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T Hla

Publications and source records attributed to T Hla.

At least 19 recordsLinked to original sources
PubMed2001

Lysophospholipids--receptor revelations.

Upon cell activation, membrane phospholipids are metabolized into potent lysophospholipid (LP) mediators, such as sphingosine 1-phosphate and lysophosphatidic acid. LPs fulfill signaling roles in organisms as diverse as yeast and humans. The recent discovery of G protein-coupled receptors for LPs in higher eukaryotes, and their involvement in regulating diverse processes such as angiogenesis, cardiac development, neuronal survival, and immunity, has stimulated growing interest in these lipid mediators. LP receptor biology has generated insights into fundamental cellular mechanisms and may provide therapeutic targets for drug development.

Animals↗
PubMed2001

Role of the sphingosine 1-phosphate receptor EDG-1 in vascular smooth muscle cell proliferation and migration.

Sphingosine 1-phosphate (S1P), a platelet-derived ligand for the EDG-1 family of G protein-coupled receptors (GPCRs), has recently emerged as a regulator of vascular development. Although S1P has potent effects on endothelial cells and vascular smooth muscle cells (VSMCs), the functions of the specific S1P receptors in the latter cell type are not known. Here we show that pup-intimal VSMCs express higher levels of EDG-1 mRNA than adult-medial VSMCs. Stable transfection of EDG-1 into adult-medial VSMCs enhanced their proliferative response to S1P, concomitant with induction of p70 S6 kinase activity and expression of cyclin D1. Pertussis toxin treatment inhibited S1P-induced p70 S6 kinase activation, cyclin D1 expression and proliferation, suggesting that EDG-1-coupling to the G(i) pathway is critical. Furthermore, blocking p70 S6 kinase phosphorylation with rapamycin inhibited cyclin D1 expression and proliferation, suggesting that activation of p70 S6 kinase is critical in EDG-1/G(i)-mediated cell proliferation. EDG-1 expression also profoundly enhanced the migratory response of adult-medial VSMCs to S1P. S1P-induced migration of adult-medial VSMCs expressing exogenous EDG-1 required G(i) activation but not p70 S6 kinase. These results suggest that enhanced expression of EDG-1 in VSMCs dramatically stimulates both the proliferative and migratory responses to S1P. Since EDG-1 is expressed in the pup-intimal phenotype of VSMCs, S1P signaling via EDG-1 may play a role in vascular diseases in which the proliferation and migration of VSMCs are dysregulated.

Animals↗
PubMed2001

Feedback control of the arachidonate cascade in rheumatoid synoviocytes by 15-deoxy-Delta(12,14)-prostaglandin J2.

Rheumatoid arthritis (RA) is a chronic polyarticular joint disease associated with massive synovial proliferation, inflammation, and angiogenesis. PPAR-gamma ligands, both 15-deoxy-Delta(12,14)-prostaglandin J2 (15d- PGJ2) and troglitazone (TRO), can inhibit the growth of RA synoviocytes in vitro, and suppress the chronic inflammation of adjuvant-induced arthritis in rats, but the potency of 15d-PGJ2 is higher than TRO. Prostaglandin (PG) E2 plays important roles in joint erosion and synovial inflammation. In the present study, 15d-PGJ2, but not TRO and other prostanoids, suppressed interleukin (IL)-1beta-induced PGE2 synthesis in rheumatoid synovial fibroblasts (RSFs) through the inhibition of cyclooxygenase (COX-2) and cytosolic phospholipase A2 (cPLA2) expression. Furthermore, the inhibition was not affected by pretreatment with anti-PPAR-gamma antibody. It means that this anti-inflammatory effect of 15d-PGJ2 for PG synthesis may be independent of PPAR-gamma and 15d-PGJ2 is a key regulator of negative feedback of the arachidonate cascade on the COX pathway. These findings provide new insight into the feedback mechanism of the arachidonate cascade.

Arachidonic Acid↗
PubMed2001

Overexpression of cyclooxygenase-2 is sufficient to induce tumorigenesis in transgenic mice.

