MAP kinase subtypes and Akt regulate diosgenin-induced apoptosis of rheumatoid synovial cells in association with COX-2 expression and prostanoid production

In the present study, we investigated the signalling pathways involved in diosgenin-induced apoptosis in human rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) in vitro with particular interest on Akt and MAPKs activation in relation to arachidonic acid metabolism via COX-2 pathway. MAPK activation was measured by ELISA quantification in diosgenin-treated human RA FLS. Expression of Akt and phospho-Akt was analyzed by Western blot analysis. Nuclear factor-kappaB (NF-kappaB) translocation was evaluated by electromobility shift assay. The prostanoid production (COX-2 activity) was measured by quantitative ELISA. Diosgenin-induced apoptosis in the presence of MAPK or Akt inhibitors was detected by a quantitative determination of DNA fragmentation. Treatment of human RA FLS with 40 microM diosgenin caused an activation of p38 and JNK and an inhibition of ERK phosphorylation. Akt and NF-kappaB are potentially required for diosgenin-induced apoptosis in human RA FLS because 40 microM diosgenin abrogated Akt phosphorylation which correlated with an inhibition of NF-kappaB nuclear translocation. SB203580 and SP600125 (p38 and JNK inhibitors) reduced diosgenin-induced DNA fragmentation whereas U0126 and LY294002 (MEK and PI3 kinase/Akt inhibitors) caused an amplification of proapoptotic effect of diosgenin. Diosgenin increased COX-2 activity resulting in PGE2 and 6-keto-PGF1alpha overproduction in human RA FLS. All MAPK inhibitors markedly reduced diosgenin-induced PGE2 and 6-keto-PGF1alpha synthesis except for SP600125 on 6-keto-PGF1alpha production. These results provide, for the first time, strong evidence that a combined association implicating a MEK inhibitor (U0126) and diosgenin is the most effective in inducing very strong apoptosis with down-regulation of COX-2 expression and activity in human RA FLS.     

Molecular mechanisms of lipopolysaccharide-induced cyclooxygenase-2 expression in human neutrophils: involvement of the mitogen-activated protein kinase pathway and regulation by anti-inflammatory cytokines

Neutrophils are an important cellular source of proinflammatory mediators, whose regulation may be of potential benefit for the treatment of a number of inflammatory diseases. However, the mechanisms of lipopolysaccharide (LPS)-induced neutrophil activation and its regulation by anti-inflammatory cytokines have not yet been fully elucidated. Recent studies have revealed that mitogen-activated protein kinases (MAPK) play a crucial role in the generation of proinflammatory mediators in some cell types. Therefore, we conducted this study to determine whether MAPK activation could be involved in prostaglandin E(2) (PGE(2)) production and cyclooxygenase (COX)-2 expression in LPS-stimulated human neutrophils. PD98059 (MEK1 inhibitor) and SB203580 (p38(MAPK) inhibitor) reduced PGE(2) production as well as COX-2 expression in LPS-stimulated neutrophils. In addition, both extracellular signal-regulated protein kinase (ERK) and p38(MAPK) were phosphorylated and activated in time- and dose-dependent manners. Since we previously showed that IL-10 and IL-4 similarly inhibited COX-2 expression in LPS-stimulated neutrophils, we next tested the effects of IL-10 and IL-4 on the phosphorylation and activation of both kinases. IL-10 inhibited the phosphorylation and activation of p38(MAPK), but not ERK. In addition, IL-4 caused a marginal inhibition in the activation of p38(MAPK). Taken together, these results suggest that both ERK and p38(MAPK) pathways are involved in LPS-induced COX-2 expression and PGE(2) production in neutrophils, and IL-10 and IL-4 inhibit neutrophil prostanoid synthesis by down-regulating the activation of p38(MAPK).     

Close association of p38 and JNK with plasminogen-dependent upregulation of PAI-1 in rat astrocytes in vitro

