Involvement of p38 MAP kinase in TGF-beta-stimulated VEGF synthesis in aortic smooth muscle cells

Although it is known that transforming growth factor (TGF)-beta induces vascular endothelial growth factor (VEGF) synthesis in vascular smooth muscle cells, the underlying mechanisms are still poorly understood. In the present study, we examined whether the mitogen-activated protein (MAP) kinase superfamily is involved in TGF-beta-stimulated VEGF synthesis in aortic smooth muscle A10 cells. TGF-beta stimulated the phosphorylation of p42/p44 MAP kinase and p38 MAP kinase, but not that of SAPK (stress-activated protein kinase)/JNK (c-Jun N-terminal kinase). The VEGF synthesis induced by TGF-beta was not affected by PD98059 or U0126, specific inhibitors of the upstream kinase that activates p42/p44 MAP kinase. We confirmed that PD98059 or U0126 did actually suppress the phosphorylation of p42/p44 MAP kinase by TGF-beta in our preparations. PD169316 and SB203580, specific inhibitors of p38 MAP kinase, significantly reduced the TGF-beta-stimulated synthesis of VEGF (each in a dose-dependent manner). PD169316 or SB203580 attenuated the TGF-beta-induced phosphorylation of p38 MAP kinase. These results strongly suggest that p38 MAP kinase plays a part in the pathway by which TGF-beta stimulates the synthesis of VEGF in aortic smooth muscle cells.     

Involvement of MAP kinases in TGF-beta-stimulated vascular endothelial growth factor synthesis in osteoblasts

Transforming growth factor-beta (TGF-beta) reportedly induces vascular endothelial growth factor (VEGF) synthesis in osteoblast-like MC3T3-E1 cells. We have recently shown that TGF-beta activates p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase in these cells. In the present study, we investigated the exact mechanism of TGF-beta behind the synthesis of VEGF in MC3T3-E1 cells. PD98059 and U-0126, specific inhibitors of MEK, suppressed the VEGF synthesis induced by TGF-beta. U-0126 inhibited the TGF-beta-induced p44/p42 MAP kinase phosphorylation. SB203580 and PD169316, inhibitors of p38 MAP kinase, reduced the TGF-beta-stimulated VEGF synthesis. SB202474, a negative control for p38 MAP kinase inhibitor, did not affect the VEGF synthesis. A combination with PD98059 and SB203580 almost completely suppressed the TGF-beta-induced VEGF synthesis. Retinoic acid, which alone failed to affect VEGF synthesis, markedly enhanced the VEGF synthesis stimulated by TGF-beta. Retinoic acid enhanced the TGF-beta-increased levels of VEGF mRNA. The amplifications by retinoic acid of TGF-beta-increased VEGF synthesis and levels of VEGF mRNA were reduced by PD98059 or SB203580. The combination of PD98059 and SB203580 almost completely suppressed the enhancement by retinoic acid of VEGF synthesis induced by TGF-beta. Taken together, our results strongly suggest that both p44/p42 MAP kinase and p38 MAP kinase take part in TGF-beta-stimulated VEGF synthesis in osteoblasts, and that retinoic acid upregulates the VEGF synthesis.     

p38 MAP kinase regulates BMP-4-stimulated VEGF synthesis via p70 S6 kinase in osteoblasts

We previously reported that p70 S6 kinase takes part in bone morphogenetic protein-4 (BMP-4)-stimulated vascular endothelial growth factor (VEGF) synthesis in osteoblast-like MC3T3-E1 cells. Recently, we showed that BMP-4-induced osteocalcin synthesis is regulated by p44/p42 MAP kinase and p38 MAP kinase in these cells. In the present study, we investigated whether the MAP kinases are involved in the BMP-4-stimulated synthesis of VEGF in MC3T3-E1 cells. PD-98059 and U-0126, inhibitors of the upstream kinase of p44/p42 MAP kinase, failed to affect BMP-4-stimulated VEGF synthesis. SB-203580 and PD-169316, inhibitors of p38 MAP kinase, significantly reduced VEGF synthesis, whereas SB-202474, a negative control for p38 MAP kinase inhibitor, had little effect on VEGF synthesis. The BMP-4-stimulated phosphorylation of p38 MAP kinase was not affected by rapamycin, an inhibitor of p70 S6 kinase. On the contrary, SB-203580 and PD-169316 reduced the BMP-4-stimulated phosphorylation of p70 S6 kinase. In addition, anisomycin, an activator of p38 MAP kinase, phosphorylates p70 S6 kinase, and the phosphorylation was suppressed by SB-203580. LY-294002, an inhibitor of phosphatidylinositol 3-kinase, failed to suppress the phosphorylation of p38 MAP kinase induced by BMP-4. Not BMP-4 but anisomycin weakly induced the phosphorylation of phosphoinositide-dependent kinase-1. However, anisomycin had little effect on phosphorylation of either Akt or the mammalian target of rapamycin. Taken together, our results suggest that p38 MAP kinase functions in BMP-4-stimulated VEGF synthesis as a positive regulator at a point upstream from p70 S6 kinase in osteoblasts.     

