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Philip Cohen - One of the best experts on this subject based on the ideXlab platform.
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The phosphorylation of CapZ-interacting protein (CapZIP) by stress-activated protein kinases triggers its dissociation from CapZ.
Biochemical Journal, 2005Co-Authors: Claire E. Eyers, Ana Cuenda, Axel Knebel, Nick Morrice, Helen Mcneill, Simon Arthur, Philip CohenAbstract:A protein expressed in immune cells and muscle was detected in muscle extracts as a substrate for several SAPKs (stress-activated protein kinases). It interacted specifically with the F-actin capping protein CapZ in splenocytes, and was therefore termed 'CapZIP' (CapZ-interacting protein). Human CapZIP was phosphorylated at Ser-179 and Ser-244 by MAPKAP-K2 (mitogen-activated protein kinase-activated protein kinase 2) or MAPKAP-K3 in vitro. Anisomycin induced the phosphorylation of CapZIP at Ser-179 in Jurkat cells, which was prevented by SB 203580, consistent with phosphorylation by MAPKAP-K2 and/or MAPKAP-K3. However, osmotic shock-induced phosphorylation of Ser-179 was unaffected by SB 203580. These and other results suggest that CapZIP is phosphorylated at Ser-179 in cells by MAPKAP-K2/MAPKAP-K3, and at least one other protein kinase. Stress-activated MAP kinase family members phosphorylated human CapZIP at many sites, including Ser-68, Ser-83, Ser-108 and Ser-216. Ser-108 became phosphorylated when Jurkat cells were exposed to osmotic shock, which was unaffected by SB 203580 and/or PD 184352, or in splenocytes from mice that do not express either SAPK3/p38gamma or SAPK4/p38delta. Our results suggest that CapZIP may be phosphorylated by JNK (c-Jun N-terminal kinase), which phosphorylates CapZIP to >5 mol/mol within minutes in vitro. Osmotic shock or anisomycin triggered the dissociation of CapZIP from CapZ in Jurkat cells, suggesting that phosphorylation of CapZIP may regulate the ability of CapZ to remodel actin filament assembly in vivo.
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TAB3, a new binding partner of the protein kinase TAK1.
Biochemical Journal, 2004Co-Authors: Peter C. F. Cheung, Angel R. Nebreda, Philip CohenAbstract:We have identified a new binding partner of the TGFbeta (transforming growth factor-beta)-activated protein kinase (TAK1), termed TAB3 (TAK1-binding protein-3), which shares 48% amino acid sequence identity with TAB2. Our results indicate that two distinct TAK1 complexes are present in cells. One comprises TAK1 complexed with TAB1 and TAB2, and the other TAK1 complexed with TAB1 and TAB3. Both complexes are activated in response to tumour necrosis factor-alpha or interleukin-1 in human epithelial KB cells or bacterial lipopolysaccharide in RAW264.7 macrophages, and are subject to feedback control by stress-activated protein kinase 2a (SAPK2a; also called p38alpha). The electrophoretic mobility of TAB2 and TAB3 decreases in response to these agonists or osmotic shock, and is reversed by treatment with protein phosphatase-1. The decrease in mobility of TAB3 is prevented if the cells are incubated with SB 203580 before stimulation, but treatment with SB 203580 produces forms of TAB2 with a mobility intermediate between that observed for TAB2 in unstimulated and stimulated cells. Similar results were obtained in embryonic fibroblasts from mice deficient in SAPK2a/p38alpha. Our results indicate that TAB3 is phosphorylated via the SAPK2a/p38alpha pathway, whereas TAB2 is phosphorylated at two or more sites by both an SAPK2a/p38alpha-dependent and an SB 203580-independent kinase. The SAPK2a/p38alpha-mediated phosphorylation of TAB2 and TAB3 may contribute to the SAPK2a/p38alpha-mediated feedback control of TAK1 activity that also involves the phosphorylation of TAB1. We also show that the agonist-induced activation of TAK1 complexes requires the phosphorylation of the TAK1 catalytic subunit at a serine/threonine residue(s).