The cyclooxygenase (COX)-2 gene encodes an inducible prostaglandin synthase enzyme that is overexpressed in adenocarcinomas and other tumors. Deletion of the murine Cox-2 gene in Min mice reduced the incidence of intestinal tumors, suggesting that it is required for tumorigenesis. However, it is not known if overexpression of Cox-2 is sufficient to induce tumorigenic transformation. We have derived transgenic mice that overexpress the human COX-2 gene in the mammary glands using the murine mammary tumor virus promoter. The human Cox-2 mRNA and protein are expressed in mammary glands of female transgenic mice and were strongly induced during pregnancy and lactation. Female virgin Cox-2 transgenic mice showed precocious lobuloalveolar differentiation and enhanced expression of the beta-casein gene, which was inhibited by the Cox inhibitor indomethacin. Mammary gland involution was delayed in Cox-2 transgenic mice with a decrease in apoptotic index of mammary epithelial cells. Multiparous but not virgin females exhibited a greatly exaggerated incidence of focal mammary gland hyperplasia, dysplasia, and transformation into metastatic tumors. Cox-2-induced tumor tissue expressed reduced levels of the proapoptotic proteins Bax and Bcl-x(L) and an increase in the anti-apoptotic protein Bcl-2, suggesting that decreased apoptosis of mammary epithelial cells contributes to tumorigenesis. These data indicate that enhanced Cox-2 expression is sufficient to induce mammary gland tumorigenesis. Therefore, inhibition of Cox-2 may represent a mechanism-based chemopreventive approach for carcinogenesis.

Animals↗
PubMed2001

Sphingosine 1-phosphate activates Akt, nitric oxide production, and chemotaxis through a Gi protein/phosphoinositide 3-kinase pathway in endothelial cells.

Sphingosine 1-phosphate (SPP) binds to members of the endothelial differentiation gene family (EDG) of receptors and leads to diverse signaling events including cell survival, growth, migration and differentiation. However, the mechanisms of how SPP activates these proangiogenic pathways are poorly understood. Here we show that SPP signals through the EDG-1 receptor to the heterotrimeric G protein G(i), leading to activation of the serine/threonine kinase Akt and phosphorylation of the Akt substrate, endothelial nitric-oxide synthase (eNOS). Inhibition of G(i) signaling, and phosphoinositide 3-kinase (PI 3-kinase) activity resulted in a decrease in SPP-induced endothelial cell chemotaxis. SPP also stimulates eNOS phosphorylation and NO release and these effects are also attenuated by inhibition of G(i) signaling, PI 3-kinase, and Akt. However, inhibition of NO production did not influence SPP-induced chemotaxis but effectively blocked the chemotactic actions of vascular endothelial growth factor. Thus, SPP signals through G(i) and PI 3-kinase leading to Akt activation and eNOS phosphorylation.

Animals↗
PubMed2001

Sphingosine 1-phosphate-induced endothelial cell migration requires the expression of EDG-1 and EDG-3 receptors and Rho-dependent activation of alpha vbeta3- and beta1-containing integrins.

Sphingosine 1-phosphate (SPP), a platelet-derived bioactive lysophospholipid, is a regulator of angiogenesis. However, molecular mechanisms involved in SPP-induced angiogenic responses are not fully defined. Here we report the molecular mechanisms involved in SPP-induced human umbilical vein endothelial cell (HUVEC) adhesion and migration. SPP-induced HUVEC migration is potently inhibited by antisense phosphothioate oligonucleotides against EDG-1 as well as EDG-3 receptors. In addition, C3 exotoxin blocked SPP-induced cell attachment, spreading and migration on fibronectin-, vitronectin- and Matrigel-coated surfaces, suggesting that endothelial differentiation gene receptor signaling via the Rho pathway is critical for SPP-induced cell migration. Indeed, SPP induced Rho activation in an adherence-independent manner, whereas Rac activation was dispensible for cell attachment and focal contact formation. Interestingly, both EDG-1 and -3 receptors were required for Rho activation. Since integrins are critical for cell adhesion, migration, and angiogenesis, we examined the effects of blocking antibodies against alpha(v)beta(3), beta(1), or beta(3) integrins. SPP induced Rho-dependent integrin clustering into focal contact sites, which was essential for cell adhesion, spreading and migration. Blockage of alpha(v)beta(3)- or beta(1)-containing integrins inhibited SPP-induced HUVEC migration. Together our results suggest that endothelial differentiation gene receptor-mediated Rho signaling is required for the activation of integrin alpha(v)beta(3) as well as beta(1)-containing integrins, leading to the formation of initial focal contacts and endothelial cell migration.

Base Sequence↗
PubMed2001

Sphingosine 1-phosphate receptors.

