Overexpression of fucosyltransferase IV promotes A431 cell proliferation through activating MAPK and PI3K/Akt signaling pathways

Lewis Y (LeY) is a carbohydrate tumor‐asssociated antigen. The majority of cancer cells derived from epithelial tissue express LeY type difucosylated oligosaccharide. Fucosyltransferase IV (FUT4) is an essential enzyme that catalyzes the synthesis of LeY oligosaccharide. Our previous studies have shown that FUT4 overexpression promotes A431 cell proliferation, but the mechanism is still largely unknown. Herein, we investigated the role of the mitogen‐activated protein kinases (MAPKs) and phosphoinositide‐3 kinase (PI3K)/Akt signaling pathways on FUT4‐induced cell proliferation. Results show that overexpression of FUT4 increases the phosphorylation of ERK1/2, p38 MAPK, and PI3K/Akt. Inhibitors of PI3K (LY294002 and Wortmannin) prevented the phosphorylation of ERK1/2, p38 MAPK, and Akt PI3K). Moreover, phosphorylation of Akt is abolished by inhibitors of ERK1/2 (PD98059) and p38 MAPK (SB203580). These data suggested that FUT4 not only activates MAPK and PI3K/Akt signals, but also promotes the crosstalk among these signaling pathways. In addition, FUT4‐induced stimulation of cell proliferation correlates with increased cell cycle progression by promoting cells into S‐phase. The mechanism involves in increased expression of cyclin D1, cyclin E, CDK 2, CDK 4, and pRb, and decreased level of cyclin‐dependent kinases inhibitors p21 and p27, which are blocked by the inhibitors of upstream signal molecules, MAPK and PI3K/Akt. In conclusion, these studies suggest that FUT4 regulates A431 cell growth through controlling cell cycle progression via MAPK and PI3K/Akt signaling pathways. J. Cell. Physiol. 225: 612–619, 2010. © 2010 Wiley‐Liss, Inc.     

PI3 kinase/Akt activation mediates estrogen and IGF-1 nigral DA neuronal neuroprotection against a unilateral rat model of Parkinson's disease

Recently, using the medial forebrain bundle (MFB) 6-hydroxydopmaine (6-OHDA) lesion rat model of Parkinson's disease (PD), we have demonstrated that blockade of central IGF-1 receptors (IGF-1R) attenuated estrogen neuroprotection of substantia nigra pars compacta (SNpc) DA neurons, but exacerbated 6-OHDA lesions in IGF-1 only treated rats (Quesada and Micevych [2004]: J Neurosci Res 75:107-116). This suggested that the IGF-1 system is a central mechanism through which estrogen acts to protect the nigrostriatal DA system. Moreover, these results also suggest that IGF-1R-induced intracellular signaling pathways are involved in the estrogen mechanism that promotes neuronal survival. In vitro, two convergent intracellular signaling pathways used by estrogen and IGF-1, the mitogen-activated protein kinase (MAPK/ERK), and phosphatidyl-inositol-3-kinase/Akt (PI3K/Akt), have been demonstrated to be neuroprotective. Continuous central infusions of MAPK/ERK and PI3K/Akt inhibitors were used to test the hypothesis that one or both of these signal transduction pathways mediates estrogen and/or IGF-1 neuroprotection of SNpc DA neurons after a unilateral administration of 6-OHDA into the MFB of rats. Motor behavior tests and tyrosine hydroxylase immunoreactivity revealed that the inhibitor of the PI3K/Akt pathway (LY294002) blocked the survival effects of both estrogen and IGF-1, while an inhibitor of the MAPK/ERK signaling (PD98059) was ineffective. Western blot analyses showed that estrogen and IGF-1 treatments increased PI3K/Akt activation in the SN; however, MAPK/ERK activation was decreased in the SN. Indeed, continuous infusions of inhibitors blocked phosphorylation of PI3K/Akt and MAPK/ERK. These findings indicate that estrogen and IGF-1-mediated SNpc DA neuronal protection is dependent on PI3K/Akt signaling, but not on the MAPK/ERK pathway.     

