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Biomedical subjects

R H Fertel

Publications and source records attributed to R H Fertel.

At least 19 recordsLinked to original sources
PubMed1992

Tumor necrosis factor-alpha-dependent production of reactive nitrogen intermediates mediates IFN-gamma plus IL-2-induced murine macrophage tumoricidal activity.

We have previously established that IFN-gamma plus IL-2 induces murine macrophage tumoricidal activity. The purpose of this study was to identify the effector molecules that account for the IFN-gamma plus IL-2-induced macrophage cytotoxicity against P815 mastocytoma cells. ANA-1 macrophages and normal thioglycollate-elicited mouse peritoneal macrophages produced little or no detectable nitrite (NO2-) after incubation with IFN-gamma alone or IL-2 alone; however, IL-2 synergized with IFN-gamma for the production of NO2-. IFN-gamma plus IL-2 did not induce NO2- production or tumoricidal activity in ANA-1 macrophages that were cultured in medium devoid of L-arginine or in ANA-1 macrophages that were incubated with NG-monomethyl-L-arginine. As observed previously with ANA-1 macrophage tumoricidal activity, IL-4 inhibited IFN-gamma plus IL-2-induced, but not IFN-gamma plus LPS-induced, NO2- production. IL-4 also selectively decreased the ability of IFN-gamma and/or IL-2 to augment TNF-alpha mRNA expression in ANA-1 macrophages. Lastly, incubation of ANA-1 macrophages with anti-TNF mAb selectively inhibited the ability of IFN-gamma plus IL-2 to induce NO2- production and tumoricidal activity. These results indicate that IFN-gamma plus IL-2-induced tumoricidal activity is dependent upon the metabolism of L-arginine to reactive nitrogen intermediates, and they establish a role for TNF-alpha as a required intermediate for IL-2-dependent NO2- production and tumoricidal activity.

Animals↗
PubMed1992

Effects of brain-gut related peptides on cAMP levels in myenteric ganglia of guinea-pig small intestine.

This study was designed to test the hypothesis that stimulation of adenylate cyclase and elevation of cAMP is involved in the signal transduction process for substance P, calcitonin gene-related peptide, vasoactive intestinal peptide, cholecystokinin or gastrin releasing peptide in myenteric ganglia. Enzymatically dissociated ganglia from the myenteric plexus of the guinea-pig small intestine were used to study changes in levels of cAMP in response to application of the brain-gut peptides in the presence and absence of forskolin. Application of substance P and calcitonin gene-related peptide were found to increase intraganglionic cAMP in a dose-dependent fashion when a phosphodiesterase inhibitor was present. The ED50 values for substance P and calcitonin gene-related peptide were 5 microM and 0.75 microM, respectively. The presence of forskolin in the incubation medium resulted in significant upward shifts of the dose-response curves for both peptides. Neither vasoactive intestinal peptide, cholecystokinin nor gastrin releasing peptide stimulated increases in intraganglionic cAMP under the same experimental conditions used for substance P and calcitonin gene-related peptide.

1-Methyl-3-isobutylxanthine↗
PubMed1992

Catecholamines modulate epidermal growth factor-induced prostaglandin E2 production in amnion-like (WISH) cells by means of a cyclic adenosine monophosphate-dependent pathway.

OBJECTIVE: The purpose of this study was to test the hypothesis that catecholamines can modulate epidermal growth factor-induced prostaglandin E2 production in amnion-derived cells via a cyclic adenosine monophosphate-dependent pathway. STUDY DESIGN: Human amnion-derived WISH cells were used as the model system to study the regulation of prostaglandin E2 production. The concentrations of prostaglandin E2 and cyclic adenosine monophosphate were measured by radioimmunoassay. Statistical significance was determined with the Student t test. RESULTS: Preexposure of WISH cells to either epinephrine, norepinephrine, or dopamine inhibited epidermal growth factor-induced prostaglandin E2 production. In addition, propranolol blocked both the increase in cyclic adenosine monophosphate accumulation and the inhibition of prostaglandin E2 production caused by epinephrine. CONCLUSIONS: These results indicate that epidermal growth factor-induced prostaglandin E2 production can be attenuated by preexposure of amnion cells to catecholamines and that the inhibitory effect of catecholamines on epidermal growth factor response may be mediated via a beta-adrenergic receptor--coupled adenylate cyclase.

1-Methyl-3-isobutylxanthine↗
PubMed1992

Proinflammatory cytokines interact synergistically with epidermal growth factor to stimulate PGE2 production in amnion-derived cells.