Sphingosine-1-phosphate (SPP) is a bioactive lipid produced from the metabolism of sphingomyelin. It is an important constituent of serum and regulates cell growth, survival, migration, differentiation and gene expression. Its mode of action has been enigmatic; however, recent findings have shown that a family of G-protein-coupled receptors (GPCR) of the endothelial differentiation gene (EDG) family serve as plasma membrane-localized receptors for SPP. Furthermore, the EDG receptors appear to be SPP receptor subtypes with distinct signaling characteristics. In vascular endothelial cells, SPP acts on EDG-1 and EDG-3 subtypes of receptors to induce cell survival and morphogenesis. Such pathways appear to be critical for SPP-induced angiogenic response in vivo. In addition, the EDG-1 gene is essential for vascular maturation in development. Moreover, developmental studies in Zebrafish have indicated that SPP signaling via the EDG-5 like receptor Miles Apart (Mil) is essential for heart development. These data strongly suggest that a physiological role of SPP is in the formation of the cardiovascular system. Despite these recent findings, much needs to be clarified with respect to the physiological role of SPP synthesis and action. This review will focus on the recent findings on SPP receptors and the effects on the cardiovascular system.

Lysophospholipids↗
PubMed2001

Akt-mediated phosphorylation of the G protein-coupled receptor EDG-1 is required for endothelial cell chemotaxis.

The role of the protein kinase Akt in cell migration is incompletely understood. Here we show that sphingosine-1-phosphate (S1P)-induced endothelial cell migration requires the Akt-mediated phosphorylation of the G protein-coupled receptor (GPCR) EDG-1. Activated Akt binds to EDG-1 and phosphorylates the third intracellular loop at the T(236) residue. Transactivation of EDG-1 by Akt is not required for G(i)-dependent signaling but is indispensable for Rac activation, cortical actin assembly, and chemotaxis. Indeed, T236AEDG-1 mutant sequestered Akt and acted as a dominant-negative GPCR to inhibit S1P-induced Rac activation, chemotaxis, and angiogenesis. Transactivation of GPCRs by Akt may constitute a specificity switch to integrate rapid G protein-dependent signals into long-term cellular phenomena such as cell migration.

Actins↗
PubMed2000

Serum withdrawal-induced post-transcriptional stabilization of cyclooxygenase-2 mRNA in MDA-MB-231 mammary carcinoma cells requires the activity of the p38 stress-activated protein kinase.

Overexpression of the cyclooxygenase-2 (COX-2) gene is observed in several neoplastic diseases. However, molecular mechanisms involved in the regulation of expression of COX-2 are not well understood. In this report, we describe a unique post-transcriptional regulatory mechanism of COX-2 mRNA stabilization in MDA-MB-231 cells, a highly metastatic cell line derived from a human mammary tumor. High levels of COX-2 mRNA, protein, and enzyme activity were induced by serum withdrawal, which were potently inhibited by the addition of serum or >100-kDa serum factor. Nuclear run-on analysis and actinomycin D chase experiments indicate that regulation is primarily at the level of post-transcriptional mRNA stability. Interestingly, SB203580, an inhibitor of the p38 stress-activated protein kinase (SAPK), and overexpression of the dominant-negative p38alpha construct potently inhibited the serum withdrawal-induced COX-2 mRNA levels. Indeed, the half-life of COX-2 mRNA decreased from 9 to 4.5 h after SB203580 treatment, suggesting that signal transduction by the p38 SAPK pathway is required for COX-2 mRNA stability.

Apoptosis↗
PubMed2000

Expression of cyclooxygenase-2 in prostate carcinoma.

BACKGROUND: Nonsteroidal antiinflammatory drugs inhibiting cyclooxygenase (COX) enzyme activity in both its constitutive (COX-1) and inducible (COX-2) isoforms were shown also to inhibit the development of colon carcinoma in animal models. COX-2 is an inducer of angiogenesis of new blood vessels. The expression of COX-1 and COX-2 in prostate tissues from patients with prostate carcinoma was investigated using reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. METHODS: Tumor specimens were obtained from 28 prostate carcinoma (PC) patients, 8 benign prostatic hyperplasia (BPH) patients, 1 prostatic intraepithelial neoplasia (PIN) patient, and 8 specimens of normal prostate tissue (NP). Affinity-purified COX-1 and COX-2 antibodies were used in immunochemistry. RESULTS: Very weak expression of COX-1 and marked expression of immunoreactive COX-2 in tumor cells was obtained. In contrast, expression of both isoforms was very weak in all cases of BPH and in the NP tissues. Immunoreactive COX-1 also was very weak in all cases of benign tissues. The extent and intensity of immunoreactive COX-2 polypeptides in tumor cells was statistically much greater than those of cells from BPH. Immunostaining with normal rabbit immunoglobulin G was completely negative. By RT-PCR analysis, enhanced expression of COX-2, but not COX-1, was observed in PC tissue. BPH displayed faint expression of COX-2. CONCLUSIONS: The results of the current study demonstrated that human prostate carcinoma cells generated COX-2, and that COX-2 might play an important role in the proliferation of prostate carcinoma cells. These findings suggest that inhibition of COX-2 development may lead not only to inhibition of the proliferation and metastasis of prostate carcinoma but also to the inhibition of prostate carcinogenesis.