Sonic hedgehog promotes proliferation and differentiation of adult muscle cells: Involvement of MAPK/ERK and PI3K/Akt pathways

Sonic hedgehog (Shh) has been reported to act as a mitogen and survival factor for muscle satellite cells. However, its role in their differentiation remains ambiguous. Here, we provide evidence that Shh promotes the proliferation and differentiation of primary cultures of chicken adult myoblasts (also termed satellite cells) and mouse myogenic C2 cells. These effects are reversed by cyclopamine, a specific chemical inhibitor of the Shh pathway. In addition, we show that Shh and its downstream molecules are expressed in adult myoblast cultures and localize adjacent to Pax7 in muscle sections. These gene expressions are regulated during postnatal muscle growth in chicks. Most importantly, we report that Shh induces MAPK/ERK and phosphoinositide 3-kinase (PI3K)-dependent Akt phosphorylation and that activation of both signaling pathways is essential for Shh's signaling in muscle cells. However, the effect of Shh on Akt phosphorylation is more robust than that on MAPK/ERK, and data suggest that Shh influences these pathways in a manner similar to IGF-I. By exploiting specific chemical inhibitors of the MAPK/ERK and PI3K/Akt signaling pathways, UO126 and Ly294002, respectively, we demonstrate that Shh-induced Akt phosphorylation, but not that of MAPK/ERK, is required for its promotive effects on muscle cell proliferation and differentiation. Taken together, we suggest that Shh acts in an autocrinic manner in adult myoblasts, and provide first evidence of a role for PI3K/Akt in Shh signaling during myoblast differentiation.     

Cartducin stimulates mesenchymal chondroprogenitor cell proliferation through both extracellular signal-regulated kinase and phosphatidylinositol 3-kinase/Akt pathways

Cartducin, a paralog of Acrp30/adiponectin, is a secretory protein produced by both chondrogenic precursors and proliferating chondrocytes, and belongs to a novel C1q family of proteins. We have recently shown that cartducin promotes the growth of both mesenchymal chondroprogenitor cells and chondrosarcoma-derived chondrocytic cells in vitro. However, the cartducin-signaling pathways responsible for the regulation of cell proliferation have not been documented. In this study, we examined whether cartducin exists in serum and further investigated the intracellular signaling pathways stimulated by cartducin in mesenchymal chondroprogenitor cells. Western blot analysis showed that, unlike Acrp30/adiponectin, cartducin was undetectable in mouse serum. Next, mesenchymal chondroprogenitor N1511 cells were stimulated with cartducin, and three major groups of mitogen-activated protein kinase (MAPK) pathways and the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway were examined. Cartducin activated extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt, but not c-jun N-terminal kinase (JNK) nor p38 MAPK. The MEK1/2 inhibitor, U0126, blocked cartducin-stimulated ERK1/2 phosphorylation and suppressed the DNA synthesis induced by cartducin in N1511 cells. The PI3K inhibitor, LY294002, blocked cartducin-stimulated Akt phosphorylation and a decrease in cartducin-induced DNA synthesis in N1511 cells was also observed. These data suggest that cartducin is a peripheral skeletal growth factor, and that the proliferation of mesenchymal chondroprogenitor cells stimulated by cartducin is associated with activations of the ERK1/2 and PI3K/Akt signaling pathways.     

Interplay between PI3K/Akt and MAPK signaling pathways in DNA-damaging drug-induced apoptosis

In order to elucidate the role of the mitogen-activated protein kinases, including JNK, p38 MAPK and ERK, as well as the survival-associated PI3K/Akt signaling pathway, in the response to chemotherapy, we have conducted a comparative study regarding the effects of doxorubicin on these pathways. Doxorubicin was determined to elicit the apoptosis of NIH3T3 cells in a dose-dependent manner. Prior to cell death, both Akt and p38 MAPK were transiently activated, and subsequently inactivated almost wholly, whereas ERK and JNK evidenced sustained activations in response to the drug treatment. The inhibition of PI3K/Akt and p38 MAPK both accelerated and enhanced doxorubicin-induced apoptosis and ERK inhibition apparently exerted negative effect on apoptosis. The modulation of PI3K/Akt activation by treatment of LY294002 or expression of Akt mutants such as Akt-DN or Myr-Akt exerted a significant effect on the activation of ERK1/2. We also observed that PI3K/Akt and sustained ERK activation were associated intimately with the etoposide-induced apoptosis. Taken together, our results clearly suggest that the differential regulation of the PI3K/Akt, ERK1/2, and p38 MAPK signaling pathways are crucial in the context of DNA-damaging drug-induced apoptosis, and this has compelled us to propose that the sustained activation of ERK1/2 pathway may be generally involved in the apoptosis induced by anticancer DNA-damaging drugs, including doxorubicin and etoposide.     