Activation of p44/p42 mitogen-activated protein kinase limits triiodothyronine-stimulated alkaline phosphatase activity in osteoblasts

It has been shown that thyroid hormone stimulates the activity of alkaline phosphatase, a marker of mature osteoblast phenotype, in osteoblasts. In the present study, we investigated whether p44/p42 mitogen-activated protein (MAP) kinase is involved in the thyroid hormone-stimulated alkaline phosphatase activity in osteoblast-like MC3T3-E1 cells. Triiodothyronine (T(3)) markedly induced the phosphorylation of p44/p42 MAP kinase. PD98059 and U0126, inhibitors of the upstream kinase that activates p44/p42 MAP kinase, significantly enhanced the T(3)-induced alkaline phosphatase activity in a dose-dependent manner. The phosphorylation of p44/p42 MAP kinase induced by T(3) was reduced by U0126. These results strongly suggest that p44/p42 MAP kinase takes part in the thyroid hormone-stimulated alkaline phosphatase activity in osteoblasts and that p44/p42 MAP kinase plays an inhibitory role in the thyroid hormone-effect.     

Differential roles of MAP kinases in atorvastatin-induced VEGF release in cardiac myocytes

Statins, specific inhibitors of 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, are now widely used for treatment of patients with hypercholesterolemia. In addition to the reduction of cholesterol biosynthesis, accumulating evidence indicates that statins have several pleiotropic effects especially on cardiovascular system. However, the exact role of statin in cardiac myocytes remains unclear. In the present study, we investigated whether atorvastatin induces vascular endothelial growth factor (VEGF) release in cardiac myocytes, and the underlying mechanism. We observed that atorvastatin significantly stimulated VEGF release in a dose-dependent manner. It induced the phosphorylation of p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase but not SAPK (stress-activated protein kinase)/JNK (c-Jun N-terminal kinase). The atorvastatin-induced VEGF release was enhanced by PD98059, which is a specific inhibitor of the upstream kinase that activates p44/p42 MAP kinase (MEK). Further, it was significantly reduced by SB203580, a specific inhibitor of p38 MAP kinase. Furthermore, the atorvastatin-induced phosphorylation of p38 MAP kinase was attenuated by SB203580, whereas it was enhanced by PD98059. Taken together, these results suggest that the atorvastatin-induced VEGF release in cardiac myocytes is positively regulated by p38 MAP kinase and negatively regulated byp44/p42 MAP kinase and that the atorvastatin-induced phosphorylation of p38 MAP kinase is regulated by p44/p42 MAP kinase in these cells.     

Up-regulation by zinc of FGF-2-induced VEGF release through enhancing p44/p42 MAP kinase activation in osteoblasts

We previously reported that basic fibroblast growth factor (FGF-2) activates stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) and p44/p42 mitogen-activated protein (MAP) kinase resulting in the stimulation of vascular endothelial growth factor (VEGF) release in osteoblast-like MC3T3-E1 cells. In the present study, we investigated whether zinc affects the VEGF release by FGF-2 in MC3T3-E1 cells. The FGF-2-induced VEGF release was significantly enhanced by ZnSO(4) but not Na(2)SO(4). The enhancing effect of ZnSO(4) was dose-dependent between 1 and 100 muM. ZnSO(4) markedly enhanced the FGF-2-induced phosphorylation of p44/p42 MAP kinase while having little effect on the SAPK/JNK phosphorylation. PD98059 significantly reduced the amplification by ZnSO(4) of the FGF-2-stimulated VEGF release. Taken together, our findings strongly suggest that zinc enhances FGF-2-stimulated VEGF release resulting from up-regulating activation of p44/p42 MAP kinase in osteoblasts.     