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Feedback control of the protein kinase TAK1 by SAPK2a/p38α
The EMBO Journal, 2003Co-Authors: Peter C. F. Cheung, David G. Campbell, Angel R. Nebreda, Philip CohenAbstract:TAB1, a subunit of the kinase TAK1, was phosphorylated by SAPK2a/p38α at Ser423, Thr431 and Ser438 in vitro. TAB1 became phosphorylated at all three sites when cells were exposed to cellular stresses, or stimulated with tumour necrosis factor-α (TNF-α), interleukin-1 (IL-1) or lipopolysaccharide (LPS). The phosphorylation of Ser423 and Thr431 was prevented if cells were pre-incubated with SB 203580, while the phosphorylation of Ser438 was partially inhibited by PD 184352. Ser423 is the first residue phosphorylated by SAPK2a/p38α that is not followed by proline. The activation of TAK1 was enhanced by SB 203580 in LPS-stimulated macrophages, and by proinflammatory cytokines or osmotic shock in epithelial KB cells or embryonic fibroblasts. The activation of TAK1 by TNF-α, IL-1 or osmotic shock was also enhanced in embryonic fibroblasts from SAPK2a/p38α-deficient mice, while incubation of these cells with SB 203580 had no effect. Our results suggest that TAB1 participates in a SAPK2a/p38α-mediated feedback control of TAK1, which not only limits the activation of SAPK2a/p38α but synchronizes its activity with other signalling pathways that lie downstream of TAK1 (JNK and IKK).
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Inhibition of SAPK2a/p38 prevents hnRNP A0 phosphorylation by MAPKAP-K2 and its interaction with cytokine mRNAs.
The EMBO Journal, 2002Co-Authors: Simon Rousseau, Nick Morrice, Mark Peggie, David G. Campbell, Matthias Gaestel, Philip CohenAbstract:Lipopolysaccharide (LPS) stimulates production of inflammatory mediators, partly by stabilizing [interleukin-6 (IL-6), cyclooxygenase 2 (COX-2)] and/or stimulating translation [tumour necrosis factor-α (TNF-α)] of their mRNAs. Such regulation depends on AU-rich elements (AREs) within the 3′-untranslated regions and is partially suppressed by SB 203580 (which inhibits SAPK2a/p38). The LPS-induced production of TNF-α and IL-6 is suppressed in MAPKAP-K2-deficient mice (a kinase activated by SAPK2a/p38). Here, we identify 18 macrophage proteins that bind to AREs and show that hnRNP A0 is a major substrate for MAPKAP-K2 in this fraction. MAPKAP-K2 phosphorylated hnRNP A0 at Ser84 in vitro and this residue became phosphorylated in LPS-stimulated cells. Phosphorylation was prevented by SB 203580 and suppressed in macrophages derived from MAPKAP-K2-deficient mice. The mRNAs encoding TNF-α, COX-2 and macrophage inflammatory protein-2 (MIP-2) bound to hnRNP A0 in LPS-stimulated macrophages, an interaction prevented by SB 203580. The LPS-induced stabilization of MIP-2 mRNA and production of MIP-2 protein were abolished when macrophages were incubated with SB 203580 plus PD 184352 (which inhibits the classical MAP kinase cascade). Our data suggest that LPS-induced binding of hnRNP A0 to AREs may contribute to the post-transcriptional regulation of specific mRNAs.
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Stress-induced regulation of eukaryotic elongation factor 2 kinase by SB 203580-sensitive and -insensitive pathways.
The Biochemical journal, 2002Co-Authors: Axel Knebel, Claire E Haydon, Nick Morrice, Philip CohenAbstract:Eukaryotic elongation factor 2 (eEF2) kinase, the enzyme that inactivates eEF2, is controlled by phosphorylation. Previous work showed that stress-activated protein kinase 4 (SAPK4, also called p38delta) inhibits eEF2 kinase in vitro by phosphorylating Ser-359, while ribosomal protein S6 kinases inhibit eEF2 kinase by phosphorylating Ser-366 [Knebel, Morrice and Cohen (2001) EMBO J. 20, 4360-4369; Wang, Li, Williams, Terada, Alessi and Proud (2001) EMBO J. 20, 4370-4379]. In the present study we have examined the effects of the protein synthesis inhibitor anisomycin and tumour necrosis factor-alpha (TNF-alpha) on the phosphorylation of eEF2 kinase. We demonstrate that Ser-359, Ser-366 and two novel sites (Ser-377 and Ser-396) are all phosphorylated in human epithelial KB cells, but only the phosphorylation of Ser-359 and Ser-377 increases in response to these agonists and correlates with the dephosphorylation (activation) of eEF2. Ser-377 is probably a substrate of MAPKAP-K2/K3 (mitogen-activated protein kinase-activated protein kinase 2/kinase 3) in cells, because eEF2 kinase is phosphorylated efficiently by these protein kinases in vitro and phosphorylation of this site, induced by TNF-alpha and low (but not high) concentrations of anisomycin, is prevented by SB 203580, which inhibits SAPK2a/p38, their "upstream" activator. The phosphorylation of Ser-359 induced by high concentrations of anisomycin is probably catalysed by SAPK4/p38delta in cells, because no other stress-activated, proline-directed protein kinase tested phosphorylates this site in vitro and phosphorylation is insensitive to SB 203580. Interestingly, the phosphorylation of Ser-359 induced by TNF-alpha or low concentrations of anisomycin is suppressed by SB 203580, indicating that phosphorylation is also mediated by a novel pathway. Since the phosphorylation of Ser-377 does not inhibit eEF2 kinase in vitro, our results suggest that anisomycin or TNF-alpha inhibit eEF2 kinase via the phosphorylation of Ser-359.
