Biomedical subjects

D Ghigo

Publications and source records attributed to D Ghigo.

At least 19 recordsLinked to original sources

PubMed2003

Insulin stimulates glucose transport via nitric oxide/cyclic GMP pathway in human vascular smooth muscle cells.

OBJECTIVE: In cultured human vascular smooth muscle cells, insulin increases cyclic GMP production by inducing nitric oxide (NO) synthesis. The aim of the present study was to determine whether in these cells the insulin-stimulated NO/cyclic GMP pathway plays a role in the regulation of glucose uptake. METHODS AND RESULTS: Glucose transport in human vascular smooth muscle cells was measured as uptake of 2-deoxy-d-[3H]glucose, cyclic GMP synthesis was checked by radioimmunoassay, and GLUT4 recruitment into the plasma membrane was determined by immunofluorescence. Insulin-stimulated glucose transport and GLUT4 recruitment were blocked by an inhibitor of NO synthesis and mimicked by NO-releasing drugs. Insulin- and NO-elicited glucose uptake were blocked by inhibitors of soluble guanylate cyclase and cyclic GMP-dependent protein kinase; furthermore, glucose transport was stimulated by an analog of cyclic GMP. CONCLUSIONS: Our results suggest that insulin-elicited glucose transport (and the corresponding GLUT4 recruitment into the plasma membrane) in human vascular smooth muscle cells is mediated by an increased synthesis of NO, which stimulates the production of cyclic GMP and the subsequent activation of a cyclic GMP-dependent protein kinase.

Biological Transport↗

PubMed2001

Iron inhibits the nitric oxide synthesis elicited by asbestos in murine macrophages.

Crocidolite fibers stimulated nitric oxide synthase (NOS) activity and expression in glial and alveolar murine macrophages: this effect was inhibited by iron supplementation and enhanced by iron chelation. We suggest that in these cells crocidolite stimulates NOS expression by decreasing the iron bioavailability and activating an iron-sensitive transcription factor.

Animals↗

PubMed2001

The concentration of nitrite in seminal plasma does not correlate with sperm concentration, sperm motility, leukocytospermia, or sperm culture.

OBJECTIVE: To correlate the concentration of nitrite (the stable metabolite of nitric oxide) in seminal plasma with sperm number and motility, leukocytospermia, and sperm culture. DESIGN: Prospective study. SETTING: Academic research institution. PATIENT(S): Seventy normozoospermic or dyspermic men enrolled in an artificial insemination/in vitro fertilization program. INTERVENTION(S): Semen samples (n = 70) were checked for sperm concentration, total sperm count, sperm motility, seminal leukocyte concentration, and sperm culture; similarly, the concentration of nitrite in seminal plasma was measured by Griess reaction. MAIN OUTCOME MEASURE(S): Measurement of nitrite concentration in seminal plasma and its correlation with sperm concentration, total sperm count, sperm motility, leukocytospermia, and sperm culture. RESULT(S): The concentration of nitrite in seminal plasma does not correlate with sperm concentration, total sperm count, or with the proportion of immotile or rapid-forward motile spermatozoa. Moreover, the concentration of nitrite in seminal plasma is not significantly increased when sperm culture is positive, nor does it correlate with leukocyte concentration in semen. CONCLUSION(S): Our results do not support the hypothesis that in vivo nitric oxide synthesis affects sperm function; alternatively, our results could suggest that nitrite in the seminal plasma is not a sensitive marker of in vivo nitric oxide synthesis.

Fertilization in Vitro↗

PubMed2001

Signaling pathway of nitric oxide-induced acrosome reaction in human spermatozoa.

