Src family protein tyrosine kinase signaling mediates monosodium urate crystal-induced IL-8 expression by monocytic THP-1 cells.

Neutrophil-dependent inflammation dependent on monosodium urate (MSU) crystal-induced IL-8 expression occurs in gout. MSU crystals activate phagocyte Src family tyrosine kinases and the serine/threonine kinase p70s6k. Thus, using monocytic THP-1 cells, we assessed the potential for Src family kinases and p70s6k to mediate MSU-induced IL-8 expression. MSU crystals induced phosphorylation of p70s6k and the Src kinases c-Src, Lyn, Hck, and Fyn. IL-8 expression was attenuated more by the Src kinase inhibitor PP1 than by the p70s6k inhibitor rapamycin. PP1 inhibited crystal-induced phosphorylation of ERK1/2 and IkappaBalpha and suppressed IkappaB kinase (IKK) activation and NF-kappaB binding to the IL-8 promoter, signals that mediate MSU-induced IL-8 expression. Transfection of the native Src inhibitor, C-terminal Src kinase (Csk), also suppressed crystal-induced c-Src, ERK1/2, and IkappaBalpha phosphorylation and IL-8 expression. We conclude that Src family tyrosine kinase signaling plays a significant role in MSU crystal-induced IL-8 expression via stimulation of ERK1/2 pathway and NF-kappaB activation.     

Ceramide-mediated stimulation of inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF) accumulation in murine macrophages requires tyrosine kinase activity

In macrophages, bacterial lipopolysaccharide (LPS) has been noted to mimic certain effects of the sphingolipid ceramide, suggesting that ceramide may be involved in macrophage activation by LPS and/or that LPS utilizes ceramide-related signaling pathways. Putative downstream targets of ceramide include a ceramide-activated (serine/threonine) protein kinase (CAPK) and phosphatase (CAPP). However, the potential role of tyrosine phosphorylation pathways in macrophage response to ceramide has not been examined. Herein we report that cell-permeable analogs of ceramide up-regulate both inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF) production in RAW 264.7 murine macrophages. Herbimycin A and genistein, potent natural inhibitors of protein tyrosine (but not serine/threonine) phosphorylation, block ceramide-induced iNOS and TNF production. Furthermore, the highly src-family selective pyrazolopyrimidine inhibitor PP1 also blocks ceramide-induced iNOS and TNF production in RAW 264.7 cells. We found that PP1 also inhibits ceramide-mediated tyrosine phosphorylation of the src-family kinase hck. These data indicate that src-related tyrosine kinases play a critical role in macrophage activation by ceramide.     

Src-family protein tyrosine kinase negatively regulates cerebellar long-term depression

Protein phosphorylation is a major mechanism for the regulation of synaptic transmission. Previous studies have shown that several serine/threonine kinases are involved in the induction of long-term depression (LTD) at excitatory synapses on a Purkinje neuron (PN) in the cerebellum. Here, we show that Src-family protein tyrosine kinases (SFKs) are involved in the regulation of the LTD induction. Intracellular application of c-Src suppressed LTD. We also show that application of a SFK-selective inhibitor PP2 recovered LTD from the suppression caused by the inhibition of mGluR1 activity. These results indicate that SFKs negatively regulate the LTD induction at excitatory synapses on a cerebellar PN.     

Toll-like receptor-mediated tyrosine phosphorylation of paxillin via MyD88-dependent and -independent pathways

Toll-like receptor (TLR)-mediated recognition of pathogens represents one of the most important mechanisms of innate immunity. A proximal signaling event of TLR is the direct binding of an adaptor protein MyD88 to TLR and recruitment of the IL-1R-associated kinase (IRAK). In the present study, we examined the effect of several TLR ligands on protein tyrosine phosphorylation in rat macrophages. Macrophage-activating lipopeptide-2 kDa (MALP2) and lipoarabinomannan were used as activators of TLR2, while lipopolysaccharides (LPS) and lipoteichoic acid were used as TLR4 ligands. All these ligands induced tyrosine phosphorylation of proline-rich tyrosine kinase 2 (Pyk2) and its substrate paxillin, an integrin-associated focal adhesion adaptor protein, in the macrophages. PP2, an inhibitor of Src family tyrosine kinases, prevented the TLR-induced phosphorylation of paxillin and Pyk2 without affecting TLR-induced IRAK activation. MALP2 failed to induce paxillin phosphorylation in the macrophages from MyD88-knockout mice. In contrast, the effect of LPS weakened, but was still observed even in the MyD88-deficient cells. Thus, TLR regulate the function of paxillin in an Src family-dependent mechanism through both MyD88-dependent and MyD88-independent pathways.     