As reported previously, stimulation of astrocytes with plasminogen (PLGn) remarkably enhances their production/release of plasminogen activator inhibitor-1 (PAI-1). In addition, both p38 mitogen-activated protein kinase (p38MAPK) and c-Jun N-terminal kinase (JNK) are activated in these astrocytes. However, it remains to be determined whether the MAPK activation is associated with the PAI-1 induction in PLGn-stimulated astrocytes. In the present study, we investigated the relationship between MAPK activity and PAI-1 induction in PLGn-stimulated astrocytes. PLGn stimulation led to definitive phosphorylation of three MAPKs: external signal regulated kinase (ERK), JNK and p38. These results suggest that all of these MAPKs, either alone or in combination, are involved in PAI-1 induction. To verify this association, an inhibition experiment was carried out by using inhibitors specific for each MAPK. The results of the immunoblotting analysis indicated that 20 μM SB203580 (the p38 inhibitor) or SP600125 (the JNK inhibitor) suppressed approximately 85% or 40% of PLGn-inducible PAI-1, respectively. Only 20% inhibition was achieved by pretreatment of astrocytes with 20 μM PD98059 (the inhibitor of MEK1/2, an upstream kinase of ERK). In conclusion, p38 and JNK were shown to be the major MAPKs involved in the signaling cascade leading to PAI-1 induction in astrocytes stimulated with PLGn.     

Low concentrations of nitric oxide (NO) induced cell death in PC12 cells through activation of p38 mitogen-activated protein kinase (p38 MAPK) but not via extracellular signal-regulated kinases (ERK1/2) or c-Jun N-terminal protein kinase (JNK)

Nitric oxide (NO), a highly reactive gaseous molecule, has been previously reported to induce apoptosis-like cell death even at a low concentration in PC12 cells. In this study, we examined NO-induced activation of members of the mitogen-activated protein kinase (MAPK) family, i.e., p38 MAPK, extracellular signal-regulated kinases (ERK1/2), and c-Jun N-terminal protein kinase (JNK). Following the exposure of PC12 cells to an NO donor, (+)-(E)-4-ethyl-2-[hydroxyimino]-5-nitro-3-hexenamide (NOR3; 100 muM), the phosphorylation level of p38 MAPK increased time dependently from 2 to 6 h, but that of both ERK1/2 and JNK did not. Treatment with a p38 MAPK inhibitor SB203580 partially blocked the NOR3-induced cell death. Neither PD98059, U0126 (inhibitors of ERK1/2) nor SP600125 (a specific inhibitor of JNK) treatments had any significant effect on the NOR3-induced cell death. These findings suggest that the activation of a p38 MAPK pathway, but not that of ERK1/2 or JNK, plays an essential role in the apoptosis-like cell death induced by low concentrations of NO.     

Role of the Pyk2–MAP Kinase–cPLA2 Signaling Pathway in Shear-Dependent Platelet Aggregation

Mechanisms of shear-induced platelet aggregation are not established. Data that ristocetin-induced von Willebrand factor (VWF) binding to glycoprotein (Gp) Ibalpha activates proline-rich tyrosine kinase 2 (Pyk2) and extracellular-regulated kinase (ERK) has led to speculation that these events are coupled and that a MAP kinase may activate cytosolic phospholipase A2 (cPLA2)-mediated arachidonic acid (AA) release. To test this hypothesis and clarify the role of AA metabolism in shear-induced VWF-dependent platelet aggregation, we examined Pyk2, ERK1/2, and p38 phosphorylation, and arachidonic acid release and metabolism in platelets subjected to pathological shear stress in vitro. We observe tyrosine phosphorylation of Pyk2, p38, and ERK1/2 but no measurable increase in free AA, 12-hydroxyeicosatetraenoic acid, or thromboxane A2. Inhibitors of ERK, p38, or cyclooxygenase activation fail to affect shear-induced platelet aggregation. When washed platelets are aspirin-pretreated, arachidonic acid release becomes measurable and aggregation at 60 and 120 s is attenuated. These data indicate that shear-induced VWF binding to platelet GpIb-IX-V activates Pyk2, ERK1/2, p38, and cPLA2, but that the magnitude of these responses is below the threshold needed to enhance shear-induced VWF-dependent platelet aggregation in the presence of plasma. These results provide a mechanistic basis for the long-standing observation that shear-dependent platelet aggregation is unaffected by the antiplatelet drug aspirin.     

Role of PKC-delta during hypoxia in heart-derived H9c2 cells

In the present study, we investigated the role of protein kinase C (PKC) isoforms during hypoxia in heart-derived H9c2 cells. Hypoxia caused a rapid translocation of PKC-delta from soluble to particulate fraction and a downregulation of PKC-epsilon and PKC-zeta, whereas PKC-alpha and PKC-beta I remained unaltered. When H9c2 cells were pretreated with PKC-delta inhibitor rottlerin (3 microM), hypoxia-induced apoptotic and necrotic cell death were significantly increased. Hypoxic insult also caused an activation of extracellular signal-regulated protein kinase (ERK) and p38 MAPK with no change in c-Jun NH(2)-terminal protein kinase (JNK) phosphorylation. Hypoxia-induced cell death was increased by treatment with ERK1/2 inhibitor U0126 (10 microM), but attenuated by p38 MAPK inhibitor SB202190 (10 microM). Treatment with rottlerin completely blocked the hypoxia-induced ERK phosphorylation, whereas it significantly increased p38 MAPK phosphorylation. The hypoxia-induced translocation of PKC-delta was not altered by U0126 and/or SB202190. From these results, it is suggested that hypoxia causes a rapid translocation of PKC-delta and subsequently ERK activation and p38 inactivation, rendering H9c2 cells resistant to hypoxia-induced cell death.     