Nitric oxide (NO) has been recently shown to modulate in vitro motility, viability, the acrosome reaction (AR), and metabolism of spermatozoa in various mammalian species, but the mechanism or mechanisms through which it influences sperm functions has not been clarified. In human capacitated spermatozoa, both the intracellular cGMP level and the percentage of AR-positive cells were significantly increased after 4 h of incubation with the NO donor, sodium nitroprusside (SNP). SNP-induced AR was significantly reduced in the presence of the soluble guanylate cyclase (sGC) inhibitors, LY83583 and ODQ; this block was bypassed by adding 8-bromo-cGMP, a cell-permeating cGMP analogue, to the incubation medium. Finally, Rp-8-Br-cGMPS and Rp-8-pCPT-cGMPS, two inhibitors of the cGMP-dependent protein kinases (PKGs), inhibited the SNP-induced AR. Furthermore, SNP-induced AR did not occur in Ca2+ -free medium or in the presence of the protein kinase C (PKC) inhibitor, calphostin C. This study suggests that the AR-inducing effect of exogenous NO on capacitated human spermatozoa is accomplished via stimulation of an NO-sensitive sGC, cGMP synthesis, and PKG activation. In this effect the activation of PKC is also involved, and the presence of extracellular Ca2+ is required.

Acrosome Reaction↗

PubMed2000

In vitro and in vivo vasodilating activity of nitroso derivatives gem -substituted with electron-withdrawing groups.

A series of nitroso compounds gem -substituted with electron-withdrawing groups (R(2)C(X)NO, R=alkyl, X=NO(2), CN, Cl), were studied for their in vitro and in vivo vasodilating properties as well as for their ability to activate soluble guanylate cyclase (sGC) in RFL-6 cells. All the compounds, with the sole exception of chloro derivative, display good in vitro vasodilating action and are able to increase the basal level of cGMP. Their potencies as vasodilators decrease in the presence of oxyhaemoglobin, a scavenger of nitric oxide (NO). The haemodynamic profile of the most interesting compounds, assessed in anaesthetized pigs, is also in line with a release of NO from these compounds.

Action Potentials↗

PubMed1999

Follicular fluid proteins stimulate nitric oxide (NO) synthesis in human sperm: a possible role for NO in acrosomal reaction.

Nitric oxide (NO) is a free radical involved in the regulation of several functions of the male genitourinary system. It is produced by neurons and the endothelium and epithelia of reproductive system; it mediates penile erection and regulates sperm motility, viability, and metabolism. Here we show that human spermatozoa exhibit a detectable NO synthase (NOS) activity, measured both as ability of the intact sperm and cell lysate to convert L-[3H]arginine into L-[3H]citrulline and as 24 h accumulation of extracellular nitrite in intact sperm suspensions. NOS activity (identified as an endothelial isoform) was inhibited by L-canavanine and N(G)-monomethyl-L-arginine, and nitrite accumulation was inhibited by the NO scavenger hemoglobin; both enzyme activity and nitrite production were increased by a 24 h incubation of spermatozoa with protein-enriched extracts of human follicular fluid (PFF); a significant increase of citrulline synthesis was observed only after a 4 h incubation with 40% PFF, a time period during which acrosomal reactivity was significantly increased. PFF-induced acrosomal reaction was inhibited by L-canavanine and hemoglobin, and the NO donors sodium nitroprusside (SNP), S-nitroso-N-acetyl-penicillamine (SNAP), and DETA NONOate were able to increase the percentage of reacted spermatozoa. Our results suggest that NO synthesized by human sperm may play a role in follicular fluid-induced acrosomal reaction.

Acrosome Reaction↗

PubMed1999

Human vascular smooth muscle cells express a constitutive nitric oxide synthase that insulin rapidly activates, thus increasing guanosine 3':5'-cyclic monophosphate and adenosine 3':5'-cyclic monophosphate concentrations.

