are common types of adult nerve system tumors. Although most cases are considered benign, due to its high rate of recurrence and easy malignant progression to anaplastic meningioma they present a puzzle for the current treatment. The HER-2 oncogene has important value for meningioma cells development and progression. So far, little is known about the effect on the exact underlying signal pathway and molecular mechanisms of HER-2-positive meningioma cells. The goal of the present study was to determine the effects of HER-2 gene and possible involvement of MAPK signal pathway in human malignant meningioma. We applied q-PCR analysis, immunofluorescence (IF) staining, western blot analysis, animal model, MAPK inhibition, MTT assay and cell invasion analysis for the investigation. The results demonstrated that the downregulation of the expression of HER-2 significantly inhibited cell motility and proliferation of human meningioma cells in vivo. Accordingly, in the HER-2-overexpression meningioma cells with the inhibition of ERK1/2, ERK5, JNK, in the cells with the ERK1/2, ERK5 inhibition, protein expression was markedly suppressed as well as the cell proliferation resistance. No difference was observed in the HER-2-overexpression meningioma cells with the inhibition of JNK. These findings suggest that HER-2 gene can affect the proliferation ability of human meningioma cells in vivo and MAPK signal pathway may contribute to the carcinogenesis and development of human meningiomas combinating with HER-2. PMID:27279438

  12. Cross-talk between Smad and p38 MAPK signalling in transforming growth factor {beta} signal transduction in human glioblastoma cells

    SciTech Connect

    Dziembowska, Magdalena; Danilkiewicz, Malgorzata; Wesolowska, Aleksandra; Zupanska, Agata; Chouaib, Salem; Kaminska, Bozena . E-mail: bozenakk@nencki.gov.pl

    2007-03-23

    Transforming growth factor-beta (TGF-{beta}) is a multifunctional cytokine involved in the regulation of cell proliferation, differentiation, and survival. Malignant tumour cells often do not respond to TGF-{beta} by growth inhibition, but retain responsiveness to cytokine in regulating extracellular matrix deposition, cell adhesion, and migration. We demonstrated that TGF-{beta}1 does not affect viability or proliferation of human glioblastoma T98G, but increases transcriptional responses exemplified by induction of MMP-9 expression. TGF-{beta} receptors were functional in T98G glioblastoma cells leading to SMAD3/SMAD4 nuclear translocation and activation of SMAD-dependent promoter. In parallel, a selective activation of p38 MAPK, and phosphorylation of its substrates: ATF2 and c-Jun proteins were followed by a transient activation of AP-1 transcription factor. Surprisingly, an inhibition of p38 MAPK with a specific inhibitor, SB202190, abolished TGF-inducible activation of Smad-dependent promoter and decreased Smad2 phosphorylation. It suggests an unexpected interaction between Smad and p38 MAPK pathways in TGF-{beta}1-induced signalling.

  13. SB203580 Modulates p38 MAPK Signaling and Dengue Virus-Induced Liver Injury by Reducing MAPKAPK2, HSP27, and ATF2 Phosphorylation

    PubMed Central

    Sreekanth, Gopinathan Pillai; Chuncharunee, Aporn; Sirimontaporn, Aunchalee; Panaampon, Jutatip; Noisakran, Sansanee; Yenchitsomanus, Pa-thai; Limjindaporn, Thawornchai

    2016-01-01

    Dengue virus (DENV) infection causes organ injuries, and the liver is one of the most important sites of DENV infection, where viral replication generates a high viral load. The molecular mechanism of DENV-induced liver injury is still under investigation. The mitogen activated protein kinases (MAPKs), including p38 MAPK, have roles in the hepatic cell apoptosis induced by DENV. However, the in vivo role of p38 MAPK in DENV-induced liver injury is not fully understood. In this study, we investigated the role of SB203580, a p38 MAPK inhibitor, in a mouse model of DENV infection. Both the hematological parameters, leucopenia and thrombocytopenia, were improved by SB203580 treatment and liver transaminases and histopathology were also improved. We used a real-time PCR microarray to profile the expression of apoptosis-related genes. Tumor necrosis factor α, caspase 9, caspase 8, and caspase 3 proteins were significantly lower in the SB203580-treated DENV-infected mice than that in the infected control mice. Increased expressions of cytokines including TNF-α, IL-6 and IL-10, and chemokines including RANTES and IP-10 in DENV infection were reduced by SB203580 treatment. DENV infection induced the phosphorylation of p38MAPK, and its downstream signals including MAPKAPK2, HSP27 and ATF-2. SB203580 treatment did not decrease the phosphorylation of p38 MAPK, but it significantly reduced the phosphorylation of MAPKAPK2, HSP27, and ATF2. Therefore, SB203580 modulates the downstream signals to p38 MAPK and reduces DENV-induced liver injury. PMID:26901653

  14. Tgfβ-Smad and MAPK signaling mediate scleraxis and proteoglycan expression in heart valves.

    PubMed

    Barnette, Damien N; Hulin, Alexia; Ahmed, A S Ishtiaq; Colige, Alain C; Azhar, Mohamad; Lincoln, Joy

    2013-12-01

    Mature heart valves are complex structures consisting of three highly organized extracellular matrix layers primarily composed of collagens, proteoglycans and elastin. Collectively, these diverse matrix components provide all the necessary biomechanical properties for valve function throughout life. In contrast to healthy valves, myxomatous valve disease is the most common cause of mitral valve prolapse in the human population and is characterized by an abnormal abundance of proteoglycans within the valve tri-laminar structure. Despite the clinical significance, the etiology of this phenotype is not known. Scleraxis (Scx) is a basic-helix-loop-helix transcription factor that we previously showed to be required for establishing heart valve structure during remodeling stages of valvulogenesis. In this study, we report that remodeling heart valves from Scx null mice express decreased levels of proteoglycans, particularly chondroitin sulfate proteoglycans (CSPGs), while overexpression in embryonic avian valve precursor cells and adult porcine valve interstitial cells increases CSPGs. Using these systems we further identify that Scx is positively regulated by canonical Tgfβ2 signaling during this process and this is attenuated by MAPK activity. Finally, we show that Scx is increased in myxomatous valves from human patients and mouse models, and overexpression in human mitral valve interstitial cells modestly increases proteoglycan expression consistent with myxomatous mitral valve phenotypes. Together, these studies identify an important role for Scx in regulating proteoglycans in embryonic and mature valve cells and suggest that imbalanced regulation could influence myxomatous pathogenesis.

  15. Tgfβ-Smad and MAPK signaling mediate scleraxis and proteoglycan expression in heart valves

    PubMed Central

    Barnette, Damien N.; Hulin, Alexia; Ahmed, A.S. Ishtiaq; Colige, Alain C.; Azhar, Mohammed; Lincoln, Joy

    2013-01-01

    Mature heart valves are complex structures consisting of three highly organized extracellular matrix layers primarily composed of collagens, proteoglycans and elastin. Collectively, these diverse matrix components provide all the necessary biomechanical properties for valve function throughout life. In contrast to healthy valves, myxomatous valve disease is the most common cause of mitral valve prolapse in the human population and is characterized by an abnormal abundance of proteoglycans within the valve tri-laminar structure. Despite the clinical significance, the etiology of this phenotype is not known. Scleraxis (Scx) is a basic-helix-loop-helix transcription factor that we previously showed to be required for establishing heart valve structure during remodeling stages of valvulogenesis. In this study, we report that remodeling heart valves from Scx null mice express decreased levels of proteoglycans, particularly chondroitin sulfate proteoglycans (CSPGs), while overexpression in embryonic avian valve precursor cells and adult porcine valve interstitial cells increases CSPGs. Using these systems we further identify that Scx is positively regulated by canonical Tgfβ2 signaling during this process and this is attenuated by MAPK activity. Finally, we show that Scx is increased in myxomatous valves from human patients and mouse models, and overexpression in human mitral valve interstitial cells modestly increases proteoglycan expression consistent with myxomatous mitral valve phenotypes. Together, these studies identify an important role for Scx in regulating proteoglycans in embryonic and mature valve cells and suggest that imbalanced regulation could influence myxomatous pathogenesis. PMID:24157418

  16. Glucocorticoid receptor-induced MAPK phosphatase-1 (MPK-1) expression inhibits paclitaxel-associated MAPK activation and contributes to breast cancer cell survival.

    PubMed

    Wu, Wei; Pew, Travis; Zou, Min; Pang, Diana; Conzen, Suzanne D

    2005-02-11

    Glucocorticoid receptor (GR) activation has recently been shown to inhibit apoptosis in breast epithelial cells. We have previously described a group of genes that is rapidly up-regulated in these cells following dexamethasone (Dex) treatment. In an effort to dissect the mechanisms of GR-mediated breast epithelial cell survival, we now examine the molecular events downstream of GR activation. Here we show that GR activation leads to both the rapid induction of MAPK phosphatase-1 (MKP-1) mRNA and its sustained expression. Induction of the MKP-1 protein in the MCF10A-Myc and MDA-MB-231 breast epithelial cell lines was also seen. Paclitaxel treatment resulted in MAPK activation and apoptosis of MDA-MB-231 breast cancer cells, and both processes were inhibited by Dex pretreatment. Furthermore, induction of MKP-1 correlated with the inhibition of extracellular signal-regulated kinase (ERK1/2) and c-Jun N-terminal kinase (JNK) activity, whereas p38 activity was minimally affected. Blocking Dex-induced MKP-1 induction using small interfering RNA increased ERK1/2 and JNK phosphorylation and decreased cell survival. ERK1/2 and JNK inactivation was associated with Ets-like transcription factor-1 (ELK-1) dephosphorylation. To explore the gene expression changes that occur downstream of ELK-1 dephosphorylation, we used a combination of temporal gene expression data and promoter element analyses. This approach revealed a previously unrecognized transcriptional target of ELK-1, the human tissue plasminogen activator (tPA). We verified the predicted ELK-1--> tPA transcriptional regulatory relationship using a luciferase reporter assay. We conclude that GR-mediated MAPK inactivation contributes to cell survival and that the potential transcriptional targets of this inhibition can be identified from large scale gene array analysis.

  17. Identification, Nomenclature, and Evolutionary Relationships of Mitogen-Activated Protein Kinase (MAPK) Genes in Soybean

    PubMed Central

    Neupane, Achal; Nepal, Madhav P.; Piya, Sarbottam; Subramanian, Senthil; Rohila, Jai S.; Reese, R. Neil; Benson, Benjamin V.

    2013-01-01

    Mitogen-activated protein kinase (MAPK) genes in eukaryotes regulate various developmental and physiological processes including those associated with biotic and abiotic stresses. Although MAPKs in some plant species including Arabidopsis have been identified, they are yet to be identified in soybean. Major objectives of this study were to identify GmMAPKs, assess their evolutionary relationships, and analyze their functional divergence. We identified a total of 38 MAPKs, eleven MAPKKs, and 150 MAPKKKs in soybean. Within the GmMAPK family, we also identified a new clade of six genes: four genes with TEY and two genes with TQY motifs requiring further investigation into possible legume-specific functions. The results indicated the expansion of the GmMAPK families attributable to the ancestral polyploidy events followed by chromosomal rearrangements. The GmMAPK and GmMAPKKK families were substantially larger than those in other plant species. The duplicated GmMAPK members presented complex evolutionary relationships and functional divergence when compared to their counterparts in Arabidopsis. We also highlighted existing nomenclatural issues, stressing the need for nomenclatural consistency. GmMAPK identification is vital to soybean crop improvement, and novel insights into the evolutionary relationships will enhance our understanding about plant genome evolution. PMID:24137047

  18. Alterations in left ventricular function during intermittent hypoxia: Possible involvement of O-GlcNAc protein and MAPK signaling.

    PubMed

    Guo, Xueling; Shang, Jin; Deng, Yan; Yuan, Xiao; Zhu, Die; Liu, Huiguo

    2015-07-01

    Obstructive sleep apnea, characterized by recurrent episodes of hypoxia [intermittent hypoxia (IH)], has been identified as a risk factor for cardiovascular diseases. The O-linked β-N-acetylglucosamine (O-GlcNAc) modification (O-GlcNAcylation) of proteins has important regulatory implications on the pathophysiology of cardiovascular disorders. In this study, we examined the role of O-GlcNAcylation in cardiac architecture and left ventricular function following IH. Rats were randomly assigned to a normoxia and IH group (2 min 21% O2; 2 min 6-8% O2). Left ventricular function, myocardial morphology and the levels of signaling molecules were then measured. IH induced a significant increase in blood pressure, associated with a gradually abnormal myocardial architecture. The rats exposed to 2 or 3 weeks of IH presented with augmented left ventricular systolic and diastolic function, which declined at week 4. Consistently, the O-GlcNAc protein and O-GlcNAcase (OGA) levels in the left ventricular tissues steadily increased following IH, reaching peak levels at week 3. The O-GlcNAc transferase (OGT), extracellular signal-regulated kinase 1/2 (ERK1/2) and the p38 mitogen-activated protein kinase (p38 MAPK) phosphorylation levels were affected in an opposite manner. The phosphorylation of calcium/calmodulin-dependent protein kinase II (CaMKII) remained unaltered. In parallel, compared with exposure to normoxia, 4 weeks of IH augmented the O-GlcNAc protein, OGT, phosphorylated ERK1/2 and p38 MAPK levels, accompanied by a decrease in OGA levels and an increase in the levels of myocardial nuclear factor-κB (NF-κB), inflammatory cytokines, caspase-3 and cardiomyocyte apoptosis. Taken together, our suggest a possible involvement of O-GlcNAc protein and MAPK signaling in the alterations of left ventricular function and cardiac injury following IH.

  19. Multiple MAPK cascades regulate the transcription of IME1, the master transcriptional activator of meiosis in Saccharomyces cerevisiae.

    PubMed

    Kahana-Edwin, Smadar; Stark, Michal; Kassir, Yona

    2013-01-01

    The choice between alternative developmental pathways is primarily controlled at the level of transcription. Induction of meiosis in budding yeasts in response to nutrient levels provides a system to investigate the molecular basis of cellular decision-making. In Saccharomyces cerevisiae, entry into meiosis depends on multiple signals converging upon IME1, the master transcriptional activator of meiosis. Here we studied the regulation of the cis-acting regulatory element Upstream Activation Signal (UAS)ru, which resides within the IME1 promoter. Guided by our previous data acquired using a powerful high-throughput screening system, here we provide evidence that UASru is regulated by multiple stimuli that trigger distinct signal transduction pathways as follows: (i) The glucose signal inhibited UASru activity through the cyclic AMP (cAMP/protein kinase A (PKA) pathway, targeting the transcription factors (TFs), Com2 and Sko1; (ii) high osmolarity activated UASru through the Hog1/mitogen-activated protein kinase (MAPK) pathway and its corresponding TF Sko1; (iii) elevated temperature increased the activity of UASru through the cell wall integrity pathway and the TFs Swi4/Mpk1 and Swi4/Mlp1; (iv) the nitrogen source repressed UASru activity through Sum1; and (v) the absence of a nitrogen source was detected and transmitted to UASru by the Kss1 and Fus3 MAPK pathways through their respective downstream TFs, Ste12/Tec1 and Ste12/Ste12 as well as by their regulators Dig1/2. These signaling events were specific to UASru; they did not affect the mating and filamentation response elements that are regulated by MAPK pathways. The complex regulation of UASru through all the known vegetative MAPK pathways is unique to S. cerevisiae and is specific for IME1, likely because it is the master regulator of gametogenesis. PMID:24236068

  20. Multiple MAPK Cascades Regulate the Transcription of IME1, the Master Transcriptional Activator of Meiosis in Saccharomyces cerevisiae

    PubMed Central

    Kahana-Edwin, Smadar; Stark, Michal; Kassir, Yona

    2013-01-01

    The choice between alternative developmental pathways is primarily controlled at the level of transcription. Induction of meiosis in budding yeasts in response to nutrient levels provides a system to investigate the molecular basis of cellular decision-making. In Saccharomyces cerevisiae, entry into meiosis depends on multiple signals converging upon IME1, the master transcriptional activator of meiosis. Here we studied the regulation of the cis-acting regulatory element Upstream Activation Signal (UAS)ru, which resides within the IME1 promoter. Guided by our previous data acquired using a powerful high-throughput screening system, here we provide evidence that UASru is regulated by multiple stimuli that trigger distinct signal transduction pathways as follows: (i) The glucose signal inhibited UASru activity through the cyclic AMP (cAMP/protein kinase A (PKA) pathway, targeting the transcription factors (TFs), Com2 and Sko1; (ii) high osmolarity activated UASru through the Hog1/mitogen-activated protein kinase (MAPK) pathway and its corresponding TF Sko1; (iii) elevated temperature increased the activity of UASru through the cell wall integrity pathway and the TFs Swi4/Mpk1 and Swi4/Mlp1; (iv) the nitrogen source repressed UASru activity through Sum1; and (v) the absence of a nitrogen source was detected and transmitted to UASru by the Kss1 and Fus3 MAPK pathways through their respective downstream TFs, Ste12/Tec1 and Ste12/Ste12 as well as by their regulators Dig1/2. These signaling events were specific to UASru; they did not affect the mating and filamentation response elements that are regulated by MAPK pathways. The complex regulation of UASru through all the known vegetative MAPK pathways is unique to S. cerevisiae and is specific for IME1, likely because it is the master regulator of gametogenesis. PMID:24236068

  1. Role of protein kinase C in the TBT-induced inhibition of lytic function and MAPK activation in human natural killer cells

    PubMed Central

    Abraha, Abraham B.; Rana, Krupa; Whalen, Margaret M.

    2010-01-01

    Human natural killer (NK) cells are lymphocytes that destroy tumor and virally infected cells. Previous studies have shown that exposures of NK cells to tributyltin (TBT) greatly diminish their ability to destroy tumor cells (lytic function) while activating mitogen-activated protein kinases (MAPK) (p44/42, p38, and JNK) in the NK cells. The signaling pathway that regulates NK lytic function appears to include activation of protein kinase C (PKC) as well as MAPK activity. The TBT-induced activation of MAPKs would trigger a portion of the NK lytic signaling pathway, which would then leave the NK cell unable to trigger this pathway in response to a subsequent encounter with a target cell. In the present study we evaluated the involvement of PKC in the inhibition of NK lysis of tumor cells and activation of MAPKs caused by TBT exposures. TBT caused a 2–3 fold activation of PKC at concentrations ranging from 50–300 nM (16–98 ng/mL), indicating that activation of PKC occurs in response to TBT exposures. This would then leave the NK cell unable to respond to targets. Treatment with the PKC inhibitor, bisindolylmaleimide I, caused an 85% decrease in the ability of NK cells to lyse tumor cells validating the involvement of PKC in the lytic signaling pathway. The role of PKC in the activation of MAPKs by TBT was also investigated using bisindolylmaleimide I. The results indicated that in NK cells where PKC activation was blocked there was no activation of the MAPK, p44/42 in response to TBT. However, TBT-induced activation of the MAPKs, p38 and JNK did not require PKC activation. These results indicate the pivotal role of PKC in the TBT-induced loss of NK lytic function including the activation of p44/42 by TBT in NK cells. PMID:20390410

  2. Oligonol Ameliorates CCl₄-Induced Liver Injury in Rats via the NF-Kappa B and MAPK Signaling Pathways.

    PubMed

    Bak, Jeonghyeon; Je, Nam Kyung; Chung, Hae Young; Yokozawa, Takako; Yoon, Sik; Moon, Jeon-Ok

    2016-01-01

    Oxidative stress is thought to be a key risk factor in the development of hepatic diseases. Blocking or retarding the reactions of oxidation and the inflammatory process by antioxidants could be a promising therapeutic intervention for prevention or treatment of liver injuries. Oligonol is a low molecular weight polyphenol containing catechin-type monomers and oligomers derived from lychee fruit. In this study, we investigated the anti-inflammatory effect of oligonol on carbon tetrachloride- (CCl4-) induced acute hepatic injury in rats. Oral administration of oligonol (10 or 50 mg/kg) reduced CCl4-induced abnormalities in liver histology and serum AST and serum ALT levels. Oligonol treatment attenuated the CCl4-induced production of inflammatory mediators, including TNF-α, IL-1β, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) mRNA levels. Western blot analysis showed that oligonol suppressed proinflammatory nuclear factor-kappa B (NF-κB) p65 activation, phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 mitogen-activated protein kinases (MAPKs) as well as Akt. Oligonol exhibited strong antioxidative activity in vitro and in vivo, and hepatoprotective activity against t-butyl hydroperoxide-induced HepG2 cells. Taken together, oligonol showed antioxidative and anti-inflammatory effects in CCl4-intoxicated rats by inhibiting oxidative stress and NF-κB activation via blockade of the activation of upstream kinases including MAPKs and Akt.

  3. Oligonol Ameliorates CCl4-Induced Liver Injury in Rats via the NF-Kappa B and MAPK Signaling Pathways

    PubMed Central

    Bak, Jeonghyeon; Je, Nam Kyung; Chung, Hae Young; Yokozawa, Takako; Yoon, Sik; Moon, Jeon-Ok

    2016-01-01

    Oxidative stress is thought to be a key risk factor in the development of hepatic diseases. Blocking or retarding the reactions of oxidation and the inflammatory process by antioxidants could be a promising therapeutic intervention for prevention or treatment of liver injuries. Oligonol is a low molecular weight polyphenol containing catechin-type monomers and oligomers derived from lychee fruit. In this study, we investigated the anti-inflammatory effect of oligonol on carbon tetrachloride- (CCl4-) induced acute hepatic injury in rats. Oral administration of oligonol (10 or 50 mg/kg) reduced CCl4-induced abnormalities in liver histology and serum AST and serum ALT levels. Oligonol treatment attenuated the CCl4-induced production of inflammatory mediators, including TNF-α, IL-1β, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) mRNA levels. Western blot analysis showed that oligonol suppressed proinflammatory nuclear factor-kappa B (NF-κB) p65 activation, phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 mitogen-activated protein kinases (MAPKs) as well as Akt. Oligonol exhibited strong antioxidative activity in vitro and in vivo, and hepatoprotective activity against t-butyl hydroperoxide-induced HepG2 cells. Taken together, oligonol showed antioxidative and anti-inflammatory effects in CCl4-intoxicated rats by inhibiting oxidative stress and NF-κB activation via blockade of the activation of upstream kinases including MAPKs and Akt. PMID:26798422

  4. Caffeine inhibits UV-mediated NF-kappaB activation in A2058 melanoma cells: an ATM-PKCdelta-p38 MAPK-dependent mechanism.

    PubMed

    Ravi, Dashnamoorthy; Muniyappa, Harish; Das, Kumuda C

    2008-01-01

    Mammalian ultraviolet (UV) radiation response is a gene induction cascade activated by several transcription factors, including NF-kappaB. Although NF-kappaB is induced by UV radiation, the signal transduction mechanism remains relatively unclear. In the present study, we show that UV-induced NF-kappaB activation is mediated by the activation of Ataxia telangiecia mutated (ATM) and protein kinase C (PKC). We also show that caffeine specifically inhibits UV-mediated NF-kappaB activation, but not TNFalpha-mediated NF-kappaB activation. In addition, our study shows that ATM, but not ATM-Rad3-related (ATR) or DNA-dependent protein kinase (DNA-PK) is involved in UV-induced NF-kappaB activation. Because SB203580 (a p38 MAPK inhibitor), or Calphostin C or rottlerin (PKC inhibitors) was able to inhibit UV-mediated NF-kappaB activation, we evaluated whether caffeine could inhibit p38 MAPK or PKC activity. Caffeine or rottlerin inhibited UV-induced phosphorylation of p38 MAPK, but not anisomycin-induced phosphorylation of p38 MAPK, suggesting that p38 MAPK is downstream of PKC. Additionally, caffeine could effectively inhibit UV-induced increases in PKC activity. Taken together, our study demonstrates that caffeine is a potent inhibitor of UV-induced NF-kappaB activation. Additionally, this inhibition occurs due to the inhibitory action of caffeine on ATM and PKC, resulting in the inhibition of p38 MAPK activation.

  5. C5a Induces the Synthesis of IL-6 and TNF-α in Rat Glomerular Mesangial Cells through MAPK Signaling Pathways

    PubMed Central

    Ji, Mingde; Lu, Yanlai; Zhao, Chenhui; Gao, Wenxing; He, Fengxia; Zhang, Jing; Zhao, Dan; Qiu, Wen; Wang, Yingwei

    2016-01-01

    Inflammatory response has been reported to contribute to the renal lesions in rat Thy-1 nephritis (Thy-1N) as an animal model of human mesangioproliferative glomerulonephritis (MsPGN). Besides C5b-9 complex, C5a is also a potent pro-inflammatory mediator and correlated to severity of various nephritic diseases. However, the role of C5a in mediating pro-inflammatory cytokine production in rats with Thy-1N is poorly defined. In the present studies, the levels of C5a, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were first determined in the renal tissues of rats with Thy-1N. Then, the expression of IL-6 and TNF-α was detected in rat glomerular mesangial cells (GMC) stimulated with our recombinant rat C5a in vitro. Subsequently, the activation of mitogen-activated protein kinase (MAPK) signaling pathways (p38 MAPK, ERK1/2 and JNK) and their roles in the regulation of IL-6 and TNF-α production were examined in the GMC induced by C5a. The results showed that the levels of C5a, IL-6 and TNF-α were markedly increased in the renal tissues of Thy-1N rats. Rat C5a stimulation in vitro could up-regulate the expression of IL-6 and TNF-α in rat GMC, and the activation of MAPK signaling pathways was involved in the induction of IL-6 and TNF-α. Mechanically, p38 MAPK activation promoted IL-6 production, while either ERK1/2 or JNK activation promoted TNF-α production in the GMC with exposure to C5a. Taken together, these data implicate that C5a induces the synthesis of IL-6 and TNF-α in rat GMC through the activation of MAPK signaling pathways. PMID:27583546

  6. Methanol Extract of Croton Pycnanthus Benth. Inhibits Osteoclast Differentiation by Suppressing the MAPK and NF-κB Signaling Pathways

    PubMed Central

    Lee, Jiyeon

    2014-01-01

    Background Osteoclasts are differentiated from monocytes/macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-kappa B (NF-κB) ligand (RANKL). Croton pycnanthus Benth. (CPB) is a herbal plant that belongs to Euphorbiaceae family. The aim of this study was to investigate the effects of CPB on osteoclastogenesis and RANKL-dependent signaling pathways. Methods Methanol extract of CPB was obtained from International Biological Material Research Center. Osteoclast differentiation was achieved by culturing mouse bone marrow-derived macrophages (BMMs) with M-CSF and RANKL. Osteoclast numbers were evaluated by counting multinuclear cells positive for tartrate-resistant acid phosphatase (TRAP). mRNA and protein levels were analyzed by real-time polymerase chain reaction (PCR) and Western blotting, respectively. The activation of signaling molecules were assessed after acute stimulation of cells with high dose of RANKL by Western blotting with phospho-specific antibodies. Results CPB reduced the generation of TRAP-positive multinucleated cells and the activation of mitogen-activated protein kinase (MAPK) and NF-κB signaling pathways. The induction of the expression of c-Fos, nuclear factor-activated T cells c1 (NFATc1) and dendritic cell-specific transmembrane protein (DC-STAMP) by RANKL was also suppressed. Conclusions CPB exerts negative effects on osteoclast differentiation in response to the RANKL. The inhibitory mechanism involves the suppression of MAPK and NF-κB signaling pathways and subsequently the down-regulation of c-Fos and NFATc1 transcription factors. PMID:25489576

  7. 11-epi-Sinulariolide Acetate Reduces Cell Migration and Invasion of Human Hepatocellular Carcinoma by Reducing the Activation of ERK1/2, p38MAPK and FAK/PI3K/AKT/mTOR Signaling Pathways

    PubMed Central

    Lin, Jen-Jie; Su, Jui-Hsin; Tsai, Chi-Chu; Chen, Yi-Jen; Liao, Ming-Hui; Wu, Yu-Jen

    2014-01-01

    Cancer metastasis is one of the major causes of death in cancer. An active compound, 11-epi-sinulariolide acetate (11-epi-SA), isolated from the cultured soft coral Sinularia flexibilis has been examined for potential anti-cell migration and invasion effects on hepatocellular carcinoma cells (HCC). However, the molecular mechanism of anti-migration and invasion by 11-epi-SA on HCC, along with their corresponding effects, remain poorly understood. In this study, we investigated anti-migration and invasion effects and the underlying mechanism of 11-epi-SA in HA22T cells, and discovered by trans-well migration and invasion assays that 11-epi-SA provided a concentration-dependent inhibitory effect on the migration of human HCC HA22T cells. After treatment with 11-epi-SA for 24 h, there were suppressed protein levels of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9) and urokinase-type plasminogen activator (uPA) in HA22T cells. Meanwhile, the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase-2 (TIMP-2) were increased in a concentration-dependent manner. Further investigation revealed that 11-epi-SA suppressed the phosphorylation of ERK1/2 and p38MAPK. The 11-epi-SA also suppressed the expression of the phosphorylation of FAK/PI3K/AKT/mTOR pathways. PMID:25222667

  8. Obacunone exhibits anti-proliferative and anti-aromatase activity in vitro by inhibiting the p38 MAPK signaling pathway in MCF-7 human breast adenocarcinoma cells.