Recent evidence has implicated cytokines and growth factors in the initiation of parturition in women. In the present study, the amnion-derived cell line WISH was used to determine whether proinflammatory cytokines (interleukins 1 beta, 6, and 8, tumor necrosis factor-alpha, and granulocyte/macrophage colony stimulating factor) could amplify epidermal growth factor-induced prostaglandin E2 production. WISH cells were preincubated with cytokines (0.0001-10 ng/ml) for 60 min and then challenged with EGF (10 ng/ml) for 4 hrs after which PGE2 production was measured by radioimmunoassay. EGF, IL-1 beta and TNF-alpha alone caused a dose-dependent increase in PGE2 production, while IL-6, IL-8 and GM-CSF were ineffective over the dose range tested. When cells were preincubated with IL-1 beta or TNF-alpha, there was a dose-dependent potentiation of EGF-induced PGE2 production that was greater than the sum of EGF alone and IL-1 beta or TNF-alpha alone. In each case, the minimum dose of IL-1 beta or TNF-alpha which amplified EGF-induced PGE2 production was 0.1 ng/ml (p less than 0.05, Student's t-test). These data show that low concentrations of IL-1 beta or TNF-alpha may serve to amplify EGF-mediated PGE2 biosynthesis in amnion-derived cells and suggest that cytokines may modulate EGF function in responsive cells.

Amnion↗
PubMed1992

Binding of carbamyl-platelet-activating factor to the Raji lymphoblast platelet-activating factor receptor.

Carbamyl-platelet-activating factor (1-hexadecyl-2-N-methylcarbamyl-glycero-3-phosphocholine; CPAF) is an analog of platelet-activating factor (PAF) containing an N-methylcarbamyl moiety at the sn-2 position. CPAF was tested for effects on the Raji lymphoblast PAF receptor. Binding studies conducted at 4 degrees C demonstrated specific binding that reached saturation within 60-80 min. Scatchard analysis of CPAF binding data revealed a single class of CPAF binding sites (14,800/cell) with a K = 2.9 +/- 0.9 nM. Competition binding studies with PAF indicated that CPAF has about one-third the potency of native PAF. Unlike PAF, however, CPAF was not significantly metabolized by Raji lymphoblasts at 37 degrees C. CPAF was shown to have PAF-agonistic qualities, since 100 pM to 1 microM CPAF increased free intracellular calcium in a dose-dependent manner. The structurally dissimilar PAF receptor antagonists CV-6209 and alprazolam inhibited the CPAF-induced calcium changes at doses that competed with CPAF binding. Treatment of Raji lymphoblasts with PAF or CPAF (10 pM-1 microM) did not affect spontaneous proliferation, suggesting that the PAF receptor is not involved in the proliferative process in this cell line. These studies demonstrate that CPAF is a metabolically stable lymphoblast PAF receptor agonist that may provide a useful tool in the further elucidation of the role of PAF in lymphocyte function.

Burkitt Lymphoma↗
PubMed1991

Determination of levels of cyclic AMP in the myenteric plexus of guinea-pig small intestine.

Enzymatically dissociated ganglia from the myenteric plexus of the guinea-pig small intestine were used to investigate changes in levels of cyclic 3',5'-adenosine monophosphate (cAMP) in response to stimulation of adenylate cyclase by forskolin and inhibition of phosphodiesterase by 3-isobutyl-1-methylxanthine (IBMX). A linear relation with a positive correlation coefficient greater than 0.98 was found between: (1) amount of cAMP and number of ganglia; (2) amount of protein and number of ganglia; (3) amount of DNA and amount of protein; (4) amount of DNA and number of ganglia. Basal levels of cAMP were 2.25 +/- 0.21 fmol per ganglion for 900 ganglia. Forskolin stimulated a dose-dependent increase in cAMP over a concentration range of 0.05 to 50 microM, with a level of 18.6 +/- 4.9 fmol/ganglion at 50 microM forskolin. The inactive forskolin analog 1,9-dideoxyforskolin did not elevate cAMP. Addition of IBMX to the incubation medium stimulated a dose-dependent increase in cAMP over a concentration range of 0.1-1000 microM, with a level of 17.58 +/- 3.38 fmol/ganglion at 1000 microM IBMX. Application of 1 mM IBMX strongly potentiated the stimulating action of forskolin on cAMP levels. Our results derived from direct determination of cAMP changes in small intestinal myenteric ganglia are consistent with existing electrophysiological evidence for second messenger function of cAMP in slow synaptic modulation of excitability in AH/Type 2 neurons of the enteric nervous system.

1-Methyl-3-isobutylxanthine↗
PubMed1991

Evidence for platelet-activating factor receptors in several B lymphoblastoid cell lines.