Regulation of adhesion of AML14.3D10 cells by surface clustering of beta2-integrin caused by ERK-independent activation of cPLA2

We examined the role of cell surface clustering of beta2-integrin caused by protein kinase C (PKC)-activated-cPLA2 in adhesion of eosinophilic AML14.3D10 (AML) cells. Phorbol 12-myristate 13-acetate (PMA) caused time- and concentration-dependent adhesion of AML cells to plated bovine serum albumin (BSA), which was blocked by anti-CD11b or anti-CD18 monoclonal antibodies (mAb) directed against beta2-integrin. Inhibition of PKC with Ro-31-8220 or rottlerin blocked PMA-induced cell adhesion in a concentration-dependent fashion. Inhibition of cytosolic phospholipase A2 (cPLA2) with trifluoromethyl ketone or methyl arachidonyl fluorophosphonate also blocked PMA-induced cell adhesion. PMA caused time-dependent p42/44 mitogen-activated protein kinase (MAPK) (ERK) phosphorylation in these cells. U0126, a MAPK/extracellular signal-regulated protein kinase kinase (MEK) inhibitor, at the concentrations that blocked PMA-induced ERK phosphorylation, had no effect on PMA stimulated AML cell adhesion. Neither p38 MAPK nor c-Jun N-terminal kinase (JNK) was phosphorylated by PMA. PMA also caused increased cPLA2 activity, which was inhibited by Ro-31-8220, but not U0126. Confocal immunofluorescence microscopy showed that PMA caused clustering of CD11b on the cell surface, which was blocked by either PKC or cPLA2 inhibition. PMA stimulation also caused up-regulation of CD11b on the AML cell surface. However, this up-regulation was not affected by cPLA2- or PKC-inhibition. Using the mAb, CBRM1/5, we also demonstrated that PMA does not induce the active conformation of CD11b/CD18. Our data indicate that PMA causes AML cell adhesion through beta2-integrin by PKC activation of cPLA2. This pathway is independent of MEK/ERK and does not require change of CD11b/CD18 to its active conformation. We find that avidity caused by integrin surface clustering - rather than conformational change or up-regulation of CD11b/CD18 - causes PMA stimulated adhesion of AML cells.     

Stress-activated protein kinases mediate cell migration in human airway epithelial cells

Airway epithelial cell (AEC) repair immediately after injury requires coordinated cell spreading and migration at the site of injury. Stress-activated protein kinases such as p38 MAPK and c-Jun N-terminal Protein Kinase (JNK) modulate several responses to cell stress and injury, but their role in AEC migration is not clear. We examined migration in confluent 16HBE14o(-) human AEC lines and in primary AEC grown on collagen-IV. Wounds were created by mechanical abrasion and followed to closure using digital microscopy. Inhibitors of either p38 extracellular signal-regulated kinase (ERK)1/2 (PD98059), mitogen-activated protein kinase (MAPK) (SB203580), or JNK (SP600125) could block cell migration substantially. Inhibiting JNK but not p38 MAPK or ERK1/2 blocked extension of cells into the wound region from the original line of injury. Initial migration was associated with phosphorylation of ERK, p38 MAPK, and JNK within 5-15 min. The downstream effector of p38, heat shock protein 27, also was phosphorylated rapidly after injury; phosphorylation could be blocked by prior treatment with SB203580 but not SP600125. The downstream effector of JNK, c-Jun, likewise was phosphorylated rapidly after injury and could be blocked by inhibiting JNK. Our data demonstrate that p38 MAPK, JNK, and ERK1/2 participate in the early stages of AEC migration.     

