Details of a Researcher - KOUNO Takayuki

Papers - KOUNO Takayuki

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Poly(I:C) induced microRNA-146a regulates epithelial barrier and secretion of proinflammatory cytokines in human nasal epithelial cells

Ryo Miyata, Takuya Kakuki, Kazuaki Nomura, Tsuyoshi Ohkuni, Noriko Ogasawara, Ken-ichi Takano, Takumi Konno, Takayuki Kohno, Norimasa Sawada, Tetsuo Himi, Takashi Kojima

EUROPEAN JOURNAL OF PHARMACOLOGY ( ELSEVIER SCIENCE BV ) 761 375 - 382 2015.08 [Refereed]

Human nasal epithelial cells (HNECs) are important in the tight junctional barrier and innate immune defense protecting against pathogens invading via Toll-like receptors (TERs). MicroRNAs (miRNAs) I egulate expression of tight junctions as direct or indirect targeting genes and maintain the barrier Function. However, the roles of miRNAs in the epithelial barrier of HNECs via TLRs remain unknown. In the present study, to investigate the effects of miRNAs on the epithelial barrier of HNECs via TLRs, primary cultured HNECs transfected with human telomerase reverse transcriptase (hTERT-HNECs), were treated with the TLR3 ligand poly(I:C) and miRNA array analysis was performed. In the miRNA array of the cells treated with poly(I:C), upregulation of miR-187, -146a, -574, -4274, -4433, -4455 and -4750, and downregulation of miR-4785 by more than twofold compared to the control were observed. When control HNECs were treated with mimics and inhibitors of these miRNAs, an miR-146a mimic induced expression of tight junction proteins claudin-1, occludin and JAM-A together with an increase of the epithelial barrier function. The poly(I:C)-induced miR-146a was regulated via the distinct TLR3-mediatecl signal pathways PI3K, JNK and NE-kappa B. Furthermore, the miR-146a mimic prevented downregulation of claudin-1 and JAM-A and the secretion of proinflammatory cytokines IL-8 and TNF-alpha induced by poly (I:C) by targeting TRAFG. These findings indicate that, in HNECs, miRNA-146a plays crucial roles in maintenance of the tight junction barrier and innate immune defense protecting against invading pathogens. (C) 2015 Elsevier B.V. All rights reserved.

DOI PubMed

ヒト子宮内膜癌細胞の悪性化におけるlipolysis-stimulated lipoprotein receptorの役割

嶋田浩志, 郷久晴朗, 金野匠, 高橋俊太, 畠山翔翼, 齋藤豪, 幸野貴之, 小島隆

日本細胞生物学会大会講演要旨集 ( (一社)日本細胞生物学会 ) 67回 167 - 167 2015.06

子宮体がん基礎肥満による子宮内膜癌の悪性化におけるLSR(lipolysis-stimulated lipoprotein receptor)の役割

嶋田浩志, 小島隆, 郷久晴朗, 金野匠, 高橋俊太, 畠山翔翼, 幸野貴之, 寺本瑞絵, 田中綾一, 岩崎雅宏, 斎藤豪

日本婦人科腫瘍学会雑誌 ( (公社)日本婦人科腫瘍学会 ) 33 ( 3 ) 535 - 535 2015.06

c-Jun N-terminal kinase inhibitor SP600125 enhances barrier function and elongation of human pancreatic cancer cell line HPAC in a Ca-switch model

Takumi Konno, Takafumi Ninomiya, Takayuki Kohno, Shin Kikuchi, Norimasa Sawada, Takashi Kojima