Cbl functions downstream of Src kinases in Fc gamma RI signaling in primary human macrophages

Cbl is a cytosolic protein that is rapidly tyrosine phosphorylated in response to Fc receptor activation and binds to the adaptor proteins Grb2, CrkL, and Nck. A few reports describe Cbl interactions in primary human hematopoietic cells. We show evidence that Cbl participates in signaling initiated by Fc gammaRI receptor cross-linking in human primary macrophages, and functions downstream of Src family kinases in this pathway. Fc gammaRI stimulation in human macrophages was associated with rapid and transient tyrosine phosphorylation of the Cbl adaptor protein. Immunoprecipitated Cbl was complexed with several tyrosine phosphorylated proteins, the most prominent of which was a 38-kDa band identified as the CrkL adaptor protein. CrkL associated with tyrosine-phosphorylated Cbl and itself became tyrosine phosphorylated after Fc gammaRI cross-linking. SLP-76, a recently cloned Grb2-associated protein, was strongly tyrosine phosphorylated after Fc gammaRI stimulation and was associated with both Cbl and Grb2. Grb2 and Cbl binding to SLP-76 were inducible after Fc gammaRI stimulation of the macrophages. Nck was inducibly bound to Cbl after Fc gammaRI stimulation, whereas Grb2 was constitutively associated with it. Shc was also inducibly tyrosine phosphorylated and bound to Grb2 after Fc gammaRI stimulation of the macrophages. PP1, a specific inhibitor of Src kinases, inhibited the Fc gammaRI-induced respiratory burst, as well as the tyrosine phosphorylation of Cbl and its inducible association with CrkL. These results suggest a fundamental role for the tyrosine phosphorylation of Cbl, CrkL, SLP-76, and Shc and the association of Cbl with CrkL, SLP-76, and Nck in Fc gammaRI signaling in human macrophages. Experiments performed with PP1, the specific Src kinase inhibitor, demonstrate the first evidence that Cbl and the Cbl-Crkl interaction are downstream targets for myeloid Src kinases required for the activation of myeloid NADPH oxidase activity.     

PP2, a potent inhibitor of Src family kinases, protects against hippocampal CA1 pyramidal cell death after transient global brain ischemia

It has been indicated that Src family protein tyrosine kinases (SrcPTKs) potentiate N-methyl-D-aspartate (NMDA) receptor function by phosphorylating NR2A subunits and that postsynaptic density protein 95 (PSD-95) facilitates this regulation. In this paper, we define the role of SrcPTKs in delayed neuronal damage following transient brain ischemia and explore the underlying mechanisms involved in this event. Transient global brain ischemia was induced by the four-vessel occlusion method. A specific Src family kinase inhibitor PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyramidine) and a PP2 negative control PP3 (4-amino-7-phenylpyrazolo[3,4-d]pyramidine) were infused into rat cerebroventricule 30 min before occlusion. Hematoxylin and eosine staining showed that the number of surviving pyramidal neurons in rat hippocampal CA1 subfield increased markedly in PP2-treated rats comparing to PP3-treated groups after 5 days of reperfusion following ischemia. Additionally, immunoprecipitation and immunoblot analysis revealed that preadministration of PP2, but not PP3, attenuated not only the increased tyrosine phosphorylation of NR2A but also the enhanced interactions among Src, NR2A and PSD-95 induced by ischemia/reperfusion. In conclusion, SrcPTKs promote binding of the kinases and their substrate NR2A attributed to the scaffolding effect of PSD-95 during transient brain ischemia and reperfusion, which are responsible for the elevation of NR2A tyrosine phosphorylation and consequent delayed neuronal cell death.     

Role of Src kinases and Syk in Fcgamma receptor-mediated phagocytosis and phagosome-lysosome fusion.