Aldosterone upregulates connective tissue growth factor gene expression via p38 MAPK pathway and mineralocorticoid receptor in ventricular myocytes

The effect of aldosterone on connective tissue growth factor (CTGF) was examined in rat embryonic ventricular myocytes. Upon aldosterone treatment, CTGF expression was significantly increased in a dose and time-dependent manner. To explore the molecular mechanism for this upregulation, we examined the role of mineralocorticoid receptor. Pre-treatment of an antagonist (spironolactone) at 5-fold excess of aldosterone blocked the CTGF induction by aldosterone, suggesting that the upregulation was mediated by mineralocorticoid receptor. Aldosterone treatment resulted in activation of ERK1/2, p38 MAPK, and JNK pathways with a more transient pattern in p38 MAPK. Blocking studies using pretreatment of the inhibitor of each pathway revealed that p38 MAPK cascade may be important for aldosterone-mediated CTGF upregulation as evidenced by the blocking of CTGF induction by SB203580 (p38 MAPK inhibitor), but not by PD098059 (ERK1/2 inhibitor) and JNK inhibitor I. Interestingly, JNK inhibitor I and PD098059 decreased the basal level of CTGF expression. On the other hand, pretreatment of spironolactone abrogated the p38 MAPK activation, indicating that mineralocorticoid receptor mechanism is linked to p38 MAPK pathway. Taken together, our findings suggest that aldosterone induces CTGF expression via both p38 MAPK cascade and mineralocorticoid receptor and that cross-talk exists between the two pathways.     

Tumor necrosis factor-alpha up-regulates the expression of CCL2 and adhesion molecules of human proximal tubular epithelial cells through MAPK signaling pathways

Both circulating and urinary tumor necrosis factor (TNF)-alpha levels have been shown to increase in inflammatory chronic kidney diseases and TNF-alpha can induce secretion of other inflammatory mediators from many cell types. Chemokine, mononuclear chemoattractant protein-1 (CCL2/MCP-1), and cell surface adhesion molecules, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), in renal proximal tubular epithelial cells (PTEC) are important for promoting recruitment and adhesion of infiltrating macrophages and lymphocytes to inflamed renal tissue. This study aimed to investigate the effect of TNF-alpha on the expression of these inflammation-related molecules of human PTEC and the underlying intracellular mitogen-activated protein kinase (MAPK) regulatory signaling mechanisms. Cytokine expression profile of TNF-alpha-activated PTEC was assayed by protein array. The concentration of CCL2 was analyzed by ELISA, while the expression of cell surface ICAM-1 and VCAM-1 and intracellular phosphorylated p38 MAPK, c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) was assessed using flow cytometry. TNF-alpha could significantly induce CCL2, ICAM-1 and VCAM-1 expression of PTEC. Selective inhibitors of p38 MAPK (SB203580), JNK (SP600125) and ERK (PD98059) could suppress TNF-alpha-induced CCL2 and ICAM-1 expression, while only p38 MAPK and ERK inhibitors could suppress TNF-alpha-induced VCAM-1 expression. JNK inhibitor was found to up-regulate VCAM-1 expression but did not elicit any additive effect with TNF-alpha on VCAM-1 expression. Moreover, p38 MAPK inhibitor was found to abrogate the TNF-alpha-induced ERK phosphorylation, suggesting that there was a one-way interaction between p38 MAPK and ERK pathways during the TNF-alpha activation. TNF-alpha can play a crucial role in the immunopathogenesis of nephritis by the induction of CCL2, ICAM-1 and VCAM-1 expression via the activation of the intracellular MAPK signaling pathway, which may contribute to macrophage and lymphocyte infiltration.     

Induction of neurite outgrowth in PC12 cells by the medium-chain fatty acid octanoic acid

It has been shown that polyunsaturated fatty acids such as arachinonic and docosahexanoic acids but not monounsaturated and saturated long-chain fatty acids promote basal and nerve growth factor (NGF)-induced neurite extension of PC12 cells, a line derived from a rat pheochromocytoma. On the other hand, short-chain fatty acids and valproic acid (2-propylpentanoic acid) enhance the growth of neurite processes of the cells only in the presence of inducers. In this study, we demonstrated that straight medium-chain fatty acids (MCFAs) at millimolar concentrations alone potently induced neuronal differentiation of PC12 cells. Hexanoic, heptanoic and octanoic acids dose-dependently induced neurite outgrowth of the cells: their maximal effects determined 2 days after addition to the culture medium were more marked than the effect of NGF. PC12 cells exposed to octanoic acid expressed increased levels of the neuronal marker beta-tubulin isotype III. Nonanoic, decanoic, and dodecanoic acids also induced growth of neurite processes, but their maximal effects were less marked than that of octanoic acid. In contrast, the polyunsaturated fatty acid linoleic acid and short-chain fatty acids had only slight or almost no effects on neurite formation in the absence of NGF. The effect of octanoic acid was synergistic with or additive to the effects of NGF and dibutyryl cyclic AMP. Octanoic acid upregulated phosphorylation of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK), critical signaling molecules in neuronal differentiation, but not phosphorylation of Akt, a signaling molecule downstream of phosphatidylinositol 3-kinase (PI3K). Moreover, growth of neurites induced by octanoic acid was potently inhibited by treatment of cells with the p38 MAPK inhibitor SB203580 and the ERK kinase inhibitor PD98059 but not inhibited and only slightly inhibited by the JNK inhibitor SP600125 and the PI3K inhibitor wortmannin, respectively. Taken together, our results indicate that MCFAs, including octanoic acid, induced neurite outgrowth of PC12 cells in the absence of NGF and suggest that the activation of p38 MAPK and ERK pathways is involved in this process.     