    PubMed

    Kim, Jinhee; Jayaprakasha, G K; Patil, Bhimanagouda S

    2014-10-01

    Overexpression of the aromatase enzyme CYP19 has been implicated in the onset of estrogen-dependent breast carcinogenesis. Obacunone, a natural compound present in citrus fruits, has been demonstrated for various biological activities including anti-cancer and anti-inflammatory properties. In the present study, we have isolated obacunone and obacunone glucoside (OG) from lemon seeds, then fractionated these compounds using chromatographic techniques and characterized them by HPLC, LC-MS, and 2D NMR spectral analysis. To investigate the mechanism of anti-cancer and anti-aromatase activities of limonoids, their cytotoxic effect was tested on human breast cancer (MCF-7) and non-malignant (MCF-12F) breast cells. MTT assays confirmed that obacunone was strongly inhibited MCF-7 cell proliferation without affecting non-malignant breast cells. Treatment with obacunone increased apoptosis by up-regulating expression of the pro-apoptotic protein Bax and down-regulating the anti-apoptotic protein Bcl2, as well as inducing G1 cell cycle arrest. In addition, obacunone significantly inhibited aromatase activity in an in vitro enzyme assay. Exposure of MCF-7 breast cancer cells to obacunone down-regulated expression of inflammatory molecules including nuclear factor-kappa B (NF-κB) and cyclooxygenase-2 (COX-2). Furthermore, we found that obacunone inhibited COX-2 and NF-κB by activation of the p38 mitogen-activated protein kinase (MAPK). Finally, the uptake level of obacunone into MCF-7 cells was measured by HPLC and its structure was confirmed by LC-HR-MS. This study demonstrated that obacunone may have the potential to prevent estrogen-responsive breast cancer through inhibition of the aromatase enzyme and inflammatory pathways, as well as activation of apoptosis. PMID:24927687

  9. Hypoxia-inducible factor-2 alpha promotes the proliferation of human placenta-derived mesenchymal stem cells through the MAPK/ERK signaling pathway

    PubMed Central

    Zhu, Chengxing; Yu, Jiong; Pan, Qiaoling; Yang, Jinfeng; Hao, Guangshu; Wang, Yingjie; Li, Lanjuan; Cao, Hongcui

    2016-01-01

    Human placenta-derived mesenchymal stem cells (hPMSCs) reside in a physiologically low-oxygen microenvironment. Hypoxia influences a variety of stem cell cellular activities, frequently involving hypoxia-inducible factor-2 alpha (HIF-2α). This research showed that hPMSCs cultured in hypoxic conditions (5% O2) exhibited a more naïve morphology and had a higher proliferative capability and higher HIF-2α expression than hPMSCs cultured in normoxic conditions (21% O2). Similar to the hypoxic cultures, hPMSCs over-expressing HIF-2α showed higher proliferative potential and higher expression of CCND1 (CyclinD1), MYC (c-Myc), POU5F1 (Oct4) and the components of the MAPK/ERK pathway. In contrast, these genes were down-regulated in the HIF-2α-silenced hPMSCs. After adding the MAPK/ERK inhibitor PD0325901, cell growth and the expression of CCND1 and MYC were inhibited. Furthermore, the chromatin immunoprecipitation (ChIP) assay and electrophoretic mobility shift assay (EMSA) showed that HIF-2α bound to the MAPK3 (ERK1) promoter, indicative of its direct regulation of MAPK/ERK components at the transcriptional level during hPMSC expansion. Taken together, our results suggest that HIF-2α facilitated the preservation of hPMSC stemness and promoted their proliferation by regulating CCND1 and MYC through the MAPK/ERK signaling pathway. PMID:27765951

  10. Crosstalk Between MAPK/ERK and PI3K/AKT Signal Pathways During Brain Ischemia/Reperfusion

    PubMed Central

    Zhou, Jing; Du, Ting; Li, Baoman; Rong, Yan; Verkhratsky, Alexei

    2015-01-01

    The epidermal growth factor receptor (EGFR) is linked to the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) and Raf/mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK1/2) signaling pathways. During brain ischemia/reperfusion, EGFR could be transactivated, which stimulates these intracellular signaling cascades that either protect cells or potentiate cell injury. In the present study, we investigated the activation of EGFR, PI3K/AKT, and Raf/MAPK/ERK1/2 during ischemia or reperfusion of the brain using the middle cerebral artery occlusion model. We found that EGFR was phosphorylated and transactivated during both ischemia and reperfusion periods. During ischemia, the activity of PI3K/AKT pathway was significantly increased, as judged from the strong phosphorylation of AKT; this activation was suppressed by the inhibitors of EGFR and Zn-dependent metalloproteinase. Ischemia, however, did not induce ERK1/2 phosphorylation, which was dependent on reperfusion. Coimmunoprecipitation of Son of sevenless 1 (SOS1) with EGFR showed increased association between the receptor and SOS1 in ischemia, indicating the inhibitory node downstream of SOS1. The inhibitory phosphorylation site of Raf-1 at Ser259, but not its stimulatory phosphorylation site at Ser338, was phosphorylated during ischemia. Furthermore, ischemia prompted the interaction between Raf-1 and AKT, while both the inhibitors of PI3K and AKT not only abolished AKT phosphorylation but also restored ERK1/2 phosphorylation. All these findings suggest that Raf/MAPK/ERK1/2 signal pathway is inhibited by AKT via direct phosphorylation and inhibition at Raf-1 node during ischemia. During reperfusion, we observed a significant increase of ERK1/2 phosphorylation but no change in AKT phosphorylation. Inhibitors of reactive oxygen species and phosphatase and tensin homolog restored AKT phosphorylation but abolished ERK1/2 phosphorylation, suggesting that the reactive oxygen species

  11. Mitogen-activated protein kinase cascades in signaling plant growth and development.

    PubMed

    Xu, Juan; Zhang, Shuqun

    2015-01-01

    Mitogen-activated protein kinase (MAPK) cascades are ubiquitous signaling modules in eukaryotes. Early research of plant MAPKs has been focused on their functions in immunity and stress responses. Recent studies reveal that they also play essential roles in plant growth and development downstream of receptor-like protein kinases (RLKs). With only a limited number of MAPK components, multiple functional pathways initiated from different receptors often share the same MAPK components or even a complete MAPK cascade. In this review, we discuss how MAPK cascades function as molecular switches in response to spatiotemporal-specific ligand-receptor interactions and the availability of downstream substrates. In addition, we discuss other possible mechanisms governing the functional specificity of plant MAPK cascades, a question central to our understanding of MAPK functions.

  12. Role of amygdala MAPK activation on immobility behavior of forced swim rats.

    PubMed

    Huang, Tung-Yi; Lin, Chih-Hung

    2006-10-01

    The role of amygdala mitogen-activated protein kinase (MAPK) in rats during a forced swim test was investigated. The variation of amygdala MAPK level was studied in control rats and early-life maternally deprived rats. A forced swim test was carried out to estimate the immobility level. The data showed that the immobility time of rats that received maternal deprivation in early life was longer than that of control rats and Western blot analysis also showed that the amygdala phospho-MAPK level in maternally deprived rats was almost two times higher than in control rats. Intra-amygdala infusion of PD098059 or U0126, MEK inhibitors, suppressed immobility behavior during the forced swim test in both rats. Western blot analysis also showed that the amygdala MAPK activities in both rats infused with MEK inhibitors were also suppressed in parallel with expression of immobility behavior. The suppressed MAPK activities as well as the restoration of immobility time returned to the original level 48 h later. These results suggest that amygdala MAPK activation might play a role in the regulation of immobility behavior in rats during the forced swim test. Moreover, it could provide a hint that amygdala MAPK activation might be involved in the formation of depression-like behavior.

  13. Hesperetin Inhibits Vascular Formation by Suppressing of the PI3K/AKT, ERK, and p38 MAPK Signaling Pathways.

    PubMed

    Kim, Gi Dae

    2014-12-01

    Hesperetin has been shown to possess a potential anti-angiogenic effect, including vascular formation by endothelial cells. However, the mechanisms underlying the potential anti-angiogenic activity of hesperetin are not fully understood. In the present study, we evaluated whether hesperetin has anti-angiogenic effects in human umbilical vascular endothelial cells (HUVECs). HUVECs were treated with 50 ng/mL vascular endothelial growth factor (VEGF) to induce proliferation as well as vascular formation, followed by treatment with several doses of hesperetin (25, 50, and 100 μM) for 24 h. Cell proliferation and vascular formation were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assay, respectively. In addition, cell signaling related to cell proliferation and vascular formation was analyzed by western blot. Furthermore, a mouse aorta ring assay was performed to confirm the effect of hesperetin on vascular formation. Hesperetin treatment did not cause differences in HUVECs proliferation. However, hesperetin significantly inhibited VEGF-induced cell migration and tube formation of HUVECs (P<0.05). Moreover, hesperetin suppressed the expression of ERK, p38 MAPK, and PI3K/AKT in the VEGF-induced HUVECs. In an ex vivo model, hesperetin also suppressed microvessel sprouting of mouse aortic rings. Taken together, the findings suggest that hesperetin inhibited vascular formation by endothelial cells via the inhibition of the PI3K/AKT, ERK and p38 MAPK signaling.

  14. Zearalenone Mycotoxin Affects Immune Mediators, MAPK Signalling Molecules, Nuclear Receptors and Genome-Wide Gene Expression in Pig Spleen

    PubMed Central

    Pistol, Gina Cecilia; Braicu, Cornelia; Motiu, Monica; Gras, Mihail Alexandru; Marin, Daniela Eliza; Stancu, Mariana; Calin, Loredana; Israel-Roming, Florentina; Berindan-Neagoe, Ioana; Taranu, Ionelia

    2015-01-01

    The toxicity of zearalenone (ZEA) was evaluated in swine spleen, a key organ for the innate and adaptative immune response. Weaned pigs were fed for 18 days with a control or a ZEA contaminated diet. The effect of ZEA was assessed on wide genome expression, pro- (TNF-α, IL-8, IL-6, IL-1β, IFN-γ) and anti-inflammatory (IL-10, IL-4) cytokines, other molecules involved in inflammatory processes (MMPs/TIMPs), as well as signaling molecules, (p38/JNK1/JNK2-MAPKs) and nuclear receptors (PPARγ/NFkB/AP-1/STAT3/c-JUN). Microarray analysis showed that 46% of total number of differentially expressed genes was involved in cellular signaling pathway, 13% in cytokine network and 10% in the inflammatory response. ZEA increased expression and synthesis of pro- inflammatory (TNF-α, IL-8, IL-6, IL-1β) and had no effect on IFN-γ, IL-4 and IL-10 cytokines in spleen. The inflammatory stimulation might be a consequence of JNK pathway activation rather than of p-38MAPK and NF-kB involvement whose gene and protein expression were suppressed by ZEA action. In summary, our findings indicated the role of ZEA as an immune disruptor at spleen level. PMID:26011631

  15. Importin-7 mediates memory consolidation through regulation of nuclear translocation of training-activated MAPK in Drosophila.

    PubMed

    Li, Qian; Zhang, Xuchen; Hu, Wantong; Liang, Xitong; Zhang, Fang; Wang, Lianzhang; Liu, Zhong-Jian; Zhong, Yi

    2016-03-15

    Translocation of signaling molecules, MAPK in particular, from the cytosol to nucleus represents a universal key element in initiating the gene program that determines memory consolidation. Translocation mechanisms and their behavioral impact, however, remain to be determined. Here, we report that a highly conserved nuclear transporter, Drosophila importin-7 (DIM-7), regulates import of training-activated MAPK for consolidation of long-term memory (LTM). We show that silencing DIM-7 functions results in impaired LTM, whereas overexpression of DIM-7 enhances LTM. This DIM-7-dependent regulation of LTM is confined to a consolidation time window and in mushroom body neurons. Image data show that bidirectional alteration in DIM-7 expression results in proportional changes in the intensity of training-activated MAPK accumulated within the nuclei of mushroom body neurons during LTM consolidation. Such DIM-7-regulated nuclear accumulation of activated MAPK is observed only in the training specified for LTM induction and determines the amplitude, but not the time course, of memory consolidation. PMID:26929354

  16. Importin-7 mediates memory consolidation through regulation of nuclear translocation of training-activated MAPK in Drosophila

    PubMed Central

    Li, Qian; Zhang, Xuchen; Liang, Xitong; Zhang, Fang; Wang, Lianzhang; Zhong, Yi

    2016-01-01

    Translocation of signaling molecules, MAPK in particular, from the cytosol to nucleus represents a universal key element in initiating the gene program that determines memory consolidation. Translocation mechanisms and their behavioral impact, however, remain to be determined. Here, we report that a highly conserved nuclear transporter, Drosophila importin-7 (DIM-7), regulates import of training-activated MAPK for consolidation of long-term memory (LTM). We show that silencing DIM-7 functions results in impaired LTM, whereas overexpression of DIM-7 enhances LTM. This DIM-7–dependent regulation of LTM is confined to a consolidation time window and in mushroom body neurons. Image data show that bidirectional alteration in DIM-7 expression results in proportional changes in the intensity of training-activated MAPK accumulated within the nuclei of mushroom body neurons during LTM consolidation. Such DIM-7–regulated nuclear accumulation of activated MAPK is observed only in the training specified for LTM induction and determines the amplitude, but not the time course, of memory consolidation. PMID:26929354

  17. Resistin increases platelet P-selectin levels via p38 MAPK signal pathway.

    PubMed

    Qiu, Wenbing; Chen, Naping; Zhang, Qin; Zhuo, Liyuan; Wang, Xihong; Wang, Dongming; Jin, Hong

    2014-03-01

    Resistin, an adipokine associated with the metabolic syndrome, is believed to have a role in thrombotic conditions. This work analyses the effects of resistin on P-selectin expression using a combination of ex vivo human studies, in vivo animal models and in vitro cell cultures. Human platelets and vascular endothelial cells were incubated with resistin, with or without anti-Toll-like receptor 4 (TLR-4) or mitogen-activated protein kinases (MAPK) pathway inhibitors, whereas mice were treated with resistin infusion followed by analysis of P-selectin expression. Resistin increased both human and murine platelet P-selectin expression compared with controls (human: 48.02% ± 7.6% vs 35.12% ± 2.62%, p < 0.05; mouse: 8.17% ± 0.37% vs 4.44% ± 0.37%, p < 0.05), through the p38 MAPK pathway. In contrast, resistin had no effect on endothelial P-selectin production. We conclude that resistin induces platelet activation by increasing P-selectin expression through the p38 MAPK-dependent pathway. These data provide one mechanism for the prothrombotic state in individuals with the metabolic syndrome.

  18. Inhibition of EGFR/MAPK signaling reduces microglial inflammatory response and the associated secondary damage in rats after spinal cord injury

    PubMed Central

    2012-01-01

    Background Emerging evidence indicates that reactive microglia-initiated inflammatory responses are responsible for secondary damage after primary traumatic spinal cord injury (SCI); epidermal growth factor receptor (EGFR) signaling may be involved in cell activation. In this report, we investigate the influence of EGFR signaling inhibition on microglia activation, proinflammatory cytokine production, and the neuronal microenvironment after SCI. Methods Lipopolysaccharide-treated primary microglia/BV2 line cells and SCI rats were used as model systems. Both C225 and AG1478 were used to inhibit EGFR signaling activation. Cell activation and EGFR phosphorylation were observed after fluorescent staining and western blot. Production of interleukin-1beta (IL-1β) and tumor necrosis factor alpha (TNFα) was tested by reverse transcription PCR and ELISA. Western blot was performed to semi-quantify the expression of EGFR/phospho-EGFR, and phosphorylation of Erk, JNK and p38 mitogen-activated protein kinases (MAPK). Wet-dry weight was compared to show tissue edema. Finally, axonal tracing and functional scoring were performed to show recovery of rats. Results EGFR phosphorylation was found to parallel microglia activation, while EGFR blockade inhibited activation-associated cell morphological changes and production of IL-1β and TNFα. EGFR blockade significantly downregulated the elevated MAPK activation after cell activation; selective MAPK inhibitors depressed production of cytokines to a certain degree, suggesting that MAPK mediates the depression of microglia activation brought about by EGFR inhibitors. Subsequently, seven-day continual infusion of C225 or AG1478 in rats: reduced the expression of phospho-EGFR, phosphorylation of Erk and p38 MAPK, and production of IL-1β and TNFα; lessened neuroinflammation-associated secondary damage, like microglia/astrocyte activation, tissue edema and glial scar/cavity formation; and enhanced axonal outgrowth and functional

  19. Daphnetin protects oxidative stress-induced neuronal apoptosis via regulation of MAPK signaling and HSP70 expression

    PubMed Central

    Qi, Zhilin; Qi, Shimei; Gui, Lin; Shen, Lei; Feng, Zunyong

    2016-01-01

    Neurodegenerative disorders are characterized by progressive degeneration and loss of neurons in the brain. Oxidative stress is implicated in the pathogenesis of neurological disorders, although the pathological mechanism remains unelucidated. Daphnetin, an active ingredient extracted from Changbai daphne (Daphne Korean Nakai), exhibits various pharmacological effects, including anti-inflammatory, anti-oxidative and anti-tumor effects. However, the neuroprotective effects, as well as the specific mechanisms of daphnetin, remain unclear. Neuronal-like rat pheochromocytoma PC12 cells were pretreated with daphnetin for 2 h, then treated with or without H2O2 for various times. Cell morphology was detected using an inverted microscope, the apoptotic ratio was determined by Annexin V fluorescein isothiocyanate/propidium iodide assay, nuclear morphology was observed and photographed using a fluorescence microscope following 4′,6-diamidino-2-phenylindole staining. The levels of pro-caspase 3, cleavage of poly ADP-ribose polymerase and caspase 3 were detected by western blotting. In addition, the activation of mitogen-activated protein kinase (MAPK) signal pathway and the expression of HSP70 were detected by western blotting. The present study demonstrated that daphnetin attenuated hydrogen peroxide (H2O2)-induced apoptosis in a concentration-dependent manner, reduced the cleavage of poly ADP ribose polymerase and caspase 3, and inhibited the phosphorylation of p38 MAPK and c-Jun N-terminal kinases (JNK) in H2O2-induced PC12 cells. In addition, daphnetin induced the expression of HSP70 in a dose- and time-dependent manner, and daphnetin-induced HSP70 expression was reduced by extracellular signal-regulated kinase (ERK) 1/2 inhibitor U0126 in PC12 cells. Therefore, the present results indicate that daphnetin protects PC12 cells against oxidative stress injury by regulating p38 MAPK and JNK signaling and increasing the expression of HSP70 via ERK signaling. This suggests

  20. Control of the segmentation process by graded MAPK/ERK activation in the chick embryo.

    PubMed

    Delfini, Marie-Claire; Dubrulle, Julien; Malapert, Pascale; Chal, Jérome; Pourquié, Olivier

    2005-08-01

    The regular spacing of somites during vertebrate embryogenesis involves a dynamic gradient of FGF signaling that controls the timing of maturation of cells in the presomitic mesoderm (PSM). How the FGF signal is transduced by PSM cells is unclear. Here, we first show that the FGF gradient is translated into graded activation of the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway along the PSM in the chicken embryo. Using in ovo electroporation of PSM cells, we demonstrate that constitutive activation of ERK signaling in the PSM blocks segmentation by preventing maturation of PSM cells, thus phenocopying the overexpression of FGF8. Conversely, inhibition of ERK phosphorylation mimics a loss of function of FGF signaling in the PSM. Interestingly, video microscopy analysis of cell movements shows that ERK regulates the motility of PSM cells, suggesting that the decrease of cell movements along the PSM enables mesenchymal PSM cells to undergo proper segmentation. Together, our data demonstrate that ERK is the effector of the gradient of FGF in the PSM that controls the segmentation process.

  1. Stress activated protein kinases, JNKs and p38 MAPK, are differentially activated in ganglia and heart of land snail Helix lucorum (L.) during seasonal hibernation and arousal.

    PubMed

    Michaelidis, Basile; Hatzikamari, Maria; Antoniou, Vassiliki; Anestis, Andreas; Lazou, Antigone

    2009-06-01

    The present work aimed to investigate the phosphorylation and hence activation of stress activated protein kinases, p38 MAPK and JNKs in the tissues of the snail Helix lucorum during seasonal hibernation. Snails were put in large glass boxes, which were placed outdoors so that they were exposed to natural conditions of light and temperature. Phosphorylation and hence activation of JNKs and p38 MAPK was determined in both heart and ganglia. Deep hibernation caused significant increases in the levels of the phosphorylated form of JNK and p38-MAPK in both heart and ganglia. Phosphorylation of JNK remained elevated in the ganglia or increased after a transient drop in the heart, when the snails were prepared for arousal. In addition, phosphorylation of p38-MAPK was further increased in the heart during this period. These data support the conclusion that MAPK signalling cascade might contribute in the physiological and biochemical remodelling in the tissues of land snails during hibernation and upon preparation for arousal.

  2. Inhibition of MAPK signaling pathways enhances cell death induced by 5-Aminolevulinic acid-photodynamic therapy in skin squamous carcinoma cells.

    PubMed

    Ge, Xinhong; Liu, Jianping; Shi, Zhiyun; Jing, Li; Yu, Nan; Zhang, Xiujuan; Jiao, Yaning; Wang, Yili; Li, P Andy

    2016-04-01

    Combination of a photosensitizer, 5-aminolevulinic acid (ALA), with photodynamic therapy (PDT) has been widely used to treat skin squamous cell carcinoma (SCC). However, a portion of SCC patients do not respond well to PDT. The molecular reason for this resistance is not clear. We hypothesize that mitogen-activated phosphorylation kinase (MAPK) plays a key role in mediating SCC resistance to PDT. To determine whether inhibition of MAPK signaling enhances the anti-tumor effect of ALA-PDT in SCC. The human squamous carcinoma cell line, SCL-1, was either untreated or treated with various combinations of ALA, PDT light source and inhibitors of MAPK signaling components. ALA-PDT treatment significantly decreased cell viability, increased the percentage of annexin-V positive cells and resulted in formation of apoptotic bodies. ALA-PDT treated cells showed increased levels of p-MEK, p-ERK1/2, p-p38, p-Elk-1, p-JNK and p-c-Jun. Addition of inhibitors for ERK1/2 (PD98059), p38 (SB203580) and JNK (SP60125) reversed the changes and led to a more dramatic decrease in SCL-1 cell viability than seen with ALA-PDT alone. Inhibition of the MAPK pathway enhances the cytotoxic effect of ALA-PDT on SCL-1.

  3. Inhibition of MAPK signaling pathways enhances cell death induced by 5-Aminolevulinic acid-photodynamic therapy in skin squamous carcinoma cells.

    PubMed

    Ge, Xinhong; Liu, Jianping; Shi, Zhiyun; Jing, Li; Yu, Nan; Zhang, Xiujuan; Jiao, Yaning; Wang, Yili; Li, P Andy

    2016-04-01

    Combination of a photosensitizer, 5-aminolevulinic acid (ALA), with photodynamic therapy (PDT) has been widely used to treat skin squamous cell carcinoma (SCC). However, a portion of SCC patients do not respond well to PDT. The molecular reason for this resistance is not clear. We hypothesize that mitogen-activated phosphorylation kinase (MAPK) plays a key role in mediating SCC resistance to PDT. To determine whether inhibition of MAPK signaling enhances the anti-tumor effect of ALA-PDT in SCC. The human squamous carcinoma cell line, SCL-1, was either untreated or treated with various combinations of ALA, PDT light source and inhibitors of MAPK signaling components. ALA-PDT treatment significantly decreased cell viability, increased the percentage of annexin-V positive cells and resulted in formation of apoptotic bodies. ALA-PDT treated cells showed increased levels of p-MEK, p-ERK1/2, p-p38, p-Elk-1, p-JNK and p-c-Jun. Addition of inhibitors for ERK1/2 (PD98059), p38 (SB203580) and JNK (SP60125) reversed the changes and led to a more dramatic decrease in SCL-1 cell viability than seen with ALA-PDT alone. Inhibition of the MAPK pathway enhances the cytotoxic effect of ALA-PDT on SCL-1. PMID:27032574

  4. Gastrointestinal Congestion Dilates the Hepatic Artery Through the P38 MAPK Signal Transduction Pathway During Liver Transplantation.

    PubMed

    Cao, Zhongping; Tang, Xiaowen; Hou, Shike

    2016-01-01

    During the neohepatic stage of liver transplantation, hemodynamics change markedly. The current study aimed to investigate whether gastrointestinal congestion caused by inferior vena cava and hepatic portal vein clamping can dilate the hepatic artery and to determine the associated mechanisms. Ring segments of the hepatic artery were treated with the plasma from gastrointestinal congestion or the superior vena cava. The fractions in gastrointestinal congestion and the superior vena cava plasma were tested, and the effect of these fractions on the tone of the hepatic artery ring was examined. Different signal transduction blockers and different inhibitors were then used to determine the exact signal transduction pathway involved. In addition, endothelial cell structure was observed by transmission electron microscopy after treatment with the gastrointestinal congestion plasma or the superior vena cava plasma. Gastrointestinal congestion plasma contained more inflammatory cytokines than superior vena cava plasma, and these cytokines could cause hepatic artery ring dilatation. A P38 mitogen-activated protein kinase (P38 MAPK) signal transduction pathway blocker and nitric oxide (NO), prostaglandin (PGI2), nuclear factor-κB (NF-κB), and adenosine triphosphate (ATP)-sensitive K (KATP) channel inhibitors were able to significantly reverse the ring tension caused by gastrointestinal congestion plasma. The normal endothelium was also injured by treatment with gastrointestinal congestion plasma. The inflammatory cytokines in gastrointestinal congestion can cause hepatic artery ring dilatation through the P38 MAPK signal transduction pathway, and this phenomenon is also associated with NO, PGI2, NF-κB, and the KATP channel. These inflammatory cytokines can injure endothelial cells in the hepatic artery. PMID:26955003

  5. Dynamic ubiquitination of the mitogen-activated protein kinase kinase (MAPKK) Ste7 determines mitogen-activated protein kinase (MAPK) specificity.

    PubMed

    Hurst, Jillian H; Dohlman, Henrik G

    2013-06-28

    Ubiquitination is a post-translational modification that tags proteins for proteasomal degradation. In addition, there is a growing appreciation that ubiquitination can influence protein activity and localization. Ste7 is a prototype MAPKK in yeast that participates in both the pheromone signaling and nutrient deprivation/invasive growth pathways. We have shown previously that Ste7 is ubiquitinated upon pheromone stimulation. Here, we show that the Skp1/Cullin/F-box ubiquitin ligase SCF(Cdc4) and the ubiquitin protease Ubp3 regulate Ste7 ubiquitination and signal specificity. Using purified components, we demonstrate that SCF(Cdc4) ubiquitinates Ste7 directly. Using gene deletion mutants, we show that SCF(Cdc4) and Ubp3 have opposing effects on Ste7 ubiquitination. Although SCF(Cdc4) is necessary for proper activation of the pheromone MAPK Fus3, Ubp3 is needed to limit activation of the invasive growth MAPK Kss1. Finally, we show that Fus3 phosphorylates Ubp3 directly and that phosphorylation of Ubp3 is necessary to limit Kss1 activation. These results reveal a feedback loop wherein one MAPK limits the ubiquitination of an upstream MAPKK and thereby prevents spurious activation of a second competing MAPK. PMID:23645675

  6. Activation of MAPK Is Required for ROS Generation and Exocytosis in HMC-1 Cells Induced by Trichomonas vaginalis-Derived Secretory Products

    PubMed Central

    Narantsogt, Giimaa; Min, Arim; Nam, Young Hee; Lee, Young Ah; Kim, Kyeong Ah; Agvaandaram, Gurbadam; Dorjsuren, Temuulen; El-Benna, Jamel; Shin, Myeong Heon

    2015-01-01

    Trichomonas vaginalis is a flagellated protozoan parasite that causes vaginitis and cervicitis in women and asymptomatic urethritis and prostatitis in men. Mast cells have been reported to be predominant in vaginal smears and vaginal walls of patients infected with T. vaginalis. Mitogen-activated protein kinase (MAPK), activated by various stimuli, have been shown to regulate the transcriptional activity of various cytokine genes in mast cells. In this study, we investigated whether MAPK is involved in ROS generation and exocytotic degranulation in HMC-1 cells induced by T. vaginalis-derived secretory products (TvSP). We found that TvSP induces the activation of MAPK and NADPH oxidase in HMC-1 cells. Stimulation with TvSP induced phosphorylation of MAPK and p47phox in HMC-1 cells. Stimulation with TvSP also induced up-regulation of CD63, a marker for exocytosis, along the surfaces of human mast cells. Pretreatment with MAPK inhibitors strongly inhibited TvSP-induced ROS generation and exocytotic degranulation. Finally, our results suggest that TvSP induces intracellular ROS generation and exocytotic degranulation in HMC-1 via MAPK signaling. PMID:26537039

  7. Activation of MAPK Is Required for ROS Generation and Exocytosis in HMC-1 Cells Induced by Trichomonas vaginalis-Derived Secretory Products.

    PubMed

    Narantsogt, Giimaa; Min, Arim; Nam, Young Hee; Lee, Young Ah; Kim, Kyeong Ah; Agvaandaram, Gurbadam; Dorjsuren, Temuulen; El-Benna, Jamel; Shin, Myeong Heon

    2015-10-01

    Trichomonas vaginalis is a flagellated protozoan parasite that causes vaginitis and cervicitis in women and asymptomatic urethritis and prostatitis in men. Mast cells have been reported to be predominant in vaginal smears and vaginal walls of patients infected with T. vaginalis. Mitogen-activated protein kinase (MAPK), activated by various stimuli, have been shown to regulate the transcriptional activity of various cytokine genes in mast cells. In this study, we investigated whether MAPK is involved in ROS generation and exocytotic degranulation in HMC-1 cells induced by T. vaginalis-derived secretory products (TvSP). We found that TvSP induces the activation of MAPK and NADPH oxidase in HMC-1 cells. Stimulation with TvSP induced phosphorylation of MAPK and p47(phox) in HMC-1 cells. Stimulation with TvSP also induced up-regulation of CD63, a marker for exocytosis, along the surfaces of human mast cells. Pretreatment with MAPK inhibitors strongly inhibited TvSP-induced ROS generation and exocytotic degranulation. Finally, our results suggest that TvSP induces intracellular ROS generation and exocytotic degranulation in HMC-1 via MAPK signaling.

  8. Infectious bursal disease virus infection induces macrophage activation via p38 MAPK and NF-kappaB pathways.

    PubMed

    Khatri, Mahesh; Sharma, Jagdev M

    2006-06-01

    In the present study, we show that infection with infectious bursal disease virus (IBDV) causes activation of macrophages, the key cells involved in inflammatory and immune-regulatory functions. Exposure of cultured spleen macrophages (SM) from SPF chickens to IBDV resulted in the production of nitric oxide (NO). In addition, there was upregulation of mRNA expression of inducible nitric oxide synthase (iNOS), IL-8 and cyclooxygenase-2 (COX-2). The signal transduction pathways involved in macrophage activation were examined. The role of mitogen-activated protein kinases (MAPKs) and nuclear factor-kappaB (NF-kappaB) was tested by using specific pharmacological inhibitors. Addition of p38 MAPK inhibitor, SB-203580 and NF-kappaB inhibitor Bay 11-7082, suppressed IBDV-induced NO production and mRNA expression of iNOS, IL-8 and COX-2. The results suggest that IBDV uses cellular signal transduction machinery, in particular the p38 MAPK and NF-kappaB pathways, to elicit macrophage activation. The increased production of NO, IL-8 and COX-2 by macrophages may contribute to bursa inflammatory responses commonly seen during the acute IBDV infection.