Cyclophosphamide-induced apoptosis in A431 cells is inhibited by fucosyltransferase IV

Fucosyltransferase IV (FUT4) is an essential enzyme that catalyzes the synthesis of difucosylated oligosaccharide LeY which is overexpressed in the cancers derived from the epithelial tissues. Our previous studies have shown that FUT4 overexpression promotes A431 cell proliferation through the MAPK and PI3K/Akt signaling pathways, but the relationship between FUT4 and apoptosis remained unclear. Here, we investigated the effect of FUT4 overexpression on cyclophosphamide (CPA)-induced apoptosis in A431 cells. Western blot analysis showed that FUT4 overexpression decreased expression of Bax, Caspase 3, and PARP proteins, and increased anti-apoptotic Bcl-2 protein in A431 cells. The anti-apoptosis effect of FUT4 was confirmed both by Annexin-V/PI and JC-1 assays. The results showed that FUT4 overexpression up-regulated phosphorylation of ERK1/2 and Akt which was inhibited by CPA in dose-dependent manner. By blocking the ERK/MAPK and PI3K/Akt pathways with specific inhibitors, we demonstrated that these two pathways were required in mediating the anti-apoptosis effect of FUT4. We concluded that FUT4 inhibited cell apoptosis induced by CPA through decreasing the expression of apoptotic proteins Bax, Caspase 3, and PARP and increasing the expression of anti-apoptotic protein Bcl-2 via the ERK/MAPK and PI3K/Akt signaling pathways in A431 cells.     

Overexpression of miR-623 suppresses progression of hepatocellular carcinoma via regulating the PI3K/Akt signaling pathway by targeting XRCC5

It has been reported that miR-623 is deregulated in lung adenocarcinoma and inhibits tumor growth and invasion. However, it is unclear whether miR-623 has a role in the progression of hepatocellular carcinoma (HCC). Herein, we found that miR-623 was significantly downregulated in HCC, and that its expression was related to poor clinical outcomes of patients with HCC. Upregulation of miR-623 decreased cell proliferation, viability, migration, and invasion and further promoted apoptosis in 7721, Huh7, and Bel-7402 cells. Moreover, we also observed that miR-623 regulated the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt), Wnt/β-catenin, and extracellular regulated protein kinases/c-Jun N-terminal kinase (ERK/JNK) signaling pathways as well as the expression level of related proteins. Further, X-ray repair cross complementing 5 (XRCC5) was a direct target for miR-623, and the suppression of PI3K/Akt, Wnt/β-catenin, and ERK/JNK signaling pathways and cell proliferation and invasion abilities caused by miR-623 in HCC cells was significantly reversed by the upregulation of XRCC5. Collectively, our data suggested that miR-623 suppressed the progression of HCC by regulating the PI3K/Akt, Wnt/β-catenin, and ERK/JNK pathways by targeting XRCC5 in HCC in vitro, indicating that miR-623 may be a target for the therapy of HCC.     

Halofuginone inhibits Smad3 phosphorylation via the PI3K/Akt and MAPK/ERK pathways in muscle cells: Effect on myotube fusion

Halofuginone, a novel inhibitor of Smad3 phosphorylation, has been shown to inhibit muscle fibrosis and to improve cardiac and skeletal muscle functions in the mdx mouse model of Duchenne muscular dystrophy. Here, we demonstrate that halofuginone promotes the phosphorylation of Akt and mitogen-activated protein kinase (MAPK) family members in a C2 muscle cell line and in primary myoblasts derived from wild-type and mdx mice diaphragms. Halofuginone enhanced the association of phosphorylated Akt and MAPK/extracellular signal-regulated protein kinase (ERK) with the non-phosphorylated form of Smad3, accompanied by a reduction in Smad3 phosphorylation levels. This reduction was reversed by inhibitors of the phosphoinositide 3′-kinase/Akt (PI3K/Akt) and MAPK/ERK pathways, suggesting their specific role in mediating halofuginone's inhibitory effect on Smad3 phosphorylation. Halofuginone enhanced Akt, MAPK/ERK and p38 MAPK phosphorylation and inhibited Smad3 phosphorylation in myotubes, all of which are crucial for myotube fusion. In addition, halofuginone increased the association Akt and MAPK/ERK with Smad3. As a consequence, halofuginone promoted myotube fusion, as reflected by an increased percentage of C2 and mdx myotubes containing high numbers of nuclei, and this was reversed by specific inhibitors of the PI3K and MAPK/ERK pathways. Together, the data suggest a role, either direct or via inhibition of Smad3 phosphorylation, for Akt or MAPK/ERK in halofuginone-enhanced myotube fusion, a feature which is crucial to improving muscle function in muscular dystrophies.     