Evidence implicating a mid-region sequence of IGFBP-3 in its specific IGF-independent actions

In the present study, we investigated whether the mitogen-activated protein (MAP) kinase superfamily is involved in the bone morphogenetic protein (BMP)-4-stimulated synthesis of osteocalcin in osteoblast-like MC3T3-E1 cells. BMP-4 dose-dependently stimulated osteocalcin synthesis. BMP-4 markedly induced the phosphorylation of p44/p42 MAP kinase and p38 MAP kinase, while having little effect on SAPK (stress-activated protein kinase)/JNK (c-Jun N terminal kinase) phosphorylation. SB203580 and PD169316, specific inhibitors of p38 MAP kinase, significantly reduced the osteocalcin synthesis stimulated by BMP-4. In contrast, PD98059 and U0126, inhibitors of upstream kinase of p44/p42 MAP kinase, markedly enhanced the BMP-4-stimulated osteocalcin synthesis. The BMP-4-induced phosphorylation of p44/p42 MAP kinase was suppressed by PD98059, which did not, however, affect the BMP-4-induced phosphorylation of p38 MAP kinase. Taken together, our results strongly suggest that p38 MAP kinase takes part in BMP-4-stimulated osteocalcin synthesis as a positive regulator in osteoblasts, whereas p44/p42 MAP kinase acts as a negative regulator in the synthesis.     

SAPK/JNK plays a role in transforming growth factor-beta-induced VEGF synthesis in osteoblasts.

We previously reported that transforming growth factor-beta (TGF-beta) activates p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase, resulting in the stimulation of vascular endothelial growth factor (VEGF) synthesis in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the involvement of stress-activated protein kinase/c- Jun N-terminal kinase (SAPK/JNK), another member of the MAP kinase superfamily, in TGF-beta-induced VEGF synthesis in these cells. TGF-beta markedly induced SAPK/JNK phosphorylation. SP600125, a specific inhibitor of SAPK/JNK, markedly reduced TGF-beta-induced VEGF synthesis. SP600125 suppressed TGF-beta-induced SAPK/JNK phosphorylation. PD98059, an inhibitor of upstream kinase of p44/p42 MAP kinase and SB203580, an inhibitor of p38 MAP kinase, each failed to reduce TGF-beta-induced SAPK/JNK phosphorylation. A combination of SP600125 and PD98059 or SP600125 and SB203580 suppressed TGF-beta-stimulated VEGF synthesis in an additive manner. These results strongly suggest that TGF-beta activates SAPK/JNK in osteoblasts, and that SAPK/JNK plays a role in addition to p42/p44 MAP kinase and p38 MAP kinase in TGF-beta-induced VEGF synthesis.     

Platelet-derived growth factor-BB amplifies PGF2alpha-stimulated VEGF synthesis in osteoblasts: function of phosphatidylinositol 3-kinase

We have reported that prostaglandin F2alpha (PGF2alpha) stimulates the synthesis of vascular endothelial growth factor (VEGF) via p44/p42 mitogen-activated protein (MAP) kinase in osteoblast-like MC3T3-E1 cells. In addition, we recently showed that phosphatidylinositol 3 (PI3)-kinase activated by platelet-derived growth factor-BB (PDGF-BB) negatively regulates the interleukin-6 synthesis in these cells. In the present study, we investigated the effect of PDGF-BB on the PGF2alpha-induced VEGF synthesis in MC3T3-E1 cells. PDGF-BB, which alone did not affect the levels of VEGF, significantly enhanced the PGF2alpha-stimulated VEGF synthesis. The amplifying effect of PDGF-BB was dose dependent in the range between 10 and 70 ng/ml. LY294002 or wortmannin, specific inhibitors of PI3-kinase, which by itself failed to affect the PGF2alpha-stimulated VEGF synthesis, significantly suppressed the amplification by PDGF-BB. PD98059, a specific inhibitor of MEK1/2, suppressed the amplification by PDGF-BB of the PGF2alpha-stimulated VEGF synthesis similar to the levels of PGF2alpha with PD98059. PDGF-BB itself induced the phosphorylation of p44/p42 MAP kinase in these cells, and the effects of PDGF-BB and PGF2alpha on the phosphorylation of p44/p42 MAP kinase were additive. Moreover, LY294002 had little effect on the phosphorylation of p44/p42 MAP kinase induced by PGF2alpha with PDGF-BB. These results strongly suggest that PGF2alpha-stimulated VEGF synthesis is amplified by PI3-kinase-mediating PDGF-BB signaling in osteoblasts, and that the effect is exerted at a point downstream from p44/p42 MAP kinase.     