Previous studies have shown that Raji, an Epstein-Barr virus (EBV)-immortalized Burkitt lymphoma B cell line, contains functional platelet-activating factor (PAF) receptors. Twelve other lymphoid cell lines, including Burkitt and non-Burkitt B cell lines, T cell lines, and a non B, non T cell line were tested for the presence of PAF receptors. Radioligand binding studies conducted at 4 degrees C revealed that six lymphoid cell lines of B cell origin (Raji, P3HR-1, BJAB, BJA/HR-1, Dakiki and PB-1) could specifically bind [3H]PAF. Treatment of four (Raji, P3HR-1, Dakiki and PB-1) of the above lymphoid cell lines with PAF resulted in an increase in free intracellular calcium, indicating that these specific PAF binding sites were functional PAF receptors. Other B cell lines (Daudi, B95-8, sfBT, CB-1), T cell lines (MOLT-4, CCRF-CEM) and a non B, non T cell line (NALM-6) had no PAF binding sites and showed no PAF-induced increase in intracellular calcium levels. These studies demonstrate evidence for the presence of PAF receptors on several B lymphocyte cell lines.

Animals↗
PubMed1990

Metabolism of 15-hydroxy-5,8,11,13-eicosatetraenoic acid by MOLT-4 cells and blood T-lymphocytes.

MOLT-4 lymphocytes metabolize 15-hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE) via beta-oxidation with retention of the hydroxyl group at the omega 6-carbon atom. 15-HETE oxidation is accompanied by the time-dependent accumulation of both beta-hydroxy acids and metabolites produced by repetitive cycles of the beta-oxidation spiral. Detection of 7-hydroxy-5-dodecenoic acid shows that these cells continue to beta-oxidize the substrate when the conjugated diene is allylic to a hydroxyl group. When 15-HETE was the substrate, it was also possible to detect 12-hydroxy-5,8,10-heptadecatrien-1-al and 3,15-dihydroxy-8,11,13-eicosatrienoic acid. The former product may be produced by alpha-oxidation of 13-hydroxy-6,9,11-octadecatrienoic acid followed by its decarboxylation. Detection of a 20-carbon metabolite, lacking a double bond at position 5, suggests that an intermediate of beta-oxidation was used as a substrate for chain elongation. When 13-hydroxy-6,9,11-octadecatrienoic acid was used as a substrate, it was indeed possible to detect 3,15-dihydroxy-8,11,13-eicosatrienoic acid as well as 15-hydroxy-8,11,13-eicosatrienoic acid. In addition, 13-hydroxy-6,9,11-octadecatrienoic acid was a precursor for the biosynthesis of both 14-hydroxy-7,10,12-nonadecatrien-1-al and 1,14-dihydroxy-7,10,12-nonadecatriene. These studies with MOLT-4 cells as well as with T-lymphocytes isolated from blood show that products of the 15-lipoxygenase pathway are metabolized with the accumulation of a variety of compounds. Since 15-HETE has been implicated as a modulator of T-cell function, these findings raise the possibility that the newly described metabolites may be involved in regulating lymphocyte function.

Adult↗
PubMed1990

The metabolism of platelet-activating factor in human T-lymphocytes.

The metabolism of 1-[3H]alkyl-2-acetyl-sn-glycero-3-phosphocholine (1-[3H]alkyl-2-acetyl-GPC; platelet-activating factor; PAF) was investigated in purified human peripheral blood T-lymphocytes and a human leukemia cell line of T-cell origin (MOLT-4). The major metabolic products of T-lymphocyte PAF metabolism are 1-alkyl-2-acyl-GPC, 1-alkyl-2-lyso-GPC and neutral lipid. The pattern of PAF metabolism in peripheral blood T-lymphocytes and MOLT-4 lymphoblasts was similar, although MOLT-4 lymphoblasts transformed PAF to 1-alkyl-2-acyl-GPC faster than peripheral blood T-lymphocytes (67% vs. 21% of added label after 64 min at 37 degrees C, respectively). Pre-exposure of MOLT-4 lymphoblasts to 1 mM of the serine hydrolase inhibitor phenylmethylsulfonyl fluoride resulted in an inhibition of PAF metabolism. Our results indicate that intact T-lymphocytes actively metabolize this biologically active phospholipid by the deacetylation-transacylation pathway.

Cell Line↗
PubMed1990

Evidence of a role for protein kinase C in epidermal growth factor-induced prostaglandin E2 synthesis in amnion cells.