Hypoxia-induced ischemic tolerance in neonatal rat brain involves enhanced ERK1/2 signaling

Hypoxic preconditioning (HP) 24 h before hypoxic-ischemic (HI) injury confers significant neuroprotection in neonatal rat brain. Recent studies have shown that the mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K) intracellular signaling pathways play a role in the induction of tolerance to ischemic injury in heart and brain. To study the role of MAPK (ERK1/2, JNK, p38MAPK) and PI3K/Akt/GSK3beta signaling pathways in hypoxia-induced ischemic tolerance, we examined the brains of newborn rats at different time points after exposure to sublethal hypoxia (8% O(2) for 3 h). Immunoblot analysis showed that HP had no effect on the levels of phosphorylated Akt, GSK3beta, JNK and p38MAPK. In contrast, significantly increased levels of phosphorylated ERK1/2 were observed 0.5 h after HP. Double immunofluorescence staining showed that hypoxia-induced ERK1/2 phosphorylation was found mainly in microvessels throughout the brain and in astrocytes in white matter tracts. Inhibition of hypoxia-induced ERK1/2 pathway with intracerebral administration of U0126 significantly attenuated the neuroprotection afforded by HP against HI injury. These findings suggest that activation of ERK1/2 signaling may contribute to hypoxia-induced tolerance in neonatal rat brain in part by preserving vascular and white matter integrity after HI.     

Extracellular ATP-induced proliferation of adventitial fibroblasts requires phosphoinositide 3-kinase, Akt, mammalian target of rapamycin, and p70 S6 kinase signaling pathways

Extracellular nucleotides are increasingly recognized as important regulators of growth in a variety of cell types. Recent studies have demonstrated that extracellular ATP is a potent inducer of fibroblast growth acting, at least in part, through an ERK1/2-dependent signaling pathway. However, the contributions of additional signaling pathways to extracellular ATP-mediated cell proliferation have not been defined. By using both pharmacologic and genetic approaches, we found that in addition to ERK1/2, phosphatidylinositol 3-kinase (PI3K), Akt, mammalian target of rapamycin (mTOR), and p70 S6K-dependent signaling pathways are required for ATP-induced proliferation of adventitial fibroblasts. We found that extracellular ATP acting in part through G(i) proteins increased PI3K activity in a time-dependent manner and transient phosphorylation of Akt. This PI3K pathway is not involved in ATP-induced activation of ERK1/2, implying activation of independent parallel signaling pathways by ATP. Extracellular ATP induced dramatic increases in mTOR and p70 S6K phosphorylation. This activation of the mTOR/p70 S6 kinase (p70 S6K) pathway in response to ATP is because of independent contributions of PI3K/Akt and ERK1/2 pathways, which converge on the level of p70 S6K. ATP-dependent activation of mTOR and p70 S6K also requires additional signaling inputs perhaps from pathways operating through Galpha or Gbetagamma subunits. Collectively, our data demonstrate that ATP-induced adventitial fibroblast proliferation requires activation and interaction of multiple signaling pathways such as PI3K, Akt, mTOR, p70 S6K, and ERK1/2 and provide evidence for purinergic regulation of the protein translational pathways related to cell proliferation.     

IGF-I/PI3K/Akt and IGF-I/MAPK/ERK pathways in vivo in skeletal muscle are regulated by nutrition and contribute to somatic growth in the fine flounder

The insulin-like growth factor-I (IGF-I) is a key regulator of skeletal muscle growth in vertebrates, promoting mitogenic and anabolic effects through the activation of the MAPK/ERK and the PI3K/Akt signaling pathways. Nutrition also affects skeletal muscle growth, activating intracellular pathways and inducing protein synthesis and accretion. Thus, both hormonal and nutritional signaling regulate muscle mass. In this context, plasma IGF-I levels and the activation of both pathways in response to food were evaluated in the fine flounder using fasting and refeeding trials. The present study describes for the first time in a nonmammalian species that the MAPK/ERK and PI3K/Akt are activated by exogenous circulating IGF-I, as well as showing that the MAPK/ERK pathway activation is modulated by the nutritional status. Also, these results show that there is a time-dependent regulation of IGF-I plasma levels and its signaling pathways in muscle. Together, these results suggest that the nutritionally managed IGF-I could be regulating the activation of the MAPK/ERK and the PI3K/Akt signaling pathways differentially according to the nutritional status, triggering different effects in growth parameters and therefore contributing to somatic growth in fish. This study contributes to the understanding of the nutrient regulation of IGF-I and its signaling pathways in skeletal muscle growth in nonmammalian species, therefore providing insight concerning the events controlling somatic growth in vertebrates.     