Endothelin-1 induces vascular endothelial growth factor synthesis in osteoblasts: involvement of p38 mitogen-activated protein kinase

We previously reported that endothelin-1 (ET-1) stimulates p44/p42 mitogen-activated protein (MAP) kinase and p38 MAP kinase in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the effect of ET-1 on the synthesis of vascular endothelial growth factor (VEGF) in these cells. ET-1 significantly stimulated VEGF secretion time-dependently 18 hours after the stimulation. The stimulatory effect was dose-dependent in the range between 0.1 nM and 0.1 micro;M. BQ123, an antagonist of endothelin(A) (ET(A)) receptor, inhibited the ET-1-induced VEGF secretion. The ET-1-induced VEGF secretion was suppressed by SB203580 and PD169316, inhibitors of p38 MAP kinase, but not PD98059, an inhibitor of the upstream kinase that activates p44/p42 MAP kinase. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a protein kinase C (PKC)-activating phorbol ester, stimulated VEGF secretion. Calphostin C, a specific PKC inhibitor, suppressed the VEGF secretion by ET-1. TPA-induced VEGF secretion was suppressed by SB203580. Taken together, our results strongly suggest that ET-1 stimulates VEGF synthesis via ET(A) receptor in osteoblasts and that p38 MAP kinase is involved at a point downstream from PKC in the VEGF synthesis.     

Nitrosylation: the next phosphorylation?

Vitamin D3 plays an important role in the regulation of mineral homeostasis, cell differentiation, and proliferation. However, the exact role of vitamin D3 in vascular smooth muscle cells remains unclear. In the present study, we investigated whether vitamin D3 induces vascular endothelial growth factor (VEGF) release in aortic smooth muscle A10 cells. 1,25-Dihydroxyvitamin D3 (1,25(OH)2VD3), an active form of vitamin D3, stimulated the VEGF release while 24,25-dihydroxyvitamin D3 (24,25(OH)2VD3), an inactive form of vitamin D3, had little effect on the release. The stimulatory effect of 1,25(OH)2VD3 was dose dependent in the range between 10 pM and 10 nM. 1,25(OH)2VD3 induced the phosphorylation of p38 mitogen-activated protein (MAP) kinase but 24,25(OH)2VD3 did not. PD169316 and SB203580, specific inhibitors of p38 MAP kinase, significantly reduced the 1,25(OH)2VD3-stimulated release of VEGF. On the contrary, SB202474, a negative control for p38 MAP kinase inhibitor, had little effect on the VEGF release. PD169316 attenuated the 1,25(OH)2VD3-induced phosphorylation of p38 MAP kinase. These results strongly suggest that 1,25(OH)2VD3 stimulates the release of VEGF in aortic smooth muscle cells via p38 MAP kinase activation.     

Lipoteichoic acid-stimulated p42/p44 MAPK activation via Toll-like receptor 2 in tracheal smooth muscle cells

Lipoteichoic acid (LTA), the principal component of the cell wall of gram-positive bacteria, triggers several inflammatory responses. However, the mechanisms underlying its action on human tracheal smooth muscle cells (HTSMCs) were largely unknown. This study was to investigate the mechanisms underlying LTA-stimulated p42/p44 mitogen-activated protein kinase (MAPK) using Western blotting assay. LTA stimulated phosphorylation of p42/p44 MAPK via a Toll-like receptor 2 (TLR2). Pretreatment with pertussis toxin attenuated the LTA-induced responses. LTA-stimulated phosphorylation of p42/p44 MAPK was attenuated by inhibitors of tyrosine kinase (genistein), phosphatidylcholine-phospholipase C (PLC; D609), phosphatidylinositol (PI)-PLC (U-73122), PKC (staurosporine, G?-6976, rottlerin, or Ro-318220), MEK1/2 (U-0126), PI 3-kinase (LY-294002 and wortmannin), and an intracellular Ca(2+) chelator (BAPTA-AM). LTA directly evoked initial transient peak of [Ca(2+)](i), supporting the involvement of Ca(2+) mobilization in LTA-induced responses. These results suggest that in HTSMCs, LTA-stimulated p42/p44 MAPK phosphorylation is mediated through a TLR2 receptor and involves tyrosine kinase, PLC, PKC, Ca(2+), MEK, and PI 3-kinase.     

Reactive oxidant and p42/44 MAP kinase signaling is necessary for mechanical strain-induced proliferation in pulmonary epithelial cells.