Aged↗
PubMed2000

Characterization of a novel sphingosine 1-phosphate receptor, Edg-8.

Three G protein-coupled receptors (Edg-1, Edg-3, and Edg-5) for the lysolipid phosphoric acid mediator sphingosine 1-phosphate have been described by molecular cloning. Using a similar sequence that we found in the expressed sequence tag data base, we cloned and characterized of a fourth, high affinity, rat brain sphingosine 1-phosphate receptor, Edg-8. When HEK293T cells were co-transfected with Edg-8 and G protein DNAs, prepared membranes showed sphingosine 1- phosphate-dependent increases in [(35)S]guanosine 5'-(3-O-thio)triphosphate binding with an EC(50) of 90 nm. In a rat hepatoma Rh7777 cell line that exhibits modest endogenous responses to sphingosine 1-phosphate, this lipid mediator inhibited forskolin-driven rises in cAMP by greater than 90% when the cells were transfected with Edg-8 DNA (IC(50) 0.7 nm). This response is blocked fully by prior treatment of cultures with pertussis toxin, thus implicating signaling through G(i/o)alpha proteins. Furthermore, Xenopus oocytes exhibit a calcium response to sphingosine 1-phosphate after injection of Edg-8 mRNA, but only when oocytes are co-injected with chimeric G(q/i)alpha protein mRNA. Membranes from HEK293T and Rh7777 cell cultures expressing Edg-8 exhibited high affinity (K(D) = 2 nm) binding for radiolabeled sphingosine 1-phosphate. Rat Edg-8 RNA is expressed in spleen and throughout adult rat brain where in situ hybridization revealed it to be associated with white matter. Together our data demonstrate that Edg-8 is a high affinity sphingosine 1-phosphate receptor that couples to G(i/o)alpha proteins and is expressed predominantly by oligodendrocytes and/or fibrous astrocytes in the rat brain.

Amino Acid Sequence↗
PubMed2000

Inhibition of human lung cancer cell growth by the peroxisome proliferator-activated receptor-gamma agonists through induction of apoptosis.

Peroxisome proliferator-activated receptors (PPARs), members of the nuclear hormone receptors superfamily, have an important regulatory role in adipogenesis and inflammation. PPAR-gamma ligands induce terminal differentiation and growth inhibition of human breast cancer cells and prostatic cancer cells. In this study, we demonstrated that PPAR-gamma, but not PPAR-alpha, was expressed in human lung cancer cell lines by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. We also found that the synthetic PPAR-gamma agonist thiazolidinedione compounds (troglitazone) and the endogenous PPAR-gamma ligand, 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)), inhibited the growth of human lung cancer cells through the induction of apoptosis. However, PPAR-alpha agonist (bezafibrate) and other prostanoids (PGE(2), PGF(2alpha)) did not induce apoptosis. These findings suggest that PPAR-gamma may play an important role in the pathogenesis of lung cancer and that PPAR-gamma agonist may be useful therapeutic agents in the treatment of human lung cancer.

Apoptosis↗
PubMed2000

Selective inhibition of cyclooxygenase-2 with antisense oligodeoxynucleotide restricts induction of rat adjuvant-induced arthritis.

The effects of cyclooxygenase (COX)-2 antisense oligodeoxynucleotide (ODN) in induction of adjuvant-induced arthritis were investigated. Female Lewis rats were injected with Mycobacterium butyricum intradermally at the base of tails to induce arthritis. Synthetic 18 mer phosphorothioate ODNs corresponding to the translation initiation site of rat COX-2 mRNA were prepared. The antisense (AS), sense (S), and "scrambled" (Sc) ODNs were intraperitoneally administered. Arthropathy was evaluated with arthritis score, paw edema, and histological examination. Expression of COX-1 and -2 protein and mRNA were examined with immunostaining and reverse-transcription polymerase chain reaction, respectively. COX-2 AS ODN significantly suppressed induction of arthritis in a dose-dependent manner without severe adverse effects, whereas S and Sc ODNs did not show significant inhibitory effects. COX-2 mRNA and protein expression were also suppressed only by COX-2 AS ODN without any alteration of COX-1 expression. These data suggest that selective inhibition of COX-2 with AS ODN may have a therapeutic potency in the treatment of rheumatoid arthritis.