Identification of signaling molecules mediating corticotropin-releasing hormone-R1alpha-mitogen-activated protein kinase (MAPK) interactions: the critical role of phosphatidylinositol 3-kinase in regulating ERK1/2 but not p38 MAPK activation

In most target cells, activation of the type 1 CRH receptor (CRH-R1) by CRH or urocortin (UCN I) leads to stimulation of the Gs-protein/adenylyl cyclase/protein kinase A cascade. Signal transduction of CRH-R1 also involves alternative pathways such as phosphorylation of ERK1/2 and p38 MAPK, two members of the MAPK family that mediate important pathophysiological responses. The intracellular pathways by which CRH-R1 activates these MAPK are only partially understood; here we characterized further signaling mechanisms and molecules involved in CRH-R1-mediated ERK1/2 and p38 MAPK activation. In human embryonic kidney 293 cells overexpressing recombinant CRH-R1alpha, UCN I induced ERK1/2 and p38 MAPK activation was dependent on signaling molecules involved in agonist-induced CRH-R1alpha trafficking and endocytosis. Furthermore, time course studies and use of selective inhibitors demonstrated that ERK1/2 activation occured within 5 min, was sustained for at least 60 min, and was dependent on both phosphatidylinositol 3-kinase (PI3-K)/Akt activation and epidermoid growth factor receptor transactivation involving matrix metelloproteinases. UCN I effect on p38 MAPK phosphorylation was more transient, returned to basal within 40 min and was dependent on epidermoid growth factor receptor transactivation, but not PI3-K/Akt activation. Overexpression of G(alpha-)transducin, showed that G(betagamma)-subunit activation is only partially required for ERK1/2 phosphorylation and does not play a role in p38 MAPK phosphorylation, whereas overexpression of a dominant-negative Ras (Ras N17) attenuated both ERK and p38 MAPK activation. In conclusion, a complex signaling network appears to mediate CRH-R1alpha-MAPK interactions; PI3-K might play a critical role in the regulation of CRH-R1alpha signaling selectivity and cellular responses.     

PI3K/Akt-sensitive MEK-independent compensatory circuit of ERK activation in ER-positive PI3K-mutant T47D breast cancer cells

We explored the crosstalk between cell survival (phosphatidylinositol 3-kinase (PI3K)/Akt) and mitogenic (Ras/Raf/MEK/extracellular signal-regulated kinase (ERK)) signaling pathways activated by an epidermal growth factor (EGF) and analyzed their sensitivity to small molecule inhibitors in the PI3K-mutant estrogen receptor (ER)-positive MCF7 and T47D breast cancer cells. In contrast to MCF7 cells, ERK phosphorylation in T47D cells displayed resistance to MEK inhibition by several structurally different compounds, such as U0126, PD 098059 and PD 198306, MEK suppression by small interfering RNA (siRNA) and was also less sensitive to PI3K inhibition by wortmannin. Similar effect was observed in PI3K-wild type ER-positive BT-474 cells, albeit to a much lesser extent.MEK-independent ERK activation was induced only by ErbB receptor ligands and was resistant to inhibition of several kinases and phosphatases that are known to participate in the regulation of Ras/mitogen-activated protein kinase (MAPK) cascade. Although single agents against PDK1 or Akt did not affect EGF-induced ERK phosphorylation, a combination of PI3K/Akt and MEK inhibitors synergistically suppressed ERK activation and cellular growth. siRNA-mediated silencing of class I PI3K or Akt1/2 genes also significantly decreased U0126-resistant ERK phosphorylation.Our data suggest that in T47D cells ErbB family ligands induce a dynamic, PI3K/Akt-sensitive and MEK-independent compensatory ERK activation circuit that is absent in MCF7 cells. We discuss candidate proteins that can be involved in this activation circuitry and suggest that PDZ-Binding Kinase/T-LAK Cell-Originated Protein Kinase (PBK/TOPK) may play a role in mediating MEK-independent ERK activation.     