Mechanical strain is necessary for normal lung growth and development. Individuals with respiratory failure are supported with mechanical ventilation, leading to altered lung growth and injury. Understanding signaling pathways initiated by mechanical strain in lung epithelial cells will help guide development of strategies aimed at optimizing strain-induced lung growth while mitigating ventilator-induced lung injury. To study strain-induced proliferative signaling, focusing on the role of reactive oxidant species (ROS) and p42/44 mitogen-activated protein (MAP) kinase, human pulmonary epithelial H441 and MLE15 cells were exposed to equibiaxial cyclic mechanical strain. ROS were increased within 15 min of strain. N-acetylcysteine inactivated strain-induced ROS and inhibited p42/44 MAP kinase phosphorylation and strain-induced proliferation. PD98059 and UO126, p42/44 MAP kinase inhibitors, blocked strain-induced proliferation. To verify the specificity of p42/44 MAP kinase inhibition, cells were transfected with dominant-negative mitogen-activated protein kinase kinase-1 plasmid DNA. Transfected cells did not proliferate in response to mechanical strain. To determine whether strain-induced tyrosine kinase activity is necessary for strain-induced ROS-p42/44 MAP kinase signaling, genistein, a tyrosine kinase inhibitor, was used. Genistein did not block strain-induced ROS production or p42/44 MAP kinase phosphorylation. Gadolinium, a mechanosensitive calcium channel blocker, blocked strain-induced ROS production and p42/44 MAP kinase phosphorylation but not strain-induced tyrosine phosphorylation. These data support ROS production and p42/44 MAP kinase phosphorylation being involved in a common strain-induced signaling pathway, necessary for strain-induced proliferation in pulmonary epithelial cells, with a parallel strain-induced tyrosine kinase pathway.     

Sustained phosphorylation of mitogen-activated protein kinase is required for basic fibroblast growth factor-mediated axonal branch formation in cultured rat hippocampal neurons

Basic fibroblast growth factor (bFGF) has been reported to promote the formation of axonal branches in cultured brain neurons. In the present study, we investigated whether the mitogen-activated protein kinase (MAPK) cascade was involved in this action of bFGF in cultured rat hippocampal neurons. Recombinant human bFGF (0.1-10 ng/ml) induced phosphorylation of p44/42 MAPK in a concentration and time-dependent manner. The phosphorylation of p44/42 MAPK occurred rapidly within 5 min after addition of bFGF, and lasted for 48 h. The bFGF-induced phosphorylation of p44/42 MAPK and axonal branch formation were both blocked by simultaneous addition of U0126 and PD98059, specific inhibitors of MAPK kinases. Furthermore, when U0126 and PD98059 were added 24 h after bFGF, phosphorylation of p44/42 m MAPK was decreased, and axonal branch formation was stopped. These results suggest that sustained activation of the MAPK cascade is required for bFGF-mediated axonal branch formation.     

Bovine milk lactoferrin induces synthesis of the angiogenic factors VEGF and FGF2 in osteoblasts via the p44/p42 MAP kinase pathway

Lactoferrin (LF) belongs to the transferrin family and is present in several physiological fluids, including milk and colostrum. LF has recently been identified as an anabolic factor for bone. Here we investigated whether bovine LF (bLF) induces synthesis of angiogenic factors by osteoblasts. If so, we examined the underlying mechanism. We found that bLF purified from milk increased the mRNA expression of vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF2) in murine osteoblast-like MC3T3-E1 cells and primary murine osteoblasts in a time- and dose-dependent manner. Furthermore, bLF increased VEGF and FGF2 protein levels in MC3T3-E1 cells. In addition, treatment of MC3T3-E1 cells with bLF rapidly induced phosphorylation of p44/p42 mitogen-activated protein (MAP) kinase. The bLF-mediated increases in VEGF and FGF2 mRNA and protein were inhibited by U0126, a specific inhibitor of the upstream kinase that activates p44/p42 MAP kinase (MEK). Taken together, our results strongly suggest that bLF induces VEGF and FGF2 synthesis in a p44/p42 MAP kinase-dependent manner in MC3T3-E1 cells.     