Phagocytosis is increased by Fcgamma receptors (FcgammaRs), and studies with syk(-/-) macrophages demonstrated that Syk kinase is required for FcgammaR phagocytosis. Similar studies with macrophages lacking the Src family kinases Hck, Fgr, and Lyn showed that these kinases are not required for phagocytosis but that they enhance the rate of particle engulfment. In this report we show that both wild-type and hck(-/-)fgr(-/-) macrophages expressed Fyn, Src, and Yes and that these kinases were activated on ingestion of immunoglobulin G (IgG)-coated particles and redistributed, together with Syk, to actin-rich phagocytic cups and the phagosomal membrane. At doses blocking IgG-dependent phagocytosis, the tyrosine kinase inhibitors PP1 and piceatannol inhibited both Src family kinase and Syk activities, as well as their redistribution to actin-rich phagocytic cups. Hck, Fgr, and Lyn were dispensable for lysosome-phagosome fusion (PLF) induced by IgG-coated particles. However, PP1 or piceatannol hampered unopsonized yeast-induced PLF despite the fact that they did not block yeast internalization.     

胰腺癌BxPC3细胞中Src激酶对Notch-1活化的调节

目的 探讨在胰腺癌细胞BxPC3中,Src激酶对Notch-1活化的影响.方法 用siRNA干扰的方法分别抑制Notch-1和c-Src的表达;加入Src激酶抑制剂PP2抑制Src激酶活性;MTT法检测细胞的生长;Western blot检测Notch-1蛋白活性形式NICD水平的变化.结果 抑制Notch-1表达及抑制Src激酶活性可明显抑制BxPC3细胞生长;抑制Src激酶活性及抑制c-Src蛋白表达可下调Notch-1 NICD水平.结论 Src激酶在胰腺癌细胞BxPC3中促进Notch-1的活化,促进BxPc3细胞的生长.     

CD137 ligand‐mediated reverse signals increase cell viability and cytokine expression in murine myeloid cells: Involvement of mTOR/p70S6 kinase and Akt

Cross‐linking of CD137 ligand (CD137L), a member of the TNF family, with recombinant CD137‐Fc (rCD137‐Fc) protein enhanced adherence of bone marrow‐derived macrophages, and increased the expression of ICAM‐1, IL‐1β, IL‐6, M‐CSF and phosphotyrosine proteins. In RAW264.7 cells, a murine myeloid cell line, rCD137‐Fc not only increased adherence but also cell multiplication, in a manner comparable to LPS or M‐CSF. In addition, it up‐regulated expression of IL‐1β, IL‐1 receptor antagonist, IL‐6, COX2, tenascin C, neuropeptide Y and M‐CSF mRNA. Neutralization of M‐CSF by incubating the RAW264.7 cells with anti‐M‐CSF mAb did not prevent the CD137L signal‐induced viability. Viability was blocked by PP2, an Src tyrosine kinase inhibitor, rapamycin, an mTOR inhibitor and LY294002, a PI3K inhibitor, but not by Wortmannin, another PI3K inhibitor. Cross‐linking of CD137L increased phosphorylation of Akt and p70S6 kinase. The latter was blocked by PP2, rapamycin or LY294002, but not by Wortmannin, whereas phosphorylation of Akt was blocked by LY294002 or Wortmannin. These findings demonstrate that reverse signals evoked by CD137L regulate immune functions in macrophages.     

Modulation of TLR signalling by the C-terminal Src kinase (Csk) in macrophages

In macrophages and monocytes, lipopolysaccharide (LPS) triggers the production of pro-inflammatory cytokine through Toll-like receptor (TLR) 4. Although major TLR signalling pathways are mediated by serine or threonine kinases including IKK, TAK1, p38 and JNKs, a number of reports suggested that tyrosine phosphorylation of intracellular proteins is involved in LPS signalling. Here, we identified several tyrosine-phosphorylated proteins using mass spectrometric analysis in response to LPS stimulation. Among these proteins, we characterized C-terminal Src kinase (Csk), which negatively regulates Src-like kinases in RAW 264.7 cells using RNAi knockdown technology. Unexpectedly, LPS-induced CD40 activation and the secretion of pro-inflammatory cytokine such as IL-6 and TNF-alpha, was down-regulated in Csk knockdown cells. Furthermore, overall cellular tyrosine phosphorylation and TLR4-mediated activation of IkappaB-alpha, Erk and p38 but not of JNK, were also down-regulated in Csk knockdown cells. The protein expression levels of a tyrosine kinase, Fgr, were reduced in Csk knockdown cells, suggesting that Csk is a critical regulator of TLR4-mediated signalling by modifying the levels of Src-like kinases.     