Possible pathophysiological roles of mitogen-activated protein kinases (MAPKs) in endometriosis

PROBLEM: Endometriosis accompanies local inflammatory reactions in the peritoneal cavity. We examined the phosphorylation of mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated kinase (ERK), p38 MAPK (p38) and c-Jun N-terminal kinase (JNK) in endometriotic stromal cells, and their possible pathophysiological roles in endometriosis in relation to proinflammatory substances. METHOD OF STUDY: Endometriotic stromal cells were isolated from endometriomas and were cultured for the experiments. Phosphorylation of MAPKs in endometriotic stromal cells treated with interleukin (IL)-1beta, tumor necrosis factor (TNF)alpha and H(2)O(2) were examined by Western blot analysis. Effects of PD98059, SB202190 and SP600125 (inhibitors of ERK, p38 and JNK, respectively) on IL-1beta-induced secretion of IL-6 and IL-8, and on IL-1beta-induced expression of cyclo-oxygenase-2 (COX-2) in endometriotic cells were studied. In addition, eutopic endometrial tissues were collected, and the phosphorylation rate of p38 in eutopic endometrial tissues and endometriotic tissues were determined. RESULTS: IL-1beta, TNFalpha and H(2)O(2) stimulated the phosphorylation of ERK, p38 and JNK, while the total amounts of proteins of the respective MAPKs were virtually the same compared with those in the unstimulated controls. Both SB202190 and SP600125 suppressed IL-1beta-induced secretion of IL-6 and IL-8, and PD98059 suppressed IL-1beta-induced secretion of IL-8. Both SB202190 and PD98059 suppressed IL-1beta-induced expression of COX-2 in endometriotic cells. The p38 phosphorylation rates in the endometriotic tissues were significantly higher than those in the eutopic endometrial tissues of the same patients. CONCLUSIONS: Given the current theory that inflammatory changes are involved in the progression of endometriosis, MAPKs could play as pivotal intracellular signal transducers in endometriotic cells, and thus have a pathophysiological role in the disease.     

IL-5-induced integrin adhesion of human eosinophils caused by ERK1/2-mediated activation of cPLA2

We examined the mechanism by which interleukin (IL)-5 causes beta(2)-integrin adhesion of human eosinophils. IL-5 caused time-dependent activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38alpha in eosinophils as detected by their phosphorylation. Preincubation of eosinophils with U0126, a mitogen-activated protein kinase/ERK kinase inhibitor, suppressed IL-5-induced activation of cytosolic phospholipase A(2) (cPLA(2)) and eosinophil adhesion, and p38 inhibition by SB203580 had neither effect. ERK1/2 phosphorylation and eosinophil adhesion were blocked by inhibition of the src-family tyrosine kinase, Janus tyrosine kinase (JAK)2, or phosphoinositide-3 kinase (PI3K). Coimmunoprecipitation assay demonstrated that Lyn, a src-family tyrosine kinase, was constitutively associated with PI3K. Inhibition of src-tyrosine kinase but not JAK2 suppressed PI3K activation. Our data suggest that IL-5 induces beta(2)-integrin adhesion of human eosinophils by regulation of cPLA(2) activation caused by ERK1/2 phosphorylation. This phosphorylation results from activation of PI3K and protein tyrosine kinases. We also find that src-family tyrosine kinase, possibly Lyn, is the upstream kinase causing PI3K activation.     