  9. Comparative analysis of MAPK and PI3K/AKT pathway activation and inhibition in human and canine melanoma.

    PubMed

    Fowles, J S; Denton, C L; Gustafson, D L

    2015-09-01

    The lack of advanced animal models of human cancers is considered a barrier to developing effective therapeutics. Canine and human melanomas are histologically disparate but show similar disease progression and response to therapies. The purpose of these studies was to compare human and canine melanoma tumours and cell lines regarding MAPK and PI3K/AKT signalling dysregulation, and response to select molecularly targeted agents. Pathway activation was investigated via microarray and mutational analysis. Growth inhibition and cell cycle effects were assessed for pathway inhibitors AZD6244 (MAPK) and rapamycin (PI3K/AKT) in human and canine melanoma cells. Human and canine melanoma share similar differential gene expression patterns within the MAPK and PI3K/AKT pathways. Constitutive pathway activation and similar sensitivity to AZD6244 and rapamycin was observed in human and canine cells. These results show that human and canine melanoma share activation and sensitivity to inhibition of cancer-related signalling pathways despite differences in activating mutations. PMID:23745794

  10. Endothelial lipase is upregulated by interleukin-6 partly via the p38 MAPK and p65 NF-κB signaling pathways

    PubMed Central

    Yue, Xin; Wu, Minghui; Jiang, Hong; Hao, Jing; Zhao, Qinghao; Zhu, Qing; Saren, Gaowa; Zhang, Yun; Zhang, Xiaoli

    2016-01-01

    To investigate the effects of inflammatory factor interleukin (IL)-6 on the expression of endothelial lipase (EL) and its potential signaling pathways in atherosclerosis, a primary culture of human umbilical vein endothelial cells (HUVECs) was established and treated as follows: i) Control group without any treatment; ii) recombinant human (rh)IL-6 treatment (10 ng/ml) for 0, 4, 8, 12 and 24 h; iii) p38 mitogen-activated protein kinases (MAPKs) inhibitor (SB203580, 10 µmol/l) pretreatment for 1 h prior to rhIL-6 (10 ng/ml) treatment; iv) nuclear factor (NF)-κB activation inhibitor (pyrrolidine dithiocarbamate, 10 mmol/l) pretreatment for 1 h prior to rhIL-6 (10 ng/ml) treatment. EL levels were detected by immunocytochemical staining and western blot analysis. Proliferation of HUVECs was detected by immunostaining of proliferating cell nuclear antigen (PCNA) and an MTT assay. p38 MAPK and NF-κB p65 levels were detected by western blotting. The results showed that rhIL-6 treatment increased EL expression and proliferation of HUVECs. NF-κB p65 and MAPK p38 protein levels also increased in a time-dependent manner in HUVECs after rhIL-6 treatment. NF-κB inhibitor and MAPK p38 inhibitor prevented the effects of rhIL-6 on EL expression. In conclusion, inflammatory factor IL-6 may participate in the pathogenesis of atherosclerosis by increasing EL expression and the proliferation of endothelial cells via the p38 MAPK and NF-κB signaling pathways. PMID:27430252

  11. Endothelial lipase is upregulated by interleukin-6 partly via the p38 MAPK and p65 NF-κB signaling pathways.

    PubMed

    Yue, Xin; Wu, Minghui; Jiang, Hong; Hao, Jing; Zhao, Qinghao; Zhu, Qing; Saren, Gaowa; Zhang, Yun; Zhang, Xiaoli

    2016-09-01

    To investigate the effects of inflammatory factor interleukin (IL)‑6 on the expression of endothelial lipase (EL) and its potential signaling pathways in atherosclerosis, a primary culture of human umbilical vein endothelial cells (HUVECs) was established and treated as follows: i) Control group without any treatment; ii) recombinant human (rh)IL‑6 treatment (10 ng/ml) for 0, 4, 8, 12 and 24 h; iii) p38 mitogen‑activated protein kinases (MAPKs) inhibitor (SB203580, 10 µmol/l) pretreatment for 1 h prior to rhIL‑6 (10 ng/ml) treatment; iv) nuclear factor (NF)‑κB activation inhibitor (pyrrolidine dithiocarbamate, 10 mmol/l) pretreatment for 1 h prior to rhIL‑6 (10 ng/ml) treatment. EL levels were detected by immunocytochemical staining and western blot analysis. Proliferation of HUVECs was detected by immunostaining of proliferating cell nuclear antigen (PCNA) and an MTT assay. p38 MAPK and NF‑κB p65 levels were detected by western blotting. The results showed that rhIL‑6 treatment increased EL expression and proliferation of HUVECs. NF‑κB p65 and MAPK p38 protein levels also increased in a time‑dependent manner in HUVECs after rhIL‑6 treatment. NF‑κB inhibitor and MAPK p38 inhibitor prevented the effects of rhIL‑6 on EL expression. In conclusion, inflammatory factor IL‑6 may participate in the pathogenesis of atherosclerosis by increasing EL expression and the proliferation of endothelial cells via the p38 MAPK and NF-κB signaling pathways. PMID:27430252

  12. Neospora caninum Activates p38 MAPK as an Evasion Mechanism against Innate Immunity

    PubMed Central

    Mota, Caroline M.; Oliveira, Ana C. M.; Davoli-Ferreira, Marcela; Silva, Murilo V.; Santiago, Fernanda M.; Nadipuram, Santhosh M.; Vashisht, Ajay A.; Wohlschlegel, James A.; Bradley, Peter J.; Silva, João S.; Mineo, José R.; Mineo, Tiago W. P.

    2016-01-01

    Due to the high prevalence and economic impact of neosporosis, the development of safe and effective vaccines and therapies against this parasite has been a priority in the field and is crucial to limit horizontal and vertical transmission in natural hosts. Limited data is available regarding factors that regulate the immune response against this parasite and such knowledge is essential in order to understand Neospora caninum induced pathogenesis. Mitogen-activated protein kinases (MAPKs) govern diverse cellular processes, including growth, differentiation, apoptosis, and immune-mediated responses. In that sense, our goal was to understand the role of MAPKs during the infection by N. caninum. We found that p38 phosphorylation was quickly triggered in macrophages stimulated by live tachyzoites and antigen extracts, while its chemical inhibition resulted in upregulation of IL-12p40 production and augmented B7/MHC expression. In vivo blockade of p38 resulted in an amplified production of cytokines, which preceded a reduction in latent parasite burden and enhanced survival against the infection. Additionally, the experiments indicate that the p38 activation is induced by a mechanism that depends on GPCR, PI3K and AKT signaling pathways, and that the phenomena here observed is distinct that those induced by Toxoplasma gondii’s GRA24 protein. Altogether, these results showed that N. caninum manipulates p38 phosphorylation in its favor, in order to downregulate the host’s innate immune responses. Additionally, those results infer that active interference in this signaling pathway may be useful for the development of a new therapeutic strategy against neosporosis. PMID:27679624

  13. Neospora caninum Activates p38 MAPK as an Evasion Mechanism against Innate Immunity

    PubMed Central

    Mota, Caroline M.; Oliveira, Ana C. M.; Davoli-Ferreira, Marcela; Silva, Murilo V.; Santiago, Fernanda M.; Nadipuram, Santhosh M.; Vashisht, Ajay A.; Wohlschlegel, James A.; Bradley, Peter J.; Silva, João S.; Mineo, José R.; Mineo, Tiago W. P.

    2016-01-01

    Due to the high prevalence and economic impact of neosporosis, the development of safe and effective vaccines and therapies against this parasite has been a priority in the field and is crucial to limit horizontal and vertical transmission in natural hosts. Limited data is available regarding factors that regulate the immune response against this parasite and such knowledge is essential in order to understand Neospora caninum induced pathogenesis. Mitogen-activated protein kinases (MAPKs) govern diverse cellular processes, including growth, differentiation, apoptosis, and immune-mediated responses. In that sense, our goal was to understand the role of MAPKs during the infection by N. caninum. We found that p38 phosphorylation was quickly triggered in macrophages stimulated by live tachyzoites and antigen extracts, while its chemical inhibition resulted in upregulation of IL-12p40 production and augmented B7/MHC expression. In vivo blockade of p38 resulted in an amplified production of cytokines, which preceded a reduction in latent parasite burden and enhanced survival against the infection. Additionally, the experiments indicate that the p38 activation is induced by a mechanism that depends on GPCR, PI3K and AKT signaling pathways, and that the phenomena here observed is distinct that those induced by Toxoplasma gondii’s GRA24 protein. Altogether, these results showed that N. caninum manipulates p38 phosphorylation in its favor, in order to downregulate the host’s innate immune responses. Additionally, those results infer that active interference in this signaling pathway may be useful for the development of a new therapeutic strategy against neosporosis.

  14. Neospora caninum Activates p38 MAPK as an Evasion Mechanism against Innate Immunity.

    PubMed

    Mota, Caroline M; Oliveira, Ana C M; Davoli-Ferreira, Marcela; Silva, Murilo V; Santiago, Fernanda M; Nadipuram, Santhosh M; Vashisht, Ajay A; Wohlschlegel, James A; Bradley, Peter J; Silva, João S; Mineo, José R; Mineo, Tiago W P

    2016-01-01

    Due to the high prevalence and economic impact of neosporosis, the development of safe and effective vaccines and therapies against this parasite has been a priority in the field and is crucial to limit horizontal and vertical transmission in natural hosts. Limited data is available regarding factors that regulate the immune response against this parasite and such knowledge is essential in order to understand Neospora caninum induced pathogenesis. Mitogen-activated protein kinases (MAPKs) govern diverse cellular processes, including growth, differentiation, apoptosis, and immune-mediated responses. In that sense, our goal was to understand the role of MAPKs during the infection by N. caninum. We found that p38 phosphorylation was quickly triggered in macrophages stimulated by live tachyzoites and antigen extracts, while its chemical inhibition resulted in upregulation of IL-12p40 production and augmented B7/MHC expression. In vivo blockade of p38 resulted in an amplified production of cytokines, which preceded a reduction in latent parasite burden and enhanced survival against the infection. Additionally, the experiments indicate that the p38 activation is induced by a mechanism that depends on GPCR, PI3K and AKT signaling pathways, and that the phenomena here observed is distinct that those induced by Toxoplasma gondii's GRA24 protein. Altogether, these results showed that N. caninum manipulates p38 phosphorylation in its favor, in order to downregulate the host's innate immune responses. Additionally, those results infer that active interference in this signaling pathway may be useful for the development of a new therapeutic strategy against neosporosis. PMID:27679624

  15. Maternal Inheritance of Twist and Analysis of MAPK Activation in Embryos of the Polychaete Annelid Platynereis dumerilii

    PubMed Central

    Pfeifer, Kathrin; Schaub, Christoph; Domsch, Katrin; Dorresteijn, Adriaan; Wolfstetter, Georg

    2014-01-01

    In this study, we aimed to identify molecular mechanisms involved in the specification of the 4d (mesentoblast) lineage in Platynereis dumerilii. We employ RT-PCR and in situ hybridization against the Platynereis dumerilii twist homolog (Pdu-twist) to reveal mesodermal specification within this lineage. We show that Pdu-twist mRNA is already maternally distributed. After fertilization, ooplasmatic segregation leads to relocation of Pdu-twist transcripts into the somatoblast (2d) lineage and 4d, indicating that the maternal component of Pdu-twist might be an important prerequisite for further mesoderm specification but does not represent a defining characteristic of the mesentoblast. However, after the primordial germ cells have separated from the 4d lineage, zygotic transcription of Pdu-twist is exclusively observed in the myogenic progenitors, suggesting that mesodermal specification occurs after the 4d stage. Previous studies on spiral cleaving embryos revealed a spatio-temporal correlation between the 4d lineage and the activity of an embryonic organizer that is capable to induce the developmental fates of certain micromeres. This has raised the question if specification of the 4d lineage could be connected to the organizer activity. Therefore, we aimed to reveal the existence of such a proposed conserved organizer in Platynereis employing antibody staining against dpERK. In contrast to former observations in other spiralian embryos, activation of MAPK signaling during 2d and 4d formation cannot be detected which questions the existence of a conserved connection between organizer function and specification of the 4d lineage. However, our experiments unveil robust MAPK activation in the prospective nephroblasts as well as in the macromeres and some micromeres at the blastopore in gastrulating embryos. Inhibition of MAPK activation leads to larvae with a shortened body axis, defects in trunk muscle spreading and improper nervous system condensation, indicating a

  16. Frankincense and myrrh suppress inflammation via regulation of the metabolic profiling and the MAPK signaling pathway

    PubMed Central

    Su, Shulan; Duan, Jinao; Chen, Ting; Huang, Xiaochen; Shang, Erxin; Yu, Li; Wei, Kaifeng; Zhu, Yue; Guo, Jianming; Guo, Sheng; Liu, Pei; Qian, Dawei; Tang, Yuping

    2015-01-01

    Frankincense and myrrh are highly effective in treatment of inflammatary diseases, but lacking of the therapy mechanisms. We undertook this stuty to evaluate the effects on Adjuvant-induced Arthritis (AIA) rats and to explore the underlying mechanisms by analyzing the metabolic profiling and signaling pathway evaluated by expression of inflammatory cytokines, c-jun and c-fos and corresponding phosphorylation levels. The results stated the elevated expression levels of TNFα, PGE2, IL-2, NO, and MDA in serum and swelling paw of AIA rats were significantly decreased after treatment, which exerted more remarkable inhibitive effects of combined therapy. The metbolic profiling of plasma and urine were clearly improved and twenty-one potential biomarkers were identified. Moreover, the inhibited effects of five bioactive components on cytokine transcription in PHA stimulated-PBMC showed the MAPK pathway might account for this phenomenon with considerable reduction in phosphorylated forms of all the three MAPK (ERK1/2, p38 and JNK) and down regulation of c-jun and c-fos. PMID:26329643

  17. [RAS/MAPK signal transduction pathway and its role in the pathogenesis of Noonan syndrome].

    PubMed

    Gos, Monika; Leszkiewicz, Monika; Abramowicz, Anna

    2012-01-01

    Noonan syndrome (NS) is one of the most frequent dysmorphic syndromes in children with a frequency of 1/1000-1/2500 of newborns. Noonan syndrome is a multi-organ disease with a broad spectrum of clinical symptoms. The most characteristic features of NS are: craniofacial dysmorphy, short stature, cardiovascular defects, bone and skeletal defects and delayed puberty (cryptorchidism in males). Noonan syndrome has a genetic background and is inherited in autosomal dominant manner. The recent studies have shown that it is due to the presence of mutation in one of the genes encoding proteins of RAS/MAPK signalling pathway responsible for cell proliferation and differentiation. Till now, NS causing mutations were identified in PTPN11, SOS1, RAF1, KRAS, BRAF, SHOC2 and NRAS genes, and this may partially explain the broad phenotypic spectrum observed in patients. Noonan syndrome is one of the RAS-opathies, therefore the molecular analysis of RAS/ MAPK genes might be a very useful tool in clinical differentiation of the disease.

  18. Mitogen-Activated Protein Kinase Signaling in Plant-Interacting Fungi: Distinct Messages from Conserved Messengers[W

    PubMed Central

    Hamel, Louis-Philippe; Nicole, Marie-Claude; Duplessis, Sébastien; Ellis, Brian E.

    2012-01-01

    Mitogen-activated protein kinases (MAPKs) are evolutionarily conserved proteins that function as key signal transduction components in fungi, plants, and mammals. During interaction between phytopathogenic fungi and plants, fungal MAPKs help to promote mechanical and/or enzymatic penetration of host tissues, while plant MAPKs are required for activation of plant immunity. However, new insights suggest that MAPK cascades in both organisms do not operate independently but that they mutually contribute to a highly interconnected molecular dialogue between the plant and the fungus. As a result, some pathogenesis-related processes controlled by fungal MAPKs lead to the activation of plant signaling, including the recruitment of plant MAPK cascades. Conversely, plant MAPKs promote defense mechanisms that threaten the survival of fungal cells, leading to a stress response mediated in part by fungal MAPK cascades. In this review, we make use of the genomic data available following completion of whole-genome sequencing projects to analyze the structure of MAPK protein families in 24 fungal taxa, including both plant pathogens and mycorrhizal symbionts. Based on conserved patterns of sequence diversification, we also propose the adoption of a unified fungal MAPK nomenclature derived from that established for the model species Saccharomyces cerevisiae. Finally, we summarize current knowledge of the functions of MAPK cascades in phytopathogenic fungi and highlight the central role played by MAPK signaling during the molecular dialogue between plants and invading fungal pathogens. PMID:22517321

  19. Heat stress prevents lipopolysaccharide-induced apoptosis in pulmonary microvascular endothelial cells by blocking calpain/p38 MAPK signalling

    PubMed Central

    Liu, Zhi-feng; Zheng, Dong; Fan, Guo-chang; Peng, Tianqing; Su, Lei

    2016-01-01

    Pulmonary microvascular endothelial cells (PMECs) injury including apoptosis plays an important role in the pathogenesis of acute lung injury during sepsis. Our recent study has demonstrated that calpain activation contributes to apoptosis in PMECs under septic conditions. This study investigated how calpain activation mediated apoptosis and whether heat stress regulated calpain activation in lipopolysaccharides (LPS)-stimulated PMECs. In cultured mouse primary PMECs, incubation with LPS (1 µg/ml, 24 h) increased active caspase-3 fragments and DNA fragmentation, indicative of apoptosis. These effects of LPS were abrogated by pre-treatment with heat stress (43 °C for 2 h). LPS also induced calpain activation and increased phosphorylation of p38 MAPK. Inhibition of calpain and p38 MAPK prevented apoptosis induced by LPS. Furthermore, inhibition of calpain blocked p38 MAPK phosphorylation in LPS-stimulated PMECs. Notably, heat stress decreased the protein levels of calpain-1/2 and calpain activities, and blocked p38 MAPK phosphorylation in response to LPS. Additionally, forced up-regulation of calpain-1 or calpain-2 sufficiently induced p38 MAPK phosphorylation and apoptosis in PMECs, both of which were inhibited by heat stress. In conclusion, heat stress prevents LPS-induced apoptosis in PMECs. This effect of heat stress is associated with down-regulation of calpain expression and activation, and subsequent blockage of p38 MAPK activation in response to LPS. Thus, blocking calpain/p38 MAPK pathway may be a novel mechanism underlying heat stress-mediated inhibition of apoptosis in LPS-stimulated endothelial cells. PMID:27325431

  20. Pioglitazone Protected against Cardiac Hypertrophy via Inhibiting AKT/GSK3β and MAPK Signaling Pathways

    PubMed Central

    Wei, Wen-Ying; Ma, Zhen-Guo; Xu, Si-Chi; Zhang, Ning; Tang, Qi-Zhu

    2016-01-01

    Peroxisome proliferator activated receptor γ (PPARγ) has been closely involved in the process of cardiovascular diseases. This study was to investigate whether pioglitazone (PIO), a PPARγ agonist, could protect against pressure overload-induced cardiac hypertrophy. Mice were orally given PIO (2.5 mg/kg) from 1 week after aortic banding and continuing for 7 weeks. The morphological examination and biochemical analysis were used to evaluate the effects of PIO. Neonatal rat ventricular cardiomyocytes were also used to verify the protection of PIO against hypertrophy in vitro. The results in our study demonstrated that PIO remarkably inhibited hypertrophic response induced by aortic banding in vivo. Besides, PIO also suppressed cardiac fibrosis in vivo. PIO treatment also inhibited the activation of protein kinase B (AKT)/glycogen synthase kinase-3β (GSK3β) and mitogen-activated protein kinase (MAPK) in the heart. In addition, PIO alleviated angiotensin II-induced hypertrophic response in vitro. In conclusion, PIO could inhibit cardiac hypertrophy via attenuation of AKT/GSK3β and MAPK pathways. PMID:27110236

  1. SIRT1 protects rat lung tissue against severe burn-induced remote ALI by attenuating the apoptosis of PMVECs via p38 MAPK signaling

    PubMed Central

    Bai, Xiaozhi; Fan, Lei; He, Ting; Jia, Wenbin; Yang, Longlong; Zhang, Jun; Liu, Yang; Shi, Jihong; Su, Linlin; Hu, Dahai

    2015-01-01

    Silent information regulator type-1 (SIRT1) has been reported to be involved in the cardiopulmonary protection. However, its role in the pathogenesis of burn-induced remote acute lung injury (ALI) is currently unknown. The present study aims to investigate the role of SIRT1 in burn-induced remote ALI and the involved signaling pathway. We observed that SIRT1 expression in rat lung tissue after burn injury appeared an increasing trend after a short period of suppression. The upregulation of SIRT1 stimulated by resveratrol exhibited remission of histopathologic changes, reduction of cell apoptosis, and downregulation of pro-inflammatory cytokines in rat pulmonary tissues suffering from severe burn. We next used primary pulmonary microvascular endothelial cells (PMVECs) challenged by burn serum (BS) to simulate in vivo rat lung tissue after burn injury, and found that BS significantly suppressed SIRT1 expression, increased cell apoptosis, and activated p38 MAPK signaling. The use of resveratrol reversed these effects, while knockdown of SIRT1 by shRNA further augmented BS-induced increase of cell apoptosis and activation of p38 MAPK. Taken together, these results indicate that SIRT1 might protect lung tissue against burn-induced remote ALI by attenuating PMVEC apoptosis via p38 MAPK signaling, suggesting its potential therapeutic effects on the treatment of ALI. PMID:25992481

  2. Inhibitory effects of enalaprilat on rat cardiac fibroblast proliferation via ROS/P38MAPK/TGF-β1 signaling pathway.

    PubMed

    Yu, Min; Zheng, Yang; Sun, Hong-Xia; Yu, Du-Juan

    2012-01-01

    Enalaprilat (Ena.), an angiotensin II (Ang II) converting enzyme inhibitor (ACEI), can produce some therapeutic effects on hypertension, ventricular hypertrophy and myocardial remodeling in clinic, but its precise mechanism, especially its signaling pathways remain elusive. In this study, cardiac fibroblasts (CFb) was isolated by the trypsin digestion method; a BrdU proliferation assay was adopted to determine cell proliferation; an immunofluorescence assay was used to measure intracellular reactive oxygen species (ROS); immunocytochemistry staining and Western blotting assay were used to detect phosphorylated p38 mitogen activated protein kinase (p-p38MAPK) and transforming growth factor-β(1) (TGF-β(1)) protein expression, respectively. The results showed that Ang II (10(-7) M) stimulated the cardiac fibroblast proliferation which was inhibited by NAC (an antioxidant), SB203580 (a p38MAPK inhibitor) or enalaprilat; Ang II caused an burst of intracellular ROS level within thirty minutes, an increase in p-p38MAPK (3.6-fold of that in the control group), as well as an elevation of TGF-β(1) meantime; NAC, an antioxidant, and enalaprilat treatment attenuated cardiac fibroblast proliferation induced by Ang II and decreased ROS and p-p38MAPK protein levels in rat cardiac fibroblast; SB203580 lowered TGF-β(1) protein expression in rats' CFb in a dose-dependent manner. It could be concluded that enalaprilat can inhibit the cardiac fibroblast proliferation induced by Ang II via blocking ROS/P38MAPK/TGF-β(1) signaling pathways and the study provides a theoretical proof for the application of ACEIs in treating myocardial fibrosis and discovering the primary mechanism through which ACEIs inhibit CFb proliferation. PMID:22395404

  3. Molecular Changes Involving MEK3-p38 MAPK Activation in Chronic Masticatory Myalgia.

    PubMed

    Meng, H; Gao, Y; Kang, Y F; Zhao, Y P; Yang, G J; Wang, Y; Cao, Y; Gan, Y H; Xie, Q F

    2016-09-01

    The exact mechanism underlying chronic masticatory myalgia (CMM), a conspicuous symptom in temporomandibular disorders, remains unclear. This investigation compared gene expression profiles between CMM patients and healthy subjects. Peripheral blood leukocytes were collected in 8 cases and 8 controls and subjected to whole genome microarray analyses. Data were analyzed with Gene Ontology and interactive pathways analyses. According to Gene Ontology analysis, categories such as ion transport, response to stimuli, and metabolic process were upregulated. The pathway analysis suggested overexpression of the mitogen-activated protein kinase (MAPK) pathway in CMM patients and to a higher degree in a pathway network. Overexpression of representative members of the MAPK pathway-including MAPK kinase 3 (MEK3), calcium voltage-gated channel auxiliary subunit gamma 2 (CACNG2), and growth arrest and DNA damage-inducible gamma (GADD45G)-was validated with real-time polymerase chain reaction. The upregulation of MEK3 was negatively correlated with the age of the CMM group. In the next step, the authors focused on MEK3, the gene that exhibited the greatest degree of differential expression, and its downstream target protein p38 MAPK. The results revealed upregulation of MEK3, as well as phosphorylated MEK3 and phosphorylated p38 MAPK, in CMM patients. These results provide a "fingerprint" for mechanistic studies of CMM in the future and highlight the importance of MEK3-p38 MAPK activation in CMM. PMID:27418173

  4. Activation of neutral-sphingomyelinase, MAPKs, and p75 NTR-mediating caffeic acid phenethyl ester–induced apoptosis in C6 glioma cells

    PubMed Central

    2014-01-01

    Background Caffeic acid phenethyl ester (CAPE), a component of propolis, is reported to possess anti-inflammatory, anti-bacterial, anti-viral, and anti-tumor activities. Previously, our laboratory demonstrated the in vitro and in vivo bioactivity of CAPE and addressed the role of p53 and the p38 mitogen-activated protein kinase (MAPK) pathway in regulating CAPE-induced apoptosis in C6 glioma cells. Results C6 cancer cell lines were exposed to doses of CAPE; DNA fragmentation and MAPKs and NGF/P75NTR levels were then determined. SMase activity and ceramide content measurement as well as western blotting analyses were performed to clarify molecular changes. The present study showed that CAPE activated neutral sphingomyelinase (N-SMase), which led to the ceramide-mediated activation of MAPKs, including extracellular signal-regulated kinase (ERK), Jun N-terminus kinase (JNK), and p38 MAPK. In addition, CAPE increased the expression of nerve growth factor (NGF) and p75 neurotrophin receptor (p75NTR). The addition of an N-SMase inhibitor, GW4869, established that NGF/p75NTR was the downstream target of N-SMase/ceramide. Pretreatment with MAPK inhibitors demonstrated that MEK/ERK and JNK acted upstream and downstream, respectively, of NGF/p75NTR. Additionally, CAPE-induced caspase 3 activation and poly [ADP-ribose] polymerase cleavage were reduced by pretreatment with MAPK inhibitors, a p75NTR peptide antagonist, or GW4869. Conclusions Taken together, N-SMase activation played a pivotal role in CAPE-induced apoptosis by activation of the p38 MAPK pathway and NGF/p75NTR may explain a new role of CAPE induced apoptosis in C6 glioma. PMID:24997497

  5. MAPK pathway activation by chronic lead-exposure increases vascular reactivity through oxidative stress/cyclooxygenase-2-dependent pathways

    SciTech Connect

    Simões, Maylla Ronacher; Aguado, Andrea; Fiorim, Jonaína; Silveira, Edna Aparecida; Azevedo, Bruna Fernandes; Toscano, Cindy Medice; Zhenyukh, Olha; Briones, Ana María; Alonso, María Jesús; Vassallo, Dalton Valentim; Salaices, Mercedes

    2015-03-01

    Chronic exposure to low lead concentration produces hypertension; however, the underlying mechanisms remain unclear. We analyzed the role of oxidative stress, cyclooxygenase-2-dependent pathways and MAPK in the vascular alterations induced by chronic lead exposure. Aortas from lead-treated Wistar rats (1st dose: 10 μg/100 g; subsequent doses: 0.125 μg/100 g, intramuscular, 30 days) and cultured aortic vascular smooth muscle cells (VSMCs) from Sprague Dawley rats stimulated with lead (20 μg/dL) were used. Lead blood levels of treated rats attained 21.7 ± 2.38 μg/dL. Lead exposure increased systolic blood pressure and aortic ring contractile response to phenylephrine, reduced acetylcholine-induced relaxation and did not affect sodium nitroprusside relaxation. Endothelium removal and L-NAME left-shifted the response to phenylephrine more in untreated than in lead-treated rats. Apocynin and indomethacin decreased more the response to phenylephrine in treated than in untreated rats. Aortic protein expression of gp91(phox), Cu/Zn-SOD, Mn-SOD and COX-2 increased after lead exposure. In cultured VSMCs lead 1) increased superoxide anion production, NADPH oxidase activity and gene and/or protein levels of NOX-1, NOX-4, Mn-SOD, EC-SOD and COX-2 and 2) activated ERK1/2 and p38 MAPK. Both antioxidants and COX-2 inhibitors normalized superoxide anion production, NADPH oxidase activity and mRNA levels of NOX-1, NOX-4 and COX-2. Blockade of the ERK1/2 and p38 signaling pathways abolished lead-induced NOX-1, NOX-4 and COX-2 expression. Results show that lead activation of the MAPK signaling pathways activates inflammatory proteins such as NADPH oxidase and COX-2, suggesting a reciprocal interplay and contribution to vascular dysfunction as an underlying mechanisms for lead-induced hypertension. - Highlights: • Lead-exposure increases oxidative stress, COX-2 expression and vascular reactivity. • Lead exposure activates MAPK signaling pathway. • ROS and COX-2 activation by

  6. Nitric oxide induces prion protein via MEK and p38 MAPK signaling.

    PubMed

    Wang, Vinchi; Chuang, Tzu-Chao; Hsu, Yaw-Don; Chou, Wei-Yuan; Kao, Ming-Ching

    2005-07-22

    The prion diseases or transmissible spongiform encephalopathy, such as human Creutzfeldt-Jakob disease (CJD) and so-called mad cow disease, are attributed to the causative agent, the scrapie variant of prion protein (PrP(Sc)) which causes fatal neurodegeneration. To investigate if stresses such as nitric oxide (NO) induced the cellular isoform of prion protein (PrP(C)), lipopolysaccharide, and sodium nitroprusside were used to treat N2a and NT2 cells, which resulted in elevated levels of the PRNP mRNA and prion protein. The signaling pathway for the NO-induced PrP(C) production involved guanylyl cyclase, MEK, and p38 MAPK as shown by the effect of specific pharmacological inhibitors ODQ, PD98059, and SB203580, respectively. Knowing the PrP induction by the biologically existing stimulus, this study provides useful information about the possible cellular mechanism and strategies for the treatment of CJD. PMID:15936714

  7. Distinct docking mechanisms mediate interactions between the Msg5 phosphatase and mating or cell integrity mitogen-activated protein kinases (MAPKs) in Saccharomyces cerevisiae.

    PubMed

    Palacios, Lorena; Dickinson, Robin J; Sacristán-Reviriego, Almudena; Didmon, Mark P; Marín, María José; Martín, Humberto; Keyse, Stephen M; Molina, María

    2011-12-01

    MAPK phosphatases (MKPs) are negative regulators of signaling pathways with distinct MAPK substrate specificities. For example, the yeast dual specificity phosphatase Msg5 dephosphorylates the Fus3 and Slt2 MAPKs operating in the mating and cell wall integrity pathways, respectively. Like other MAPK-interacting proteins, most MKPs bind MAPKs through specific docking domains. These include D-motifs, which contain basic residues that interact with acidic residues in the common docking (CD) domain of MAPKs. Here we show that Msg5 interacts not only with Fus3, Kss1, and Slt2 but also with the pseudokinase Slt2 paralog Mlp1. Using yeast two-hybrid and in vitro interaction assays, we have identified distinct regions within the N-terminal domain of Msg5 that differentially bind either the MAPKs Fus3 and Kss1 or Slt2 and Mlp1. Whereas a canonical D-site within Msg5 mediates interaction with the CD domains of Fus3 and Kss1, a novel motif ((102)IYT(104)) within Msg5 is involved in binding to Slt2 and Mlp1. Furthermore, mutation of this site prevents the phosphorylation of Msg5 by Slt2. This motif is conserved in Sdp1, another MKP that dephosphorylates Slt2, as well as in Msg5 orthologs from other yeast species. A region spanning amino acids 274-373 within Slt2 and Mlp1 mediates binding to this Msg5 motif in a CD domain-independent manner. In contrast, Slt2 uses its CD domain to bind to its upstream activator Mkk1. This binding flexibility may allow MAPK pathways to exploit additional regulatory controls in order to provide fine modulation of both pathway activity and specificity. PMID:22006927

  8. Dexamethasone in the presence of desipramine enhances MAPK/ERK1/2 signaling possibly via its interference with β-arrestin.

    PubMed

    Lucki, Anat; Klein, Ehud; Karry, Rachel; Ben-Shachar, Dorit

    2014-01-01

    Antidepressant medication is the standard treatment for major depression disorder (MDD). However, the response to these treatments is often incomplete and many patients remain refractory. In the present study, we show that the glucocorticoid receptor (GR) agonist dexamethasone (DEX) increased MAPK/ERK1/2 signaling in the presence of the noradrenergic antidepressant, desipramine (DMI), while no such effect was induced by DEX or DMI alone in human neuroblastoma SH-SY5Y cells. This enhancement was dependent on the activation of both α(2) adrenergic receptors (AR) and GR. The timing of MAPK/ERK1/2 activation as well as DEX-induced reduction in membranous α(2) AR suggests the involvement of a β-arrestin-dependent mechanism. In line with the latter, DEX increased cytosolic and decreased membranous levels of β-arrestin. Concomitantly, DEX induced a time-dependent increase in cytosolic α(2) AR-β-arrestin interaction and a decrease in β-arrestin interaction with Mdm2 E3 ubiquitin ligase. All of these effects of DEX were prevented by the GR antagonist RU486. Our data suggest an additional intracellular role for DEX, in which activation of GR interferes with the trafficking and degradation of β-arrestin-α2c-AR complex. We suggest that such an interaction in the presence of DMI can enhance MAPK/ERK1/2 signaling, a key player in neural plasticity and neurogenesis processes, which is impaired in MDD, while stimulated by antidepressants.

  9. Differential Effects of E2 on MAPK Activity in the Brain and Heart of Aged Female Rats

    PubMed Central

    Shults, Cody L.; Rao, Yathindar S.; Pak, Toni R.