HISTOCHEMISTRY AND CELL BIOLOGY ( SPRINGER ) 143 ( 5 ) 471 - 479 2015.05 [Refereed]

c-Jun N-terminal kinase (JNK), known as a stress-activated protein kinase, regulates normal epithelial biological processes, including assembly of adherens and tight junctions, and it is involved in the development of several cancers. The JNK inhibitor SP600125 enhances epithelial barrier function through modulation of tight junction molecules in normal human pancreatic epithelial cells. Furthermore, this JNK inhibitor suppresses the growth of human pancreatic cancer cells. However, the effects of SP600125 on the epithelial barrier in human pancreatic cancer cells remain unknown. In the present study, the JNK inhibitor SP600125 markedly enhanced the barrier function and cell elongation of well-differentiated human pancreatic cancer cell line HPAC in a Ca-switch model. The epithelial barrier function induced by SP600125 was regulated by phosphorylated beta-catenin without changes in the tight junction molecules. The cell elongation induced by SP600125 was closely related to the expression of the F-actin-binding protein DrebrinE. These findings suggest that JNK is involved in the regulation of the epithelial barrier function and cell shape during remodeling of pancreatic cancer cells. The JNK inhibitor SP600125 may have potential as a therapeutic drug for pancreatic cancer via induction of differentiation.

DOI PubMed

Irsogladine Maleate Regulates Gap Junctional Intercellular Communication-Dependent Epithelial Barrier in Human Nasal Epithelial Cells

Ryo Miyata, Kazuaki Nomura, Takuya Kakuki, Ken-ichi Takano, Takayuki Kohno, Takumi Konno, Norimasa Sawada, Tetsuo Himi, Takashi Kojima

JOURNAL OF MEMBRANE BIOLOGY ( SPRINGER ) 248 ( 2 ) 327 - 336 2015.04 [Refereed]

The airway epithelium of the human nasal mucosa acts as the first physical barrier that protects against inhaled substances and pathogens. Irsogladine maleate (IM) is an enhancer of gastric mucosal protective factors via upregulation of gap junctional intercellular communication (GJIC). GJIC is thought to participate in the formation of functional tight junctions. However, the effects of IM on GJIC and the epithelial barrier in human nasal epithelial cells (HNECs) remain unknown. To investigate the effects of IM on GJIC and the tight junctional barrier in HNECs, primary cultures of HNECs transfected with human telomerase reverse transcriptase (hTERT-HNECs) were treated with IM and the GJIC inhibitors oleamide and 18 beta-GA. Some cells were pretreated with IM before treatment with TLR3 ligand poly(I:C) to examine whether IM prevented the changes via TLR3-mediated signal pathways. In hTERT-HNECs, GJIC blockers reduced the expression of tight junction molecules claudin-1, -4, -7, occludin, tricellulin, and JAM-A. IM induced GJIC activity and enhanced the expression of claudin-1, -4, and JAM-A at the protein and mRNA levels with an increase of barrier function. GJIC blockers prevented the increase of the tight junction proteins induced by IM. Furthermore, IM prevented the reduction of JAM-A but not induction of IL-8 and TNF-alpha induced by poly(I:C). In conclusion, IM can maintain the GJIC-dependent tight junctional barrier via regulation of GJIC in upper airway nasal epithelium. Therefore, it is possible that IM may be useful as a nasal spray to prevent the disruption of the epithelial barrier by viral infections and exposure to allergens in human nasal mucosa.

DOI PubMed

Pseudomonas aeruginosa elastase causes transient disruption of tight junctions and downregulation of PAR-2 in human nasal epithelial cells

Kazuaki Nomura, Kazufumi Obata, Takashi Keira, Ryo Miyata, Satoshi Hirakawa, Ken-ichi Takano, Takayuki Kohno, Norimasa Sawada, Tetsuo Himi, Takashi Kojima

RESPIRATORY RESEARCH ( BIOMED CENTRAL LTD ) 15 2014.02 [Refereed]

