Profiling of prostaglandin biosynthesis in biopsy fragments of human lung carcinomas and normal human lung by capillary gas chromatography-negative ion chemical ionization mass spectrometry

Methods for the profiling of prostaglandin F2 alpha (PGF2 alpha), prostaglandin D2 (PGD2), prostaglandin E2 (PGE2), thromboxane B2 (TXB2) and 6-keto-prostaglandin F1 alpha (6KPGF1 alpha) biosynthesis in tissue samples of clinical origin by capillary gas chromatography-negative ion chemical ionization mass spectrometry (CGC-NICIMS) are detailed. Aliquots (25 microliter 1) of incubates (1 ml volume) of human lung carcinoma and normal human lung tissue fragments (total protein content = 0.2 to 2.0 mg) were derivatized for vapor phase analysis in the presence of 0.75 to 1.60 ng of tetradeuterated analogs of PGE2, PGF2 alpha and 6KPGF1 alpha without prior extraction and/or chromatography. The derivatized analytes and internal standards were detected by simultaneous monitoring of ions at six different masses characteristic for each of the derivatized prostanoids. The inter-sample and intra-sample coefficients of variation for the assay method were typically less than 12%. The analysis of biological samples was completed with less than 2.5% of each derivatized sample per injection. The samples were of adequate purity for the identification and quantitation of each of the eicosanoids. The methods described in this report are highly selective and highly sensitive with detection limits of 0.1 to 0.2 picograms per injection. The analytical procedures provide the basis for comparisons of the qualitative and quantitative profiles of prostaglandin biosynthesis and should be adaptable for use in a variety of biological and clinical studies.     

Simultaneous quantification of seven prostanoids using liquid chromatography/tandem mass spectrometry: the effects of arachidonic acid on prostanoid production in mouse bone marrow-derived mast cells

We have developed a method for the simultaneous estimation of the levels of the prostanoids 6-keto prostaglandin (PG) Flalpha, PGB2, PGD2, PGE2, PGF2(alpha), PGJ2, and thromboxane (TX) B2 in blood- or serum-containing medium using liquid chromatography-tandem mass spectrometry. These prostanoids and their deuterium derivatives, which were used as internal standards, were subjected to solid-phase extraction using Empore C18 HD disk cartridges and analyzed in the selected reaction-monitoring mode. A linear response curve starting at 10 pg of prostanoid/tube was observed for each prostanoid. The accuracy of the method was demonstrated with samples containing known amounts of the prostanoids. Furthermore, we used this method to analyze the prostanoids produced in mouse bone marrow-derived mast cells stimulated with arachidonic acid, which resulted in the production of PGD2, PGE2, PGF2alpha, and TXB2. The results suggest that this simultaneous quantification method is useful for the analysis of the production of biomedically important prostanoids.     

The effects of prostanoids on estrogen-dominated rat myometrial longitudinal muscle in vitro

The purpose of these experiments was to characterize the contractile response of longitudinal muscle from the estrogen-dominated rat uterus to natural and synthetic prostanoids. The biological significance is 1) to provide evidence for or against a physiological role for each natural prostanoid in the regulation of myometrial activity, 2) to determine if each prostanoid has pharmacological potential for the manipulation of myometrial activity, and 3) to understand the structural requirements for prostanoid action on the myometrium. All analogs tested produced excitation of the myometrium in vitro through what appeared to be a direct action on the muscle. The order of potency of the natural prostanoids was prostaglandin (PG) F2 alpha = PGD2 = PGE2 = PGE1 greater than PGA2 = PGB2 = 6-keto-PGF1 alpha. This order of potency was not consistent with any single currently recognized prostanoid receptor. Furthermore, PGF2 alpha had an EC50 (effective concentration that produces 50% of the maximal response) of 0.5 microM, which was low in comparison to other PGF2 alpha-sensitive tissues. There were large differences in the maximum tension developed in response to the prostanoids tested, only PGF2 alpha, PGE2 and 6-keto PGF1 alpha were full agonists. Although the simplest explanation of these data was that the rat uterus contains a single novel type of prostanoid receptor, the existence of multiple receptor subtypes could not be disproved. Evidence from the effect of synthetic analogs suggested that neither thromboxane A2 nor PGI2 are physiological regulators of activity in this tissue.     