    2016-01-01

    Aging and the coincident loss of circulating estrogens at menopause lead to increased risks for neurological and cardiovascular pathologies. Clinical studies show that estrogen therapy (ET) can be beneficial in mitigating these negative effects, in both the brain and heart, when it is initiated shortly after the perimenopausal transition. However, this same therapy is detrimental when initiated >10 years postmenopause. Importantly, the molecular mechanisms underlying this age-related switch in ET efficacy are unknown. Estrogen receptors (ERs) mediate the neuroprotective and cardioprotective functions of estrogens by modulating gene transcription or, non-genomically, by activating second messenger signaling pathways, such as mitogen activated protein kinases (MAPK). These kinases are critical regulators of cell signaling pathways and have widespread downstream effects. Our hypothesis is that age and estrogen deprivation following menopause alters the expression and activation of the MAPK family members p38 and ERK in the brain and heart. To test this hypothesis, we used a surgically induced model of menopause in 18 month old rats through bilateral ovariectomy (OVX) followed by an acute dose of 17β-estradiol (E2) administered at varying time points post-OVX (1 week, 4 weeks, 8 weeks, or 12 weeks). Age and E2 treatment differentially regulated kinase activity in both the brain and heart, and the effects were also brain region specific. MAPK signaling plays an integral role in aging, and the aberrant regulation of those signaling pathways might be involved in age-related disorders. Clinical studies show benefits of ET during early menopause but detrimental effects later, which might be reflective of changes in kinase expression and activation status. PMID:27487271

  10. Differential Effects of E2 on MAPK Activity in the Brain and Heart of Aged Female Rats.

    PubMed

    Pinceti, Elena; Shults, Cody L; Rao, Yathindar S; Pak, Toni R

    2016-01-01

    Aging and the coincident loss of circulating estrogens at menopause lead to increased risks for neurological and cardiovascular pathologies. Clinical studies show that estrogen therapy (ET) can be beneficial in mitigating these negative effects, in both the brain and heart, when it is initiated shortly after the perimenopausal transition. However, this same therapy is detrimental when initiated >10 years postmenopause. Importantly, the molecular mechanisms underlying this age-related switch in ET efficacy are unknown. Estrogen receptors (ERs) mediate the neuroprotective and cardioprotective functions of estrogens by modulating gene transcription or, non-genomically, by activating second messenger signaling pathways, such as mitogen activated protein kinases (MAPK). These kinases are critical regulators of cell signaling pathways and have widespread downstream effects. Our hypothesis is that age and estrogen deprivation following menopause alters the expression and activation of the MAPK family members p38 and ERK in the brain and heart. To test this hypothesis, we used a surgically induced model of menopause in 18 month old rats through bilateral ovariectomy (OVX) followed by an acute dose of 17β-estradiol (E2) administered at varying time points post-OVX (1 week, 4 weeks, 8 weeks, or 12 weeks). Age and E2 treatment differentially regulated kinase activity in both the brain and heart, and the effects were also brain region specific. MAPK signaling plays an integral role in aging, and the aberrant regulation of those signaling pathways might be involved in age-related disorders. Clinical studies show benefits of ET during early menopause but detrimental effects later, which might be reflective of changes in kinase expression and activation status. PMID:27487271

  11. Effects of Maternal Chromium Restriction on the Long-Term Programming in MAPK Signaling Pathway of Lipid Metabolism in Mice.

    PubMed

    Zhang, Qian; Sun, Xiaofang; Xiao, Xinhua; Zheng, Jia; Li, Ming; Yu, Miao; Ping, Fan; Wang, Zhixin; Qi, Cuijuan; Wang, Tong; Wang, Xiaojing

    2016-01-01

    It is now broadly accepted that the nutritional environment in early life is a key factor in susceptibility to metabolic diseases. In this study, we evaluated the effects of maternal chromium restriction in vivo on the modulation of lipid metabolism and the mechanisms involved in this process. Sixteen pregnant C57BL mice were randomly divided into two dietary treatments: a control (C) diet group and a low chromium (L) diet group. The diet treatment was maintained through gestation and lactation period. After weaning, some of the pups continued with either the control diet or low chromium diet (CC or LL), whereas other pups switched to another diet (CL or LC). At 32 weeks of age, serum lipid metabolism, proinflammatory indexes, oxidative stress and anti-oxidant markers, and DNA methylation status in adipose tissue were measured. The results indicated that the maternal low chromium diet increased body weight, fat pad weight, serum triglyceride (TG), low-density lipoprotein cholesterol (LDL), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), and oxidized glutathione (GSSG). There was a decrease in serum reduced/oxidized glutathione (GSH/GSSG) ratio at 32 weeks of age in female offspring. From adipose tissue, we identified 1214 individual hypomethylated CpG sites and 411 individual hypermethylated CpG sites in the LC group when compared to the CC group. Pathway analysis of the differential methylation genes revealed a significant increase in hypomethylated genes in the mitogen-activated protein kinase (MAPK) signaling pathway in the LC group. Our study highlights the importance of the MAPK signaling pathway in epigenetic changes involved in the lipid metabolism of the offspring from chromium-restricted dams. PMID:27517955

  12. Effects of Maternal Chromium Restriction on the Long-Term Programming in MAPK Signaling Pathway of Lipid Metabolism in Mice

    PubMed Central

    Zhang, Qian; Sun, Xiaofang; Xiao, Xinhua; Zheng, Jia; Li, Ming; Yu, Miao; Ping, Fan; Wang, Zhixin; Qi, Cuijuan; Wang, Tong; Wang, Xiaojing

    2016-01-01

    It is now broadly accepted that the nutritional environment in early life is a key factor in susceptibility to metabolic diseases. In this study, we evaluated the effects of maternal chromium restriction in vivo on the modulation of lipid metabolism and the mechanisms involved in this process. Sixteen pregnant C57BL mice were randomly divided into two dietary treatments: a control (C) diet group and a low chromium (L) diet group. The diet treatment was maintained through gestation and lactation period. After weaning, some of the pups continued with either the control diet or low chromium diet (CC or LL), whereas other pups switched to another diet (CL or LC). At 32 weeks of age, serum lipid metabolism, proinflammatory indexes, oxidative stress and anti-oxidant markers, and DNA methylation status in adipose tissue were measured. The results indicated that the maternal low chromium diet increased body weight, fat pad weight, serum triglyceride (TG), low-density lipoprotein cholesterol (LDL), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), and oxidized glutathione (GSSG). There was a decrease in serum reduced/oxidized glutathione (GSH/GSSG) ratio at 32 weeks of age in female offspring. From adipose tissue, we identified 1214 individual hypomethylated CpG sites and 411 individual hypermethylated CpG sites in the LC group when compared to the CC group. Pathway analysis of the differential methylation genes revealed a significant increase in hypomethylated genes in the mitogen-activated protein kinase (MAPK) signaling pathway in the LC group. Our study highlights the importance of the MAPK signaling pathway in epigenetic changes involved in the lipid metabolism of the offspring from chromium-restricted dams. PMID:27517955

  13. Formin-induced actin cables are required for polarized recruitment of the Ste5 scaffold and high level activation of MAPK Fus3.

    PubMed

    Qi, Maosong; Elion, Elaine A

    2005-07-01

    Little is known about how a mitogen-activated protein kinase (MAPK) cascade is targeted to specific sites at the plasma membrane during receptor stimulation. In budding yeast, the Ste5 scaffold is recruited to a receptor-coupled G protein during mating pheromone stimulation, allowing the tethered MAPK cascade to be activated by Ste20, a Cdc42-anchored kinase. Here we show that stable recruitment of Ste5 at cortical sites requires the formin Bni1, Bni1-induced actin cables, Rho1 and Myo2. Rho1 directs recruitment of Bni1 via the Rho-binding domain, and Bni1 mediates localization of Ste5 through actin cables and Myo2, which co-immunoprecipitates with Ste5 during receptor stimulation. Bni1 is also required for polarized recruitment and full activation of MAPK Fus3, which must bind Ste5 to be activated, and polarized recruitment of Cdc24, the guanine exchange factor that binds Ste5 and promotes its recruitment to the G protein. In contrast, Bni1 is not important for activation of MAPK Kss1, which can be activated while not bound to Ste5 and does not accumulate at cortical sites. These findings reveal that Bni1 mediates the formation of a Ste5 scaffold/Fus3 MAPK signaling complex at polarized sites, and suggests that a pool of Ste5 may translocate along formin-induced actin cables to the cell cortex. PMID:15961405

  14. Tension Force Downregulates Matrix Metalloproteinase Expression and Upregulates the Expression of Their Inhibitors through MAPK Signaling Pathways in MC3T3-E1 cells

    PubMed Central

    Karasawa, Yoko; Tanaka, Hideki; Nakai, Kumiko; Tanabe, Natsuko; Kawato, Takayuki; Maeno, Masao; Shimizu, Noriyoshi

    2015-01-01

    Objective: Matrix metalloproteinases (MMPs), produced by osteoblasts, catalyze the turnover of extracellular matrix (ECM) molecules in osteoid, and the regulation of MMP activity depends on interactions between MMPs and tissue inhibitors of metalloproteinases (TIMPs). We focused on the degradation process of ECM in osteoid that was exposed to mechanical strain, and conducted an in vitro study using MC3T3-E1 osteoblastic cells to examine the effects of tension force (TF) on the expression of MMPs and TIMPs, and activation of mitogen-activated protein kinase (MAPK) pathways. Design: Cells were incubated on flexible-bottomed culture plates and stimulated with or without cyclic TF for 24 hours. The expression of MMPs and TIMPs was examined at mRNA and protein levels by real-time RT-PCR and Western blotting, respectively. The phosphorylation of extracellular signal-regulated kinase (ERK) 1/2, p38 MAPK, and stress-activated protein kinases/c-jun N-terminal kinases (SAPK/JNK) were examined by Western blotting. Results: TF decreased the expression of MMP-1, -3, -13 and phosphorylated ERK1/2. In contrast, TF increased the expression of TIMP-2, -3 and phosphorylated SAPK/JNK. The expression of MMP-2, -14, TIMP-1, -4 and phosphorylated p38 MAPK was unaffected by TF. MMP-1, -3 and -13 expression decreased in cells treated with the ERK inhibitor PD98059 compared with untreated control cells. The JNK inhibitor SP600125 inhibited the TF-induced upregulation of TIMP-2 and -3. Conclusions: The results suggest that TF suppresses the degradation process that occurs during ECM turnover in osteoid via decreased production of MMP-1, -3 and -13, and increased production of TIMP-2 and -3 through the MAPK signaling pathways in osteoblasts. PMID:26640410

  15. Nucleophosmin Mutants Promote Adhesion, Migration and Invasion of Human Leukemia THP-1 Cells through MMPs Up-regulation via Ras/ERK MAPK Signaling

    PubMed Central

    Xian, Jingrong; Shao, Huiyuan; Chen, Xianchun; Zhang, Shuaishuai; Quan, Jing; Zou, Qin; Jin, Hongjun; Zhang, Ling

    2016-01-01

    Acute myeloid leukemia (AML) with mutated nucleophosmin (NPM1) has been defined as a unique subgroup in the new classification of myeloid neoplasm, and the AML patients with mutated NPM1 frequently present extramedullary infiltration, but how NPM1 mutants regulate this process remains elusive. In this study, we found that overexpression of type A NPM1 gene mutation (NPM1-mA) enhanced the adhesive, migratory and invasive potential in THP-1 AML cells lacking mutated NPM1. NPM1-mA had up-regulated expression and gelatinolytic matrix metalloprotease-2 (MMP-2)/MMP-9 activity, as assessed by real-time PCR, western blotting and gelatin zymography. Following immunoprecipitation analysis to identify the interaction of NPM1-mA with K-Ras, we focused on the effect of NPM1-mA overexpression on the Ras/Mitogen-activated protein kinase (MAPK) signaling axis and showed that NPM1-mA increased the MEK and ERK phosphorylation levels, as evaluated by western blotting. Notably, a specific inhibitor of the ERK/MAPK pathway (PD98059), but not p38/MAPK, JNK/MAPK or PI3-K/AKT inhibitors, markedly decreased the cell invasion numbers in a transwell assay. Further experiments demonstrated that blocking the ERK/MAPK pathway by PD98059 resulted in reduced MMP-2/9 protein levels and MMP-9 activity. Additionally, NPM1-mA overexpression had down-regulated gene expression and protein production of tissue inhibitor of MMP-2 (TIMP-2) in THP-1 cells. Furthermore, evaluation of gene expression data from The Cancer Genome Atlas (TCGA) dataset revealed that MMP-2 was overexpressed in AML patient samples with NPM1 mutated and high MMP-2 expression associated with leukemic skin infiltration. Taken together, our results reveal that NPM1 mutations contribute to the invasive potential of AML cells through MMPs up-regulation via Ras/ERK MAPK signaling pathway activation and offer novel insights into the potential role of NPM1 mutations in leukemogenesis. PMID:26884713

  16. The dynamics of MAPK inactivation at fertilization in mouse eggs

    PubMed Central

    Gonzalez-Garcia, Jose Raul; Bradley, Josephine; Nomikos, Michail; Paul, Laboni; Machaty, Zoltan; Lai, F. Anthony; Swann, Karl

    2014-01-01

    ABSTRACT Egg activation at fertilization in mammals is initiated by prolonged Ca2+ oscillations that trigger the completion of meiosis and formation of pronuclei. A fall in mitogen-activated protein kinase (MAPK) activity is essential for pronuclear formation, but the precise timing and mechanism of decline are unknown. Here, we have measured the dynamics of MAPK pathway inactivation during fertilization of mouse eggs using novel chemiluminescent MAPK activity reporters. This reveals that the MAPK activity decrease begins during the Ca2+ oscillations, but MAPK does not completely inactivate until after pronuclear formation. The MAPKs present in eggs are Mos, MAP2K1 and MAP2K2 (MEK1 and MEK2, respectively) and MAPK3 and MAPK1 (ERK1 and ERK2, respectively). Notably, the MAPK activity decline at fertilization is not explained by upstream destruction of Mos, because a decrease in the signal from a Mos–luciferase reporter is not associated with egg activation. Furthermore, Mos overexpression does not affect the timing of MAPK inactivation or pronuclear formation. However, the late decrease in MAPK could be rapidly reversed by the protein phosphatase inhibitor, okadaic acid. These data suggest that the completion of meiosis in mouse zygotes is driven by an increased phosphatase activity and not by a decline in Mos levels or MEK activity. PMID:24741069

  17. p38 Mitogen-Activated Protein Kinase (MAPK) Increases Arginase Activity and Contributes to Endothelial Dysfunction in Corpora Cavernosa from Angiotensin-II Treated Mice

    PubMed Central

    Toque, Haroldo A.; Romero, Maritza J.; Tostes, Rita C.; Shatanawi, Alia; Chandra, Surabhi; Carneiro, Zidonia N.; Inscho, Edward W.; Webb, R. Clinton; Caldwell, Ruth B.; Caldwell, R. William

    2010-01-01

    Introduction Angiotensin II (AngII) activates p38 mitogen-activated protein kinase (MAPK) and elevates arginase activity in endothelial cells. Upregulation of arginase activity has been implicated in endothelial dysfunction by reducing NO bioavailability. However, signaling pathways activated by AngII in the penis are largely unknown. Aim We hypothesized that activation of p38 MAPK increases arginase activity and thus impairs penile vascular function in AngII-treated mice. Methods Male C57BL/6 mice were implanted with osmotic minipumps containing saline or AngII (42 μg/kg/h) for 14 days and co-treated with p38 MAPK inhibitor, SB 203580 (5 μg/kg/day), beginning 2 days before minipump implantation. Systolic blood pressure (SBP) was measured. Corpus cavernosum (CC) tissue was used for vascular functional studies and protein expression levels of p38 MAPK, arginase and constitutive NOS, and arginase activity. Main Outcome Measures Arginase expression and activity; expression of phospho-p38 MAPK, -eNOS and nNOS proteins; endothelium-dependent and nitrergic nerve-mediated relaxations were determined in CC from control and AngII-infused mice. Results AngII increased SBP (22%) and increased CC arginase activity and expression (~2-fold), and phosphorylated P38 MAPK levels (30%) over control. Treatment with SB 203580 prevented these effects. Endothelium-dependent NO-mediated relaxation to acetylcholine was significantly reduced by AngII and this effect was prevented by SB 203580 (P<0.01). AngII (2-week) did not alter nitrergic function. However, SB 203580 significantly increased nitrergic relaxation in both control and AngII tissue at lower frequencies. Maximum contractile responses for phenylephrine and electrical field stimulation were increased by AngII (56% and 171%, respectively), and attenuated by SB 203580 treated. AngII treatment also decreased eNOS phosphorylation at Ser-1177 compared to control. Treatment with SB 203580 prevented all these changes. Conclusion p38

  18. p38 mitogen-activated protein kinase (p38 MAPK)-mediated autoimmunity: lessons to learn from ANCA vasculitis and pemphigus vulgaris.

    PubMed

    Mavropoulos, Athanasios; Orfanidou, Timoklia; Liaskos, Christos; Smyk, Daniel S; Billinis, Charalambos; Blank, Miri; Rigopoulou, Eirini I; Bogdanos, Dimitrios P

    2013-03-01

    Evidence is beginning to accumulate that p38 mitogen activated protein kinase (p38 MAPK) signaling pathway plays an important role in the regulation of cellular and humoral autoimmune responses. The exact mechanisms and the degree by which the p38 MAPK pathway participates in the immune-mediated induction of diseases have started to emerge. This review discusses the recent advances in the molecular dissection of the p38 MAPK pathway and the findings generated by reports investigating its role in the pathogenesis of autoimmune diseases such as rheumatoid arthritis, systemic lupus erythematosus, and autoimmune hepatitis. Application of newly-developed protocols based on sensitive flow cytometric detection has proven to be a useful tool in the investigation of the phosphorylation of p38 MAPK within different peripheral blood mononuclear cell populations and may help us to better understand the enigmatic role of this signaling cascade in the induction of autoimmunity as well as its role in immunosuppressive-induced remission. Special attention is paid to reported data proposing a specific role for autoantibody-induced activation of p38 MAPK-mediated immunopathology in the pathogenesis of autoimmune blistering diseases and anti-neutrophilic antibody-mediated vasculitides.

  19. Trimethyltin-Induced Microglial Activation via NADPH Oxidase and MAPKs Pathway in BV-2 Microglial Cells

    PubMed Central

    Kim, Da Jung; Kim, Yong Sik

    2015-01-01

    Trimethyltin (TMT) is known as a potent neurotoxicant that causes neuronal cell death and neuroinflammation, particularly in the hippocampus. Microglial activation is one of the prominent pathological features of TMT neurotoxicity. Nevertheless, it remains unclear how microglial activation occurs in TMT intoxication. In this study, we aimed to investigate the signaling pathways in TMT-induced microglial activation using BV-2 murine microglial cells. Our results revealed that TMT generates reactive oxygen species (ROS) and increases the expression of CD11b and nuclear factor-κB- (NF-κB-) mediated nitric oxide (NO) and tumor necrosis factor- (TNF-) α in BV-2 cells. We also observed that NF-κB activation was controlled by p38 and JNK phosphorylation. Moreover, TMT-induced ROS generation occurred via nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in BV-2 cells. Interestingly, treatment with the NADPH oxidase inhibitor apocynin significantly suppressed p38 and JNK phosphorylation and NF-κB activation and ultimately the production of proinflammatory mediators upon TMT exposure. These findings indicate that NADPH oxidase-dependent ROS generation activated p38 and JNK mitogen-activated protein kinases (MAPKs), which then stimulated NF-κB to release proinflammatory mediators in the TMT-treated BV-2 cells. PMID:26221064

  20. Ethanol Negatively Regulates Hepatic Differentiation of hESC by Inhibition of the MAPK/ERK Signaling Pathway In Vitro

    PubMed Central

    Ma, Xiaocui; Tschudy-Seney, Benjamin; Chen, Jiamei; Magner, Nataly L.; Revzin, Alexander; Nolta, Jan A.; Zern, Mark A.; Duan, Yuyou

    2014-01-01

    Background Alcohol insult triggers complex events in the liver, promoting fibrogenic/inflammatory signals and in more advanced cases, aberrant matrix deposition. It is well accepted that the regenerative capacity of the adult liver is impaired during alcohol injury. The liver progenitor/stem cells have been shown to play an important role in liver regeneration -in response to various chronic injuries; however, the effects of alcohol on stem cell differentiation in the liver are not well understood. Methods We employed hepatic progenitor cells derived from hESCs to study the impact of ethanol on hepatocyte differentiation by exposure of these progenitor cells to ethanol during hepatocyte differentiation. Results We found that ethanol negatively regulated hepatic differentiation of hESC-derived hepatic progenitor cells in a dose-dependent manner. There was also a moderate cell cycle arrest at G1/S checkpoint in the ethanol treated cells, which is associated with a reduced level of cyclin D1 in these cells. Ethanol treatment specifically inhibited the activation of the ERK but not JNK nor the p38 MAP signaling pathway. At the same time, the WNT signaling pathway was also reduced in the cells exposed to ethanol. Upon evaluating the effects of the inhibitors of these two signaling pathways, we determined that the Erk inhibitor replicated the effects of ethanol on the hepatocyte differentiation and attenuated the WNT/β-catenin signaling, however, inhibitors of WNT only partially replicated the effects of ethanol on the hepatocyte differentiation. Conclusion Our results demonstrated that ethanol negatively regulated hepatic differentiation of hESC-derived hepatic progenitors through inhibiting the MAPK/ERK signaling pathway, and subsequently attenuating the WNT signaling pathway. Thus, our finding provides a novel insight into the mechanism by which alcohol regulates cell fate selection of hESC-derived hepatic progenitor cells, and the identified pathways may provide

  1. AMF/PGI-mediated tumorigenesis through MAPK-ERK signaling in endometrial carcinoma.

    PubMed

    Li, Yiran; Jia, Yuanhui; Che, Qi; Zhou, Qian; Wang, Kai; Wan, Xiao-Ping

    2015-09-22

    Autocrine motility factor (AMF), which is also known as phosphoglucose isomerase (PGI), enhances tumor cell growth and motility. In this study, we found that AMF and its receptor were both highly expressed in Endometrial Carcinoma (EC) tissues compared to normal tissues. Levels of AMF were increased in serum of endometrial cancer patients. Downregulation of AMF by shRNA inhibited invasion, migration and proliferation as well as growth in a three-dimensional culture. AMF cytokine function, but not enzymatic activity of PGI, regulated tumorigenic activities of AMF. The MAPK-ERK1/2 pathway contributed to AMF-induced effects in EC cells. In agreement, Mek inhibitor decreased AMF-induced invasion, migration and proliferation of EC cells. In addition, in two mouse tumor metastasis models (EC cells delivered through left ventricle or intraperitoneally) AMF-silenced EC cells showed decreased tumor proliferative and metastatic capacities. We suggest that AMF/PGI is a potential therapeutic target in endometrial carcinoma.

  2. Nicotine enhances invasion and metastasis of human colorectal cancer cells through the nicotinic acetylcholine receptor downstream p38 MAPK signaling pathway.

    PubMed

    Xiang, Tao; Fei, Rushan; Wang, Zhe; Shen, Zhonglei; Qian, Jing; Chen, Wenbin

    2016-01-01

    Nicotine as a cigarette component is an established risk factor for colorectal cancer tumorigenesis. The downstream signaling pathways of nicotinic acetylcholine receptors (nAchRs) are believed to be responsible for the cellular effects. In the present study, we evaluated the effects and novel mechanisms for nicotine on the capacity for colorectal cancer cell invasion and metastasis. LOVO and SW620 colorectal cancer cells were stimulated with nicotine in vitro. A Transwell chamber model was applied to detect the capacity for tumor cell invasion. Assays for gelatin zymography and western blotting were applied to detect the activity and expression of metastasis-related matrix metalloproteinases (MMPs), respectively. Signal transduction was assessed by immunoblotting for the phosphorylation of relevant signal molecules and the application of pharmaceutical inhibitors. We showed that nicotine increased LOVO and SW620 colorectal cancer cell invasion along with enhanced activity and expression of MMP-1, -2 and -9. Nicotine increased phosphorylation of p38, ERK, Akt and PI3K p85 but had no effect on phosphorylation of JNK, or NF-κB. Of the pharmaceutical inhibitors of U0126 (ERK1/2 inhibitor), LY294002 (Akt activation inhibitor), SB239063 (p38 MAPK activation inhibitor) and hexamethonium (Hex) (nAchRs inhibitor), the cellular and molecular effects were reduced by the applications of SB239063 and Hex. We concluded that nicotine stimulates the invasion and metastasis of colon cancer cells in vitro via activation of the nAchRs and the p38 MAPK downstream signaling pathway. Therefore, p38 MAPK may have potential as a therapeutic target for smoking-related human colorectal cancer metastasis.

  3. Requirement of ERα and basal activities of EGFR and Src kinase in Cd-induced activation of MAPK/ERK pathway in human breast cancer MCF-7 cells

    SciTech Connect

    Song, Xiulong Wei, Zhengxi; Shaikh, Zahir A.

    2015-08-15

    Cadmium (Cd) is a common environmental toxicant and an established carcinogen. Epidemiological studies implicate Cd with human breast cancer. Low micromolar concentrations of Cd promote proliferation of human breast cancer cells in vitro. The growth promotion of breast cancer cells is associated with the activation of MAPK/ERK pathway. This study explores the mechanism of Cd-induced activation of MAPK/ERK pathway. Specifically, the role of cell surface receptors ERα, EGFR, and Src kinase was evaluated in human breast cancer MCF-7 cells treated with 1–3 μM Cd. The activation of ERK was studied using a serum response element (SRE) luciferase reporter assay. Receptor phosphorylation was detected by Western blot analyses. Cd treatment increased both the SRE reporter activity and ERK1/2 phosphorylation in a concentration-dependent manner. Cd treatment had no effect on reactive oxygen species (ROS) generation. Also, blocking the entry of Cd into the cells with manganese did not diminish Cd-induced activation of MAPK/ERK. These results suggest that the effect of Cd was likely not caused by intracellular ROS generation, but through interaction with the membrane receptors. While Cd did not appear to activate either EGFR or Src kinase, their inhibition completely blocked the Cd-induced activation of ERK as well as cell proliferation. Similarly, silencing ERα with siRNA or use of ERα antagonist blocked the effects of Cd. Based on these results, it is concluded that not only ERα, but also basal activities of EGFR and Src kinase are essential for Cd-induced signal transduction and activation of MAPK/ERK pathway for breast cancer cell proliferation. - Highlights: • Low micromolar concentrations of Cd rapidly activate ERK1/2 in MCF-7 cells. • Signal transduction and resulting cell proliferation require EGFR, ERα, and Src. • These findings implicate Cd in promotion of breast cancer.

  4. Neuroprotective effect of sulfated polysaccharide isolated from sea cucumber Stichopus japonicus on 6-OHDA-induced death in SH-SY5Y through inhibition of MAPK and NF-κB and activation of PI3K/Akt signaling pathways.

    PubMed

    Cui, Chao; Cui, Ningshan; Wang, Peng; Song, Shuliang; Liang, Hao; Ji, Aiguo

    2016-02-01

    The purpose of this study is to investigate the protective effect and molecular mechanism of the sulfated polysaccharide (SJP) isolated from the sea cucumber Stichopus japonicus against 6-OHDA-induced toxicity in SH-SY5Y cells. The results showed that SJP could protect SH-SY5Y cells against 6-OHDA-induced cell injury. We found that SJP effectively improves cell viability, decreases LDH leakage, and reverses morphological damage. Moreover, SJP significantly increases SOD activity but decreases MDA levels and ROS generation. Effect of SJP on 6-OHDA-induced cell death in SH-SY5Y cells is associated with an arrest in the G1/S phase of the cell cycle and inhibits the expression of Cyclin D3. 6-OHDA-induced intracellular generation of ROS and mitochondrial dysfunctions, release of cytochrome c, imbalance of Bax/Bcl-2, cleaved caspase-9/caspase-9 and cleaved caspase-3/caspase-3 ratio, and p-p53 activation were strikingly attenuated by SJP pretreatment. Meanwhile, SJP counteracted NF-κB activation, thereby preventing up-regulation of iNOS and intracellular NO release. The data provide the first evidence that SJP protects SH-SY5Y cells against 6-OHDA toxicity possibly by inhibiting MAPK and NF-κB and activating PI3K/Akt signaling pathways. Thus, SJP is a candidate for further evaluation of its protective effects against neurodegeneration in PD.

  5. Neuroprotective effect of sulfated polysaccharide isolated from sea cucumber Stichopus japonicus on 6-OHDA-induced death in SH-SY5Y through inhibition of MAPK and NF-κB and activation of PI3K/Akt signaling pathways.

    PubMed

    Cui, Chao; Cui, Ningshan; Wang, Peng; Song, Shuliang; Liang, Hao; Ji, Aiguo

    2016-02-01

    The purpose of this study is to investigate the protective effect and molecular mechanism of the sulfated polysaccharide (SJP) isolated from the sea cucumber Stichopus japonicus against 6-OHDA-induced toxicity in SH-SY5Y cells. The results showed that SJP could protect SH-SY5Y cells against 6-OHDA-induced cell injury. We found that SJP effectively improves cell viability, decreases LDH leakage, and reverses morphological damage. Moreover, SJP significantly increases SOD activity but decreases MDA levels and ROS generation. Effect of SJP on 6-OHDA-induced cell death in SH-SY5Y cells is associated with an arrest in the G1/S phase of the cell cycle and inhibits the expression of Cyclin D3. 6-OHDA-induced intracellular generation of ROS and mitochondrial dysfunctions, release of cytochrome c, imbalance of Bax/Bcl-2, cleaved caspase-9/caspase-9 and cleaved caspase-3/caspase-3 ratio, and p-p53 activation were strikingly attenuated by SJP pretreatment. Meanwhile, SJP counteracted NF-κB activation, thereby preventing up-regulation of iNOS and intracellular NO release. The data provide the first evidence that SJP protects SH-SY5Y cells against 6-OHDA toxicity possibly by inhibiting MAPK and NF-κB and activating PI3K/Akt signaling pathways. Thus, SJP is a candidate for further evaluation of its protective effects against neurodegeneration in PD. PMID:26773499

  6. Extracellular Matrix Stiffness Regulates Osteogenic Differentiation through MAPK Activation

    PubMed Central

    Hwang, Jun-Ha; Byun, Mi Ran; Kim, A. Rum; Kim, Kyung Min; Cho, Hang Jun; Lee, Yo Han; Kim, Juwon; Jeong, Mi Gyeong; Hwang, Eun Sook; Hong, Jeong-Ho

    2015-01-01

    Mesenchymal stem cell (MSC) differentiation is regulated by the extracellular matrix (ECM) through activation of intracellular signaling mediators. The stiffness of the ECM was shown to be an important regulatory factor for MSC differentiation, and transcriptional coactivator with PDZ-binding motif (TAZ) was identified as an effector protein for MSC differentiation. However, the detailed underlying mechanism regarding the role of ECM stiffness and TAZ in MSC differentiation is not yet fully understood. In this report, we showed that ECM stiffness regulates MSC fate through ERK or JNK activation. Specifically, a stiff hydrogel matrix stimulates osteogenic differentiation concomitant with increased nuclear localization of TAZ, but inhibits adipogenic differentiation. ERK and JNK activity was significantly increased in cells cultured on a stiff hydrogel. TAZ activation was induced by ERK or JNK activation on a stiff hydrogel because exposure to an ERK or JNK inhibitor significantly decreased the nuclear localization of TAZ, indicating that ECM stiffness-induced ERK or JNK activation is important for TAZ-driven osteogenic differentiation. Taken together, these results suggest that ECM stiffness regulates MSC differentiation through ERK or JNK activation. PMID:26262877

  7. Extracellular Matrix Stiffness Regulates Osteogenic Differentiation through MAPK Activation.

    PubMed

    Hwang, Jun-Ha; Byun, Mi Ran; Kim, A Rum; Kim, Kyung Min; Cho, Hang Jun; Lee, Yo Han; Kim, Juwon; Jeong, Mi Gyeong; Hwang, Eun Sook; Hong, Jeong-Ho

    2015-01-01

    Mesenchymal stem cell (MSC) differentiation is regulated by the extracellular matrix (ECM) through activation of intracellular signaling mediators. The stiffness of the ECM was shown to be an important regulatory factor for MSC differentiation, and transcriptional coactivator with PDZ-binding motif (TAZ) was identified as an effector protein for MSC differentiation. However, the detailed underlying mechanism regarding the role of ECM stiffness and TAZ in MSC differentiation is not yet fully understood. In this report, we showed that ECM stiffness regulates MSC fate through ERK or JNK activation. Specifically, a stiff hydrogel matrix stimulates osteogenic differentiation concomitant with increased nuclear localization of TAZ, but inhibits adipogenic differentiation. ERK and JNK activity was significantly increased in cells cultured on a stiff hydrogel. TAZ activation was induced by ERK or JNK activation on a stiff hydrogel because exposure to an ERK or JNK inhibitor significantly decreased the nuclear localization of TAZ, indicating that ECM stiffness-induced ERK or JNK activation is important for TAZ-driven osteogenic differentiation. Taken together, these results suggest that ECM stiffness regulates MSC differentiation through ERK or JNK activation.