Emilia Galdiero - One of the best experts on this subject based on the ideXlab platform.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways
Medical Microbiology and Immunology, 2005Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-α and IL-1β mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways.
Medical microbiology and immunology, 2003Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-alpha and IL-1beta mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
Michel Goedert - One of the best experts on this subject based on the ideXlab platform.
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Paradoxical activation of Raf by a novel Raf inhibitor.
Chemistry & Biology, 1999Co-Authors: Clare A Hall-jackson, Michel Goedert, Philip Cohen, Patrick A. Eyers, F Tom Boyle, Neil Hewitt, Helen Plant, Philip HedgeAbstract:Background Raf is a proto-oncogene that is activated in response to growth factors or phorbol esters, and is thought to activate MAP kinase kinase-1 (MKK1) and hence the classical MAP kinase (MAPK) cascade. Results The compound ZM 336372 is identified as a potent and specific inhibitor of Raf isoforms in vitro . Paradoxically, exposure of cells to ZM 336372 induces > 100-fold activation of c-Raf (measured in the absence of compound), but without triggering any activation of MKK1 or p42 MAPK/ERK2. The ZM 336372-induced activation of c-Raf occurs without any increase in the GTP-loading of Ras and is not prevented by inhibition of the MAPK cascade, protein kinase C or phosphatidylinositide 3-kinase. ZM 336372 does not prevent growth factor or phorbol ester induced activation of MKK1 or p42 MAPK/ERK2, or reverse the phenotype of Ras- or Raf-transformed cell lines. The only other protein kinase inhibited by ZM 336372 out of 20 tested was SAPK2/p38. Although ZM 336372 is structurally unrelated to SB 203580, a potent inhibitor of SAPK2/p38, the mutation of Thr106→Met made SAPK2/p38 insensitive to ZM 336372 as well as to SB 203580. Conclusions Raf appears to suppress its own activation by a novel feedback loop, such that inhibition is always counterbalanced by reactivation. These observations imply that some agonists reported to trigger the cellular activation of c-Raf might actually be inhibitors of this enzyme, and that compounds which inhibit the kinase activity of Raf might not be useful as anticancer drugs. The binding sites for ZM 336372 and SB 203580 on Raf and SAPK2/p38 are likely to overlap.
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Use of a drug‐resistant mutant of stress‐activated protein kinase 2a/p38 to validate the in vivo specificity of SB 203580
FEBS Letters, 1999Co-Authors: Patrick A. Eyers, Michel Goedert, Paul R.l.a. Van Den Ijssel, Roy A. Quinlan, Philip CohenAbstract:Stress-activated protein kinase 2a, also called p38, is inhibited by SB 203580 and this drug has been used widely to implicate this enzyme in the regulation of many physiological processes. Here, we introduce a novel method of general application, which can be used to establish whether the effects of SB 203580 are mediated via inhibition of stress-activated protein kinase 2a/p38 or whether they result from ‘non-specific’ effects. Four events thought to occur upon activation of stress-activated protein kinase 2a/p38 have been established unequivocally. These are the activation of mitogen-activated protein kinase-activated protein kinase-2 and mitogen- and stress-activated protein kinase-1 and the phosphorylation of their presumed substrates, heat shock protein 27 and the transcription factor cyclic AMP response element binding protein, respectively. In contrast, the SB 203580-induced activation of c-Raf is independent of stress-activated protein kinase 2a/p38 inhibition.
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use of a drug resistant mutant of stress activated protein kinase 2a p38 to validate the in vivo specificity of SB 203580
FEBS Letters, 1999Co-Authors: Patrick A. Eyers, Michel Goedert, Paul R.l.a. Van Den Ijssel, Roy A. Quinlan, Philip CohenAbstract:Stress-activated protein kinase 2a, also called p38, is inhibited by SB 203580 and this drug has been used widely to implicate this enzyme in the regulation of many physiological processes. Here, we introduce a novel method of general application, which can be used to establish whether the effects of SB 203580 are mediated via inhibition of stress-activated protein kinase 2a/p38 or whether they result from ‘non-specific’ effects. Four events thought to occur upon activation of stress-activated protein kinase 2a/p38 have been established unequivocally. These are the activation of mitogen-activated protein kinase-activated protein kinase-2 and mitogen- and stress-activated protein kinase-1 and the phosphorylation of their presumed substrates, heat shock protein 27 and the transcription factor cyclic AMP response element binding protein, respectively. In contrast, the SB 203580-induced activation of c-Raf is independent of stress-activated protein kinase 2a/p38 inhibition.