Animals↗
PubMed2000

The nuclear receptor PPAR gamma and immunoregulation: PPAR gamma mediates inhibition of helper T cell responses.

The peroxisome proliferator-activated receptors (PPARs) are a family of transcription factors belonging to the nuclear receptor superfamily. Until recently, the genes regulated by PPARs were those believed to be predominantly associated with lipid metabolism. Recently, an immunomodulatory role for PPAR gamma has been described in cells critical to the innate immune system, the monocyte/macrophage. In addition, evidence for an antiinflammatory role of the PPAR gamma ligand, 15-deoxy-Delta 12,14-PGJ2 (15d-PGJ2) has been found. In the present studies, we demonstrate, for the first time, that murine helper T cell clones and freshly isolated splenocytes express PPAR gamma 1. The PPAR gamma expressed is of functional significance in that two ligands for PPAR gamma, 15d-PGJ2 and a thiazolidinedione, ciglitazone, mediate significant inhibition of proliferative responses of both the T cell clones and the freshly isolated splenocytes. This inhibition is mediated directly at the level of the T cell and not at the level of the macrophage/APC. Finally, we demonstrate that the two ligands for PPAR gamma mediate inhibition of IL-2 secretion by the T cell clones while not inhibiting IL-2-induced proliferation of such clones. The demonstration of the expression and function of PPAR gamma in T cells reveals a new level of immunoregulatory control for PPARs and significantly increases the role and importance of PPAR gamma in immunoregulation.

Adjuvants, Immunologic↗
PubMed2000

Bisphenol A diglycidyl ether (BADGE) is a PPARgamma agonist in an ECV304 cell line.

Peroxisome proliferator activated receptors (PPAR)s are nuclear transcription factors of the steroid receptor super-family. One member, PPARgamma, a critical transcription factor in adipogenesis, is expressed in ECV304 cells, and when activated participates in the induction of cell death by apoptosis. Here we describe a clone of ECV304 cells, ECV-ACO.Luc, which stably expresses a reporter gene for PPAR activation. ECV-ACO.Luc respond to the PPARgamma agonists, 15-deoxy-Delta(12,14) PGJ(2), and ciglitizone, by inducing luciferase expression. Furthermore, using ECV-ACO.Luc, we demonstrate that a newly described PPARgamma antagonist, bisphenol A diglycidyl ether (BADGE) has agonist activities. Similar to 15-deoxy-Delta(12,14) PGJ(2), BADGE induces PPARgamma activation, nuclear localization of the receptor, and induces cell death.

Benzhydryl Compounds↗
PubMed2000

Sphingosine-1-phosphate signaling via the EDG-1 family of G-protein-coupled receptors.

The bioactive lipid sphingosine-1-phosphate (SPP) is abundantly formed and released during the activation of platelets by thrombotic stimuli. Once exported, SPP interacts with the G-protein-coupled receptors (GPCR) of the EDG-1 family. SPP binds to EDG-1 with the dissociation constant of approximately 8 nM and induces signal transduction events such as mitogen-activated protein kinase (MAP kinase) activation, decrease of cAMP levels, remodeling of the actin cytoskeleton, among others. EDG-1 is a prototypical member of a large family of GPCRs that interact with glycero- and sphingolysolipid phosphates, namely, SPP and lysophosphatidic acid (LPA). Three other GPCRs, trivially termed EDG-3, EDG-5, and EDG-8, are also high-affinity receptors for SPP. The four SPP receptor subtypes regulate different intracellular signal transduction pathways. In vascular endothelial cells, cooperative signaling between EDG-1 and EDG-3 subtypes of SPP receptors results in adherens junction assembly, cell survival, morphogenesis into capillary-like networks, and angiogenesis. SPP acts distinctly, albeit cooperatively, with polypeptide angiogenic factors, resulting in the formation of mature neovessels. Thus SPP signaling as an extracellular mediator via the EDG-1 family of GPCRs may be a heretofore unrecognized mechanism for the regulation of angiogenesis and vascular endothelial cell function.

Evolution, Molecular↗