  8. Fisetin induces apoptosis and endoplasmic reticulum stress in human non-small cell lung cancer through inhibition of the MAPK signaling pathway.

    PubMed

    Kang, Kyoung Ah; Piao, Mei Jing; Madduma Hewage, Susara Ruwan Kumara; Ryu, Yea Seong; Oh, Min Chang; Kwon, Taeg Kyu; Chae, Sungwook; Hyun, Jin Won

    2016-07-01

    Fisetin (3,3',4',7-tetrahydroxyflavone), a dietary flavonoid compound, is currently being investigated for its anticancer effect in various cancer models, including lung cancer. Recent studies show that fisetin induces cell growth inhibition and apoptosis in the human non-small cell lung cancer line NCI-H460. In this study, we investigated whether fisetin can induce endoplasmic reticulum (ER) stress-mediated apoptosis in NCI-H460 cells. Fisetin induced mitochondrial reactive oxygen species (ROS) and characteristic signs of ER stress: ER staining; mitochondrial Ca(2+) overload; expression of ER stress-related proteins; glucose-regulated protein (GRP)-78, phosphorylation of protein kinase RNA (PKR)-like endoplasmic reticulum kinase (PERK) and phosphorylation of eukaryotic initiation factor-2 α subunit; cleavage of activating transcription factor-6; phosphorylation of inositol-requiring kinase-1 and splicing of X-box transcription factor-1; induction of C/EBP homologous protein and cleaved caspase-12. siRNA-mediated knockdown of CHOP and ATF-6 attenuated fisetin-induced apoptotic cell death. In addition, fisetin induced phosphorylation of ERK, JNK, and p38 MAPK. Moreover, silencing of the MAPK signaling pathway prevented apoptotic cell death. In summary, our results indicate that, in NCI-H460 cells, fisetin induces apoptosis and ER stress that is mediated by induction of the MAPK signaling pathway.

  9. The Rho-GTPase Rnd1 Suppresses Mammary Tumorigenesis and EMT by Restraining Ras-MAPK signaling

    PubMed Central

    Okada, Tomoyo; Sinha, Surajit; Esposito, Ilaria; Schiavon, Gaia; López-Lago, Miguel A.; Su, Wenjing; Pratilas, Christine A.; Abele, Cristina; Hernandez, Jonathan M.; Ohara, Masahiro; Okada, Morihito; Viale, Agnes; Heguy, Adriana; Socci, Nicholas D.; Sapino, Anna; Seshan, Venkatraman E.; Long, Stephen; Inghirami, Giorgio; Rosen, Neal; Giancotti, Filippo G.

    2015-01-01

    SUMMARY We identified the Rho-GTPase Rnd1 as a candidate metastasis suppressor through bioinformatics analysis and showed that its depletion disrupt epithelial adhesion and polarity, induced Epithelial-to-Mesenchymal Transition (EMT), and cooperated with deregulated expression of c-Myc or loss of p53 to cause neoplastic conversion. Mechanistic studies revealed that Rnd1 suppresses Ras signalling by activating the GAP domain of Plexin B1, which inhibits Rap1. Rap1 inhibition in turn led to derepression of p120-RasGAP, which was able to inhibit Ras. Inactivation of Rnd1 in mammary epithelial cells induced highly undifferentiated and invasive tumors in mice. Conversely, Rnd1 expression inhibited spontaneous and experimental lung colonization in mouse models of metastasis. Genomic studies indicated that gene deletion in combination with epigenetic silencing or, more rarely, point mutation inactivates RND1 in human breast cancer. These results reveal a previously unappreciated mechanism through which Rnd1 restrains activation of Ras-MAPK signaling and breast tumor initiation and progression. PMID:25531777

  10. 17β-Estradiol induces odontoblastic differentiation via activation of the c-Src/MAPK pathway in human dental pulp cells.

    PubMed

    Woo, Su Mi; Seong, Kyung Joo; Oh, Sang Jin; Park, Hong Ju; Kim, Sun Hun; Kim, Won Jae; Jung, Ji Yeon

    2015-12-01

    The present study is aimed at investigating the effects of the exogenous estrogen 17β-estradiol (E2) on odontoblastic differentiation in human dental pulp cells (HDPCs) immotalized with hTERT gene and their molecular mechanism. Proliferation was detected by BrdU assay, and odontoblast differentiation induction was evaluated by the expression of dentin sialophosphoprotein (DSPP), dentin sialoprotein (DSP) and dentin matrix protein1 (DMP1), and alkaline phosphatase (ALP) activity and mineralization. Estrogen receptor-α (ER-α), c-Src, and mitogen-activated protein kinases (MAPKs) were examined and their inhibitors were used to determine the roles on odontogenic induction. E2 significantly promoted the HDPC proliferation, which was mediated by extracellular signal-related kinase 1/2. E2 upregulated DSPP, DSP, and DMP1 as the odontogenic differentiation markers and enhanced ALP activity and mineralization. E2 increased phosphorylation of ER-α and fulvestrant, an ER downregulator, significantly downregulated DSPP, DMP1, and DSP induced by E2. Moreover, E2 treatment activated c-Src and MAPKs upon odontogenic induction, whereas chemical inhibition of c-Src and MAPKs decreased expression of DSPP, DMP1, and DSP and mineralization augmented by E2. Moreover, fulvestrant reduced E2-induced phosphorylation of c-Src and MAPK and inhibition of c-Src by PP2 attenuated activation of MAPKs during E2-induced odontoblastic differentiation. Taken together, these results indicated that E2 stimulates odontoblastic differentiation of HDPCs via coordinated regulation of ER-α, c-Src, and MAPK signaling pathways, which may play a key role in the regeneration of dentin.

  11. Inhibitory effect of panduratin A on UV-induced activation of mitogen-activated protein kinases (MAPKs) in dermal fibroblast cells.

    PubMed

    Shim, Jae-Seok; Kwon, Yi-Young; Han, Young-Sun; Hwang, Jae-Kwan

    2008-10-01

    Exposure of the skin to ultraviolet (UV) induces photoaging associated with up-regulated matrix metalloproteinases (MMPs) activities and decreased collagen synthesis. We previously reported that panduratin A, a chalcone compound isolated from KAEMPFERIA PANDURATA Roxb ., decreased MMP-1 expression in UV-irradiated human skin fibroblasts. Here, we have investigated the effect of panduratin A on UV-induced activation of mitogen-activated protein kinases (MAPKs) signaling modules such as extracellular-regulated protein kinase (ERK), Jun-N-terminal kinase (JNK) and p38 kinase. Treatment with panduratin A in the range of 0.001 - 0.1 microM significantly inhibited UV-induced ERK, JNK and p38 activation. Moreover, inhibition of ERK, JNK and p38 by panduratin A resulted in decreased c-Fos expression and c-Jun phosphorylation induced by UV, which led to inhibition of activator protein-1 (AP-1) DNA binding activity. Panduratin A showed stronger activity than epigallocatechin 3- O-gallate (EGCG) known as a natural anti-aging agent. The results suggest that panduratin A can down-regulate UV-induced MMP-1 expression by inhibiting the MAPKs pathways and AP-1 activation. AP-1:activator protein-1 EGCG:epigallocatechin 3- O-gallate ERK:extracellular-regulated protein kinase JNK:c-Jun N-terminal kinase MAPK:mitogen-activated protein kinase MMP:matrix metalloproteinase UV:ultraviolet.

  12. Ginkgo biloba extract mitigates liver fibrosis and apoptosis by regulating p38 MAPK, NF-κB/IκBα, and Bcl-2/Bax signaling

    PubMed Central

    Wang, Yuanyuan; Wang, Rong; Wang, Yujie; Peng, Ruqin; Wu, Yan; Yuan, Yongfang

    2015-01-01

    Background Liver fibrosis is the consequence of diverse liver injuries and can eventually develop into liver cirrhosis. Ginkgo biloba extract (GBE) is an extract from dried ginkgo leaves that has many pharmacological effects because of its various ingredients and has been shown to be hepatoprotective. Purpose and methods Aimed to investigate the underlying protective mechanisms of GBE on carbon tetrachloride (CCl4)-induced liver fibrosis in rats. Male Sprague Dawley rats were randomly divided into four groups: control group (C), model group (M), low-dose group (L), and high-dose group (H). Liver fibrosis was induced by CCl4 groups M, L, and H: group C was administered saline. In addition, GBE at different doses was used to treat groups L and H. Results The results of hematoxylin and eosin staining, Masson’s trichrome staining, a liver function index, and a liver fibrosis index showed that GBE application noticeably mitigated fibrosis and improved the function of the liver. The western blotting and immunohistochemistry analyses indicated that GBE reduced liver fibrosis not only by inhibiting p38 MAPK and NF-κBp65 via inhibition of IκBα degradation but also by inhibiting hepatocyte apoptosis via downregulation of Bax, upregulation of Bcl-2, and subsequent inhibition of caspase-3 activation. Inflammation-associated factors and hepatic stellate cell (HSC)-activation markers further demonstrated that GBE could effectively inhibit HSC activation and inflammation as a result of its regulation of p38 MAPK and nuclear factor-kappa B/IκBα signaling. Conclusion Our findings indicated a novel role for GBE in the treatment of liver fibrosis. The potential mechanisms may be associated with the following signaling pathways: 1) the p38 MAPK and nuclear factor-kappa B/IκBα signaling pathways (inhibiting inflammation and HSCs activation) and 2) the Bcl-2/Bax signaling pathway (inhibiting the apoptosis of hepatocytes). PMID:26664050

  13. FGF signaling via MAPK is required early and improves Activin A-induced definitive endoderm formation from human embryonic stem cells

    SciTech Connect

    Sui, Lina; Mfopou, Josue K.; Geens, Mieke; Sermon, Karen; Bouwens, Luc

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer Deep study the FGF signaling role during DE specification in the context of hESCs. Black-Right-Pointing-Pointer DE differentiation from hESCs has an early dependence on FGF signaling. Black-Right-Pointing-Pointer A serum-free DE protocol is developed based on the findings. Black-Right-Pointing-Pointer The DE cells showed potential to differentiate into pancreatic progenitor cells. -- Abstract: Considering their unlimited proliferation and pluripotency properties, human embryonic stem cells (hESCs) constitute a promising resource applicable for cell replacement therapy. To facilitate this clinical translation, it is critical to study and understand the early stage of hESCs differentiation wherein germ layers are defined. In this study, we examined the role of FGF signaling in Activin A-induced definitive endoderm (DE) differentiation in the absence of supplemented animal serum. We found that activated FGF/MAPK signaling is required at the early time point of Activin A-induced DE formation. In addition, FGF activation increased the number of DE cells compared to Activin A alone. These DE cells could further differentiate into PDX1 and NKX6.1 positive pancreatic progenitors in vitro. We conclude that Activin A combined with FGF/MAPK signaling efficiently induce DE cells in the absence of serum. These findings improve our understanding of human endoderm formation, and constitute a step forward in the generation of clinical grade hESCs progenies for cell therapy.

  14. Dehydrocorydaline promotes myogenic differentiation via p38 MAPK activation.

    PubMed

    Yoo, Miran; Lee, Sang-Jin; Kim, Yong Kee; Seo, Dong-Wan; Baek, Nam-In; Ryu, Jae-Ha; Kang, Jong-Sun; Bae, Gyu-Un

    2016-10-01

    Muscle regeneration is a coordinated process that involves proliferation and differentiation of muscle progenitor cells. Activation of MyoD is a key event in myogenic differentiation, which is regulated by p38 mitogen‑activated protein kinases (MAPK). In a screen of natural compounds for the enhancement of MyoD activity, dehydrocorydaline (DHC) from the Corydalis tuber was identified. Treatment of C2C12 myoblasts with DHC increased the expression levels of muscle‑specific proteins, including MyoD, myogenin and myosin heavy chain. In addition, C2C12 myoblasts exhibited enhanced multinucleated myotube formation without any cytotoxicity. Treatment with DHC elevated p38 MAPK activation and the interaction of MyoD with an E protein, which is likely to result in activation of MyoD and promotion of myoblast differentiation. Furthermore, defects in differentiation‑induced p38 MAPK activation and myoblast differentiation induced by depletion of the promyogenic receptor protein Cdo in C2C12 myoblasts were restored by DHC treatment. In conclusion, these results indicated that DHC stimulates p38 MAPK activation, which can enhance heterodimerization of MyoD and E proteins, thus resulting in MyoD activation and myoblast differentiation. These findings suggested that DHC may be considered a potential therapeutic compound for the improvement of muscle stem cell regenerative capacity in injured muscle.

  15. Dehydrocorydaline promotes myogenic differentiation via p38 MAPK activation.

    PubMed

    Yoo, Miran; Lee, Sang-Jin; Kim, Yong Kee; Seo, Dong-Wan; Baek, Nam-In; Ryu, Jae-Ha; Kang, Jong-Sun; Bae, Gyu-Un

    2016-10-01

    Muscle regeneration is a coordinated process that involves proliferation and differentiation of muscle progenitor cells. Activation of MyoD is a key event in myogenic differentiation, which is regulated by p38 mitogen‑activated protein kinases (MAPK). In a screen of natural compounds for the enhancement of MyoD activity, dehydrocorydaline (DHC) from the Corydalis tuber was identified. Treatment of C2C12 myoblasts with DHC increased the expression levels of muscle‑specific proteins, including MyoD, myogenin and myosin heavy chain. In addition, C2C12 myoblasts exhibited enhanced multinucleated myotube formation without any cytotoxicity. Treatment with DHC elevated p38 MAPK activation and the interaction of MyoD with an E protein, which is likely to result in activation of MyoD and promotion of myoblast differentiation. Furthermore, defects in differentiation‑induced p38 MAPK activation and myoblast differentiation induced by depletion of the promyogenic receptor protein Cdo in C2C12 myoblasts were restored by DHC treatment. In conclusion, these results indicated that DHC stimulates p38 MAPK activation, which can enhance heterodimerization of MyoD and E proteins, thus resulting in MyoD activation and myoblast differentiation. These findings suggested that DHC may be considered a potential therapeutic compound for the improvement of muscle stem cell regenerative capacity in injured muscle. PMID:27573543

  16. Tanreqing Injection Attenuates Lipopolysaccharide-Induced Airway Inflammation through MAPK/NF-κB Signaling Pathways in Rats Model

    PubMed Central

    Liu, Wei; Jiang, Hong-li; Cai, Lin-li; Yan, Min; Dong, Shou-jin; Mao, Bing

    2016-01-01

    Background. Tanreqing injection (TRQ) is a commonly used herbal patent medicine for treating inflammatory airway diseases in view of its outstanding anti-inflammatory properties. In this study, we explored the signaling pathways involved in contributions of TRQ to LPS-induced airway inflammation in rats. Methods/Design. Adult male Sprague Dawley (SD) rats randomly divided into different groups received intratracheal instillation of LPS and/or intraperitoneal injection of TRQ. Bronchoalveolar Lavage Fluid (BALF) and lung samples were collected at 24 h, 48 h, and 96 h after TRQ administration. Protein and mRNA levels of tumor necrosis factor- (TNF-) α, Interleukin- (IL-) 1β, IL-6, and IL-8 in BALF and lung homogenate were observed by ELISA and real-time PCR, respectively. Lung sections were stained for p38 MAPK and NF-κB detection by immunohistochemistry. Phospho-p38 MAPK, phosphor-extracellular signal-regulated kinases ERK1/2, phospho-SAPK/JNK, phospho-NF-κB p65, phospho-IKKα/β, and phospho-IκB-α were measured by western blot analysis. Results. The results showed that TRQ significantly counteracted LPS-stimulated release of TNF-α, IL-1β, IL-6, and IL-8, attenuated cells influx in BALF, mitigated mucus hypersecretion, suppressed phosphorylation of NF-κB p65, IκB-α, ΙKKα/β, ERK1/2, JNK, and p38 MAPK, and inhibited p38 MAPK and NF-κB p65 expression in rat lungs. Conclusions. Results of the current research indicate that TRQ possesses potent exhibitory effects in LPS-induced airway inflammation by, at least partially, suppressing the MAPKs and NF-κB signaling pathways, in a general dose-dependent manner. PMID:27366191

  17. Hepatocyte growth factor increases the invasive potential of PC-3 human prostate cancer cells via an ERK/MAPK and Zeb-1 signaling pathway

    PubMed Central

    HAN, YILI; LUO, YONG; WANG, YONGXING; CHEN, YATONG; LI, MINGCHUAN; JIANG, YONGGUANG

    2016-01-01

    Hepatocyte growth factor (HGF) has been implicated in epithelial-mesenchymal transition (EMT) in numerous types of cancer. However, to the best of our knowledge, there has been no previous evidence that HGF has a role in prostate cancer. The present study aimed to investigate the effect of HGF on EMT and invasive potential, as well as the underlying molecular mechanisms, in a human prostate cancer cell line. Therefore, PC-3 cells were treated with various concentrations of HGF for varying durations. EMT-associated proteins, including E-cadherin and vimentin, were examined by western blot analysis. The effects of HGF on cell proliferation, migration, invasion and tumorigenicity were assessed using MTT, wound-healing, Transwell and soft-agar assays. Subsequently, the role of c-Met in the mediation of EMT-like changes was investigated using reverse transcription-polymerase chain reaction, western blot analysis and gene knockdown by small interfering RNA. Finally, western blot analysis was used to quantify the expression of a downstream transcription factor and extracellular signal-related kinase/mitogen activated protein kinase (ERK/MAPK) signaling pathway proteins. The results indicated that treatment with HGF induced EMT-like changes and enhanced the invasive potential of PC-3 cells. There was an increase in the expression of ERK, phosphorylated-ERK and zinc finger E-box binding homeobox-1 (Zeb-1), suggesting that EMT-like changes may be mediated through the ERK/MAPK and Zeb-1 signaling pathway. Furthermore, HGF-mediated EMT-like changes were associated with c-Met activation, and these changes were able to be blocked by c-Met knockdown. The present study demonstrated that HGF-induced EMT increased the invasive potential of PC-3 human prostate cancer cells through activating the ERK/MAPK and Zeb-1 signaling pathway. PMID:26870279

  18. Hydrogen peroxide increases [3H]-2-deoxyglucose uptake via MAPKs, cPLA2, and NF-kappaB signaling pathways in mouse embryonic stem cells.

    PubMed

    Na, Sun Im; Lee, Min Young; Heo, Jung Sun; Han, Ho Jae

    2007-01-01

    Hydrogen peroxide (H(2)O(2)) has been shown to act as a signaling molecule that is involved in many cellular functions. This study investigated the effect of H(2)O(2) on the [3H]-2-deoxyglucose (2-DG) uptake and its related signaling pathways in mouse embryonic stem (ES) cells. H(2)O(2) significantly increased the level of 2-DG uptake in a time- (> 4 hr) and concentration- (>10-4 M) dependent manner due to an increase in V(max) but not K(m). Indeed, H(2)O(2) increased the mRNA and protein level of glucose transporter 1 (GLUT 1). PD 98059 (a p44/42 MAPKs inhibitor, 10-5 M), SB 203580 (a p38 MAPK inhibitor, 10-6 M), and SP 600125 (a SAPK/JNK inhibitor, 10-6 M) blocked the H(2)O(2)-induced increase in 2-DG uptake. H(2)O(2) also increased phosphorylation of p44/42 mitogen activated protein kinases (MAPKs), p38 MAPK, and stress-activated protein kinase/Jun-N-terminal kinase (SAPK/JNK). In addition, H(2)O(2) stimulated the translocation of cytosolic phospholipase A(2) (cPLA(2)) from the cytosolic fraction to the membrane fraction, the release of arachidonic acid, and the activation of NF-kappaB. AACOCF(3) or mepacrine (cPLA(2) inhibitors, 10-6 M), SN 50 (NF-kappaB nucleus translocation inhibitor, 500 ng/ml) or Bay11-7082 (a IkappaB-alpha phosphorylation inhibitor, 2x10-5 M) blocked the H(2)O(2)-induced increase in 2-DG uptake. H(2)O(2) increased the protein level of glucose transporter 1 (GLUT 1), which was blocked by PD 98059, SB 203580, SP 600125, mepacrine, or Bay11-7082. In conclusion, H(2)O(2) increases the 2-DG uptake via MAPKs, cPLA(2), and NF-kappaB signaling pathways in mouse ES cells.

  19. Ghrelin accelerates wound healing through GHS-R1a-mediated MAPK-NF-κB/GR signaling pathways in combined radiation and burn injury in rats

    PubMed Central

    Liu, Cong; Huang, Jiawei; Li, Hong; Yang, Zhangyou; Zeng, Yiping; Liu, Jing; Hao, Yuhui; Li, Rong

    2016-01-01

    The therapeutic effect of ghrelin on wound healing was assessed using a rat model of combined radiation and burn injury (CRBI). Rat ghrelin, anti-rat tumor necrosis factor (TNF) α polyclonal antibody (PcAb), or selective antagonists of p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), and growth hormone secretagogue receptor (GHS-R) 1a (SB203580, SP600125, and [D-Lys3]-GHRP-6, respectively), were administered for seven consecutive days. Levels of various signaling molecules were assessed in isolated rat peritoneal macrophages. The results showed that serum ghrelin levels and levels of macrophage glucocorticoid receptor (GR) decreased, while phosphorylation of p38MAPK, JNK, and p65 nuclear factor (NF) κB increased. Ghrelin inhibited the serum induction of proinflammatory mediators, especially TNF-α, and promoted wound healing in a dose-dependent manner. Ghrelin treatment decreased phosphorylation of p38MAPK, JNK, and p65NF-κB, and increased GR levels in the presence of GHS-R1a. SB203580 or co-administration of SB203580 and SP600125 decreased TNF-α level, which may have contributed to the inactivation of p65NF-κB and increase in GR expression, as confirmed by western blotting. In conclusion, ghrelin enhances wound recovery in CRBI rats, possibly by decreasing the induction of TNF-α or other proinflammatory mediators that are involved in the regulation of GHS-R1a-mediated MAPK-NF-κB/GR signaling pathways. PMID:27271793

  20. Molecular cloning of the duck mitogen-activated protein kinase 1 (MAPK1) gene and the development of a quantitative real-time PCR assay to detect its expression.

    PubMed

    Cao, Shoulin; Han, Xiangan; Ding, Chan; Wang, Shaohui; Tian, Mingxing; Wang, Xiaolan; Hou, Wanwan; Yue, Jiaping; Wang, Guijun; Yu, Shengqing

    2014-09-01

    Mitogen-activated protein kinase 1 (MAPK1) acts as an integration point for multiple biochemical signals, and is involved in a wide variety of biological processes such as cell proliferation and differentiation, transcription regulation, and development. Mitogen-activated protein kinase 1 plays an important role in inducing cell death in bacterial infections. In this study, the duck MAPK1 gene was cloned for the first time from the Cherry Valley duck. Sequence analysis showed that duck MAPK1 cDNA is 1,557 bp long, with an open reading frame of 1,107 bp. It encodes 368 amino acids, with 85.4, 84.5, and 97.3% homology with the human, mouse, and chicken MAPK1 gene, respectively. Furthermore, a SYBR Green quantitative real-time PCR assay was developed to detect duck MAPK1 expression. Following Riemerella anatipestifer infection by virulent strain Yb2, MAPK1 mRNA level increased more than 200-fold in the duck spleens, suggesting that increased duck MAPK1 expression can be used as an indicator of bacterial infection. Our results provide ground work to warrant further studies of the duck MAPK1 gene in bacterial pathogenesis.

  1. Molecular cloning of the duck mitogen-activated protein kinase 1 (MAPK1) gene and the development of a quantitative real-time PCR assay to detect its expression.

    PubMed

    Cao, Shoulin; Han, Xiangan; Ding, Chan; Wang, Shaohui; Tian, Mingxing; Wang, Xiaolan; Hou, Wanwan; Yue, Jiaping; Wang, Guijun; Yu, Shengqing

    2014-09-01

    Mitogen-activated protein kinase 1 (MAPK1) acts as an integration point for multiple biochemical signals, and is involved in a wide variety of biological processes such as cell proliferation and differentiation, transcription regulation, and development. Mitogen-activated protein kinase 1 plays an important role in inducing cell death in bacterial infections. In this study, the duck MAPK1 gene was cloned for the first time from the Cherry Valley duck. Sequence analysis showed that duck MAPK1 cDNA is 1,557 bp long, with an open reading frame of 1,107 bp. It encodes 368 amino acids, with 85.4, 84.5, and 97.3% homology with the human, mouse, and chicken MAPK1 gene, respectively. Furthermore, a SYBR Green quantitative real-time PCR assay was developed to detect duck MAPK1 expression. Following Riemerella anatipestifer infection by virulent strain Yb2, MAPK1 mRNA level increased more than 200-fold in the duck spleens, suggesting that increased duck MAPK1 expression can be used as an indicator of bacterial infection. Our results provide ground work to warrant further studies of the duck MAPK1 gene in bacterial pathogenesis. PMID:24974389

  2. Regulation of SREBPs by Sphingomyelin in Adipocytes via a Caveolin and Ras-ERK-MAPK-CREB Signaling Pathway.

    PubMed

    Makdissy, Nehman; Haddad, Katia; Mouawad, Charbel; Popa, Iuliana; Younsi, Mohamed; Valet, Philippe; Brunaud, Laurent; Ziegler, Olivier; Quilliot, Didier

    2015-01-01

    Sterol response element binding protein (SREBP) is a key transcription factor in insulin and glucose metabolism. We previously demonstrated that elevated levels of membrane sphingomyelin (SM) were related to peroxisome proliferator-activated receptor-γ (PPARγ), which is a known target gene of SREBP-1 in adipocytes. However, the role of SM in SREBP expression in adipocytes remains unknown. In human abdominal adipose tissue from obese women with various concentrations of fasting plasma insulin, SREBP-1 proteins decreased in parallel with increases in membrane SM levels. An inverse correlation was found between the membrane SM content and the levels of SREBP-1c/ERK/Ras/PPARγ/CREB proteins. For the first time, we demonstrate the effects of SM and its signaling pathway in 3T3-F442A adipocytes. These cells were enriched or unenriched with SM in a range of concentrations similar to those observed in obese subjects by adding exogenous natural SMs (having different acyl chain lengths) or by inhibiting neutral sphingomyelinase. SM accumulated in caveolae of the plasma membrane within 24 h and then in the intracellular space. SM enrichment decreased SREBP-1 through the inhibition of extracellular signal-regulated protein kinase (ERK) but not JNK or p38 mitogen-activated protein kinase (MAPK). Ras/Raf-1/MEK1/2 and KSR proteins, which are upstream mediators of ERK, were down-regulated, whereas SREBP-2/caveolin and cholesterol were up-regulated. In SM-unmodulated adipocytes treated with DL-1-Phenyl-2-Palmitoylamino-3-morpholino-1-propanol (PPMP), where the ceramide level increased, the expression levels of SREBPs and ERK were modulated in an opposite direction relative to the SM-enriched cells. SM inhibited the insulin-induced expression of SREBP-1. Rosiglitazone, which is an anti-diabetic agent and potent activator of PPARγ, reversed the effects of SM on SREBP-1, PPARγ and CREB. Taken together, these findings provide novel insights indicating that excess membrane SM might

  3. Sodium houttuyfonate inhibits inflammation by blocking the MAPKs/NF-κB signaling pathways in bovine endometrial epithelial cells.

    PubMed

    Zhu, Qi; Xu, Xiaolong; Liu, Xiaoxi; Lin, Jiabao; Kan, Yao; Zhong, Yougang; Liu, Fenghua; Xu, Jianqin

    2015-06-01

    Sodium houttuyfonate (SH) has traditionally been used for the therapy of inflammatory diseases. In this research, we tried to assess the anti-inflammatory effects of SH on LPS-induced bovine endometrial epithelial cell (bEEC) inflammation. SH cell toxicity was measured using the MTT and LDH assays, and inflammatory cytokine expression was assessed by ELISA, qRT-PCR and Western blotting. We demonstrated that SH was not cytotoxic to bEECs, and that it significantly decreased the LPS-induced mRNA and protein expression of tumor necrosis factor (TNF) α, interleukin (IL)-1β, IL-6 and IL-8. Furthermore, in LPS-induced bEECs, SH inhibited IκBα degradation and NF-κB p65 phosphorylation, and suppressed the phosphorylation of the mitogen-activated protein kinases (MAPKs), p38, c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK). In conclusion, we found that SH could effectively block the NF-κB-mediated signaling pathway and reduce the inflammatory process, thereby exerting a protective effect on bEECs. PMID:25935757

  4. Geniposide suppresses LPS-induced nitric oxide, PGE2 and inflammatory cytokine by downregulating NF-κB, MAPK and AP-1 signaling pathways in macrophages.

    PubMed

    Shi, Qinghai; Cao, Jinjun; Fang, Li; Zhao, Hongyan; Liu, Zhengxiang; Ran, Jihua; Zheng, Xinchuan; Li, Xiaoling; Zhou, Yu; Ge, Di; Zhang, Hongming; Wang, Li; Ran, Ying; Fu, Jianfeng

    2014-06-01

    Inflammatory responses are important to host immune reactions, but uncontrolled inflammatory mediators may aid in the pathogenesis of other inflammatory diseases. Geniposide, an iridoid glycoside found in the herb gardenia, is believed to have broad-spectrum anti-inflammatory effects in murine models but its mechanism of action is unclear. We investigated the action of this compound in murine macrophages stimulated by lipopolysaccharide (LPS), as the stimulation of macrophages by LPS is known to induce inflammatory reactions. We determined the effect of geniposide on LPS-induced production of the inflammatory mediators, nitric oxide (NO) and prostaglandin E2 (PGE2), the mRNA and protein expression of the NO and PGE2 synthases, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively, and the mRNA and protein expression of the inflammatory cytokine, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Furthermore, nuclear factor (NF)-κB, mitogen-activated protein kinase (MAPK) and activator protein (AP)-1 activity were assayed. To understand the action of geniposide on the NF-κB and MAPK pathways, we studied the effect of NF-κB and MAPK inhibitors on the LPS-induced production of NO, PGE2 and TNF-α. Our findings clearly showed that geniposide mainly exerts its anti-inflammatory effects by inhibiting the LPS-induced NF-κB, MAPK and AP-1 signaling pathways in macrophages, which subsequently reduces overexpression of the inducible enzymes iNOS and COX-2 and suppresses the expression and release of the inflammatory factors, TNF-α, IL-6, NO and PGE2. Thus, geniposide shows promise as a therapeutic agent in inflammatory diseases. PMID:24735815

  5. Signaling by p38 MAPK stimulates nuclear localization of the microprocessor component p68 for processing of selected primary microRNAs.