AIMS/HYPOTHESIS: Insulin incubation of human vascular smooth muscle cells (hVSMC) for 120 min increases both guanosine 3':5'-cyclic monophosphate (cGMP) and adenosine 3':5'-cyclic monophosphate (cAMP) and these effects are blocked by inhibiting nitric oxide synthase (NOS). These data suggest that insulin activates a constitutive Ca2+-dependent NOS (cNOS), not described at yet in hVSMC. To test this hypothesis, we evaluated in hVSMC: i) the kinetics of the insulin-induced enhancement of the two cyclic nucleotides; ii) the ability of nitric oxide (NO) to increase both cyclic nucleotides; iii) NO involvement in the short-term influence of insulin on both cyclic nucleotides; iv) the ability of insulin to increase NO production in a few minutes; v) the presence of a cNOS activity; vi) the expression of mRNA for cNOS. METHODS: In hVSMC incubated with insulin, NO donors and the Ca2+ ionophore ionomycin, we measured cAMP and cGMP (RIA); in hVSMC incubated with insulin and ionomycin we measured NO, evaluated as L-(3H)-citrulline production from L-(3H)-arginine; by northern blot hybridization, we measured the expression of cNOS mRNA. RESULTS: i) By incubating hVSMC with 2 nmol/l insulin for 0-240 min, we observed an increase of both cGMP and cAMP (ANOVA: p = 0.0001). Cyclic GMP rose from 0.74 +/- 0.01 to 2.62 +/- 0.10 pmol/10(6) cells at 30 min (p = 0.0001); cAMP rose from 0.9 +/- 0.09 to 11.65 +/- 0.74 pmol/10(6) cells at 15 min (p=0.0001). ii) Sodium nitroprusside (100 mol/l) and glyceryltrinitrate (100 micromol/l) increased both cGMP and cAMP (p = 0.0001). iii) The effects of insulin on cyclic nucleotides were blocked by NOS inhibition. iv) An increase of NO was observed by incubating hVSMC for 5 min with 2 nmol/l insulin (p = 0.0001). v) Ionomycin (1 micromol/l) enhanced NO production (p = 0.0001) and increased both cyclic nucleotides (p = 0.0001). vi) hVSMC expressed mRNA of cNOS. CONCLUSION/INTERPRETATION: Human VSMC express cNOS, which is rapidly activated by insulin with a consequent increase of both cGMP and cAMP, suggesting that insulin-induced vasodilation in vivo is not entirely endothelium-mediated.

Calcium↗

PubMed1999

NO donor and biological properties of different benzofuroxans.

PURPOSE: To investigate the effect of benzofusion on NO donor properties and related biological activities of the furoxan system. The biological properties considered were the ability to increase the cytosolic levels of cGMP in C6 cells and vasodilation. METHODS: NO donor properties were investigated either in the presence or the absence of cysteine by using the Griess reaction, chemiluminescence, and gas chromatography. Increase of cytosolic cGMP levels were evaluated by radioimmunoassay. Vasodilating activity was assessed on rat aorta strips precontracted with noradrenaline, in the presence and the absence of oxyhemoglobin (HbO2) and methylene blue (MB), respectively. RESULTS: Benzofuroxan and its methyl and cyano derivatives were unable to release NO under the experimental conditions. Generally these compounds displayed feeble vasodilating properties and were able to weakly stimulate soluble guanylate cyclase (sGC). By contrast, benzodifuroxan and benzotrifuroxan were able to produce both NO* and its reduced form NO- , the nitroxyl anion. They displayed potent vasodilating properties and were able to increase cytosolic levels of cGMP in a concentration-dependent manner. CONCLUSIONS: The simple benzofuroxans considered here are devoid of the capability to release NO, they weakly stimulate sGC as well as manifest feeble vasodilating properties by a mechanism that does not involve a thiol-induced NO production. By contrast, benzodifuroxan and benzotrifuroxan behave as typical NO donor furoxans.

Animals↗

PubMed1998

Chloroquine stimulates nitric oxide synthesis in murine, porcine, and human endothelial cells.