Effects of fluticasone propionate inhalation on levels of arachidonic acid metabolites in patients with chronic obstructive pulmonary disease

BACKGROUND: In smoking COPD patients the bronchoalveolar lavage (BAL) fluid contains high numbers of inflammatory cells. These cells might produce arachidonic acid (AA) metabolites, which contribute to inflammation and an increased bronchomotor tone. AIMS: To investigate levels of AA metabolites in BAL fluid, before and after inhaled glucocorticoid therapy: fluticasone propionate (FP) 1 mg per day, or placebo. METHODS: A double-blind placebo controlled trial lasting six months. COPD patients were selected by clinical criteria and the presence of bronchial hyper-responsiveness (BHR). Lung function was recorded and in BAL fluid we counted cell numbers and measured LTB4, LTC4/D4/E4, PGE2, 6kPGF1alpha, PGF2alpha and TxB2. A control group consisted of asymptomatic smokers (n=6). RESULTS: Paired data were obtained from 9 FP treated and 11 placebo patients. BAL cells were almost exclusively alveolar macrophages. In patients and controls both cellularity and levels of AA metabolites were equal Cell numbers did not change after treatment. Statistically significant decreases after FP therapy were noticed for PGE2 (30%), 6kPGF1alpha (41%) and PGF2alpha (54%). CONCLUSIONS: In COPD, the capability of inflammatory cells to produce certain AA metabolites was decreased after inhaled FP treatment. This result is discussed in its relation to clinical effects, the influence of smoking, and the results of an earlier, similar study in asthma patients.     

Effects of selective and non-selective COX inhibitors on antigen-induced release of prostanoid mediators and bronchoconstriction in the isolated perfused and ventilated guinea pig lung

The contribution of cycloxygenase (COX)-1 and COX-2 in antigen-induced release of mediators and ensuing bronchoconstriction was investigated in the isolated perfused guinea pig lung (IPL). Antigen challenge with ovalbumin (OVA) of lungs from actively sensitised animals induced release of thromboxane (TX)A(2), prostaglandin (PG)D(2), PGF(2)(alpha), PGI(2) and PGE(2), measured in the lung effluent as immunoreactive TXB(2), PGD(2)-MOX, PGF(2)(alpha), 6-keto PGF(1)(alpha) and PGE(2), respectively. This release was abolished by the non-selective COX inhibitor flurbiprofen (10 microM). In contrast, neither the selective COX-1 inhibitor FR122047 nor the selective COX-2 inhibitor celecoxib (10 microM each) significantly inhibited the OVA-induced bronchoconstriction or release of COX products, except for PGD(2). Another non-selective COX inhibitor, diclofenac (10 microM) also significantly inhibited antigen-induced bronchoconstriction. The data suggest that both COX isoenzymes, COX-1 and COX-2 contribute to the immediate antigen-induced generation of prostanoids in IPL and that the COX-1 and COX-2 activities are not associated with different profiles of prostanoid end products.     

High performance liquid chromatography and UV detection for the separation and quantitation of prostaglandins

A fast and reliable method for the separation and quantitation of arachidonic acid metabolites PGF1 alpha, PGF2 alpha, PGD2, PGE1, PGE2, PGB2, PGA2, 6-keto PGE1, 6-keto PGF1 alpha, TxB2 and 15-keto PGE2 by high-performance liquid chromatography has been developed. Utilizing a single reverse-phase column and a UV spectrophotometer, sensitivity as little as 30 nanograms of each of these prostaglandins can be separated and subsequently detected. Although this study was performed using standards, it is highly promising for future application to biological fluids.     