    PubMed

    Hong, Sungguan; Noh, Hyangsoon; Chen, Haoming; Padia, Ravi; Pan, Zhixing K; Su, Shi-Bing; Jing, Qing; Ding, Han-Fei; Huang, Shuang

    2013-03-12

    The importance of microRNAs (miRNAs) in biological and disease processes necessitates a better understanding of the mechanisms that regulate miRNA abundance. We showed that the activities of the mitogen-activated protein kinase (MAPK) p38 and its downstream effector kinase MAPK-activated protein kinase 2 (MK2) were necessary for the efficient processing of a subset of primary miRNAs (pri-miRNAs). Through yeast two-hybrid screening, we identified p68 (also known as DDX5), a key component of the Drosha complex that processes pri-miRNAs, as an MK2-interacting protein, and we found that MK2 phosphorylated p68 at Ser(197) in cells. In wild-type mouse embryonic fibroblasts (MEFs) treated with a p38 inhibitor or in MK2-deficient (MK2(-/-)) MEFs, expression of a phosphomimetic mutant p68 fully restored pri-miRNA processing, suggesting that MK2-mediated phosphorylation of p68 was essential for this process. We found that, whereas p68 was present in the nuclei of wild-type MEFs, it was found mostly in the cytoplasm of MK2(-/-) MEFs. Nuclear localization of p68 depended on MK2-mediated phosphorylation of Ser(197). In addition, inhibition of p38 MAPK promoted the growth of wild-type MEFs and breast cancer MCF7 cells by enhancing the abundance of c-Myc through suppression of the biogenesis of the miRNA miR-145, which targets c-Myc. Because pri-miRNA processing occurs in the nucleus, our findings suggest that the p38 MAPK-MK2 signaling pathway promotes miRNA biogenesis by facilitating the nuclear localization of p68.

  6. Geniposide suppresses LPS-induced nitric oxide, PGE2 and inflammatory cytokine by downregulating NF-κB, MAPK and AP-1 signaling pathways in macrophages.

    PubMed

    Shi, Qinghai; Cao, Jinjun; Fang, Li; Zhao, Hongyan; Liu, Zhengxiang; Ran, Jihua; Zheng, Xinchuan; Li, Xiaoling; Zhou, Yu; Ge, Di; Zhang, Hongming; Wang, Li; Ran, Ying; Fu, Jianfeng

    2014-06-01

    Inflammatory responses are important to host immune reactions, but uncontrolled inflammatory mediators may aid in the pathogenesis of other inflammatory diseases. Geniposide, an iridoid glycoside found in the herb gardenia, is believed to have broad-spectrum anti-inflammatory effects in murine models but its mechanism of action is unclear. We investigated the action of this compound in murine macrophages stimulated by lipopolysaccharide (LPS), as the stimulation of macrophages by LPS is known to induce inflammatory reactions. We determined the effect of geniposide on LPS-induced production of the inflammatory mediators, nitric oxide (NO) and prostaglandin E2 (PGE2), the mRNA and protein expression of the NO and PGE2 synthases, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively, and the mRNA and protein expression of the inflammatory cytokine, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Furthermore, nuclear factor (NF)-κB, mitogen-activated protein kinase (MAPK) and activator protein (AP)-1 activity were assayed. To understand the action of geniposide on the NF-κB and MAPK pathways, we studied the effect of NF-κB and MAPK inhibitors on the LPS-induced production of NO, PGE2 and TNF-α. Our findings clearly showed that geniposide mainly exerts its anti-inflammatory effects by inhibiting the LPS-induced NF-κB, MAPK and AP-1 signaling pathways in macrophages, which subsequently reduces overexpression of the inducible enzymes iNOS and COX-2 and suppresses the expression and release of the inflammatory factors, TNF-α, IL-6, NO and PGE2. Thus, geniposide shows promise as a therapeutic agent in inflammatory diseases.

  7. Sphingosine kinase inhibitor suppresses IL-18-induced interferon-gamma production through inhibition of p38 MAPK activation in human NK cells

    SciTech Connect

    Cheon, Soyoung; Song, Seok Bean; Jung, Minkyung; Park, Yoorim; Bang, Jung-Wook; Kim, Tae Sung; Park, Hyunjeong; Kim, Cherl-hyun; Yang, Yool-hee; Bang, Sa Ik; Cho, Daeho

    2008-09-12

    Natural killer (NK) cells play an important role in the innate immune response. Interleukin-18 (IL-18) is a well-known interferon-gamma (IFN-{gamma} inducing factor, which stimulates immune response in NK and T cells. Sphingosine kinase (SPHK) catalyzes the formation of sphingosine 1-phosphate (S1P), which acts as a second messenger to function as an anti-apoptotic factor and proliferation stimulator of immune cells. In this study, to elucidate whether SPHK is involved in IL-18-induced IFN-{gamma} production, we measured IL-18-induced IFN-{gamma} production after pre-treatment with SPHK inhibitor (SKI) in NK-92MI cells. We found that IL-18-induced IFN-{gamma} expression was blocked by SKI pre-treatment in both mRNA and protein levels. In addition, the increased IFN-{gamma} production by stimulation with IL-18 is mediated through both SPHK and p38 MAPK. To determine the upstream signals of SKI and p38 MAPK in IL-18-induced IFN-{gamma} production, phosphorylation levels of p38 MAPK was measured after SKI pre-treatment. As a result, inhibition of SPHK by SKI blocked phosphorylation of p38 MAPK, showing that SPHK activation by IL-18 is an upstream signal of p38 MAPK activation. Inhibition of SPHK by SKI also inhibited IL-18-induced IFN-{gamma} production in human primary NK cells. In conclusion, SPHK activation is an essential factor for IL-18-induced IFN-{gamma} production via p38 MAPK.

  8. Astaxanthin Inhibits Acetaldehyde-Induced Cytotoxicity in SH-SY5Y Cells by Modulating Akt/CREB and p38MAPK/ERK Signaling Pathways

    PubMed Central

    Yan, Tingting; Zhao, Yan; Zhang, Xia; Lin, Xiaotong

    2016-01-01

    Excessive alcohol consumption can lead to brain tissue damage and cognitive dysfunction. Acetaldehyde, the most toxic metabolite of ethanol, mediates the brain tissue damage and cognitive dysfunction induced by chronic excessive alcohol consumption. In this study, the effect of astaxanthin, a marine bioactive compound, on acetaldehyde-induced cytotoxicity was investigated in SH-SY5Y cells. It was found that astaxanthin protected cells from apoptosis by ameliorating the effect of acetaldehyde on the expression of Bcl-2 family proteins, preventing the reduction of anti-apoptotic protein Bcl-2 and the increase of pro-apoptotic protein Bak induced by acetaldehyde. Further analyses showed that astaxanthin treatment inhibited acetaldehyde-induced reduction of the levels of activated Akt and cyclic AMP-responsive element binding protein (CREB). Astaxanthin treatment also prevented acetaldehyde-induced increase of the level of activated p38 mitogen-activated protein kinase (MAPK) and decrease of the level of activated extracellular signal-regulated kinases (ERKs). Activation of Akt/CREB pathway promotes cell survival and is involved in the upregulation of Bcl-2 gene. P38MAPK plays a critical role in apoptotic events while ERKs mediates the inhibition of apoptosis. Thus, astaxanthin may inhibit acetaldehyde-induced apoptosis through promoting the activation of Akt/CREB and ERKs and blocking the activation of p38MAPK. In addition, astaxanthin treatment suppressed the oxidative stress induced by acetaldehyde and restored the antioxidative capacity of SH-SY5Y cells. Therefore, astaxanthin may protect cells against acetaldehyde-induced cytotoxicity through maintaining redox balance and modulating apoptotic and survival signals. The results suggest that astaxanthin treatment may be beneficial for preventing neurotoxicity associated with acetaldehyde and excessive alcohol consumption. PMID:26978376

  9. Endoplasmic reticulum reorganizations and Ca2+ signaling in maturing and fertilized oocytes of marine protostome worms: the roles of MAPKs and MPF.

    PubMed

    Stricker, Stephen A; Smythe, Toni L

    2003-07-01

    Before a proper Ca(2+) response is produced at fertilization, oocytes typically undergo a maturation process during which their endoplasmic reticulum (ER) is restructured. In marine protostome worms belonging to the phylum Nemertea, the ER of maturing oocytes forms numerous distinct clusters that are about 5 micro m in diameter. After fertilization, mature oocytes with such aggregates generate a normal series of Ca(2+) oscillations and eventually disassemble their ER clusters at around the time that the oscillations cease. Immature oocytes, however, lack prominent ER clusters and fail to exhibit repetitive Ca(2+) oscillations upon insemination, collectively suggesting that cell cycle-related changes in ER structure may play a role in Ca(2+) signaling. To assess the effects of meiotic regulators on the morphology of the ER and the type of Ca(2+) response that is produced at fertilization, nemertean oocytes were treated with pharmacological modulators of mitogen-activated protein kinases (MAPKs) or maturation-promoting factor (MPF) prior to confocal microscopic analyses. Based on such imaging studies and correlative assays of kinase activities, MAPKs of the ERK1/2 type (extracellular signal regulated kinases 1/2) do not seem to be essential for either structural reorganizations of the ER or repetitive Ca(2+) signaling at fertilization. Conversely, MPF levels appear to modulate both ER structure and the capacity to produce normal Ca(2+) oscillations. The significance of these findings is discussed with respect to other reports on ER structure, MPF cycling and Ca(2+) signaling in oocytes of deuterostome animals.

  10. Glycosphingolipid synthesis inhibitor represses cytokine-induced activation of the Ras-MAPK pathway in embryonic neural precursor cells.

    PubMed

    Yanagisawa, Makoto; Nakamura, Kazuo; Taga, Tetsuya

    2005-09-01

    Neuronal and glial cells in the central nervous system are generated from common neural precursor cells during development. To evaluate the functions of glycosphingolipids (GSLs) in neural precursor cells, neuroepithelial cells (NECs) were prepared from mouse embryos (E14.5), and the effects of an inhibitor of glucosylceramide synthesis, D-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), on NECs was investigated. In PDMP-treated NECs, the expression of GD3, a major ganglioside of NECs, disappeared. We found that basic fibroblast growth factor (bFGF)-induced proliferation and extracellular signal-regulated kinase (ERK) activation were repressed in PDMP-treated NECs. Leukemia inhibitory factor (LIF)-induced ERK activation was also abolished in PDMP-treated NECs, suggesting that PDMP specifically represses the Ras-MAPK pathway. bFGF-induced activation of the Ras-MAPK pathway in NECs is dependent on GSL-enriched microdomains, lipid rafts. The organization of lipid rafts and the distribution of Ras and Grb2-SOS in the microdomains were not affected. However, Ras activation was repressed in PDMP-treated NECs. In PDMP-treated NECs, some neuronal genes were up-regulated and glial genes were down-regulated. These results suggest that GSLs might be involved in the proliferation, survival, signal transduction and differentiation of NECs.

  11. Adiponectin stimulates human osteoblasts proliferation and differentiation via the MAPK signaling pathway

    SciTech Connect

    Luo Xianghang; Guo Lijuan; Yuan Lingqing; Xie Hui; Zhou Houde; Wu Xianping; Liao Eryuan . E-mail: eyliao1207@21cn.com

    2005-09-10

    Adipocytes can highly and specifically express adiponectin, and the adiponectin receptor (AdipoR) has been detected in bone-forming cells. The present study was undertaken to investigate the action of adiponectin on osteoblast proliferation and differentiation. AdipoR1 protein was detected in human osteoblasts. Adiponectin promoted osteoblast proliferation and resulted in a dose- and time-dependent increase in alkaline phosphatase (ALP) activity, osteocalcin and type I collagen production, and an increase in mineralized matrix. Suppression of AdipoR1 with small-interfering RNA (siRNA) abolished the adiponectin-induced cell proliferation and ALP expression. Adiponectin induces activation of p38 mitogen-activated protein kinase (MAPK) and c-jun N-terminal Kinase (JNK), but not ERK1/2 in osteoblasts, and these effects were blocked by suppression of AdipoR1 with siRNA. Furthermore, pretreatment of osteoblasts with the JNK inhibitor SP600125 abolished the adiponectin-induced cell proliferation. p38 inhibitor SB203580 blocked the adiponectin-induced ALP activity. These data indicate that adiponectin induces human osteoblast proliferation and differentiation, and the proliferation response is mediated by the AdipoR/JNK pathway, while the differentiation response is mediated via the AdipoR/p38 pathway. These findings suggest that osteoblasts are the direct targets of adiponectin.

  12. Activation of MAPK Is Necessary for Long-Term Memory Consolidation Following Food-Reward Conditioning

    ERIC Educational Resources Information Center

    Ribeiro, Maria J.; Schofield, Michael G.; Kemenes, Ildiko; O'Shea, Michael; Kemenes, Gyorgy; Benjamin, Paul R.

    2005-01-01

    Although an important role for the mitogen-activated protein kinase (MAPK) has been established for memory consolidation in a variety of learning paradigms, it is not known if this pathway is also involved in appetitive classical conditioning. We address this question by using a single-trial food-reward conditioning paradigm in the freshwater…

  13. Cyclooxygenase-2 induced β1-integrin expression in NSCLC and promoted cell invasion via the EP1/MAPK/E2F-1/FoxC2 signal pathway

    PubMed Central

    Pan, Jinshun; Yang, Qinyi; Shao, Jiaofang; Zhang, Li; Ma, Juan; Wang, Yipin; Jiang, Bing-Hua; Leng, Jing; Bai, Xiaoming

    2016-01-01

    Cyclooxygenase-2 (COX-2) has been implicated in cell invasion in non-small-cell lung cancer (NSCLC). However, the mechanism is unclear. The present study investigated the effect of COX-2 on β1-integrin expression and cell invasion in NSCLC. COX-2 and β1-integrin were co-expressed in NSCLC tissues. COX-2 overexpression or Prostaglandin E2 (PGE2) treatment increased β1-integrin expression in NSCLC cell lines. β1-integrin silencing suppressed COX-2-mediated tumour growth and cancer cell invasion in vivo and in vitro. Prostaglandin E Receptor EP1 transfection or treatment with EP1 agonist mimicked the effect of PGE2 treatment. EP1 siRNA blocked PGE2-mediated β1-integrin expression. EP1 agonist treatment promoted Erk1/2, p38 phosphorylation and E2F-1 expression. MEK1/2 and p38 inhibitors suppressed EP1-mediated β1-integrin expression. E2F-1 silencing suppressed EP1-mediated FoxC2 and β1-integrin upregulation. ChIP and Luciferase Reporter assays identified that EP1 agonist treatment induced E2F-1 binding to FoxC2 promotor directly and improved FoxC2 transcription. FoxC2 siRNA suppressed β1-integrin expression and EP1-mediated cell invasion. Immunohistochemistry showed E2F-1, FoxC2, and EP1R were all highly expressed in the NSCLC cases. This study suggested that COX-2 upregulates β1-integrin expression and cell invasion in NSCLC by activating the MAPK/E2F-1 signalling pathway. Targeting the COX-2/EP1/PKC/MAPK/E2F-1/FoxC2/β1-integrin pathway might represent a new therapeutic strategy for the prevention and treatment of this cancer. PMID:27654511

  14. Mammary gland specific expression of Brk/PTK6 promotes delayed involution and tumor formation associated with activation of p38 MAPK

    PubMed Central

    2011-01-01

    Introduction Protein tyrosine kinases (PTKs) are frequently overexpressed and/or activated in human malignancies, and regulate cancer cell proliferation, cellular survival, and migration. As such, they have become promising molecular targets for new therapies. The non-receptor PTK termed breast tumor kinase (Brk/PTK6) is overexpressed in approximately 86% of human breast tumors. The role of Brk in breast pathology is unclear. Methods We expressed a WAP-driven Brk/PTK6 transgene in FVB/n mice, and analyzed mammary glands from wild-type (wt) and transgenic mice after forced weaning. Western blotting and immunohistochemistry (IHC) studies were conducted to visualize markers of mammary gland involution, cell proliferation and apoptosis, as well as Brk, STAT3, and activated p38 mitogen-activated protein kinase (MAPK) in mammary tissues and tumors from WAP-Brk mice. Human (HMEC) or mouse (HC11) mammary epithelial cells were stably or transiently transfected with Brk cDNA to assay p38 MAPK signaling and cell survival in suspension or in response to chemotherapeutic agents. Results Brk-transgenic dams exhibited delayed mammary gland involution and aged mice developed infrequent tumors with reduced latency relative to wt mice. Consistent with delayed involution, mammary glands of transgenic animals displayed decreased STAT3 phosphorylation, a marker of early-stage involution. Notably, p38 MAPK, a pro-survival signaling mediator downstream of Brk, was activated in mammary glands of Brk transgenic relative to wt mice. Brk-dependent signaling to p38 MAPK was recapitulated by Brk overexpression in the HC11 murine mammary epithelial cell (MEC) line and human MEC, while Brk knock-down in breast cancer cells blocked EGF-stimulated p38 signaling. Additionally, human or mouse MECs expressing Brk exhibited increased anchorage-independent survival and resistance to doxorubicin. Finally, breast tumor biopsies were subjected to IHC analysis for co-expression of Brk and phospho-p38 MAPK

  15. Molecular cloning and expression analysis of mulberry MAPK gene family.

    PubMed

    Wei, Congjin; Liu, Xueqin; Long, Dingpei; Guo, Qing; Fang, Yuan; Bian, Chenkai; Zhang, Dayan; Zeng, Qiwei; Xiang, Zhonghuai; Zhao, Aichun

    2014-04-01

    Mitogen-activated protein kinase (MAPK) cascades play an important role in regulating various biotic and abiotic stresses in plants. Although MAPKs have been identified and characterized in a few model plants, there is little information available for mulberry Morus sp. L., one of the most ecologically and economically important perennial trees. This study identified 47 mulberry Morus notabilis MAPK (MnMAPK) family genes: 32 MnMAPKKK, five MnMAPKK and ten MnMAPK genes, and cloned ten MnMAPK cDNA genes based on a genome-wide analysis of the morus genome database. Comparative analysis with MAPK gene families from other plants suggested that MnMAPKs could be divided into five subfamilies (groups A, B, C, D and E) and they could have similar functions in response to abiotic and biotic stresses. MnMAPK gene expression analysis of different stresses (high/low temperature, salt and drought) and signal molecules (ABA, SA, H2O2 and methyl jasmonate (MeJA)) revealed that all ten MnMAPK genes responded to high/low temperature, salt and drought stresses, and that nine of the ten MnMAPKs (MnMAPK7 excepted) could be induced by ABA, SA, H2O2 and MeJA, which suggested that MnMAPKs may play pivotal roles in signal transduction pathways. Our results indicated that almost all of the MnMAPKs may be involved in environmental stress and defense responses, which provides the basis for further characterization of the physiological functions of MnMAPKs.

  16. Histamine H3-Receptor Signaling in Cardiac Sympathetic Nerves: Identification of a Novel MAPK-PLA2-COX-PGE2-EP3R Pathway

    PubMed Central

    Levi, Roberto; Seyedi, Nahid; Schaefer, Ulrich; Estephan, Rima; Mackins, Christina J.; Tyler, Eleanor; Silver, Randi B.

    2007-01-01

    We tested the hypothesis that the histamine H3-receptor (H3R)-mediated attenuation of norepinephrine (NE) exocytosis from cardiac sympathetic nerves results not only from a Gαi-mediated inhibition of the adenylyl cyclase-cAMP-PKA pathway, but also from a Gβγi-mediated activation of the MAPK-PLA2 cascade, culminating in formation of an arachidonate metabolite with anti-exocytotic characteristics (e.g., PGE2). We report in Langendorff-perfused guinea-pig hearts and isolated sympathetic nerve endings (cardiac synaptosomes), H3R-mediated attenuation of K+-induced NE exocytosis was prevented by MAPK and PLA2 inhibitors, and by cyclooxygenase and EP3-receptor (EP3R) antagonists. Moreover, H3R activation resulted in MAPK phosphorylation in H3R-transfected SH-SY5Y neuroblastoma cells, and in PLA2 activation and PGE2 production in cardiac synaptosomes; H3R-induced MAPK phosphorylation was prevented by an anti-βγ peptide. Synergism between H3R and EP3R agonists (i.e., imetit and sulprostone, respectively) suggested PGE2 may be a downstream effector of the anti-exocytotic effect of H3R activation. Furthermore, the anti-exocytotic effect of imetit and sulprostone was potentiated by the N-type Ca2+-channel antagonist ω-conotoxin GVIA, and prevented by an anti-Gβγ peptide. Our findings suggest an EP3R Gβγi-induced decrease in Ca2+ influx through N-type Ca2+-channels is involved in PGE2/EP3R-mediated attenuation of NE exocytosis elicited by H3R activation. Conceivably, activation of the Gβγi subunit of H3R and EP3R may also inhibit Ca2+ entry directly, independent of MAPK intervention. As heart failure, myocardial ischemia and arrhythmic dysfunction are associated with excessive local NE release, attenuation of NE release by H3R activation is cardioprotective. Thus, the uncovering of a novel H3R signaling pathway may ultimately bear therapeutic significance in hyper-adrenergic states. PMID:17266940

  17. The crosstalk between α-irradiated Beas-2B cells and its bystander U937 cells through MAPK and NF-κB signaling pathways.

    PubMed

    Fu, Jiamei; Yuan, Dexiao; Xiao, Linlin; Tu, Wenzhi; Dong, Chen; Liu, Weili; Shao, Chunlin

    2016-01-01

    Although accumulated evidence suggests that α-particle irradiation induced bystander effect may relevant to lung injury and cancer risk assessment, the exact mechanisms are not yet elucidated. In the present study, a cell co-culture system was used to investigate the interaction between α-particle irradiated human bronchial epithelial cells (Beas-2B) and its bystander macrophage U937 cells. It was found that the cell co-culture amplified the detrimental effects of α-irradiation including cell viability decrease and apoptosis promotion on both irradiated cells and bystander cells in a feedback loop which was closely relevant to the activation of MAPK and NF-κB pathways in the bystander U937 cells. When these two pathways in U937 cells were disturbed by special pharmacological inhibitors before cell co-culture, it was found that a NF-κB inhibitor of BAY 11-7082 further enhanced the proliferation inhibition and apoptosis induction in bystander U937 cells, but MAPK inhibitors of SP600125 and SB203580 protected cells from viability loss and apoptosis and U0126 presented more beneficial effect on cell protection. For α-irradiated epithelial cells, the activation of NF-κB and MAPK pathways in U937 cells participated in detrimental cellular responses since the above inhibitors could largely attenuate cell viability loss and apoptosis of irradiated cells. Our results demonstrated that there are bilateral bystander responses between irradiated lung epithelial cells and macrophages through MAPK and NF-κB signaling pathways, which accounts for the enhancement of α-irradiation induced damage. PMID:26613333

  18. Influence of aging and caloric restriction on activation of Ras/MAPK, calcineurin, and CaMK-IV activities in rat T cells.

    PubMed

    Pahlavani, M A; Vargas, D M

    2000-02-01

    The signaling cascade mediated by Ras (p21ras) and MAPK (mitogen-activated protein kinase) and calcium/calmodulin regulating enzymes, calcineurin (CaN) and CaMK-IV, are considered to be essential for T-cell growth and function. In the present study, the effect of aging and caloric restriction (CR) on the induction of Ras and MAPK activation by concanavalin A (ConA) was studied. Splenic T cells were isolated from young (4-6 months) and old (22-24 months) rats that had free access to food (control group), and from caloric restricted old (22-24 months) rats that beginning at 6 weeks of age were fed 60%(40% caloric restriction) of the diet consumed by the control rats. We found that the induction of Ras activity in T cells isolated from control old rats was lower (P<0.001) than that in control young rats. However, the levels of Ras activity in T cells isolated from CR old rats were similar to the levels in the age-matched control rats. The induction of MAPK activity in T cells isolated from control old rats and CR old rats was significantly less than in T cells isolated from control young rats, and caloric restriction significantly (P<0.05) reduced the age-related decline in MAPK activation. We also measured the induction of CaN and CaMK-IV activities by ConA in T cells from control young and old and CR old rats. The induction of both CaN and CaMK-IV activity decreased with age. Caloric restriction significantly (P<0.05) reduced the age-related decline in CaN activity, but had no significant effect on CaMK-IV activity. The changes in Ras/MAPK activation and in CaN and CaMK-IV activity with age or with CR were not associated with alterations in their corresponding protein levels. Thus, caloric restriction has a differential effect on the activation of the upstream signaling molecules that are altered with age.

  19. All-Trans Retinoic Acid Inhibits Human Colorectal Cancer Cells RKO Migration via Downregulating Myosin Light Chain Kinase Expression through MAPK Signaling Pathway.

    PubMed

    Zuo, Li; Yang, Xiaoping; Lu, Man; Hu, Ruolei; Zhu, Huaqing; Zhang, Sumei; Zhou, Qing; Chen, Feihu; Gui, Shuyu; Wang, Yuan

    2016-10-01

    All-trans-retinoic acid (ATRA) inhibits the invasive and metastatic potentials of various cancer cells. However, the underlying mechanism is unclear. Here, we demonstrate that ATRA inhibited colorectal cancer cells RKO (human colon adenocarcinoma cell) migration by downregulating cell movement and increasing cell adhesion. ATRA inhibited the expression and activation of myosin light chain kinase (MLCK) in RKO cells, while the expression level of MLC phosphatase (MLCP) had no change in RKO cells treated with or without ATRA. The expression and activity of MLC was also inhibited in RKO cells exposed to ATRA. Intriguingly, ATRA increased the expression of occludin messenger RNA (mRNA) and protein and its localization on cell membrane. However, ATRA did not change the expression of zonula occludens 1 (ZO-1), but increased the accumulation of ZO-1 on RKO cells membrane. ML-7, an inhibitor of MLCK, significantly inhibited RKO cell migration. Furthermore, knockdown of endogenous MLCK expression inhibited RKO migration. Mechanistically, we showed that MAPK-specific inhibitor PD98059 enhanced the inhibitory effect of ATRA on RKO migration. In contrast, phorbol 12-myristate 13-acetate (PMA) attenuated the effects of ATRA in RKO cells. Moreover, knocking down endogenous extracellular signal-regulated kinase (ERK) expression inhibited MLCK expression in the RKO cells. In conclusion, ATRA inhibits RKO migration by reducing MLCK expression via extracellular signal-regulated kinase 1/Mitogen-activated protein kinase (ERK1/MAPK) signaling pathway. PMID:27564600

  20. The mitogen-activated protein kinase (MAPK) cascade controls phosphatase and tensin homolog (PTEN) expression through multiple mechanisms.

    PubMed

    Ciuffreda, Ludovica; Di Sanza, Cristina; Cesta Incani, Ursula; Eramo, Adriana; Desideri, Marianna; Biagioni, Francesca; Passeri, Daniela; Falcone, Italia; Sette, Giovanni; Bergamo, Paola; Anichini, Andrea; Sabapathy, Kanaga; McCubrey, James A; Ricciardi, Maria Rosaria; Tafuri, Agostino; Blandino, Giovanni; Orlandi, Augusto; De Maria, Ruggero; Cognetti, Francesco; Del Bufalo, Donatella; Milella, Michele

    2012-06-01

    The mitogen-activated protein kinase (MAPK) and PI3K pathways are regulated by extensive crosstalk, occurring at different levels. In tumors, transactivation of the alternate pathway is a frequent "escape" mechanism, suggesting that combined inhibition of both pathways may achieve synergistic antitumor activity. Here we show that, in the M14 melanoma model, simultaneous inhibition of both MEK and mammalian target of rapamycin (mTOR) achieves synergistic effects at suboptimal concentrations, but becomes frankly antagonistic in the presence of relatively high concentrations of MEK inhibitors. This observation led to the identification of a novel crosstalk mechanism, by which either pharmacologic or genetic inhibition of constitutive MEK signaling restores phosphatase and tensin homolog (PTEN) expression, both in vitro and in vivo, and inhibits downstream signaling through AKT and mTOR, thus bypassing the need for double pathway blockade. This appears to be a general regulatory mechanism and is mediated by multiple mechanisms, such as MAPK-dependent c-Jun and miR-25 regulation. Finally, PTEN upregulation appears to be a major effector of MEK inhibitors' antitumor activity, as cancer cells in which PTEN is inactivated are consistently more resistant to the growth inhibitory and anti-angiogenic effects of MEK blockade. PMID:22215152

  1. Activation of HuR downstream of p38 MAPK promotes cardiomyocyte hypertrophy.

    PubMed

    Slone, Samuel; Anthony, Sarah R; Wu, Xiaoqing; Benoit, Joshua B; Aube, Jeffrey; Xu, Liang; Tranter, Michael

    2016-11-01

    The RNA binding protein Human antigen R (HuR) interacts with specific AU-rich domains in target mRNAs and is highly expressed in many cell types, including cardiomyocytes. However, the role of HuR in cardiac physiology is largely unknown. Our results show that HuR undergoes cytoplasmic translocation, indicative of its activation, in hypertrophic cardiac myocytes. Specifically, HuR cytoplasmic translocation is significantly increased in NRVMs (neonatal rat ventricular myocytes) following treatment with phenylephrine or angiotensin II, agonists of two independent Gαq-coupled GPCRs known to induce hypertrophy. This Gq-mediated HuR activation is dependent on p38 MAP kinase, but not canonical Gq-PKC signaling. Furthermore, we show that HuR activation is necessary for Gq-mediated hypertrophic growth of NRVMs as siRNA-mediated knockdown of HuR inhibits hypertrophy as measured by cell size and expression of ANF (atrial natriuretic factor). Additionally, HuR overexpression is sufficient to induce hypertrophic cell growth. To decipher the downstream mechanisms by which HuR translocation promotes cardiomyocyte hypertrophy, we assessed the role of HuR in the transcriptional activity of NFAT (nuclear factor of activated T cells), the activation of which is a hallmark of cardiac hypertrophy. Using an NFAT-luciferase reporter assay, we show an acute inhibition of NFAT transcriptional activity following pharmacological inhibition of HuR. In conclusion, our results identify HuR as a novel mediator of cardiac hypertrophy downstream of the Gq-p38 MAPK pathway, and suggest modulation of NFAT activity as a potential mechanism.

  2. Fisetin Inhibits Human Melanoma Cell Invasion through Promotion of Mesenchymal to Epithelial Transition and by Targeting MAPK and NFκB Signaling Pathways

    PubMed Central

    Pal, Harish Chandra; Sharma, Samriti; Strickland, Leah Ray; Katiyar, Santosh K.; Ballestas, Mary E.; Athar, Mohammad; Elmets, Craig A.; Afaq, Farrukh

    2014-01-01

    Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK) signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60–70% of malignant melanomas. The BRAF-MEK-ERK (MAPK) pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5–20 µM) resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059) or of NFκB (caffeic acid phenethyl ester) also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin) and an increase in epithelial markers (E-cadherin and desmoglein). Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that fisetin

  3. Fisetin inhibits human melanoma cell invasion through promotion of mesenchymal to epithelial transition and by targeting MAPK and NFκB signaling pathways.