Human amnion cells synthesize and release prostaglandin E2 in response to epidermal growth factor. The protein kinase C activator, phorbol 12-myristate, 13-acetate also stimulates amnion cell prostaglandin E2 synthesis. We used a human amnion cell line (WISH) to conduct in vitro experiments to investigate a potential role of protein kinase C in the signal transduction pathway leading to epidermal growth factor-induced prostaglandin E2 production. Pretreatment of cultured amnion cells with a low, nonstimulating dose of phorbol 12-myristate, 13-acetate potentiated the action of epidermal growth factor in causing prostaglandin E2 production as measured by radioimmunoassay. The protein kinase C-selective inhibitor staurosporine inhibited epidermal growth factor-induced prostaglandin E2 production, further suggesting a role for protein kinase C in epidermal growth factor action. Experiments were conducted in which amnion cells were rendered protein kinase C-deficient by chronic exposure to phorbol ester, which has been shown to down-regulate the enzyme. In these cells, epidermal growth factor caused prostaglandin E2 synthesis at levels comparable to native (non-protein kinase C-deficient) cells. We conclude that protein kinase C plays a more modulatory than direct role in the epidermal growth factor signal transduction cascade that leads to prostaglandin E2 production by amnion cells. We propose a bifurcating transduction scheme in which, under conditions of protein kinase C inactivation, epidermal growth factor alone causes prostaglandin E2 synthesis. When protein kinase C is activated by as yet unknown endogenous substances, the epidermal growth factor responsiveness of the amnion cells is greatly enhanced. This pathway could have important implications in a feed-forward mechanism regulating the level of prostaglandin E2 production during the onset of labor.

Alkaloids↗
PubMed1989

Identification of functional platelet-activating factor receptors in Raji lymphoblasts.

The binding and metabolism of platelet-activating factor (PAF) were characterized in Raji, a human Burkitt's lymphoma-derived cell line. Raji lymphoblasts readily metabolized PAF by deacetylation-reacylation at 37 degrees C, but not at 4 degrees C. Binding studies conducted at 4 degrees C demonstrated specific binding that reached saturation within 80 min. This binding was only partially reversible. Scatchard analysis of PAF binding data revealed a single class of PAF binding sites (17,800 +/- 3,600/cell) with a K of 2.3 +/- 0.3 nM. These high-affinity PAF binding sites were shown to be functional receptors, as 100 pM to 1 microM PAF increased free intracellular calcium in a dose-dependent manner. The dose of PAF necessary to achieve half maximal calcium mobilization response was 6.3 nM, which was in the range of the K for the receptor calculated from the binding studies. The structurally dissimilar PAF receptor antagonists CV-3988 and BN52021 inhibited the PAF-induced calcium changes at doses that competed with PAF binding. These studies provide the first evidence for a functional PAF receptor expressed on a lymphocyte cell line.

B-Lymphocytes↗
PubMed1989

Hyperthyroid adult rat cardiomyocytes. I. Nucleotide content, beta- and alpha-adrenoreceptors, and cAMP production.

Ventricular myocytes isolated from the hypertrophied hearts of thyrotoxic adult rats have an increase in mean protein content per myocyte (6.3 +/- 0.2 vs. 4.4 +/- 0.2 ng) compared with euthyroid cells. Viability and adenine nucleotide profiles are similar in both populations, but NAD content of the hyperthyroid myocytes is depressed (4.9 +/- 0.2 vs. 5.5 +/- 0.2 nmol/mg for controls) and UTP is higher (1.2 +/- 0.09 vs. 0.9 +/- 0.04 nmol/mg). Binding of (-)-[125I]iodocyanopindolol to intact hyperthyroid myocytes is increased by 42% compared with controls, with no change in the dissociation constant (Kd). This elevation in beta-receptor number is correlated to enhanced beta-agonist-induced adenosine 3',5'-cyclic monophosphate (cAMP) production. The half-maximal effective concentration (EC50) for the euthyroid isoproterenol dose-response curve is 2.14 x 10(-7) M but is decreased to 2.51 x 10(-8) M in hyperthyroid cardiac cells. Basal adenylate cyclase activity is apparently not affected by thyroid hormones, since basal cAMP levels for both groups are identical (5 pmol/mg) and both rise roughly twofold in the presence of a phosphodiesterase inhibitor. Forskolin-induced cAMP production and cAMP-specific phosphodiesterase activity are similar as well. In contrast to beta-adrenergic response, there are no significant differences in alpha 1-antagonist [3H]prazosin binding parameters between hyperthyroid and euthyroid cardiomyocytes.

1-Methyl-3-isobutylxanthine↗
PubMed1988

Evidence that 8-methoxypsoralen (8-MOP) is a T-lymphocyte immunomodulatory agent.