Differential regulation of the phosphoinositide 3-kinase and MAP kinase pathways by hepatocyte growth factor vs. insulin-like growth factor-I in myogenic cells

Hepatocyte growth factor (HGF) promotes the proliferation of adult myoblasts and inhibits their differentiation, whereas insulin-like growth factor I (IGF-I) enhances both processes. Recent studies indicate that activation of the phosphoinositide 3'-kinase (PI3K) pathway promotes myoblast differentiation, whereas activation of the mitogen-activated protein kinase/extracellular signal-regulated protein kinase (MAPK/ERK) promotes proliferation and inhibits their differentiation. This simple model is confounded by the fact that both HGF and IGF-I have been shown to activate both pathways. In this study, we have compared the ability of HGF and IGF-I to activate PI3K and MAPK/ERK in i28 myogenic cells. We find that, although the two stimuli result in comparable recruitment of the p85alpha subunit of PI3K into complexes with tyrosine-phosphorylated proteins, the p85beta regulatory subunit and p110alpha catalytic subunit of PI3K are preferentially recruited into these complexes in response to IGF-I. In agreement with this observation, IGF-I is much more potent than HGF in stimulating phosphorylation of Akt/PKB, a protein kinase downstream of PI3K. In contrast, MAPK/ERK phosphorylation was higher in response to HGF and lasted longer, relative to IGF-I. Moreover, the specific PI3K inhibitor, Wortmannin, abolished MAPK/ERK and Elk-1 phosphorylation in HGF-treated cells, suggesting the requirement of PI3K in mediating the HGF-induced MAPK pathway. UO126, a specific MAPK pathway inhibitor, had no effect on PI3K activity or Akt phosphorylation, implying that at least in muscle cells, the MAPK/ERK pathway is not required for HGF-induced PI3K activation. These results provide a biochemical rationale for the previous observations that HGF and IGF-I have opposite effects on myogenic cells, consistent with studies linking PI3K activation to differentiation and MAPK/ERK activation to proliferation in these cells. Moreover, the finding that PI3K activity is required for HGF-induced MAPK activation suggests its additional role in proliferation, rather than exclusively in the differentiation of adult myoblasts.     

Intracellular Networks of the PI3K/AKT and MAPK Pathways for Regulating Toxoplasma gondii-Induced IL-23 and IL-12 Production in Human THP-1 Cells

Interleukin (IL)-23 and IL-12 are closely related in structure, and these cytokines regulate both innate and adaptive immunity. However, the precise signaling networks that regulate the production of each in Toxoplasma gondii-infected THP-1 monocytic cells, particularly the PI3K/AKT and MAPK signaling pathways, remain unknown. In the present study, T. gondii infection upregulated the expression of IL-23 and IL-12 in THP-1 cells, and both cytokines increased with parasite dose. IL-23 secretion was strongly inhibited by TLR2 monoclonal antibody (mAb) treatment in a dose-dependent manner and by TLR2 siRNA transfection, whereas IL-12 secretion was strongly inhibited by TLR4 mAb treatment dose-dependently and by TLR4 siRNA transfection. IL-23 production was dose-dependently inhibited by the PI3K inhibitors LY294002 and wortmannin, whereas IL-12 production increased dose-dependently. THP-1 cells exposed to live T. gondii tachyzoites underwent rapid p38 MAPK, ERK1/2 and JNK activation. IL-23 production was significantly upregulated by the p38 MAPK inhibitor SB203580 dose-dependently, whereas pretreatment with 10 μM SB203580 significantly downregulated IL-12 production. ERK1/2 inhibition by PD98059 was significantly downregulated IL-23 production but upregulated IL-12 production. JNK inhibition by SP600125 upregulated IL-23 production, but IL-12 production was significantly downregulated dose-dependently. T. gondii infection resulted in AKT activation, and AKT phosphorylation was inhibited dose-dependently after pretreatment with PI3K inhibitors. In T. gondii-infected THP-1 cells, ERK1/2 activation was regulated by PI3K; however, the phosphorylation of p38 MAPK and JNK was negatively modulated by the PI3K signaling pathway. Collectively, these results indicate that IL-23 production in T. gondii-infected THP-1 cells was regulated mainly by TLR2 and then by PI3K and ERK1/2; however, IL-12 production was mainly regulated by TLR4 and then by p38 MAPK and JNK. Our findings provide new insight concerning the intracellular networks of the PI3K/AKT and MAPK signaling cascades for regulating T. gondii-induced IL-23 and IL-12 secretion in human monocytic cells.