    PubMed

    Pal, Harish Chandra; Sharma, Samriti; Strickland, Leah Ray; Katiyar, Santosh K; Ballestas, Mary E; Athar, Mohammad; Elmets, Craig A; Afaq, Farrukh

    2014-01-01

    Malignant melanoma is responsible for approximately 75% of skin cancer-related deaths. BRAF plays an important role in regulating the mitogen-activated protein kinase (MAPK) signaling cascade in melanoma with activating mutations in the serine/threonine kinase BRAF occurring in 60-70% of malignant melanomas. The BRAF-MEK-ERK (MAPK) pathway is a key regulator of melanoma cell invasion. In addition, activation of NFκB via the MAPK pathway is regulated through MEK-induced activation of IKK. These pathways are potential targets for prevention and treatment of melanoma. In this study, we investigated the effect of fisetin, a phytochemical present in fruits and vegetables, on melanoma cell invasion and epithelial-mesenchymal transition, and delineated the underlying molecular mechanism. Treatment of multiple human malignant melanoma cell lines with fisetin (5-20 µM) resulted in inhibition of cell invasion. BRAF mutated melanoma cells were more sensitive to fisetin treatment, and this was associated with a decrease in the phosphorylation of MEK1/2 and ERK1/2. In addition, fisetin inhibited the activation of IKK leading to a reduction in the activation of the NFκB signaling pathway. Treatment of cells with an inhibitor of MEK1/2 (PD98059) or of NFκB (caffeic acid phenethyl ester) also reduced melanoma cell invasion. Furthermore, treatment of fisetin promoted mesenchymal to epithelial transition in melanoma cells, which was associated with a decrease in mesenchymal markers (N-cadherin, vimentin, snail and fibronectin) and an increase in epithelial markers (E-cadherin and desmoglein). Employing three dimensional skin equivalents consisting of A375 cells admixed with normal human keratinocytes embedded onto a collagen-constricted fibroblast matrix, we found that treatment of fisetin reduced the invasive potential of melanoma cells into the dermis and increased the expression of E-cadherin with a concomitant decrease in vimentin. These results indicate that fisetin

  4. Spatio-temporal dynamics of a cell signal pathway with negative feedbacks: the MAPK/ERK pathway.

    PubMed

    Maya-Bernal, José Luis; Ramírez-Santiago, Guillermo

    2016-03-01

    We studied the spatio-temporal dynamics of a cell signal cascade with negative feedback that quantitatively emulates the regulative process that occurs in the Mitogen Activated Protein Kinase/Extracellular Regulated Kinase (MAPK/ERK) pathway. The model consists of a set of six coupled reaction-diffusion equations that describes the dynamics of the six-module pathway. In the basic module the active form of the protein transmits the signal to the next pathway’s module. As suggested by experiments, the model considers that the fifth module's kinase down-regulates the first and third modules. The feedback parameter is defined as, μ(r)( j)= k(kin)5/k(kin)(j), (j = 1, 3). We analysed the pathway's dynamics for μ(r)( j) = 0.10, 1.0, and 10 in the kinetic regimes: i) saturation of both kinases and phosphatases, ii) saturation of the phosphatases and iii) saturation of the kinases. For a regulated pathway the Total Activated Protein Profiles (TAPPs) as a function of time develop a maximum during the transient stage in the three kinetic regimes. These maxima become higher and their positions shift to longer times downstream. This scenario also applies to the TAPP's regulatory kinase that sums up its inhibitory action to that of the phosphatases leading to a maximum. Nevertheless, when μ(r)(j)= 1.0 , the TAPPs develop two maxima, with the second maximum being almost imperceptible. These results are in qualitative agreement with experimental data obtained from NIH 3T3 mouse fibroblasts. In addition, analyses of the stationary states as a function of position indicate that in the kinetic regime i) which is of physiological interest, signal transduction occurs with a relatively large propagation length for the three values of the regulative parameter. However, for μ(r)(j)= 0.10 , the sixth module concentration profile is transmitted with approximately 45% of its full value. The results obtained for μ(r)(j) = 10 , indicate that the first five concentration profiles are

  5. Inactivation of the MAPK signaling pathway by Listeria monocytogenes infection promotes trophoblast giant cell death

    PubMed Central

    Hashino, Masanori; Tachibana, Masato; Nishida, Takashi; Hara, Hideki; Tsuchiya, Kohsuke; Mitsuyama, Masao; Watanabe, Kenta; Shimizu, Takashi; Watarai, Masahisa

    2015-01-01

    Listeria monocytogenes has a well-characterized ability to cross the placental barrier, resulting in spontaneous abortion and fetal infections. However, the mechanisms resulting in infection-associated abortion are not fully understood. In this study, we demonstrate that the dephosphorylation of MAPK family proteins caused by L. monocytogenes infection of trophoblast giant (TG) cells, which are placental immune cells, contributes to infectious abortion. Dephosphorylation of c-Jun, p38, and ERK1/2 was observed in infected TG cells, causing the downregulation of cytoprotective heme oxygenase (HO)-1. Blocking the dephosphorylation of proteins, including MAPK family proteins, inhibited the decrease in HO-1 expression. Treatment with MAPK inhibitors inhibited bacterial internalization into TG cells. Moreover, Toll-like receptor 2 involved in the expression of MAPK family proteins. Infection with a listeriolysin O-deleted mutant impaired dephosphorylation of MAPK family proteins in TG cells and did not induce infectious abortion in a mouse model. These results suggest that inactivation of the MAPK pathway by L. monocytogenes induces TG cell death and causes infectious abortion. PMID:26528279

  6. Oxidized High-Density Lipoprotein Impairs Endothelial Progenitor Cells' Function by Activation of CD36-MAPK-TSP-1 Pathways

    PubMed Central

    Wu, Jianxiang; He, Zhiqing; Gao, Xiang; Wu, Feng; Ding, Ru; Ren, Yusheng; Jiang, Qijun; Fan, Min

    2015-01-01

    Abstract Aims: High-density lipoprotein (HDL) levels inversely correlate with cardiovascular events due to the protective effects on vascular wall and stem cells, which are susceptible to oxidative modifications and then lead to potential pro-atherosclerotic effects. We proposed that oxidized HDL (ox-HDL) might lead to endothelial progenitor cells (EPCs) dysfunction and investigated underlying mechanisms. Results: ox-HDL was shown to increase apoptosis and intracellular reactive oxygen species levels, but to reduce migration, angiogenesis, and cholesterol efflux of EPCs in a dose-dependent manner. p38 mitogen-activated protein kinase (MAPK) and NF-κB were activated after ox-HDL stimulation, which also upregulated thrombospondin-1 (TSP-1) expression without affecting vascular endothelial growth factor. Effects caused by ox-HDL could be significantly attenuated by pretreatment with short hairpin RNA-mediated CD36 knockdown or probucol. Data of in vivo experiments and the inverse correlation of ox-HDL and circulating EPC numbers among patients with coronary artery diseases (CAD) or CAD and type 2 diabetes also supported it. Meanwhile, HDL separated from such patients could significantly increase cultured EPC's caspase 3 activity, further supporting our proposal. Innovation: This is the most complete study to date of how ox-HDL would impair EPCs function, which was involved with activation of CD36-p38 MAPK-TSP-1 pathways and proved by not only the inverse relationship between ox-HDL and circulating EPCs in clinic but also pro-apoptotic effects of HDL separated from patients' serum. Conclusion: Activation of CD36-p38 MAPK-TSP-1 pathways contributes to the pathological effects of ox-HDL on EPCs' dysfunction, which might be one of the potential etiological factors responsible for the disturbed neovascularization in chronic ischemic disease. Antioxid. Redox Signal. 22, 308–324. PMID:25313537

  7. Baicalein inhibits MMP-2 expression in human ovarian cancer cells by suppressing the p38 MAPK-dependent NF-κB signaling pathway.

    PubMed

    Yan, Hao; Xin, Shaobin; Wang, Hui; Ma, Jing; Zhang, Heng; Wei, Hailing

    2015-07-01

    Matrix metalloproteinases (MMPs) secreted by ovarian cancer play essential roles in tumor invasion and metastasis. In this study, we investigated the effect of baicalein, which is isolated from the traditional Chinese herbal medicine Scutellaria baicalensis Georgi, on human ovarian cancer cell lines by measuring MMP-2 expression, invasive potential, and the underlying molecular mechanisms. Analysis of MMP-2 was carried out by western blots and RT-PCR. The invasion ability of ovarian cancer cells was determined using a Transwell invasion assay. Nuclear factor-κB (NF-κB) and p38 MAPK activation was assessed by western blots. The results of the present study showed that baicalein reduced the expression of MMP-2 in a dose-dependent manner and the invasion of ovarian cancer cells was also significantly suppressed by baicalein. We also found that baicalein reduced the activation of NF-κB signaling molecules; in addition, the MMP-2 expression and invasion ability of ovarian cancer cells were abolished with the treatment of the NF-κB inhibitor, pyrrolidine dithiocarbamate. However, the addition of p38 MAPK inhibitor SB203580 significantly reduced the activation of NF-κB; meanwhile, baicalein was shown to exert an inhibitory effect on p38 activation. Furthermore, the MMP-2 expression and invasion ability of ovarian cancer cells were significantly inhibited by SB203580. In conclusion, baicalein inhibits the MMP-2 expression and invasion ability of ovarian cancer cells, possibly by the p38 MAPK-dependent NF-κB signaling pathway; these findings may provide insights into the potential of using baicalein as a therapeutic strategy against ovarian cancer.

  8. Paris saponin VII suppresses osteosarcoma cell migration and invasion by inhibiting MMP-2/9 production via the p38 MAPK signaling pathway

    PubMed Central

    Cheng, Gong; Gao, Fengguang; Sun, Xiujiang; Bi, Haiyong; Zhu, Yonglin

    2016-01-01

    Metastasis is the primary cause of mortality in osteosarcoma. Targeting metastasis is a major strategy in osteosarcoma treatment. As a traditional Chinese medicine, Trillium tschonoskii Maxim has been widely used in the therapy of various diseases, including cancer. However, currently there is no evidence regarding the anti-metastasic effect of Paris saponin VII (PS VII), which is extracted from Trillium tschonoskii Maxim, on osteosarcoma cells and its underling mechanisms. The present study aimed to examine the effect of PS VII on the migration and invasion of osteosarcoma cells. Viability and proliferation of osteosarcoma cells were examined by MTT assay. Migration and invasion of osteosarcoma cells was then detected using scratch wound healing assays and Transwell assays, respectively. Additionally, the expression of matrix metalloproteinase (MMP)-2 and -9 was determined at the mRNA and protein level following treatment with PS VII. Mitogen-activated protein kinase (MAPK) expression was also detected by western blot analysis. Finally, an inhibitor of p38 MAPK was used to verify the effect of PS VII on the expression of MMP-2 and -9, as well as the migration and invasion osteosarcoma cells. This demonstrated that the proliferation, migration and invasion of the osteosarcoma cells were suppressed following treatment with PS VII. PS VII downregulated the expression of MMP-2 and -9 in a dose- and time-dependent manner. PS VII also exerted its ability to downregulate the phosphorylation of p38 MAPKs. Furthermore, by using a p38 inhibitor, SB203580, the role of PS VII in MMP-2 and -9 expression and osteosarcoma cell invasion was revealed. Taken together, these results demonstrated that PS VII suppresses the migration and invasion of osteosarcoma cells via the p38 MAPK signaling pathway. PMID:27572907

  9. Anti-inflammatory effects of chicanine on murine macrophage by down-regulating LPS-induced inflammatory cytokines in IκBα/MAPK/ERK signaling pathways

    PubMed Central

    Chen, Haixia; Sohn, Johann; Zhang, Likang; Tian, Jingge; Chen, Shuhan; Bjeldanes, Leonard F.

    2014-01-01

    Schisandra chinensis Baill is a Chinese traditional medicine with multiple pharmacological activities. In this study, chicanine, one of the major lignan compounds of Schiandra chinesis, was investigated for suppressive effects on lipopolysaccharide (LPS)-induced inflammatory responses in murine macrophages (RAW 264.7 cells). Chicanine was found to have anti-infammatory properties with the inhibition of nitric oxide (NO) and Prostaglandin E (2) (PGE2) production and nuclear factor-κB (NF-κB) signaling in LPS-stimulated RAW 264.7 cells with no cytotoxic effects. Treatment of RAW 264.7 cells with chicanine down-regulated LPS-induced expression of pro-inflammatory cytokines including TNFα, IL-1β, MCP-1, G-CSF, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). These inhibitory effects were found with the blockage of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinases 1 and 2 (ERK 1/2), and also IκB-α phosphorylation. These results indicated that anti-inflammatory actions of chicanine in macrophages involved inhibition of LPS-induced TLR4-IκBα/MAPK/ERK signaling pathways. PMID:24361309

  10. TNFα Signaling Regulates Cystic Epithelial Cell Proliferation through Akt/mTOR and ERK/MAPK/Cdk2 Mediated Id2 Signaling

    PubMed Central

    Zhou, Julie X.; Fan, Lucy X.; Li, Xiaoyan; Calvet, James P.; Li, Xiaogang

    2015-01-01

    Tumor necrosis factor alpha (TNFα) is present in cyst fluid and promotes cyst growth in autosomal dominant polycystic kidney disease (ADPKD). However, the cross-talk between TNFα and PKD associated signaling pathways remains elusive. In this study, we found that stimulation of renal epithelial cells with TNFα or RANKL (receptor activator of NF-κB ligand), a member of the TNFα cytokine family, activated either the PI3K pathway, leading to AKT and mTOR mediated the increase of Id2 protein, or MAPK and Cdk2 to induce Id2 nuclear translocation. The effects of TNFα/RANKL on increasing Id2 protein and its nuclear translocation caused significantly decreased mRNA and protein levels of the Cdk inhibitor p21, allowing increased cell proliferation. TNFα levels increase in cystic kidneys in response to macrophage infiltration and thus might contribute to cyst growth and enlargement during the progression of disease. As such, this study elucidates a novel mechanism for TNFα signaling in regulating cystic renal epithelial cell proliferation in ADPKD. PMID:26110849

  11. Bergenin Plays an Anti-Inflammatory Role via the Modulation of MAPK and NF-κB Signaling Pathways in a Mouse Model of LPS-Induced Mastitis.

    PubMed

    Gao, Xue-jiao; Guo, Meng-yao; Zhang, Ze-cai; Wang, Tian-cheng; Cao, Yong-guo; Zhang, Nai-sheng

    2015-01-01

    Mastitis is a major disease in humans and other animals and is characterized by mammary gland inflammation. It is a major disease of the dairy industry. Bergenin is an active constituent of the plants of genus Bergenia. Research indicates that bergenin has multiple biological activities, including anti-inflammatory and immunomodulatory properties. The objective of this study was to evaluate the protective effects and mechanism of bergenin on the mammary glands during lipopolysaccharide (LPS)-induced mastitis. In this study, mice were treated with LPS to induce mammary gland mastitis as a model for the disease. Bergenin treatment was initiated after LPS stimulation for 24 h. The results indicated that bergenin attenuated inflammatory cell infiltration and decreased the concentration of NO, TNF-α, IL-1β, and IL-6, which were increased in LPS-induced mouse mastitis. Furthermore, bergenin downregulated the phosphorylation of nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinases (MAPK) signaling pathway proteins in mammary glands with mastitis. In conclusion, bergenin reduced the expression of NO, TNF-α, IL-1β, and IL-6 proinflammatory cytokines by inhibiting the activation of the NF-κB and MAPKs signaling pathways, and it may represent a novel treatment strategy for mastitis.

  12. Amarogentin, a secoiridoid glycoside, abrogates platelet activation through PLC γ 2-PKC and MAPK pathways.

    PubMed

    Yen, Ting-Lin; Lu, Wan-Jung; Lien, Li-Ming; Thomas, Philip Aloysius; Lee, Tzu-Yin; Chiu, Hou-Chang; Sheu, Joen-Rong; Lin, Kuan-Hung

    2014-01-01

    Amarogentin, an active principle of Gentiana lutea, possess antitumorigenic, antidiabetic, and antioxidative properties. Activation of platelets is associated with intravascular thrombosis and cardiovascular diseases. The present study examined the effects of amarogentin on platelet activation. Amarogentin treatment (15~60  μM) inhibited platelet aggregation induced by collagen, but not thrombin, arachidonic acid, and U46619. Amarogentin inhibited collagen-induced phosphorylation of phospholipase C (PLC) γ2, protein kinase C (PKC), and mitogen-activated protein kinases (MAPKs). It also inhibits in vivo thrombus formation in mice. In addition, neither the guanylate cyclase inhibitor ODQ nor the adenylate cyclase inhibitor SQ22536 affected the amarogentin-mediated inhibition of platelet aggregation, which suggests that amarogentin does not regulate the levels of cyclic AMP and cyclic GMP. In conclusion, amarogentin prevents platelet activation through the inhibition of PLC γ2-PKC cascade and MAPK pathway. Our findings suggest that amarogentin may offer therapeutic potential for preventing or treating thromboembolic disorders.

  13. CyHV-2 ORF104 activates the p38 MAPK pathway.

    PubMed

    Du, Mi; Chen, Mingliang; Shen, Haifeng; Wang, Wei; Li, Zengpeng; Wang, Weiyi; Huang, Jianhui; Chen, Jianming

    2015-10-01

    Cyprinid herpesvirus 2 (CyHV-2) is the pathogen responsible for herpesviral hematopoietic necrosis disease, which causes huge losses on aquaculture. So far the studies of CyHV-2 mainly focus on the identification and detection of this virus, but little is known about the role of specific CyHV-2 genes in the infection process. Based on the genomic information, CyHV-2 ORF104 encodes a kinase-like protein, which is highly conserved among the three CyHVs. Our study was initiated to investigate the role of kinase-like protein ORF104 during virus infection. Subcellular localization study showed that ORF104 was mainly expressed in the nucleus in both human HEK293T and fish EPC cells. However, deletion of the putative nuclear localization signal of ORF104 (ORF104M) resulted in the cytoplasmic distribution in HEK293T. We then examined whether MAPKs were involved in the ORF104-mediated signaling pathway by overexpressing ORF104 and ORF104M in HEK293T. Overexpression of ORF104 and ORF104M resulted in the up-regulation of p38 phosphorylation, but not JNK or ERK, indicating that ORF104 specifically activates p38 signaling pathway. In vivo study showed that CyHV-2 infection enhanced p38 phosphorylation in gibel carp (Carassius auratus gibelio). Interestingly, p38 inhibitor SB203580 strongly reduced fish death caused by CyHV-2 infection. Therefore, our study for the first time reveals the function of ORF104 during CyHV-2 infection, indicating that ORF104 is a potential vaccine candidate for CyHV-2.

  14. Dioscin inhibits colon tumor growth and tumor angiogenesis through regulating VEGFR2 and AKT/MAPK signaling pathways

    SciTech Connect

    Tong, Qingyi; Qing, Yong; Wu, Yang; Hu, Xiaojuan; Jiang, Lei; Wu, Xiaohua

    2014-12-01

    Dioscin has shown cytotoxicity against cancer cells, but its in vivo effects and the mechanisms have not elucidated yet. The purpose of the current study was to assess the antitumor effects and the molecular mechanisms of dioscin. We showed that dioscin could inhibit tumor growth in vivo and has no toxicity at the test condition. The growth suppression was accompanied by obvious blood vessel decrease within solid tumors. We also found dioscin treatment inhibited the proliferation of cancer and endothelial cell lines, and most sensitive to primary cultured human umbilical vein endothelial cells (HUVECs). What's more, analysis of HUVECs migration, invasion, and tube formation exhibited that dioscin has significantly inhibitive effects to these actions. Further analysis of blood vessel formation in the matrigel plugs indicated that dioscin could inhibit VEGF-induced blood vessel formation in vivo. We also identified that dioscin could suppress the downstream protein kinases of VEGFR2, including Src, FAK, AKT and Erk1/2, accompanied by the increase of phosphorylated P38MAPK. The results potently suggest that dioscin may be a potential anticancer drug, which efficiently inhibits angiogenesis induced by VEGFR2 signaling pathway as well as AKT/MAPK pathways. - Highlights: • Dioscin inhibits tumor growth in vivo and does not exhibit any toxicity. • Dioscin inhibits angiogenesis within solid tumors. • Dioscin inhibits the proliferation, migration, invasion, and tube formation of HUVECs. • Dioscin inhibits VEGF–induced blood vessel formation in vivo. • Dioscin inhibits VEGFR2 signaling pathway as well as AKT/MAPK pathway.

  15. IL-1α Expression in Pancreatic Ductal Adenocarcinoma Affects the Tumor Cell Migration and Is Regulated by the p38MAPK Signaling Pathway

    PubMed Central

    Tjomsland, Vegard; Bojmar, Linda; Sandström, Per; Bratthäll, Charlotte; Messmer, Davorka; Spångeus, Anna; Larsson, Marie

    2013-01-01

    The interplay between the tumor cells and the surrounding stroma creates inflammation, which promotes tumor growth and spread. The inflammation is a hallmark for pancreatic adenocarcinoma (PDAC) and is to high extent driven by IL-1α. IL-1α is expressed and secreted by the tumor cells and exerting its effect on the stroma, i.e. cancer associated fibroblasts (CAF), which in turn produce massive amount of inflammatory and immune regulatory factors. IL-1 induces activation of transcription factors such as nuclear factor-κβ (NF-κβ), but also activator protein 1 (AP-1) via the small G-protein Ras. Dysregulation of Ras pathways are common in cancer as this oncogene is the most frequently mutated in many cancers. In contrast, the signaling events leading up to the expression of IL-1α by tumor cells are not well elucidated. Our aim was to examine the signaling cascade involved in the induction of IL-1α expression in PDAC. We found p38MAPK, activated by the K-Ras signaling pathway, to be involved in the expression of IL-1α by PDAC as blocking this pathway decreased both the gene and protein expression of IL-1α. Blockage of the P38MAPK signaling in PDAC also dampened the ability of the tumor cell to induce inflammation in CAFs. In addition, the IL-1α autocrine signaling regulated the migratory capacity of PDAC cells. Taken together, the blockage of signaling pathways leading to IL-1α expression and/or neutralization of IL-1α in the PDAC microenvironment should be taken into consideration as possible treatment or complement to existing treatment of this cancer. PMID:23951028

  16. Astragalin inhibits airway eotaxin-1 induction and epithelial apoptosis through modulating oxidative stress-responsive MAPK signaling

    PubMed Central

    2014-01-01

    Background Eotaxin proteins are a potential therapeutic target in treating the peribronchial eosinophilia associated with allergic airway diseases. Since inflammation is often associated with an increased generation of reactive oxygen species (ROS), oxidative stress is a mechanistically imperative factor in asthma. Astragalin (kaempferol-3-O-glucoside) is a flavonoid with anti-inflammatory activity and newly found in persimmon leaves and green tea seeds. This study elucidated that astragalin inhibited endotoxin-induced oxidative stress leading to eosinophilia and epithelial apoptosis in airways. Methods Airway epithelial BEAS-2B cells were exposed to lipopolysaccharide (LPS) in the absence and presence of 1–20 μM astragalin. Western blot and immunocytochemical analyses were conducted to determine induction of target proteins. Cell and nuclear staining was also performed for ROS production and epithelial apoptosis. Results When airway epithelial cells were exposed to 2 μg/ml LPS, astragalin nontoxic at ≤20 μM suppressed cellular induction of Toll-like receptor 4 (TLR4) and ROS production enhanced by LPS. Both LPS and H2O2 induced epithelial eotaxin-1 expression, which was blocked by astragalin. LPS activated and induced PLCγ1, PKCβ2, and NADPH oxidase subunits of p22phox and p47phox in epithelial cells and such activation and induction were demoted by astragalin or TLR4 inhibition antagonizing eotaxin-1 induction. H2O2-upregulated phosphorylation of JNK and p38 MAPK was dampened by adding astragalin to epithelial cells, while this compound enhanced epithelial activation of Akt and ERK. H2O2 and LPS promoted epithelial apoptosis concomitant with nuclear condensation or caspase-3 activation, which was blunted by astragalin. Conclusions Astragalin ameliorated oxidative stress-associated epithelial eosinophilia and apoptosis through disturbing TLR4-PKCβ2-NADPH oxidase-responsive signaling. Therefore, astragalin may be a potent agent antagonizing endotoxin

  17. atRA-induced apoptosis of mouse embryonic palate mesenchymal cells involves activation of MAPK pathway

    SciTech Connect

    Yu Zengli . E-mail: yuzengli@263.net; Xing Ying . E-mail: xingy@zzu.edu.cn

    2006-08-15

    Our previous studies have shown that atRA treatment resulted in cell-cycle block and growth inhibition in mouse embryonic palatal mesenchymal (MEPM). In the current study, gestation day (GD) 13 MEPM cells were used to test the hypothesis that the growth inhibition by atRA is due to apoptosis. The effects of atRA on apoptosis were assessed by performing MTT assay, Cell Death Detection ELISA and flow cytometry, respectively. Data analysis confirmed that atRA treatment induced apoptosis-like cell death, as shown by decreased cell viability and increased fragmented DNA and sub-G1 fraction. atRA-induced apoptosis was associated with upregulation of bcl-2, translocation of bax protein to the mitochondria from the cytosol, activation of caspase-3 and cytochrome c release into cytosol. atRA-induced apoptosis was abrogated by z-DEVD-fmk, a caspase-3 specific inhibitor, and z-VAD-fmk, a general caspase inhibitor, suggesting that the atRA-induced cell death of MEPM cells occurs through the cytochrome c- and caspase-3-dependent pathways. In addition, atRA treatment caused a strong and sustained activation of c-Jun N-terminal kinase (JNK) and p38 kinase (p38), as well as an early but transient activation of extracellular signal-regulated kinase (ERK). Importantly, atRA-induced DNA fragmentation and capase-3 activation were prevented by pretreatment with the JNK inhibitor (SP600125) and the p38 MAPK inhibitor (SB202190), but not by pretreatment with MEK inhibitor (U0126). From these results, we suggest that mitogen-activated protein kinase-dependent pathways is involved in the atRA-induced apoptosis of MEPM cells.

  18. Astragaloside IV suppresses transforming growth factor-β1 induced fibrosis of cultured mouse renal fibroblasts via inhibition of the MAPK and NF-κB signaling pathways

    SciTech Connect

    Che, Xiajing; Wang, Qin; Xie, Yuanyuan; Xu, Weijia; Shao, Xinghua; Mou, Shan Ni, Zhaohui

    2015-09-04

    Renal fibrosis, a progressive process characterized by the accumulation of extracellular matrix (ECM) leading to organ dysfunction, is a characteristic of chronic kidney diseases. Among fibrogenic factors known to regulate the renal fibrotic process, transforming growth factor-β (TGF-β) plays a central role. In the present study, we examined the effect of Astragaloside IV (AS-IV), a component of the traditional Chinese medicinal plant Astragalus membranaceus, on the processes associated with renal fibrosis in cultured mouse renal fibroblasts treated with TGF-β1. RT-PCR, western blotting, immunofluorescence staining and collagen assays showed that AS-IV suppressed TGF-β1 induced fibroblast proliferation, transdifferentiation, and ECM production in a dose-dependent manner. Examination of the underlying mechanisms showed that the effect of AS-IV on the inhibition of fibroblast differentiation and ECM formation were mediated by its modulation of the activity of the MAPK and NF-κB signaling pathways. Taken together, our results indicate that AS-IV alleviates renal interstitial fibrosis via a mechanism involving the MAPK and NF-κB signaling pathways and demonstrate the therapeutic potential of AS-IV for the treatment of chronic kidney diseases. - Highlights: • AS-IV suppressed TGF-β1 induced renal fibroblast proliferation. • AS-IV suppressed TGF-β1 induced renal fibroblast transdifferentiation. • AS-IV suppressed TGF-β1 induced ECM production. • AS-IV alleviates renal fibrosis via the MAPK and NF-κB signaling pathways.

  19. A novel synthetic compound MCAP suppresses LPS-induced murine microglial activation in vitro via inhibiting NF-kB and p38 MAPK pathways

    PubMed Central

    Kim, Byung-Wook; More, Sandeep Vasant; Yun, Yo-Sep; Ko, Hyun-Myung; Kwak, Jae-Hwan; Lee, Heesoon; Suk, Kyoungho; Kim, In-Su; Choi, Dong-Kug

    2016-01-01

    Aim: To investigate the anti-neuroinflammatory activity of a novel synthetic compound, 7-methylchroman-2-carboxylic acid N-(2-trifluoromethyl) phenylamide (MCAP) against LPS-induced microglial activation in vitro. Methods: Primary mouse microglia and BV2 microglia cells were exposed to LPS (50 or 100 ng/mL). The expression of iNOS and COX-2, proinflammatory cytokines, NF-κB and p38 MAPK signaling molecules were analyzed by RT-PCR, Western blot and ELISA. The morphological changes of microglia and nuclear translocation of NF-ĸB were visualized using phase contrast and fluorescence microscopy, respectively. Results: Pretreatment with MCAP (0.1, 1, 10 μmol/L) dose-dependently inhibited LPS-induced expression of iNOS and COX-2 in BV2 microglia cells. Similar results were obtained in primary microglia pretreated with MCAP (0.1, 0.5 μmol/L). MCAP dose-dependently abated LPS-induced release of TNF-α, IL-6 and IL-1β, and mitigated LPS-induced activation of NF-κB by reducing the phosphorylation of IκBα in BV2 microglia cells. Moreover, MCAP attenuated LPS-induced phosphorylation of p38 MAPK, whereas SB203580, a p38 MAPK inhibitor, significantly potentiated MCAP-caused inhibition on the expression of MEF-2 (a transcription factor downstream of p38 MAPK). Conclusion: MCAP exerts anti-inflammatory effects in murine microglia in vitro by inhibiting the p38 MAPK and NF-κB signaling pathways and proinflammatory responses. MCAP may be developed as a novel agent for treating diseases involving activated microglial cells. PMID:26838070

  20. Positive regulation of the Egr-1/osteopontin positive feedback loop in rat vascular smooth muscle cells by TGF-{beta}, ERK, JNK, and p38 MAPK signaling

    SciTech Connect

    Yu, Hong-Wei; Liu, Qi-Feng; Liu, Gui-Nan

    2010-05-28

    Previous studies identified a positive feedback loop in rat vascular smooth muscle cells (VSMCs) in which early growth response factor-1 (Egr-1) binds to the osteopontin (OPN) promoter and upregulates OPN expression, and OPN upregulates Egr-1 expression via the extracellular signal-regulated protein kinase (ERK) signaling pathway. The current study examined whether transforming growth factor-{beta} (TGF-{beta}) activity contributes to Egr-1 binding to the OPN promoter, and whether other signaling pathways act downstream of OPN to regulate Egr-1 expression. ChIP assays using an anti-Egr-1 antibody showed that amplification of the OPN promoter sequence decreased in TGF-{beta} DNA enzyme-transfected VSMCs relative to control VSMCs. Treatment of VSMCs with PD98059 (ERK inhibitor), SP600125 (JNK inhibitor), or SB203580 (p38 MAPK inhibitor) significantly inhibited OPN-induced Egr-1 expression, and PD98059 treatment was associated with the most significant decrease in Egr-1 expression. OPN-stimulated VSMC cell migration was inhibited by SP600125 or SB203580, but not by PD98059. Furthermore, MTT assays showed that OPN-mediated cell proliferation was inhibited by PD98059, but not by SP600125 or SB203580. Taken together, the results of the current study show that Egr-1 binding to the OPN promoter is positively regulated by TGF-{beta}, and that the p38 MAPK, JNK, and ERK pathways are involved in OPN-mediated Egr-1 upregulation.