Background: Pseudomonas aeruginosa causes chronic respiratory disease, and the elastase enzyme that it produces increases the permeability of airway epithelial cells owing to the disruption of tight junctions. P. aeruginosa is also implicated in prolonged chronic rhinosinusitis. However, the effects of P. aeruginosa elastase (PE) against the barrier formed by human nasal epithelial cells (HNECs) remain unknown. Methods: To investigate the mechanisms involved in the disruption of tight junctions by PE in HNECs, primary cultures of HNECs transfected with human telomerase reverse transcriptase (hTERT-HNECs) were used. The hTERT-HNECs were pretreated with inhibitors of various signal transduction pathways, PKC, MAPK, p38MAPK, PI3K, JNK, NF-kappa B, EGF receptor, proteasome, COX1 and COX2 before treatment with PE. Some cells were pretreated with siRNA and agonist of protease activated receptor-2 (PAR-2) before treatment with PE. Expression and structures of tight junctions were determined by Western blotting, real-time PCR, immunostaining and freeze-fracture. Transepithelial electrical resistance (TER) was examined as the epithelial barrier function. Results: PE treatment transiently disrupted the epithelial barrier and downregulated the transmembrane proteins claudin-1 and -4, occludin, and tricellulin, but not the scaffold PDZ-expression proteins ZO-1 and -2 and adherens junction proteins E-cadherin and beta-catenin. The transient downregulation of tight junction proteins was controlled via distinct signal transduction pathways such as the PKC, MAPK, PI3K, p38 MAPK, JNK, COX-1 and -2, and NF-kappa B pathways. Furthermore, treatment with PE transiently decreased PAR-2 expression, which also regulated the expression of the tight junction proteins. Treatment with a PAR-2 agonist prevented the downregulation of the tight junction proteins after PE treatment in HNECs. Conclusions: PE transiently disrupts tight junctions in HNECs and downregulates PAR-2. The transient disruption of tight junctions by PE might occur repeatedly during chronic rhinosinusitis.

DOI PubMed

Expression of tricellulin in epithelial cells and non-epithelial cells

Takashi Kojima, Takafumi Ninomiya, Takumi Konno, Takayuki Kohno, Masahiko Taniguchi, Norimasa Sawada

HISTOLOGY AND HISTOPATHOLOGY ( F HERNANDEZ ) 28 ( 11 ) 1383 - 1392 2013.11 [Refereed]

Tricellulin is the first molecular component of tricellular tight junctions at tricellular contacts where three epithelial cells meet, and it is required for the their formation and maintenance of the epithelial barrier. Tricellulin binds other tight junction proteins, and its expression and distribution are affected by the bicellular tight junction protein occludin and lipolysis-stimulated lipoprotein receptor (LSR) which is expressed at tricellular contacts. Tricellulin is also detected in endothelial cells, neurons, microglia and astrocytes. Here, we focused tricellulin expression in various types of epithelial cells, nasal epithelial cells, pancreatic duct epithelial cells cells and hepatocytes, and non-epithelial cells, dendritic cells and Schwann cells, compared to expression of the bicellular tight junction protein occludin and LSR, and discuss the regulation and the role of tricellulin in cellular specificity.

DOI DOI2 PubMed

FLT3/ITD regulates leukaemia cell adhesion through alpha 4 beta 1 integrin and Pyk2 signalling

Akira Katsumi, Hitoshi Kiyoi, Akihiro Abe, Ryohei Tanizaki, Toshihiro Iwasaki, Miki Kobayashi, Tadashi Matsushita, Kozo Kaibuchi, Takeshi Senga, Tetsuhito Kojima, Takayuki Kohno, Michinari Hamaguchi, Tomoki Naoe

EUROPEAN JOURNAL OF HAEMATOLOGY ( WILEY-BLACKWELL ) 86 ( 3 ) 191 - 198 2011.03 [Refereed]