Differences in cell activation by Chlamydophila pneumoniae and Chlamydia trachomatis infection in human endothelial cells

Seroepidemiological studies and demonstration of viable bacteria in atherosclerotic plaques have linked Chlamydophila pneumoniae infection to the development of chronic vascular lesions and coronary heart disease. In this study, we characterized C. pneumoniae-mediated effects on human endothelial cells and demonstrated enhanced phosphorylation and activation of the endothelial mitogen-activated protein kinase (MAPK) family members extracellular receptor kinase (ERK1/2), p38-MAPK, and c-Jun-NH2 kinase (JNK). Subsequent interleukin-8 (IL-8) expression was dependent on p38-MAPK and ERK1/2 activation as demonstrated by preincubation of endothelial cells with specific inhibitors for the p38-MAPK (SB202190) or ERK (U0126) pathway. Inhibition of either MAPK had almost no effect on intercellular cell adhesion molecule 1 (ICAM-1) expression. While Chlamydia trachomatis was also able to infect endothelial cells, it did not induce the expression of endothelial IL-8 or ICAM-1. These effects were specific for a direct stimulation with viable C. pneumoniae and independent of paracrine release of endothelial cell-derived mediators like platelet-activating factor, NO, prostaglandins, or leukotrienes. Thus, C. pneumoniae triggers an early signal transduction cascade in target cells that could lead to endothelial cell activation, inflammation, and thrombosis, which in turn may result in or promote atherosclerosis.     

Selective COX-2 inhibitor regulates the MAP kinase signaling pathway in human osteoarthritic chondrocytes after induction of nitric oxide

The purpose of this study was to examine the effect of cyclooxygenase-2 (COX-2) inhibitors on the mitogen-activated protein (MAP) kinase signaling pathway and synthesis of glucosaminoglycan after nitric oxide (NO) induction in articular human chondrocytes. After NO induction, the cells were divided into three groups that were treated with either ethanol (control); a selective COX-2 inhibitor (Celecoxib), or no additive, and evaluated. There were no differences in the effect of the selective COX-2 inhibitor on mitochondrial membrane potential or Annexin V levels. However, Celecoxib significantly decreased prostaglandin E2 (PGE2) production. Celecoxib also decreased the phosphorylation state of p38 and p44/42 of MAP kinase. The ratio of chondroitin-6 sulfate (C6S)/C4S was increased in response to the exposure to Celecoxib. Celecoxib did not affect apoptosis, but decreased the activation of MAP kinase in osteoarthritic chondrocytes after NO induction. NO-induced OA chondrocytes were associated with the p38 and the p44/42 MAPK signaling pathways, in a pathway that is distinct from PGE2-mediated apoptosis.     

Reactivation of Kaposi's sarcoma-associated herpesvirus from latency requires MEK/ERK, JNK and p38 multiple mitogen-activated protein kinase pathways

Lytic replication of Kaposi's sarcoma-associated herpesvirus (KSHV) promotes the progression of Kaposi's sarcoma (KS), a dominant malignancy in patients with AIDS. While 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced KSHV reactivation from latency is mediated by the protein kinase C delta and MEK/ERK mitogen-activated protein kinase (MAPK) pathways, we have recently shown that the MEK/ERK, JNK and p38 MAPK pathways modulate KSHV lytic replication during productive primary infection of human umbilical vein endothelial cells [Pan, H., Xie, J., Ye, F., Gao, S.J., 2006. Modulation of Kaposi's sarcoma-associated herpesvirus infection and replication by MEK/ERK, JNK, and p38 multiple mitogen-activated protein kinase pathways during primary infection. J. Virol. 80 (11), 5371-5382]. Here, we report that, besides the MEK/ERK pathway, the JNK and p38 MAPK pathways also mediate TPA-induced KSHV reactivation from latency. The MEK/ERK, JNK and p38 MAPK pathways were constitutively activated in latent KSHV-infected BCBL-1 cells. TPA treatment enhanced the levels of activated ERK and p38 but not those of activated JNK. Inhibitors of all three MAPK pathways reduced TPA-induced production of KSHV infectious virions in BCBL-1 cells in a dose-dependent fashion. The inhibitors blocked KSHV lytic replication at the early stage(s) of reactivation, and reduced the expression of viral lytic genes including RTA, a key immediate-early transactivator of viral lytic replication. Activation of MAPK pathways was necessary and sufficient for activating the promoter of RTA. Furthermore, we showed that the activation of RTA promoter by MAPK pathways was mediated by their downstream target AP-1. Together, these findings suggest that MAPK pathways might have general roles in regulating the life cycle of KSHV by mediating both viral infection and switch from viral latency to lytic replication.