  1. CFTR-regulated MAPK/NF-κB signaling in pulmonary inflammation in thermal inhalation injury

    PubMed Central

    Dong, Zhi Wei; Chen, Jing; Ruan, Ye Chun; Zhou, Tao; Chen, Yu; Chen, YaJie; Tsang, Lai Ling; Chan, Hsiao Chang; Peng, Yi Zhi

    2015-01-01

    The mechanism underlying pulmonary inflammation in thermal inhalation injury remains elusive. Cystic fibrosis, also hallmarked with pulmonary inflammation, is caused by mutations in CFTR, the expression of which is temperature-sensitive. We investigated whether CFTR is involved in heat-induced pulmonary inflammation. We applied heat-treatment in 16HBE14o- cells with CFTR knockdown or overexpression and heat-inhalation in rats in vivo. Heat-treatment caused significant reduction in CFTR and, reciprocally, increase in COX-2 at early stages both in vitro and in vivo. Activation of ERK/JNK, NF-κB and COX-2/PGE2 were detected in heat-treated cells, which were mimicked by knockdown, and reversed by overexpression of CFTR or VX-809, a reported CFTR mutation corrector. JNK/ERK inhibition reversed heat-/CFTR-knockdown-induced NF-κB activation, whereas NF-κB inhibitor showed no effect on JNK/ERK. IL-8 was augmented by heat-treatment or CFTR-knockdown, which was abolished by inhibition of NF-κB, JNK/ERK or COX-2. Moreover, in vitro or in vivo treatment with curcumin, a natural phenolic compound, significantly enhanced CFTR expression and reversed the heat-induced increases in COX-2/PGE2/IL-8, neutrophil infiltration and tissue damage in the airway. These results have revealed a CFTR-regulated MAPK/NF-κB pathway leading to COX-2/PGE2/IL-8 activation in thermal inhalation injury, and demonstrated therapeutic potential of curcumin for alleviating heat-induced pulmonary inflammation. PMID:26515683

  2. Brominated Flame Retardants, Tetrabromobisphenol A and Hexabromocyclododecane, Activate Mitogen-Activated Protein Kinases (MAPKs) in Human Natural Killer Cells

    PubMed Central

    Cato, Anita; Celada, Lindsay; Kibakaya, Esther Caroline; Simmons, Nadia; Whalen, Margaret M.

    2014-01-01

    NK cells provide a vital surveillance against virally infected cells, tumor cells, and antibody-coated cells through the release of cytolytic mediators and gamma interferon (IFN-γ). Hexabromocyclododecane (HBCD) is a brominated flame retardant used primarily in expanded (EPS) and extruded (XPS) polystyrene foams for thermal insulation in the building and construction industry. Tetrabromobisphenol A (TBBPA) is used both as a reactive and an additive flame retardant in a variety of materials. HBCD and TBBPA contaminate the environment and are found in human blood samples. In previous studies, we have shown that other environmental contaminants, such as the dibutyltin (DBT) and tributyltin (TBT), decrease NK lytic function by activating mitogen-activated protein kinases (MAPKs) in the NK cells. HBCD and TBBPA also interfere with NK cell(s) lytic function. The current study evaluates whether HBCD and/or TBBPA have the capacity to activate MAPKs and MAPK kinases (MAP2Ks). The effects of concentrations of HBCD and TBBPA that inhibited lytic function on the phosphorylation state and total levels of the MAPKs (p44/42, p38, and JNK) and the phosphorylation and total levels of the MAP2Ks (MEK1/2 and MKK3/6) were examined. Results indicate that exposure of human NK cells to 10-0.5 µM HBCD or TBBPA activate MAPKs and MAP2Ks. This HBCD and TBBPA-induced activation of MAPKs may leave them unavailable for activation by virally infected or tumor target cells and thus contributes to the observed decreases in lytic function seen in NK cells exposed to HBCD and TBBPA. PMID:25341744

  3. GENE REGULATION BY MAPK SUBSTRATE COMPETITION

    PubMed Central

    Kim, Yoosik; Andreu, María José; Lim, Bomyi; Chung, Kwanghun; Terayama, Mark; Jiménez, Gerardo; Berg, Celeste A.; Lu, Hang; Shvartsman, Stanislav Y.

    2011-01-01

    SUMMARY Developing tissues are patterned by coordinated activities of signaling systems, which can be integrated by a regulatory region of a gene that binds multiple transcription factors or by a transcription factor that is modified by multiple enzymes. Based on a combination of genetic and imaging experiments in the early Drosophila embryo, we describe a signal integration mechanism that cannot be reduced to a single gene regulatory element or a single transcription factor. This mechanism relies on an enzymatic network formed by Mitogen Activated Protein Kinase (MAPK) and its substrates. Specifically, anteriorly localized MAPK substrates, such as Bicoid, antagonize MAPK-dependent downregulation of Capicua, a repressor which is involved in gene regulation along the dorsoventral axis of the embryo. MAPK substrate competition provides a basis for ternary interaction of the anterior, dorsoventral, and terminal patterning systems. A mathematical model of this interaction can explain gene expression patterns with both anteroposterior and dorsoventral polarities. PMID:21664584

  4. A novel cysteine-rich neurotrophic factor in Aplysia facilitates growth, MAPK activation, and long-term synaptic facilitation.

    PubMed

    Pu, Lu; Kopec, Ashley M; Boyle, Heather D; Carew, Thomas J

    2014-04-01

    Neurotrophins are critically involved in developmental processes such as neuronal cell survival, growth, and differentiation, as well as in adult synaptic plasticity contributing to learning and memory. Our previous studies examining neurotrophins and memory formation in Aplysia showed that a TrkB ligand is required for MAPK activation, long-term synaptic facilitation (LTF), and long-term memory (LTM) for sensitization. These studies indicate that neurotrophin-like molecules in Aplysia can act as key elements in a functionally conserved TrkB signaling pathway. Here we report that we have cloned and characterized a novel neurotrophic factor, Aplysia cysteine-rich neurotrophic factor (apCRNF), which shares classical structural and functional characteristics with mammalian neurotrophins. We show that apCRNF (1) is highly enriched in the CNS, (2) enhances neurite elongation and branching, (3) interacts with mammalian TrkB and p75(NTR), (4) is released from Aplysia CNS in an activity-dependent fashion, (5) facilitates MAPK activation in a tyrosine kinase dependent manner in response to sensitizing stimuli, and (6) facilitates the induction of LTF. These results show that apCRNF is a native neurotrophic factor in Aplysia that can engage the molecular and synaptic mechanisms underlying memory formation.

  5. Upregulation of cell proliferation via Shc and ERK1/2 MAPK signaling in SaOS-2 osteoblasts grown on magnesium alloy surface coating with tricalcium phosphate.

    PubMed

    Jiang, Tianlong; Guo, Lei; Ni, Shenghui; Zhao, Yuyan

    2015-04-01

    Magnesium (Mg) alloys have been demonstrated to be viable orthopedic implants because of mechanical and biocompatible properties similar to natural bone. In order to improve its osteogenic properties, a porous β-tricalcium phosphate (β-TCP) was coated on the Mg-3AI-1Zn alloy by alkali-heat treatment technique. The human bone-derived cells (SaOS-2) were cultured on (β-TCP)-Mg-3AI-1Zn in vitro, and the osteoblast response, the morphology and the elements on this alloy surface were investigated. Also, the regulation of key intracellular signalling proteins was investigated in the SaOS-2 cells cultured on alloy surface. The results from scanning electron microscope and immunofluorescence staining demonstrated that (β-TCP)-Mg-3AI-1Zn induced significant osteogenesis. SaOS-2 cell proliferation was improved by β-TCP coating. Moreover, the (β-TCP)-Mg-3AI-1Zn surface induced activation of key intracellular signalling proteins in SaOS-2 cells. We observed an enhanced activation of Src homology and collagen (Shc), a common point of integration between bone morphogenetic protein 2, and the Ras/mitogen-activated protein kinase (MAPK) pathway. ERK1/2 MAP kinase activation was also upregulated, suggesting a role in mediating osteoblastic cell interactions with biomaterials. The signalling pathway involving c-fos (member of the activated protein-1) was also shown to be upregulated in osteoblasts cultured on the (β-TCP)-Mg-3AI-1Zn. These results suggest that β-TCP coating may contribute to successful osteoblast function on Mg alloy surface. (β-TCP)-Mg-3AI-1Zn may upregulate cell proliferation via Shc and ERK1/2 MAPK signaling in SaOS-2 osteoblasts grown on Mg alloy surface.

  6. Inhibition of Nickel Nanoparticles-Induced Toxicity by Epigallocatechin-3-Gallate in JB6 Cells May Be through Down-Regulation of the MAPK Signaling Pathways.

    PubMed

    Gu, Yuanliang; Wang, Yafei; Zhou, Qi; Bowman, Linda; Mao, Guochuan; Zou, Baobo; Xu, Jin; Liu, Yu; Liu, Kui; Zhao, Jinshun; Ding, Min

    2016-01-01

    With the rapid development in nanotechnology, nickel nanoparticles (Ni NPs) have emerged in the application of nanomedicine in recent years. However, the potential adverse health effects of Ni NPs are unclear. In this study, we examined the inhibition effects of epigallocatechin-3-gallate (EGCG) on the toxicity induced by Ni NPs in mouse epidermal cell line (JB6 cell). MTT assay showed that Ni NPs induced cytotoxicity in a dose-dependent manner while EGCG exerted a certain inhibition on the toxicity. Additionally, EGCG could reduce the apoptotic cell number and the level of reactive oxygen species (ROS) in JB6 cells induced by Ni NPs. Furthermore, we observed that EGCG could down-regulate Ni NPs-induced activator protein-1 (AP-1) and nuclear factor-κB (NF-κB) activation in JB6 cells, which has been shown to play pivotal roles in tumor initiation, promotion and progression. Western blot indicated that EGCG could alleviate the toxicity of Ni NPs through regulating protein changes in MAPK signaling pathways. In summary, our results suggest that careful evaluation on the potential health effects of Ni NPs is necessary before being widely used in the field of nanomedicine. Inhibition of EGCG on Ni NPs-induced cytotoxicity in JB6 cells may be through the MAPK signaling pathways suggesting that EGCG might be useful in preventing the toxicity of Ni NPs. PMID:26943640

  7. Inhibition of Nickel Nanoparticles-Induced Toxicity by Epigallocatechin-3-Gallate in JB6 Cells May Be through Down-Regulation of the MAPK Signaling Pathways

    PubMed Central

    Bowman, Linda; Mao, Guochuan; Zou, Baobo; Xu, Jin; Liu, Yu; Liu, Kui; Zhao, Jinshun; Ding, Min

    2016-01-01

    With the rapid development in nanotechnology, nickel nanoparticles (Ni NPs) have emerged in the application of nanomedicine in recent years. However, the potential adverse health effects of Ni NPs are unclear. In this study, we examined the inhibition effects of epigallocatechin-3-gallate (EGCG) on the toxicity induced by Ni NPs in mouse epidermal cell line (JB6 cell). MTT assay showed that Ni NPs induced cytotoxicity in a dose-dependent manner while EGCG exerted a certain inhibition on the toxicity. Additionally, EGCG could reduce the apoptotic cell number and the level of reactive oxygen species (ROS) in JB6 cells induced by Ni NPs. Furthermore, we observed that EGCG could down-regulate Ni NPs-induced activator protein-1 (AP-1) and nuclear factor-κB (NF-κB) activation in JB6 cells, which has been shown to play pivotal roles in tumor initiation, promotion and progression. Western blot indicated that EGCG could alleviate the toxicity of Ni NPs through regulating protein changes in MAPK signaling pathways. In summary, our results suggest that careful evaluation on the potential health effects of Ni NPs is necessary before being widely used in the field of nanomedicine. Inhibition of EGCG on Ni NPs-induced cytotoxicity in JB6 cells may be through the MAPK signaling pathways suggesting that EGCG might be useful in preventing the toxicity of Ni NPs. PMID:26943640

  8. Tumor-suppressive function of long noncoding RNA MALAT1 in glioma cells by downregulation of MMP2 and inactivation of ERK/MAPK signaling.

    PubMed

    Han, Y; Wu, Z; Wu, T; Huang, Y; Cheng, Z; Li, X; Sun, T; Xie, X; Zhou, Y; Du, Z

    2016-03-03

    Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is a type of long noncoding RNA. It is associated with metastasis and is a favorable prognostic factor for lung cancer. Recent studies have shown that MALAT1 plays an important role in other malignancies. But, little is known about the role of MALAT1 in glioma. In this study, quantitative reverse transcription PCR (qRT-PCR) was used to demonstrate that the expression of MALAT1 was lower than that in normal brain tissues. Stable RNA interference-mediated knockdown of MALAT1 in human glioma cell lines (U87 and U251) significantly promoted the invasion and proliferation of the glioma cells by in vitro assays. Conversely, overexpression of MALAT1 caused significant reduction in cell proliferation and invasion in vitro, and tumorigenicity in both subcutaneous and intracranial human glioma xenograft models. Furthermore, MALAT1-mediated tumor suppression in glioma cells may be via reduction of extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling activity and expression of matrix metalloproteinase 2 (MMP2). In conclusion, overall data demonstrated the tumor-suppressive role of MALAT1 in glioma by attenuating ERK/MAPK-mediated growth and MMP2-mediated invasiveness.

  9. Tumor-suppressive function of long noncoding RNA MALAT1 in glioma cells by downregulation of MMP2 and inactivation of ERK/MAPK signaling

    PubMed Central

    Han, Y; Wu, Z; Wu, T; Huang, Y; Cheng, Z; Li, X; Sun, T; Xie, X; Zhou, Y; Du, Z

    2016-01-01

    Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is a type of long noncoding RNA. It is associated with metastasis and is a favorable prognostic factor for lung cancer. Recent studies have shown that MALAT1 plays an important role in other malignancies. But, little is known about the role of MALAT1 in glioma. In this study, quantitative reverse transcription PCR (qRT-PCR) was used to demonstrate that the expression of MALAT1 was lower than that in normal brain tissues. Stable RNA interference-mediated knockdown of MALAT1 in human glioma cell lines (U87 and U251) significantly promoted the invasion and proliferation of the glioma cells by in vitro assays. Conversely, overexpression of MALAT1 caused significant reduction in cell proliferation and invasion in vitro, and tumorigenicity in both subcutaneous and intracranial human glioma xenograft models. Furthermore, MALAT1-mediated tumor suppression in glioma cells may be via reduction of extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling activity and expression of matrix metalloproteinase 2 (MMP2). In conclusion, overall data demonstrated the tumor-suppressive role of MALAT1 in glioma by attenuating ERK/MAPK-mediated growth and MMP2-mediated invasiveness. PMID:26938295

  10. Functional microarray analysis of differentially expressed genes in granulosa cells from women with polycystic ovary syndrome related to MAPK/ERK signaling

    PubMed Central

    Lan, Chen-Wei; Chen, Mei-Jou; Tai, Kang-Yu; Yu, Danny CW; Yang, Yu-Chieh; Jan, Pey-Shynan; Yang, Yu-Shih; Chen, Hsin-Fu; Ho, Hong-Nerng

    2015-01-01

    Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age. Although its aetiology and pathogenesis remain unclear, recent studies suggest that the dysfunction of granulosa cells may partly be responsible. This study aimed to use cDNA microarray technology to compare granulosa cell gene expression profiles in women with and without PCOS to identify genes that may be aetiologically implicated in the pathogenesis of PCOS. The study cohort included 12 women undergoing in vitro fertilization, six with PCOS and six without PCOS. Differential gene expression profiles were classified by post-analyses of microarray data, followed by western blot analyses to confirm the microarray data of selected genes. In total, 243 genes were differentially expressed (125 upregulated and 118 downregulated) between the PCOS and non-PCOS granulosa cells. These genes are involved in reproductive system development, amino acid metabolism and cellular development and proliferation. Comparative analysis revealed genes involved in the mitogen-activated protein kinase/extracellular regulated kinase (MAPK/ERK) signaling pathways. Western blot analyses confirmed that mitogen-activated protein kinase kinase kinase 4 and phospho-ERK1/2 were decreased in PCOS granulosa cells. This study identified candidate genes involved in MAPK/ERK signaling pathways that may influence the function of granulosa cells in PCOS. PMID:26459919

  11. TNF-α-induced depressive-like phenotype and p38(MAPK) activation are abolished by ascorbic acid treatment.

    PubMed

    Moretti, Morgana; Budni, Josiane; Freitas, Andiara Espíndola; Neis, Vivian Binder; Ribeiro, Camille Mertins; de Oliveira Balen, Grasiela; Rieger, Débora Kurrle; Leal, Rodrigo Bainy; Rodrigues, Ana Lúcia S

    2015-06-01

    We investigated the effects of ascorbic acid on depressive-like behavior induced by tumor necrosis factor (TNF-α) in mice. Additionally, we examined the effects of combined administration of ascorbic acid and antidepressants, MK-801 and 7-nitroindazole in mice exposed or not to TNF-α and the capacity of TNF-α and ascorbic acid to modulate hippocampal and cerebrocortical phosphorylation of extracellular signal-regulated kinase (ERK), p38(MAPK) and c-Jun N-terminal kinase (JNK). In control animals, ascorbic acid reduced the immobility time in the tail suspension test (TST). Unilateral intracerebroventricular administration of TNF-α produced a depressive-like behavior in the TST, and the treatment with ascorbic acid prevented this effect. Sub-effective dose of ascorbic acid combined with sub-effective doses of fluoxetine, imipramine, bupropion, MK-801 or 7-nitroindazole produced a synergistic antidepressant-like effect in mice exposed or not to TNF-α. No treatment caused significant alterations in the locomotor activity of mice. Administration of TNF-α increased the phosphorylation of p38(MAPK) in hippocampus and cerebral cortex, and the treatment with ascorbic acid prevented this effect. Ascorbic acid increased phosphorylation of ERK1 in the hippocampus of saline- and TNF-α-treated animals, however it did not produce alterations in the cerebral cortex. No effects on phosphorylation of ERK2 or JNK were found. The observed effect of ascorbic acid seems to be associated, at least partially, with a reduced p38(MAPK) phosphorylation, activation of the monoaminergic systems as well as inhibition of N-methyl-D-aspartate (NMDA) receptors and nitric oxide (NO) synthesis.

  12. The Role of Phospholipase D and MAPK Signaling Cascades in the Adaption of Lichen Microalgae to Desiccation: Changes in Membrane Lipids and Phosphoproteome.

    PubMed

    Gasulla, Francisco; Barreno, Eva; Parages, María L; Cámara, Joaquín; Jiménez, Carlos; Dörmann, Peter; Bartels, Dorothea

    2016-09-01

    Classically, lichen phycobionts are described as poikilohydric organisms able to undergo desiccation due to the constitutive presence of molecular protection mechanisms. However, little is known about the induction of cellular responses in lichen phycobionts during drying. The analysis of the lipid composition of the desiccated lichen microalga Asterochloris erici revealed the unusual accumulation of highly polar lipids (oligogalactolipids and phosphatidylinositol), which prevents the fusion of membranes during stress, but also the active degradation of cone-shaped lipids (monogalactosyldiacylglycerol and phosphatidylethanolamine) to stabilize membranes in desiccated cells. The level of phosphatidic acid increased 7-fold during desiccation, implicating a possible role for phospholipase D (PLD) in the response to osmotic stress. Inhibition of PLD with 1-butanol markedly impaired the recovery of photosynthesis activity in A. erici upon desiccation and salt stress (2 M NaCl). These two hyperosmotic stresses caused the phosphorylation of c-Jun N-terminal kinase (JNK) and p38-like mitogen-activated protein kinase (MAPK) and the dephosphorylation of extracellular signal-regulated kinase (ERK). The incubation with 1-butanol reduced the phosphorylation of JNK-like proteins and increased the dephosphorylation of ERK-like proteins, which indicates an upstream control of MAPK cascades by PLD. The phosphoproteome showed that desiccation caused the phosphorylation of several proteins in A. erici, most of them involved in protein turnover. The results demonstrate that lichen phycobionts possess both constitutive and inducible protective mechanisms to acquire desiccation tolerance. Among others, these responses are controlled by the PLD pathway through the activation of MAPK cascades. PMID:27335354

  13. The Role of Phospholipase D and MAPK Signaling Cascades in the Adaption of Lichen Microalgae to Desiccation: Changes in Membrane Lipids and Phosphoproteome.

    PubMed

    Gasulla, Francisco; Barreno, Eva; Parages, María L; Cámara, Joaquín; Jiménez, Carlos; Dörmann, Peter; Bartels, Dorothea

    2016-09-01

    Classically, lichen phycobionts are described as poikilohydric organisms able to undergo desiccation due to the constitutive presence of molecular protection mechanisms. However, little is known about the induction of cellular responses in lichen phycobionts during drying. The analysis of the lipid composition of the desiccated lichen microalga Asterochloris erici revealed the unusual accumulation of highly polar lipids (oligogalactolipids and phosphatidylinositol), which prevents the fusion of membranes during stress, but also the active degradation of cone-shaped lipids (monogalactosyldiacylglycerol and phosphatidylethanolamine) to stabilize membranes in desiccated cells. The level of phosphatidic acid increased 7-fold during desiccation, implicating a possible role for phospholipase D (PLD) in the response to osmotic stress. Inhibition of PLD with 1-butanol markedly impaired the recovery of photosynthesis activity in A. erici upon desiccation and salt stress (2 M NaCl). These two hyperosmotic stresses caused the phosphorylation of c-Jun N-terminal kinase (JNK) and p38-like mitogen-activated protein kinase (MAPK) and the dephosphorylation of extracellular signal-regulated kinase (ERK). The incubation with 1-butanol reduced the phosphorylation of JNK-like proteins and increased the dephosphorylation of ERK-like proteins, which indicates an upstream control of MAPK cascades by PLD. The phosphoproteome showed that desiccation caused the phosphorylation of several proteins in A. erici, most of them involved in protein turnover. The results demonstrate that lichen phycobionts possess both constitutive and inducible protective mechanisms to acquire desiccation tolerance. Among others, these responses are controlled by the PLD pathway through the activation of MAPK cascades.

  14. Characterization of three mitogen-activated protein kinases (MAPK) genes reveals involvement of ERK and JNK, not p38 in defense against bacterial infection in Yesso scallop Patinopecten yessoensis.

    PubMed

    Sun, Yan; Zhang, Lingling; Zhang, Meiwei; Li, Ruojiao; Li, Yangping; Hu, Xiaoli; Wang, Shi; Bao, Zhenmin

    2016-07-01

    Mitogen-activated protein kinases (MAPKs) are protein Ser/Thr kinases that play a vital role in innate immune responses by converting extracellular stimuli into a wide range of cellular responses. Although MAPKs have been extensively studied in various vertebrates and invertebrates, our current understanding of MAPK signaling cascade in scallop is in its infancy. In this study, three MAPK genes (PyERK, PyJNK, and Pyp38) were identified from Yesso scallop Patinopecten yessoensis. The open reading frame of PyERK, PyJNK, and Pyp38 was 1104, 1227, and 1104 bp, encoding 367, 408, and 367 amino acids, respectively. Conservation in some splicing sites was revealed across the three PyMAPKs, suggesting the common descent of MAPKs genes. The expression profiles of PyMAPKs over the course of ten different developmental stages showed that they had different expression patterns. In adult scallops, PyMAPKs were primarily expressed in muscles, hemocytes, gill, and mantle. To gain insights into their role in innate immunity, we investigated their expression profiles after infection with Gram-positive bacteria (Micrococcus luteus) and Gram-negative bacteria (Vibrio anguillarum). Significant difference in gene expression was only found in PyERK and PyJNK, but not Pyp38, suggesting Pyp38 may not participate in immune response to bacterial infection. Besides, PyERK and PyJNK exhibited more drastic change against the invasion of V. anguillarum than M. luteus, suggesting they could be more sensitive to Gram-negative bacteria than Gram-positive bacteria. This study provides valuable resource for elucidating the role of MAPK signal pathway in bivalve innate immune response. PMID:27155450

  15. A Standardized Chemically Modified Curcuma longa Extract Modulates IRAK-MAPK Signaling in Inflammation and Potentiates Cytotoxicity.

    PubMed

    Rana, Minakshi; Maurya, Preeti; Reddy, Sukka S; Singh, Vishal; Ahmad, Hafsa; Dwivedi, Anil K; Dikshit, Madhu; Barthwal, Manoj K

    2016-01-01

    The TLR/IL-1R pathway is a critical signaling module that is misregulated in pathologies like inflammation and cancer. Extracts from turmeric (Curcuma longa L.) enriched in curcumin and carbonyls like turmerones have been shown to exert potent anti-inflammatory effects. The present study evaluated the anti-inflammatory activity, cytotoxic effect and the underlying mechanism of a novel chemically modified, non-carbonyl compound enriched Curcuma longa L. (C. longa) extract (CMCE). CMCE (1 or 10 μg/mL; 14 h) significantly decreased LPS (50-100 ng/mL) induced TNF-α and IL-1β production in THP-1 cells, human, and mouse whole blood as measured by ELISA. LPS-induced IRAK1, MAPK activation, TLR4 expression, TLR4-MyD88 interaction, and IκBα degradation were significantly reduced in CMCE pre-treated THP-1 cells as assessed by Western blotting. CMCE (30, 100, and 300 mg/kg; 10 days p.o.) pre-treated and LPS (10 mg/kg) challenged Swiss mice exhibited attenuated plasma TNF-α, IL-1β, nitrite, aortic iNOS expression, and vascular dysfunction. In a PI permeability assay, cell lines derived from acute myeloid leukemia were most sensitive to the cytotoxic effects of CMCE. Analysis of Sub-G1 phase, Annexin V-PI positivity, loss of mitochondrial membrane potential, increased caspase-3, and PARP-1 activation confirmed CMCE induced apoptosis in HL-60 cells. IRAK inhibition also sensitized HL-60 cells to CMCE induced cytotoxicity. The present study defines the mechanism underlying the action of CMCE and suggests a therapeutic potential for its use in sepsis and leukemia. PMID:27504095

  16. A Standardized Chemically Modified Curcuma longa Extract Modulates IRAK-MAPK Signaling in Inflammation and Potentiates Cytotoxicity

    PubMed Central

    Rana, Minakshi; Maurya, Preeti; Reddy, Sukka S.; Singh, Vishal; Ahmad, Hafsa; Dwivedi, Anil K.; Dikshit, Madhu; Barthwal, Manoj K.

    2016-01-01

    The TLR/IL-1R pathway is a critical signaling module that is misregulated in pathologies like inflammation and cancer. Extracts from turmeric (Curcuma longa L.) enriched in curcumin and carbonyls like turmerones have been shown to exert potent anti-inflammatory effects. The present study evaluated the anti-inflammatory activity, cytotoxic effect and the underlying mechanism of a novel chemically modified, non-carbonyl compound enriched Curcuma longa L. (C. longa) extract (CMCE). CMCE (1 or 10 μg/mL; 14 h) significantly decreased LPS (50-100 ng/mL) induced TNF-α and IL-1β production in THP-1 cells, human, and mouse whole blood as measured by ELISA. LPS-induced IRAK1, MAPK activation, TLR4 expression, TLR4-MyD88 interaction, and IκBα degradation were significantly reduced in CMCE pre-treated THP-1 cells as assessed by Western blotting. CMCE (30, 100, and 300 mg/kg; 10 days p.o.) pre-treated and LPS (10 mg/kg) challenged Swiss mice exhibited attenuated plasma TNF-α, IL-1β, nitrite, aortic iNOS expression, and vascular dysfunction. In a PI permeability assay, cell lines derived from acute myeloid leukemia were most sensitive to the cytotoxic effects of CMCE. Analysis of Sub-G1 phase, Annexin V-PI positivity, loss of mitochondrial membrane potential, increased caspase-3, and PARP-1 activation confirmed CMCE induced apoptosis in HL-60 cells. IRAK inhibition also sensitized HL-60 cells to CMCE induced cytotoxicity. The present study defines the mechanism underlying the action of CMCE and suggests a therapeutic potential for its use in sepsis and leukemia. PMID:27504095

  17. A Standardized Chemically Modified Curcuma longa Extract Modulates IRAK-MAPK Signaling in Inflammation and Potentiates Cytotoxicity.

    PubMed

    Rana, Minakshi; Maurya, Preeti; Reddy, Sukka S; Singh, Vishal; Ahmad, Hafsa; Dwivedi, Anil K; Dikshit, Madhu; Barthwal, Manoj K

    2016-01-01

    The TLR/IL-1R pathway is a critical signaling module that is misregulated in pathologies like inflammation and cancer. Extracts from turmeric (Curcuma longa L.) enriched in curcumin and carbonyls like turmerones have been shown to exert potent anti-inflammatory effects. The present study evaluated the anti-inflammatory activity, cytotoxic effect and the underlying mechanism of a novel chemically modified, non-carbonyl compound enriched Curcuma longa L. (C. longa) extract (CMCE). CMCE (1 or 10 μg/mL; 14 h) significantly decreased LPS (50-100 ng/mL) induced TNF-α and IL-1β production in THP-1 cells, human, and mouse whole blood as measured by ELISA. LPS-induced IRAK1, MAPK activation, TLR4 expression, TLR4-MyD88 interaction, and IκBα degradation were significantly reduced in CMCE pre-treated THP-1 cells as assessed by Western blotting. CMCE (30, 100, and 300 mg/kg; 10 days p.o.) pre-treated and LPS (10 mg/kg) challenged Swiss mice exhibited attenuated plasma TNF-α, IL-1β, nitrite, aortic iNOS expression, and vascular dysfunction. In a PI permeability assay, cell lines derived from acute myeloid leukemia were most sensitive to the cytotoxic effects of CMCE. Analysis of Sub-G1 phase, Annexin V-PI positivity, loss of mitochondrial membrane potential, increased caspase-3, and PARP-1 activation confirmed CMCE induced apoptosis in HL-60 cells. IRAK inhibition also sensitized HL-60 cells to CMCE induced cytotoxicity. The present study defines the mechanism underlying the action of CMCE and suggests a therapeutic potential for its use in sepsis and leukemia.

  18. TACE release of TNF-α mediates mechanotransduction-induced activation of p38 MAPK and myogenesis

    PubMed Central

    Zhan, Mei; Jin, Bingwen; Chen, Shuen-Ei; Reecy, James M.; Li, Yi-Ping

    2011-01-01

    Summary Skeletal muscle responds to mechanical stimulation by activating p38 MAPK, a key signal for myogenesis. However, the mechanotransduction mechanism that activates p38 is unknown. Here we show that mechanical stimulation of myoblasts activates p38 and myogenesis through stimulating TNF-α release by TNF-α converting enzyme (TACE). In C2C12 or mouse primary myoblasts cultured in growth medium, static stretch activated p38 along with ERK1/2, JNK and AKT. Disrupting TNF-α signaling by TNF-α-neutralizing antibody or knocking out TNF-α receptors blocked stretch activation of p38, but not ERK1/2, JNK or AKT. Stretch also activated differentiation markers MEF2C, myogenin, p21 and myosin heavy chain in a TNF-α- and p38-dependent manner. Stretch stimulated the cleavage activity of TACE. Conversely, TACE inhibitor TAPI or TACE siRNA abolished stretch activation of p38. In addition, conditioned medium from stretched myoblast cultures activated p38 in unstretched myoblasts, which required TACE activity in the donor myoblasts, and TNF-α receptors in the recipient myoblasts. These results indicate that posttranscriptional activation of TACE mediates the mechanotransduction that activates p38-dependent myogenesis via the release of TNF-α. PMID:17264149

  19. Attenuation of the macrophage inflammatory activity by TiO₂ nanotubes via inhibition of MAPK and NF-κB pathways.