Nitric oxide (NO) is a free radical involved in the regulation of many cell functions and in the expression of several diseases. We have found that the antimalarial and antiinflammatory drug, chloroquine, is able to stimulate NO synthase (NOS) activity in murine, porcine, and human endothelial cells in vitro: the increase of enzyme activity is dependent on a de novo synthesis of some regulatory protein, as it is inhibited by cycloheximide but is not accompanied by an increased expression of inducible or constitutive NOS isoforms. Increased NO synthesis is, at least partly, responsible for chloroquine-induced inhibition of cell proliferation: indeed, NOS inhibitors revert the drug-evoked blockage of mitogenesis and ornithine decarboxylase activity in murine and porcine endothelial cells. The NOS-activating effect of chloroquine is dependent on its weak base properties, as it is exerted also by ammonium chloride, another lysosomotropic agent. Both compounds activate NOS by limiting the availability of iron: their stimulating effects on NO synthesis and inhibiting action on cell proliferation are reverted by iron supplementation with ferric nitrilotriacetate, and are mimicked by incubation with desferrioxamine. Our results suggest that NO synthesis can be stimulated in endothelial cells by chloroquine via an impairment of iron metabolism.

Aconitate Hydratase↗

PubMed1998

Modulation of guinea-pig cardiac L-type calcium current by nitric oxide synthase inhibitors.

1. Electrophysiological (whole-cell clamp) techniques were used to study the effect of NO synthase (NOS) inhibitors on guinea-pig ventricular calcium current (ICa), and biochemical measurements (Western blot and citrulline synthesis) were made to investigate the possible mechanisms of action. 2. The two NOS inhibitors, NG-monomethyl-L-arginine (L-NMMA, 1 mM) and NG-nitro-L-arginine (L-NNA, 1 mM), induced a rapid increase in ICa when applied to the external solution. D-NMMA (1 mM), the stereoisomer of L-NMMA, which has no effect on NOS, did not enhance ICa. 3. Western blot experiments gave no indication of the presence of inducible NOS protein (iNOS) in our cell preparation, neither immediately after dissociation nor after more than 24 h. Statistically, there was no significant difference between electrophysiological experiments performed on freshly dissociated cells and experiments performed the next day. Moreover cells prepared and kept in the presence of dexamethasone (3 microM), to inhibit the expression of iNOS, gave the same response to L-NMMA as control cells. 4. The stimulatory effect of L-NMMA (1 mM) on basal ICa was reversed by competition with higher doses (5 mM) of externally applied L-arginine, the natural substrate of NOS. The effect of L-NMMA was also eliminated by L-arginine in the patch pipette solution. 5. Intracellular perfusion with GDP beta S (0.5 mM), which stabilizes the G-proteins in the inactive state, did not affect the L-NMMA-induced stimulation of ICa. 6. Carbachol (1 microM) reduced the ICa previously stimulated by L-NMMA, and intracellular cGMP (10 microM) prevented L-NMMA enhancement. 7. Simultaneous treatment with L-NMMA and isoprenaline (1 microM) induced a non-cumulative enhancement of ICa that could not be reversed by carbachol (1 microM). 8. NO synthesis, measured by the formation of [3H]citrulline from L-[3H]arginine during a 15 min incubation, showed a relatively high basal NO production, which was inhibited by L-NMMA but not affected by carbachol. 9. These results suggest that inhibitors of NOS are able to modulate the basal ventricular ICa in the absence of a receptor-mediated pathway, and that NO might be required for the muscarinic reduction of ICa under isoprenaline stimulation, even if NO production is not directly controlled by the muscarinic pathway.

Animals↗

PubMed1998

Retinoic acid-induced differentiation in a human neuroblastoma cell line is associated with an increase in nitric oxide synthesis.