Internal tandem duplication of FMS-like receptor tyrosine kinase 3 (FLT3/ITD) within its juxtamembrane domain is a frequent mutation in adult acute myeloid leukaemia (AML). This mutation causes constitutive activation of FLT3 and is associated with poor prognosis. The high relapse rate of FLT3/ITD-positive AML might be partly because of insufficient eradication of slow-cycling leukaemic stem cells in the bone marrow microenvironment. beta 1 integrin mediates haematopoietic stem and progenitor cell homing along with their retention in the bone marrow and also inhibits haematopoietic proliferation and differentiation. Here, we demonstrate that inhibition of FLT3/ITD kinase activity by a FLT3 selective inhibitor named FI-700 decreases affinity of alpha 4 beta 1 integrin to soluble VCAM-1. alpha 4 beta 1 integrin deactivation by FI-700 is independent of Rap1, which is the critical regulator of integrin inside-out signalling. In addition, selective inhibition of FLT3/ITD induces Pyk2 dephosphorylation together with the inhibition of phosphatidylinositol-3-kinase (PI3K)/Akt pathway. Both wild-type and ITD-FLT3 proteins co-immunoprecipitated with beta 1 integrin and Pyk2 indicating the signal crosstalk between FLT3, beta 1 integrin and Pyk2. These results collectively indicated that the inhibition of FLT3 kinase might contribute not only to the induction of apoptosis, but also to the leukaemia cell detachment from the bone marrow microenvironment in the treatment of AML.

DOI PubMed

Sphingosine 1-phosphate induces cyclooxygeriase-2 via Ca2+-dependent, but MAPK-independent mechanism in rat vascular smooth muscle cells

Akiko Nodai, Takuji Machida, Sachiko Izumi, Yumika Hamaya, Takayuki Kohno, Yasuyuki Igarashi, Kenji Iizuka, Masaru Minami, Masahiko Hirafuji

LIFE SCIENCES ( PERGAMON-ELSEVIER SCIENCE LTD ) 80 ( 19 ) 1768 - 1776 2007.04 [Refereed]

The effects of sphingosine 1-phosphate (S1P) on prostaglandin I-2 (PGI(2)) production and cyclooxygenase (COX) expression in cultured rat vascular smooth muscle cells (VSMCs) were investigated. S1P stimulated PGI(2) production in a concentration-dependent manner, which was completely suppressed by NS-398, a selective COX-2 inhibitor, as determined by radioimmunoassay. S1P stimulated COX-2 protein and mRNA expressions in a concentration- and time-dependent manner, while it had no effect on COX-1 expression. S1P(2) and S1P(3) receptors mRNA were abundantly expressed in rat VSMCs. Suramin, an antagonist of S1P(3) receptor, almost completely inhibited S1P-induced COX-2 expression. Pretreatment of VSMCs with pertussis toxin (PTX)partially, but significantly inhibited S1P-inducedPGI(2) production and COX-2 expression. SIP also activated extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK). However, neither PD 98059, a selective inbibitor of ERK activation, nor SB 203580, a selective inhibitor of p38 MAPK, had a significant inhibitory effect on S1P-induced COX-2 expression, suggesting that the MAPK activation does not play main roles in S1P-induced COX-2 induction. S I --P-induced COX-2 expression was inhibited by PP2, an inhibitor of Src-family tyrosine kinase, Ca2+ depletion, and GF 109203X, an inhibitor of protein kinase C (PKC). These results suggest that S 1 P stimulates COX-2 induction in rat VSMCs through mechanisms involving Ca2+-dependent PKC and Src-family tyrosine kinase activation via S I --P-3 receptor coupled to PTX-sensitive and -insensitive G proteins. (C) 2007 Elsevier Inc. All rights reserved.