This study was designed to characterize the effects of the anti-psoriatic compound 8-methoxypsoralen (8-MOP) on human lymphocyte function in vitro. Normal human peripheral blood mononuclear cells were stimulated with an optimal (1%) or a suboptimal (0.05%) concentration of phytohemagglutinin (PHA). At the optimal concentration of PHA, 8-MOP (140 microM) caused a delay in lymphocyte proliferation, interleukin-2 (IL-2) production/accumulation and IL-2 receptor expression. Addition of exogenous IL-2 to cultures stimulated with an optimal concentration of PHA did not overcome the delay of lymphocyte proliferation and IL-2 receptor expression. At the suboptimal concentration of PHA, 8-MOP (140 microM) caused a sustained inhibition of lymphocyte proliferation, IL-2 production/accumulation and IL-2 receptor expression. Addition of exogenous IL-2 under these conditions restored the magnitude of lymphocyte proliferation and IL-2 receptor expression. However, the responses displayed the delayed lymphocyte proliferation and IL-2 receptor expression typical of cells incubated with 8-MOP and an optimal concentration of PHA.

Adjuvants, Immunologic↗
PubMed1987

8-Methoxypsoralen inhibits lymphocyte proliferation in vitro in the absence of ultraviolet radiation.

8-Methoxypsoralen was assessed for its effects on in vitro lymphocyte proliferation in the absence of ultraviolet radiation. 8-Methoxypsoralen inhibited both the phytohemagglutinin and concanavalin A-induced proliferation of normal human peripheral blood lymphocytes in a time and dose-dependent manner. This inhibition did not require the photoactivation of 8-methoxypsoralen in these cells. Suppressed lymphocyte proliferation in the presence of 8-methoxypsoralen could not be overcome by addition of exogenous interleukin-2. The observed decrease in lymphocyte proliferation hyporesponsiveness to interleukin-2 correlated with the ability of 8-methoxypsoralen to induce a dose-dependent decrease in interleukin-2 receptor expression on phytohemagglutinin-stimulated lymphocytes. Since interleukin-2 receptors play a central role in lymphocyte proliferation and immune reactivity, their decrease may explain the mechanism by which 8-methoxypsoralen impairs lymphocyte function.

Cells, Cultured↗
PubMed1987

Leukocytic pyrogen effects on prostaglandins in hypothalamic tissue slices.

Some studies suggest that leukocytic pyrogen (LP) increase hypothalamic prostaglandins which, in turn, affect hypothalamic thermoregulatory neurons to produce fever. The present study used radioimmunoassays to quantitate the ability of guinea pig hypothalamic tissue slices to produce prostaglandin E2 (PGE2), prostaglandin F2 alpha (PGF2 alpha), 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), and thromboxane B2 (TxB2). Dose- and time-dependent prostaglandin increases occurred when these slices were perfused with LP media. Steady-state levels of tissue release were reached at 0-3 min for 6-keto-PGF1 alpha, at 6-9 min for PGE2 and PGF2 alpha, and at 12-15 min for TxB2. With the exception of 6-keto-PGF1 alpha, all substances showed continuous dose-response relationships for concentrations ranging from 0.001 to 0.25 LP dilutions. Tissue PGE2, for example, was 0.7 pg X min-1 X mg-1 with the 0.001 LP dilution and 8.7 pg X min-1 X mg-1 with the 0.25 LP dilution. Indomethacin blocked much of the LP-induced prostaglandin increase. Although there is a relationship between hypothalamic LP and prostaglandins in response to physiological LP levels, tissue prostaglandins are several orders of magnitude lower than concentrations necessary to produce fever by hypothalamic microinjection. This suggests that prostanoids, such as PGE2, may not be the sole mediators of fever induced by leukocytic pyrogen.

Animals↗
PubMed1986

Identification of calmodulin activity in purified retroviruses.

Several viruses have been shown to require calcium for their function, and to bind calcium at specific sites. However, the nature of the calcium binding molecule on viruses has not been established. One possibility is the ubiquitous calcium-binding protein calmodulin. Our studies were designed to determine whether feline leukemia virus contained calmodulin. Accordingly, we tested purified feline leukemia virus for the presence of calmodulin-like activity. The virus, like authentic calmodulin, activated cyclic AMP phosphodiesterase. The ability of the virus to activate the enzyme was blocked in the presence of the known calmodulin inhibitors trifluoperazine and W-7. This indirect evidence for the presence of calmodulin was confirmed by radioimmunoassay. Several other retroviruses were also tested using radioimmunoassay and found to contain calmodulin. Our results indicate that the calcium binding site in retroviruses may be calmodulin.

3',5'-Cyclic-AMP Phosphodiesterases↗