The human neuroblastoma cell line SK-N-BE, after incubation with 10 microM retinoic acid (RA) or 20 nM phorbol 12-myristate 13-acetate (PMA), underwent biochemical and morphological signs of differentiation within 10-14 days. In parallel, SK-N-BE cells produced significantly higher amounts of nitric oxide (NO) in comparison with controls, as assessed by the measurement of nitrite and nitrate in the culture supernatant and of NO synthase (NOS) activity in the cell lysates (measured as ability to convert [3H]arginine into [3H]citrulline and as NADPH diaphorase activity). Nitrite/nitrate production was abolished by adding the NO scavenger hemoglobin in the culture medium and was inhibited by aminoguanidine (AG, a selective inhibitor of the inducible NOS isoform) but not by the less selective inhibitor NG-nitro-L-arginine methylester (NAME). Western blotting experiments with monoclonal antibodies against the ncNOS and iNOS isoforms suggest that RA-elicited NOS activation is not attributable to an increased expression of the protein. NAME and AG were not able to revert inhibition of proliferation induced by RA, and the NO donor sodium nitroprusside did not mimic the effect of RA and PMA. These data indicate that increased NO synthesis does not mediate RA- or PMA-induced differentiation but may be an additional marker of differentiation into sympathetic-like neuronal cells.

Cell Differentiation↗

PubMed1997

Insulin stimulates nitric oxide synthesis in human platelets and, through nitric oxide, increases platelet concentrations of both guanosine-3', 5'-cyclic monophosphate and adenosine-3', 5'-cyclic monophosphate.

The insulin-induced platelet anti-aggregating effect is attributed to a nitric oxide (NO)-mediated increase of cyclic guanosine monophosphate (cGMP). The aim of this work, carried out in human platelets, is to show whether insulin increases NO synthesis in platelets and whether it enhances not only cGMP but also cyclic adenosine monophosphate (cAMP) in these cells. We observed that 1) insulin dose-dependently increases NO production, evaluated as citrulline synthesis from L-arginine (n = 4, P = 0.015); 2) insulin dose-dependently increases not only cGMP but also cAMP: for instance, after 8 min of insulin incubation at 1,920 pmol/l, cAMP increased from 39.8 +/- 1.4 to 121.3 +/- 12.6 pmol/10(9) platelets (n = 16, P = 0.0001); 3) when insulin is incubated for 120 min, the increase of cGMP and cAMP shows a plateau between 2 and 20 min, and while the effect on cGMP is significant until 120 min, the effect on cAMP is no more significant at 60 and 120 min; 4) insulin increases the effects on cAMP of the adenylate cyclase agonists Iloprost and forskolin (n = 5, P = 0.0001) and enhances their platelet anti-aggregating effects (n = 6 and 8, respectively; P = 0.0001); and 5) the inhibition of NO synthase by N(G)-monomethyl-L-arginine blunts both the insulin effects on basal cGMP and cAMP (n = 4) and those on the Iloprost- and forskolin-induced cAMP increase (n = 5). Thus, insulin increases NO synthesis in human platelets, and, through NO, enhances both cGMP and cAMP. The platelet anti-aggregating effect exerted by insulin is, therefore, a NO-mediated phenomenon involving both cGMP and cAMP.

Adult↗

PubMed1996

Reversal of doxorubicin resistance by the amiloride analogue EIPA in multidrug resistant human colon carcinoma cells.

Although multidrug resistance (mdr) may arise through a variety of mechanisms, the most widely studied and accepted form is associated with an increased concentration of P-glycoprotein (P-gp), a 170 kd protein found in the membrane fraction of a number of mammalian cells. Since mdr seems to be related to the ability of resistant cells to extrude drugs and the circumvention of mdr is supposed to be due to the restored ability to accumulate drugs, membrane has been regarded as the crucial site for such a regulation and an important role for membrane ion exchangers has been suggested. The aim of this work was to elucidate whether the Na+/H+ antiporter is involved in the mechanism of regulation and circumvention of mdr and if 5-(N-ethyl-N-isopropyl) amiloride (EIPA), a selective inhibitor of the Na+/H+ exchanger, can modulate the functional expression of the mdr phenotype. The effect of EIPA on doxorubicin (DX) resistant cells (LoVo/DX) obtained from a human colon adenocarcinoma cell line (LoVo) was studied. EIPA at concentrations ranging from 10 to 50 mu M was able to increase the antibiotic cytotoxicity in the resistant Lovo/DX cells. The reversal of DX resistance paralleled an increase of the ability of the cells to accumulate the drug. Both drug loading and sensitivity to the inhibitory effect of DX on cell proliferation were restored by EIPA in a dose-dependent way. These results suggest a new mechanism of mdr reversal and indicate that amiloride and its derivatives may be useful in reversing DX resistance and in enhancing the clinical effectiveness of chemotherapeutics.