DOI PubMed

Involvement of Sphingosine 1-Phosphate (S1P) Receptor Type 1 and Type 4 in Migratory Response of Mouse T Cells toward S1P

Hirofumi Matsuyuki, Yasuhiro Maeda, Kazuhiro Yano, Kunio Sugahara, Kenji Chiba, Takayuki Kohno, Yasuyuki Igarashi

CELLULAR & MOLECULAR IMMUNOLOGY ( CHIN SOCIETY IMMUNOLOGY ) 3 ( 6 ) 429 - 437 2006.12 [Refereed]

Sphingosine 1-phosphate (S1P), a pleiotropic lysophospholipid, regulates signal transduction pathway via G-protein-coupled receptors termed S1P(1-5) in several types of the cells including lymphocytes. Higher levels of S1P4 mRNA as well as S1P1 mRNA are expressed in lymphoid tissues such as the spleen, thymus, lymph nodes, and Payer's patches. In contrast to S1P1 that plays an essential role in lymphocyte egress, little is known about the role of S1P4 in immune system. In this study, we found that S1P at 10 to 100 nM significantly induced the cell migration and the significant levels of S1P1 and S1P4 mRNA were expressed in mouse CD4 T cells, D10.G4.1 mouse Th2 cells, and EL-4.IL-2 mouse thymoma cells. In D10.G4.1 and EL-4.IL-2 cells, S1P-induced migration was almost completely inhibited by pretreatment with pertussis toxin, Clostoridium difficile toxin B, and ( S)-enantiomer of FTY720-phosphate, a potent agonist at S1P1 and S1P4. The members of the Rho family small GTPase, Cdc42 and Rac were activated by S1P stimulation in these cells. The transfection with dominant negative or constitutively active forms of Cdc42 and Rac revealed that the activation of both Cdc42 and Rac is essential for S1P-induced migration of these cells. The immunoprecipitation assays using CHO cells co-expressing both S1P4 and S1P1 receptors indicated that S1P4 and S1P1 are associated on the cell surface. These results suggest that the association of S1P4 and S1P1 plays an important role in migratory response of mouse T cells toward S1P.

PubMed

Purification, characterization, cDNA cloning, and expression of asialofetuin-binding C-type lectin from eggs of shishamo smelt (Osmerus [Spirinchus] lanceolatus)

M Hosono, S Sugawara, Y Ogawa, T Kohno, M Takayanagi, K Nitta

BIOCHIMICA ET BIOPHYSICA ACTA-GENERAL SUBJECTS ( ELSEVIER SCIENCE BV ) 1725 ( 2 ) 160 - 173 2005.09 [Refereed]

A novel C-type lectin (OLABL) was isolated from the eggs of shishamo smelt [Osmerus (Spirinchus) lanceolatus] by affinity chromatography on asialofetuin-Sepharose. OLABL had a molecular mass of 29 kDa on SDS-PAGE under nonreducing conditions and two subunits with masses of 15 kDa (OLABL-H) and 14 kDa (OLABL-L) under reducing conditions. Thus, OLABL is a heterodimeric protein. cDNA sequence analysis revealed that the H- and L-subunits of OLABL were composed of 137 and 136 amino acid residues, respectively, and showed almost identical (95%) sequences, with slight differences in the N-terminal and C-terminal regions. Since each subunit contained only the characteristic motif of C-type lectin-like domain (CTLD), EPN-E-WND, OLABL is a member of group VII of the CTLD-containing protein family. Although OLABL had an EPN sequence that is known as a mannosespecific motif found in the collectin family, OLABL agglutinated rabbit erythrocytes without the addition of Ca2+ ion, and this activity was inhibited by L-rhammose and D-galactose derivatives, but not by D-mannose and D-glucose. These results indicate that OLABL has similar characteristics to AJL-2, a calcium-independent lactose specific lectin isolated from Japanese eel skin mucus. Recombinant OLABLs (rHisOLABLs), His-tagged homodimers of the H- and L-subunits, were refolded from inclusion bodies expressed by Escherichia coli. rHisOLABL-L was recovered as a soluble form, but rHisOLABL-H was hardly dissolved in a renaturing buffer. The specific activities of rHisOLABL-L, rHisOLABL-H, and native OLABL were 500, 36, and 20, respectively. These findings suggest that the combination of subunits may affect the solubility and activity of these dimeric form lectins. (c) 2005 Elsevier B.V. All rights reserved.