Enhancement by sphingosine 1‐phosphate in vasopressin‐induced phosphoinositide hydrolysis in aortic smooth‐muscle cells: Involvement of p38 MAP kinase

We previously reported that sphingosine 1‐phosphate (S‐1‐P), a sphingomyelin metabolite, activates p44/p42 mitogen‐activated protein (MAP) kinase and p38 MAP kinase in aortic smooth‐muscle A10 cells. In the present study, we investigated the effect of sphingomyelin metabolites on phospholipase C‐catalyzing phosphoinositide hydrolysis induced by arginine vasopressin (AVP) in A10 cells. C₂‐ceramide and sphingosine had little effect on inositol phosphate (IP) formation stimulated by AVP. S‐1‐P, which alone slightly stimulated the IPs formation, dose‐dependently amplified the AVP‐induced formation of IPs. Tumor necrosis factor‐α enhanced the AVP‐induced formation of IPs. However, S‐1‐P did not enhance the formation of IPs by NaF, a heterotrimeric GTP‐binding protein activator. Pertussis toxin inhibited the effect of S‐1‐P. PD98059, an inhibitor of the upstream kinase that activates p44/p42 MAP kinase, had little effect on the enhancement by S‐1‐P. SB203580, an inhibitor of p38 MAP kinase, suppressed the effect of S‐1‐P on the formation of IPs by AVP. SB203580 inhibited the AVP‐induced phosphorylation of p38 MAP kinase. Pertussis toxin suppressed the phosphorylation of p38 MAP kinase by S‐1‐P. These results indicate that S‐1‐P amplifies AVP‐induced phosphoinositide hydrolysis by phospholipase C through p38 MAP kinase in vascular smooth‐muscle cells. J. Cell. Biochem. 80:46–52, 2000. © 2000 Wiley‐Liss, Inc.     

Sphingosine 1-phosphate amplifies phosphoinositide hydrolysis stimulated by prostaglandin f2 alpha in osteoblasts: involvement of p38MAP kinase

We previously showed that sphingosine 1-phosphate phosphorylates p42/p44 mitogen-activated protein (MAP) kinase and p38 MAP kinase in osteoblast-like MC3T3-E1 cells. In the present study, we investigated the effect of sphingosine 1-phosphate on phospholipase C-catalyzing phosphoinositide hydrolysis induced by prostaglandin F2alpha (PGF2 alpha) in these cells. Sphingosine 1-phosphate significantly amplified the inositol phosphates formation by PGF2 alpha. Sphingosine 1-phosphate did not enhance the formation induced by NaF, a direct activator of heterotrimeric GTP-binding proteins. PD98059, an inhibitor of the kinase that activates p42/p44 MAP kinase, had little effect on the amplification by sphingosine 1-phosphate. SB203580, an inhibitor of p38 MAP kinase, reduced the effect of sphingosine 1-phosphate on the formation of inositol phosphates by PGF2 alpha. The phosphorylation of p42/p44 MAP kinase by PGF alpha was attenuated by PD98059. SB203580 suppressed the phosphorylation of p38 MAP kinase by PGF2 alpha. Tumor necrosis factor-alpha enhanced the PGF2 alpha-stimulated formation of inositol phosphates. These results strongly suggest that sphingosine 1-phosphate amplifies PGF2 alpha-induced phosphoinositide hydrolysis by phospholipase C through p38 MAP kinase in osteoblasts.     

Sphingosine 1-Phosphate Regulates Heat Shock Protein 27 Induction by a p38 MAP Kinase-Dependent Mechanism in Aortic Smooth Muscle Cells

In an aortic smooth muscle cell line, A10 cells, we investigated the effect of sphingosine 1-phosphate on the induction of heat shock protein 27 (HSP27), a low-molecular-weight heat shock protein. Sphingosine 1-phosphate significantly induced the accumulation of HSP27 in a pertussis toxin-sensitive manner. The effect was dose-dependent in the range between 0.1 and 30 microM. Sphingosine 1-phosphate stimulated an increase in the levels of mRNA for HSP27. Sphingosine 1-phosphate stimulated both p42/p44 mitogen-activated protein (MAP) kinase and p38 MAP kinase activation. PD98059, an inhibitor of the upstream kinase that activates p42/p44 MAP kinase, did not affect sphingosine 1-phosphate-stimulated HSP27 induction. In contrast, SB203580, an inhibitor of p38 MAP kinase, reduced sphingosine 1-phosphate-induced HSP27 induction. SB203580 reduced the levels of mRNA for HSP27 induced by sphingosine 1-phosphate. These results indicate that sphingosine 1-phosphate stimulates the induction of HSP27 via p38 MAP kinase activation in aortic smooth muscle cells.