Adenocarcinoma↗

PubMed1995

Erythrocyte stages of Plasmodium falciparum exhibit a high nitric oxide synthase (NOS) activity and release an NOS-inducing soluble factor.

Nitric oxide (NO), a highly diffusible cellular mediator involved in a wide range of biological effects, has been indicated as one of the cytotoxic agents released by leukocytes to counteract malaria infection. On the other hand, NO has been implicated as a mediator of the neuropathological symptoms of cerebral malaria. In such circumstances NO production has been thought to be induced in host tissues by host-derived cytokines. Here we provide evidence for the first time that human red blood cells infected by Plasmodium falciparum (IRBC) synthesize NO. The synthesis of NO (measured as citrulline and nitrate production) appeared to be very high in comparison with human endothelial cells; no citrulline and nitrate production was detectable in noninfected red blood cells. The NO synthase (NOS) activity was very high in the lysate of IRBC (while not measurable in that of normal red blood cells) and was inhibited in a dose-dependent way by three different NOS inhibitors (L-canavanine, NG-amino-L-arginine, and NG-nitro-L-arginine). NOS activity in P. falciparum IRBC is Ca++ independent, and the enzyme shows an apparent molecular mass < 100 kD, suggesting that the parasite expresses an isoform different from those found in mammalian cells. IRBC release a soluble factor able to induce NOS in human endothelial cells. Such NOS-inducing activity is not tissue specific, is time and dose dependent, requires de novo protein synthesis, and is probably associated with a thermolabile protein having a molecular mass > 100 kD. Our data suggest that an increased NO synthesis in P. falciparum malaria can be directly elicited by soluble factor(s) by the blood stages of the parasite, without necessarily requiring the intervention of host cytokines.

Amino Acid Oxidoreductases↗

PubMed1995

Middle T antigen-transformed endothelial cells exhibit an increased activity of nitric oxide synthase.

Endothelioma cell lines transformed by polyoma virus middle T antigen (mTa) cause cavernous hemangiomas in syngeneic mice by recruitment of host cells. The production of nitric oxide (NO), as measured by nitrite and citrulline production, was significantly higher in mTa-transformed endothelial cells in comparison with nontransformed control cells. The maximal activity of NO synthase (NOS) was about 200-fold higher in cell lysates from the tEnd.1 endothelioma cell line than in lysates from nontransformed controls, whereas the affinity for arginine did not differ. The biochemical characterization of NOS and the study of mRNA transcripts indicate that tEnd.1 cells express both the inducible and the constitutive isoforms. NOS hyperactivity is not a simple consequence of cell transformation but needs a tissue-specific mTa expression. Since tEnd.1-conditioned medium induces NOS activity in normal endothelial cells, most likely NOS hyperactivity in endothelioma cells is attributable to the release of a soluble factor. This NOS-activating factor, which seems to be an anionic protein, could stimulate tEnd.1 cells to express NOS by an autocrine way. By the same mechanism, tEnd.1 cells could induce NOS in the neighboring endothelial cells, and NO release could play a role in the hemangioma development. Such hypothesis is confirmed by our in vivo experiments, showing that the administration of the NOS inhibitor L-canavanine to endothelioma-bearing mice significantly reduced both the volume and the relapse time of the tumor.

Amino Acid Oxidoreductases↗

PubMed1995

Activation of endothelial guanylate cyclase inhibits cellular reactivity.