DOI PubMed

Sphingosine 1-phosphate analogue recognition and selectivity at S1P(4) within the endothelial differentiation gene family of receptors

Y Inagaki, TT Pham, Y Fujiwara, T Kohno, DA Osborne, Y Igarashi, G Tigyi, AL Parrill

BIOCHEMICAL JOURNAL ( PORTLAND PRESS LTD ) 389 187 - 195 2005.07 [Refereed]

Synergistic computational and experimental studies provided previously unforeseen details concerning the structural basis of S1P (sphingosine 1-phosphate) recognition by the S1P(4) G-protein-coupled receptor. Similarly to reports on the S1P(1) receptor, cationic and anionic residues in the third transmembrane domain (R3.28 and E3.29 at positions 124 and 125) form ion pairs with the phosphate and ammonium of S1P, and alanine mutations at these positions abolished specific S1P binding, S1P-induced receptor activation and cell migration. Unlike findings on the S1P(1) receptor, no cationic residue in the seventh transmembrane domain interacts with the phosphate. Additionally, two previously undiscovered interactions with the S1P polar headgroup have been identified. Trp(186) at position 4.64 in the fourth transmembrane domain interacts by a cation-pi interaction with the ammonium group of S1P. Lys(204) at position 5.38 forms an ion pair with the S1P. The S1P(4) and S1P(1) receptors show differences in binding-pocket shape and electrostatic distributions that correlate with the published structure-activity relationships. In particular, the binding pocket of mS1P(4) (mouse S1P(4)) has recognition sites for the anionic phosphate and cationic ammonium groups that are equidistant from the end of the non-polar tail. In contrast, the binding pocket of hS1P(1) (human S1P(4)) places the ammonium recognition site 2 angstrom (1 angstrom = 0.1 nm) closer to the end of the non-polar tail than the phosphate recognition site.

DOI PubMed

Down-regulation of EDG5/S1P2 during myogenic differentiation results in the specific uncoupling of sphingosine 1-phosphate signalling to phospholipase D

E Meacci, F Cencetti, C Donati, F Nuti, M Farnararo, T Kohno, Y Igarashi, P Bruni

BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR AND CELL BIOLOGY OF LIPIDS ( ELSEVIER SCIENCE BV ) 1633 ( 3 ) 133 - 142 2003.09 [Refereed]

The bioactive lipid sphingosine 1-phosphate (S1P) is known to exert powerful biological effects through the interaction with various members of the endothelial differentiation gene (EDG) receptor family, recently renamed S1P receptors. In the present study, evidence is provided that differentiation of C2C12 myoblasts into myotubes was accompanied by profound changes of EDG/S1P receptor expression. Indeed, in differentiated cells a significant increase of EDG3/S1P3 together with a large decrease of EDG5/S1P2 expression at mRNA as well as protein level was detected. Moreover, SIP was capable to initiate the signalling pathways downstream to cytosolic Ca2+ increase in myombes, similarly to that observed in myoblasts, whereas the signalling of the bioactive lipid to phospholipase D (PLD), but not that of bradykinin (BK) or lysophosphatidic acid (LPA), was found impaired in differentiated cells. Intriguingly, overexpression of EDG5/S1P2, but not EDG1/S1P1 or EDG3/S1P3, potentiated the efficacy of S1P to stimulate PLD, strongly suggesting a role for EDG5/S1P2 in the signalling to PLD. This view was also supported by the marked reduction of S1P-induced PLD activity in myoblasts loaded with antisense oligodeoxyribonucleotides (ODN) to EDG5/S1P2. Furthermore, overexpression of EDG5/S1P2 rescued the coupling of S1P signalling to PLD in C2C12 myotubes. Experimental evidence here provided supports the notion that EDG5/S1P2 plays a dominant role in the coupling of S1P to PLD in myoblasts and that the down-regulation of the receptor subtype is responsible for the specific uncoupling of S1P signalling to PLD in myotubes. (C) 2003 Elsevier B.V All rights reserved.