Role of different protein tyrosine kinases in fMLP-induced neutrophil transmigration

Protein tyrosine phosphorylation is among the early signaling events in polymorphonuclear leukocyte (PMN) responses to chemoattractant stimulation. We previously showed that tyrosine phosphorylation might serve as the downstream signaling for the modulation of PMN transmigration by CD47. Here, we further investigated the role of various tyrosine kinases in PMN transmigration and identified the potential tyrosine kinases serving as CD47-mediated signaling downstream. We observed that PMN transmigration was significantly enhanced by Src family kinase inhibitors PP1 and PP2 as well as Syk tyrosine kinase inhibitor piceatannol, suggesting that these kinases have negative regulatory roles in PMN chemotaxis. In contrast, PMN chemotaxis was reduced by LFM-A13, an inhibitor of the Tec family tyrosine kinase Btk (Bruton's tyrosine kinase). LFM-A13 also dose-dependently inhibited N-formyl-Met-Leu-Phe (fMLP)-induced PMN intracellular [Ca2+] increase. Since LFM-A13 significantly enhanced PMN chemokinesis while other inhibitors had no effect, the inhibition of PMN chemotaxis by LFM-A13 might be due to the promotion of random cell migration. Among the other inhibitors we tested, AG126 significantly inhibited PMN transmigration while the MAP kinase inhibitors SB20358 and PD98059 showed an enhancing effect. No effect of herbimycin A, erbstatin analog, lavendustin A or AG490 on PMN transmigration was observed. Treatment with PP1, PP2 or piceatannol all partially reversed the delay of PMN transmigration caused by inhibitory anti-CD47 antibody. In summary, our results demonstrate distinct roles of different tyrosine kinases in regulating PMN chemotaxis and suggest Src and/or Syk kinases are likely involved in CD47-mediated downstream signaling.     

c-Src tyrosine kinase, a critical component for 5-HT2A receptor-mediated contraction in rat aorta

Serotonin (5-hydroxytryptamine, 5-HT) receptors (5-HTRs) play critical roles in brain and cardiovascular functions. In the vasculature, 5-HT induces potent vasoconstrictions, which in aorta are mainly mediated by activation of the 5-HT(2A)R subtype. We previously proposed that one signalling mechanism of 5-HT-induced vasoconstriction could be c-Src, a member of the Src tyrosine kinase family. We now provide evidence for a central role of c-Src in 5-HT(2A)R-mediated contraction. Inhibition of Src kinase activity with 10 mum 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) prior to contraction resulted in approximately 90-99% inhibition of contractions induced by 5-HT or by alpha-methyl-5-HT (5-HT(2)R agonist). In contrast, PP2 pretreatment only partly inhibited contractions induced by angiotensin II and the thromboxane A(2) mimetic, U46619, and had no significant action on phenylephrine-induced contractions. 5-Hydroxytryptamine increased Src kinase activity and PP2-sensitive tyrosine-phosphorylated proteins. As expected for c-Src identity, PP2 pretreatment inhibited 5-HT-induced contraction with an IC(50) of approximately 1 mum. Ketanserin (10 nm), a 5-HT(2A) antagonist, but not antagonists of 5-HT(2B)R (100 nm SB204741) or 5-HT(2C)R (20 nm RS102221), prevented 5-HT-induced contractions, mimicking PP2 and implicating 5-HT(2A)R as the major receptor subtype coupled to c-Src. In HEK 293T cells, c-Src and 5-HT(2A)R were reciprocally co-immunoprecipitated and co-localized at the cell periphery. Finally, 5-HT-induced Src activity was unaffected by inhibition of Rho kinase, supporting a role of c-Src upstream of Rho kinase. Together, the results highlight c-Src activation as one of the early and pivotal mechanisms in 5-HT(2A)R contractile signalling in aorta.     