The study shows that endothelial cells from human umbilical veins have a soluble guanylate cyclase which can be activated by sodium nitroprusside (SNP), SIN-1 (3-morpholinosydnonimine) and S35b (4-methyl-3-phenylsulfonylfuroxan). Cells which were pretreated with these compounds showed an inhibition of thrombin-induced arachidonic acid release, PGI2 formation, PAF synthesis and PMNL adhesion. Endothelial guanylate cyclase can also be activated by nitric oxide (NO) which is generated in endothelial cells upon stimulation with thrombin or ionomycin. It is suggested that endogenously produced NO might control cell activation and endothelial function through a cGMP-dependent mechanism.

Arachidonic Acid↗

PubMed1995

A possible role for nitric oxide in modulating the functional cyclosporine toxicity by arginine.

The renal damage consequent to cyclosporine A (CsA) administration ranges from hemodynamic alterations to irreversible chronic lesions. The initial vasoconstriction depends upon the imbalance between the various modulators of the renal vascular tone, among which the most powerful are endothelins and nitric oxide (NO). CsA could play a crucial role by inhibiting the Ca++/calmodulin-mediated activation of the constitutive NO synthase (NOS) isoform, which converts L-arginine (L-Arg) into NO and citrulline, with a 1:1 stoichiometry. To investigate the possibility of modulating CsA nephrotoxicity with L-Arg we studied six groups (G) of Lewis rats treated with daily gavage up to eight weeks: G1, CsA 40 mg/kg; G2, G1 plus L-Arg 300 mg/kg; G3, G2 plus the competitive inhibitor of NOS, NG-nitro-L-Arg (L-NNA); G4, L-Arg alone; G5, L-NNA alone; and G6, controls receiving vehicle alone. After eight weeks L-Arg treated rats were protected against the toxic effects of CsA [creatinine (Cr) values, G2, 0.62 +/- 0.05 mg/dl vs. G1, 0.99 +/- 0.16 mg/dl, P < 0.001; proteinuria (P), G2, 7.2 +/- 1.02 mg/day vs. G1, 15.1 +/- 1.9 mg/day, P < 0.01]. The administration of L-NNA abolished the protective effect of L-Arg (G3, Cr 1.23 +/- 0.16 mg/dl; P 16.9 = 2.3; P < 0.02 and P < 0.005, respectively vs. G2). The levels of Cr in G2 rats were superimposable to control groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Oxidoreductases↗

PubMed1995

Platelet activating factor produced in vitro by Kaposi's sarcoma cells induces and sustains in vivo angiogenesis.

Imbalance in the network of soluble mediators may play a pivotal role in the pathogenesis of Kaposi's sarcoma (KS). In this study, we demonstrated that KS cells grown in vitro produced and in part released platelet activating factor (PAF), a powerful lipid mediator of inflammation and cell-to-cell communication. IL-1, TNF, and thrombin enhanced the synthesis of PAF. PAF receptor mRNA and specific, high affinity binding site for PAF were present in KS cells. Nanomolar concentration of PAF stimulated the chemotaxis and chemokinesis of KS cells, endothelial cells, and vascular smooth muscle cells. The migration response to PAF was inhibited by WEB 2170, a hetrazepinoic PAF receptor antagonist. Because neoangiogenesis is essential for the growth and progression of KS and since PAF can activate vascular endothelial cells, we examined the potential role of PAF as an instrumental mediator of angiogenesis associated with KS. Conditioned medium (CM) from KS cells (KS-CM) or KS cells themselves induced angiogenesis and macrophage recruitment in a murine model in which Matrigel was injected subcutaneously. These effects were inhibited by treating mice with WEB 2170. Synthetic PAF or natural PAF extracted from plasma of patients with classical KS also induced angiogenesis, which in turn was inhibited by WEB 2170. The action of PAF was amplified by expression of other angiogenic factors and chemokines: these included basic and acidic fibroblast growth factor, placental growth factor, vascular endothelial growth factor and its specific receptor flk-1, hepatocyte growth factor, KC, and macrophage inflammatory protein-2. Treatment with WEB 2170 abolished the expression of the transcripts of these molecules within Matrigel containing KS-CM. These results indicate that PAF may cooperate with other angiogenic molecules and chemokines in inducing vascular development in KS.

Aged↗