DOI PubMed

Synthesis of fluorescence-labeled sphingosine and sphingosine 1-phosphate; Effective tools for sphingosine and sphingosine 1-phosphate behavior

T Hakogi, T Shigenari, S Katsumura, T Sano, T Kohno, Y Igarashi

BIOORGANIC & MEDICINAL CHEMISTRY LETTERS ( PERGAMON-ELSEVIER SCIENCE LTD ) 13 ( 4 ) 661 - 664 2003.02 [Refereed]

A fluorescence-labeled sphingosine and sphingosine 1-phosphate have been successfully synthesized from the oxazolidinone methyl ester derived from glycidol via monoalkylation and the stereoselective reduction of the resulting ketone. The labeled sphingosine was converted into its phosphate by treatment with sphingosine kinase I (SPHK I) from mouse. and in platelets. and it was incorporated into the Chinese Hamster Ovarian (CHO) cells. In addition, MAPK was activated by NBD-Sph-1-P through Edg-1, Sph-1-P receptor. (C) 2003 Elsevier Science Ltd. All rights reserved.

DOI PubMed

Sphingosine 1-phosphate inhibits interleukin-1 beta induction of nitric oxide synthase via extracellular-signal regulated kinase inhibition and Rho in vascular smooth muscle cells.

Y Matsumoto, M Hirafuji, T Machida, S Izumi, A Nodai, T Kohno, Y Igarashi, M Minami

JOURNAL OF PHARMACOLOGICAL SCIENCES ( JAPANESE PHARMACOLOGICAL SOC ) 91 114P - 114P 2003 [Refereed]

Modulation of sphingosine 1-phosphate, a new lipid mediator, on nitric oxide production by vascular smooth muscle cells

Masahiko Hirafuji, Yukihiro Hamaya, Yumika Matsumoto, Takuji Machida, Masaru Minami, Takayuki Kohno, Yasuyuki Igarashi

Folia Pharmacologica Japonica 120 ( 1 ) 2002 [Refereed]

The effects of sphingosine 1-phosphate (SIP) on nitric oxide (NO) production induced by interleukin (IL)-1β in cultured rat vascular smooth muscle cells (VSMCs) have been investigated. Rat VSMCs abundantly expressed mRNA of Edg-3 and Edg-5 receptor subtypes in both unstimulated and IL-1β-stimulated cells. S1P at higher than 0.1 μM inhibited the NO production induced by IL-1β in a concentration-dependent manner, in which S1P at 10 μM caused over 90% inhibition. S1P also inhibited inducible NO synthase (iNOS) protein and mRNA expressions induced by IL-1β in a concentration-dependent manner, which effects were smaller than that in the NO production. S1P also significantly inhibited GTP cyclohydrolase I (GTPCH) mRNA expression induced by IL-1β. PTX pretreatment partially prevented the inhibitory effects of S1P on the NO production and iNOS induction, while completely prevented the inhibitory effects on iNOS and GTPCH mRNA expressions. In PTX-pretreated VSMC, S1P showed a bell-shaped enhancing and inhibitory effect on iNOS protein expression. These results suggest that SIP modulates IL-1β induction of NO production by rat VSMCs through multiple mechanisms, partially via Edg-3 and Edg-5 receptor subtypes coupled to PTX-sensitive G proteins.

PubMed

Apsaragine-linked glycosylation of Edg1, a receptor for sphingosine 1-phosphate: Significance in receptor internalization and receptor-coupled signaling.

Takayuki Kohno, Atsushi Wada, Yasuyuki Igarashi

Chem. Phys. Lipid. 107,39 2000 [Refereed]

Authorship： Lead author

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Papers - KOUNO Takayuki