Lipopolysaccharide primes macrophages to increase nitric oxide production in response to Staphylococcus aureus

Sepsis, the leading cause of death in intensive care units, is associated with overproduction of nitric oxide (NO). The mechanism concerning the NO production in the sepsis caused by both Gram-negative and Gram-positive bacteria is largely unknown. The present study examines the effect of lipopolysaccharide (LPS) on Staphylococcus aureus-induced NO production in macrophages. In the na?ve murine macrophage cell line RAW264.7, heat-killed Staphylococcus aureus (HKSa) induced a significant NO production at a high concentration (100 microg/ml). However, pretreatment of the cells with increasing concentration of LPS (10-50 ng/ml) resulted in induction of NO production by HKSa even at the doses of 1 and 10 microg/ml. The expression of inducible NO synthase (iNOS) in response to HKSa was also enhanced by LPS pretreatment, suggesting that LPS priming NO production is due to the enhancement of iNOS expression. We examined whether protein kinase C (PKC), mitogen-activated protein kinases (MAPKs) and calcineurin signaling pathways are involved in the priming effects of LPS. It was found that the PKC inhibitor G?6976, the p38 inhibitor SB203580 and the calcineurin inhibitor cyclosporine A significantly reversed the priming effects of LPS on HKSa-induced NO production and iNOS expression. In contrast, the ERK1/2 inhibitor PD98059 did not block the induction of priming by LPS. These data support the hypothesis that LPS primes macrophages for enhancement of HKSa-induced NO production, and indicate that PKC, p38 and calcineurin might be involved in the LPS-induced priming.     

Selective expansion of perforin-positive CD8+ T cells by immature dendritic cells infected with live Bacillus Calmette-Guérin mycobacteria

Tyrosine phosphorylation is thought to be critical in the regulation of neutrophil functioning, and members of the Src family of tyrosine kinases have recently been shown to be regulated in activated granulocytes. We have used a specific pharmacological inhibitor of Src kinases, pyrazolpyrimidine 1 (PP1), to evaluate the role of Src kinases in cytokine/chemoattractant-induced regulation of neutrophil function. PP1 inhibits PKB phosphorylation but not STAT5 phosphorylation or the activation of MAP kinases by fMLP or GM-CSF. Pretreatment of neutrophils with PP1 and with the PI3K inhibitor LY294002 resulted in a strong inhibition of fMLP-induced superoxide production and cytokine-mediated survival but not fMLP-induced migration. It is interesting that the kinetics of inhibition of actin polymerization and the respiratory burst are very similar. Although initiation of both processes was not affected, sustained activation was inhibited by PP1. Taken together, our results demonstrate a critical role for Src kinases in regulating neutrophil cytotoxic-effector functioning through PI3K-PKB.     

Differential inhibitory effects of baicalein and baicalin on LPS-induced cyclooxygenase-2 expression through inhibition of C/EBPbeta DNA-binding activity

To evaluate the possible mechanisms responsible for the anti-inflammatory effects of baicalein or baicalin, lipopolysaccharide (LPS)-induced inflammatory responses in cultured Raw 264.7 cells were studied. In the present study, baicalein and baicalin, a flavonoid present in the root of Scutellaria baicalensis Georgi, were examined for their effects on LPS-induced cyclooxygenase-2 (COX-2) gene expression in Raw 264.7 macrophages. Baicalein, but not baicalin, inhibited COX-2 gene expression in LPS-induced Raw 264.7 cells. However, both polyphenolic compounds inhibited LPS-induced inducible nitric oxide synthase (iNOS) protein expression, iNOS mRNA expression, and NO production in a dose-dependent manner. To investigate the mechanism by which baicalein inhibits COX-2 gene expression, we examined activation of mitogen-activated protein kinases (MAPKs) in Raw 264.7 cells. We did not observe any significant change in the phosphorylation of MAPKs between baicalein- and baicalin-treated cells. Baicalein and baicalin had no effect on LPS-induced nuclear factor-kappaB (NF-kappaB) and cAMP response element binding protein (CREB) DNA binding activity. Baicalein, but not baicalin, significantly inhibited the DNA binding activity of CCAAT/enhancer binding protein beta (C/EBPbeta) These results indicated that differential effects of baicalein and baicalin on COX-2 gene expression in LPS-induced Raw 264.7 cells were mediated through inhibition of C/EBPbeta DNA binding activity. Taken together, these results suggest that baicalein acts to inhibit inflammation through inhibition of COX-2 gene expression through blockade of C/EBPbeta DNA binding activity.     