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Effect of SB 203580 on the activity of c-Raf in vitro and in vivo.
Oncogene, 1999Co-Authors: Clare A Hall-jackson, Michel Goedert, Philip Hedge, Philip CohenAbstract:The inhibition of SAPK2a/p38 (a mitogen activated protein (MAP) kinase family member) by SB 203580 depends on the presence of threonine at residue 106. Nearly all other protein kinases are insensitive to this drug because a more bulky residue occupies this site (Eyers et al., 1998). Raf is one of the few protein kinases that possesses threonine at this position, and we show that SB 203580 inhibits c-Raf with an IC50 of 2 μM in vitro. However, SB 203580 does not suppress either growth factor or phorbol ester-induced activation of the classical MAP kinase cascade in mammalian cells. One of the reasons for this is that SB 203580 also triggers a remarkable activation of c-Raf in vivo (when measured in the absence of the drug). The SB 203580-induced activation of c-Raf occurs without any increase in the GTP-loading of Ras, is not prevented by inhibitors of the MAPK cascade, protein kinase C or phosphatidylinositide 3-kinase, and is not triggered by the binding of this drug to SAPK2a/p38. The paradoxical activation of c-Raf by SB 203580 (and by another structurally unrelated c-Raf inhibitor) suggests that inhibitors of the kinase activity of c-Raf may not be effective as anti-cancer drugs.
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Effect of SB 203580 on the activity of c-Raf in vitro and in vivo
Oncogene, 1999Co-Authors: Clare A Hall-jackson, Michel Goedert, Philip Hedge, Philip CohenAbstract:The inhibition of SAPK2a/p38 (a mitogen activated protein (MAP) kinase family member) by SB 203580 depends on the presence of threonine at residue 106. Nearly all other protein kinases are insensitive to this drug because a more bulky residue occupies this site ( Eyers et al., 1998 ). Raf is one of the few protein kinases that possesses threonine at this position, and we show that SB 203580 inhibits c-Raf with an IC_50 of 2 μ M in vitro . However, SB 203580 does not suppress either growth factor or phorbol ester-induced activation of the classical MAP kinase cascade in mammalian cells. One of the reasons for this is that SB 203580 also triggers a remarkable activation of c-Raf in vivo (when measured in the absence of the drug). The SB 203580-induced activation of c-Raf occurs without any increase in the GTP-loading of Ras, is not prevented by inhibitors of the MAPK cascade, protein kinase C or phosphatidylinositide 3-kinase, and is not triggered by the binding of this drug to SAPK2a/p38. The paradoxical activation of c-Raf by SB 203580 (and by another structurally unrelated c-Raf inhibitor) suggests that inhibitors of the kinase activity of c-Raf may not be effective as anti-cancer drugs.
Marilena Galdiero - One of the best experts on this subject based on the ideXlab platform.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways
Medical Microbiology and Immunology, 2005Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-α and IL-1β mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways.
Medical microbiology and immunology, 2003Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-alpha and IL-1beta mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
Anna Longanella - One of the best experts on this subject based on the ideXlab platform.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways
Medical Microbiology and Immunology, 2005Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-α and IL-1β mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
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Induction of cytokine mRNA expression in U937 cells by Salmonella typhimurium porins is regulated by different phosphorylation pathways.
Medical microbiology and immunology, 2003Co-Authors: Marilena Galdiero, Annalisa Tortora, Nicola Damiano, Mariateresa Vitiello, Anna Longanella, Emilia GaldieroAbstract:Lipopolysaccharide (LPS) and porins of Gram-negative outer membranes are the main pathogenic factors implicated in the clinical syndrome of septic shock. The biological activity of porins and LPS are similar, but they occur by different mechanisms. It seems that porins act through different intracellular pathways with respect to LPS. In this study we analyzed the role of several inhibitors of the MEK/ERK signal pathway on the induction of proinflammatory and immunological cytokines in U937 cell line stimulated by Salmonella typhimurium porins and compared it to the cytokine induction after LPS stimulation. We investigated the effects of p38 MAP kinase inhibitor SB-203580, MEK/ERK kinase inhibitor PD-098059 and Raf-1 inhibitor forskolin, and demonstrated that they modulate cytokine mRNA expression in a different manner as a consequence of the use of porins or LPS as stimuli. TNF-alpha and IL-1beta mRNA expression is decreased by PD-098059 after stimulation with LPS but not with porins in differentiated U937 cells. IL-10 mRNA expression is inhibited by SB-203580 and PD-098059 after stimulation with porins in U937 cells. IL-6 and IL-8 mRNA expression is not changed by PD-098059 or SB-203580, after stimulation either with porins or LPS. Furthermore, mRNA expression of the studied cytokines, except for GM-CSF, is not changed using forskolin.