The phosphatidylinositol 3-kinase/protein kinase B signaling pathway is activated by lipoteichoic acid and plays a role in Kupffer cell production of interleukin-6 (IL-6) and IL-10

Sepsis caused by gram-positive bacteria lacking lipopolysaccharide (LPS) has become a major and increasing cause of mortality in intensive-care units. We have recently demonstrated that the gram-positive-specific bacterial cell wall component lipoteichoic acid (LTA) stimulates the release of the proinflammatory cytokines in Kupffer cells in culture. In the present study, we have started to assess the signal transduction events by which LTA induces the production of tumor necrosis factor alpha (TNF-alpha), interleukin-6 (IL-6), and the anti-inflammatory cytokine IL-10 in rat Kupffer cells. LTA was found to trigger phosphorylation of mitogen-activated protein kinases (MAPK) (p38 MAPK and ERK 1/2) and protein kinase B (PKB). Compared to LPS, LTA was more potent in inducing PKB phosphorylation after 40 min, although we found that the cytokine responses were similar. For both bacterial molecules, blocking phosphatidylinositol 3-kinase (PI3-K; Ly294002) or Janus kinase 2 (JAK-2; AG490) particularly affected the induction of IL-6 and IL-10 release, whereas TNF-alpha levels were strongly reduced by inhibition of Src family tyrosine kinases (PP2). All three cytokines were reduced by inhibition of p38 MAPK (SB202190) or the broad-range tyrosine kinase inhibitor genistein, whereas IL-6 release was particularly blocked by inhibition of ERK 1/2 (PD98059). Divergences in the regulatory pathways controlling TNF-alpha, IL-10, and IL-6 production in Kupffer cells following LPS or LTA stimulation may create a basis for understanding how the balance between pro- and anti-inflammatory cytokines is regulated in the liver following infections by gram-positive or gram-negative bacteria.     

c-Src in paraventricular nucleus modulates sympathetic activity and cardiac sympathetic afferent reflex in renovascular hypertensive rats

Enhanced cardiac sympathetic afferent reflex (CSAR) contributes to sympathetic activation in renovascular hypertension. The study was to determine whether c-Src in paraventricular nucleus (PVN) is involved in the enhanced CSAR and sympathetic activation in hypertensive rats induced by two-kidney one-clip (2K1C). At the end of the fourth week after 2K1C surgery, renal sympathetic nerve activity (RSNA) was recorded in anesthetized rats with baroreceptor denervation and vagotomy. The CSAR was evaluated by the RSNA response to epicardial application of capsaicin. In the PVN, c-Src activity was higher in 2K1C rats than sham-operated (Sham) rats while c-Src expression was not. Epicardial application of capsaicin or PVN microinjection of angiotensin II (Ang II) increased c-Src activity more in 2K1C than Sham rats. PVN microinjection of selective Src family kinase inhibitor 4-Amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazol [3,4-D] pyrimidine (PP2) or 2,3-Dihydro-N,N-dimethyl-2-oxo-3-[(4,5,6,7-tetrahydro-1?H-indol-2-yl)methylene]-1?H-indole-5-sulfonamide (SU6656) abolished the CSAR and decreased RSNA more in 2K1C than Sham rats. The Ang II-induced RSNA and CSAR enhancement was abolished by PP2 or SU6656 pretreatment in 2K1C and Sham rats. NAD(P)H oxidase activity and superoxide anion level in PVN were higher in 2K1C rats, which was attenuated by PP2 but increased by epicardial application of capsaicin or PVN microinjection of Ang II. The effects of capsaicin or Ang II were abolished by PP2. These results indicate that c-Src in the PVN is involved in the enhanced CSAR and sympathetic activation in renovascular hypertension, and mediates the excitatory effects of Ang II in the PVN on the CSAR and sympathetic activity via NAD(P)H oxidase-derived generation of superoxide anions.     

Activation of src-family tyrosine kinases by LPS regulates cytokine production in dendritic cells by controlling AP-1 formation

The role of src-family tyrosine kinases in LPS-induced DC maturation has not been fully addressed. We show that LPS induces activation of c-Src and Lyn in human DC. Inhibition of these kinasesby PP1 uncoupled LPS-induced cytokine production from the up-regulation of costimulatory molecules, resulting in DC still capable of stimulating T cell proliferation but much less efficient in inducing Th1 differentiation. This is the first example of a pharmacological inhibitor able to modulate the capacity of DC to induce a particular type of immune response. Inhibition of src-family kinases impaired phosphorylation and accumulation of c-Jun, leading to reduced formation of AP-1 complexes upon LPS stimulation. Thus, src-kinases control cytokine production in LPS-induced DC maturation through a timely formation of AP-1.