Prostanoids in bronchoalveolar lavage fluid do not predict outcome in congenital diaphragmatic hernia patients

Vasoactive prostanoids may be involved in persistent pulmonary hypertension (PPH) in infants with a congenital diaphragmatic hernia (CDH). We hypothesized that increased levels of prostanoids in bronchoalveolar lavage (BAL) fluid would predict clinical outcome. We measured the concentrations of 6-keto-prostaglandin F(1alpha) (6-keto-PGF(1alpha)), thromboxane B(2) (TxB(2)), protein, albumin, total cell count, and elastase-alpha1-proteinase-inhibitor complex in BAL fluid of 18 CDH patients and of 13 control subjects without PPH. We found different concentrations of prostanoids in BAL fluid of CDH patients with PPH: infants with a poor prognosis had either high levels of both 6-keto-PGF(1alpha) and TxB(2) compared to controls, or high levels of 6-keto-PGF(1alpha) only. TxB(2) levels showed a large variability in all CDH patients irrespective of outcome. We conclude that prostanoid levels in BAL fluid do not predict clinical outcome in CDH patients.     

The relation between thromboxane and prostaglandin synthesis in human decidua tissue: a comparison of eicosanoid synthesis in minced tissue with that in a cell-free preparation

Radiotracer studies and radioimmunoassay measurements demonstrate that minced tissues of human decidua produce chiefly thromboxane B2 (TxB2) (70% of total eicosanoids) and small amounts of prostaglandin F2 alpha (PGF2 alpha) (13%) PGD2 (8%), 6-keto-PGF1 alpha (5%) and PGE2 (4%). Inhibition of thromboxane synthesis with a specific inhibitor (OKY-1581: sodium (E)-3-[4(-3-pyridylmethyl)-phenyl]-2-methyl propenoate) increased prostaglandin formation in general, with the main product being PGF2 alpha (38%), a nonenzymic derivative of PGH2. Crude particulate fractions prepared from the same tissue synthesized two major products from [3H]arachidonate, TxB2 and 6-keto-PGF1 alpha (54 and 30%, respectively) and some PGF2 alpha and PGE2 (8-8%). However, in the presence of reduced glutathione (GSH), PGE2 became the main product (81%) (TxB2, 15%; PGF2 alpha, 2%; and 6-keto-PGF1 alpha, 2%). Half-maximal stimulation of PGE2 synthesis occurred at 46 microM GSH. The GSH concentration of tissue samples was found to be 110 +/- 30 microM. We conclude that human first trimester decidua cells possess the key enzymes of prostaglandin and thromboxane synthesis. Apparently, the production of these compounds is controlled by a specific mechanism in the tissue, which keeps PGE and prostacyclin synthesis in a reversibly suppressed state, whereas the formation of thromboxane is relatively stimulated.     

Redirection of eicosanoid metabolism in mPGES-1-deficient macrophages

Microsomal prostaglandin E synthase (mPGES)-1 is one of several prostaglandin E synthases involved in prostaglandin H2 (PGH2) metabolism. In the present report, we characterize the contribution of mPGES-1 to cellular PGH2 metabolism in murine macrophages by studying the synthesis of eicosanoids and expression of eicosanoid metabolism enzymes in wild type and mPGES-1-deficient macrophages. Thioglycollate-elicited macrophages isolated from mPGES-1-/- animals and genetically matched wild type controls were stimulated with diverse pro-inflammatory stimuli. Prostaglandins were released in the following order of decreasing abundance from wild type macrophages stimulated with lipopolysaccharide: prostaglandin E2 (PGE2)>thromboxane B2 (TxB2)>6-keto prostaglandin F1alpha (PGF1alpha), prostaglandin F(2alpha) (PGF2alpha), and prostaglandin D2 (PGD2). In contrast, we detected in mPGES-1-/- macrophages a >95% reduction in PGE2 production resulting in the following altered prostaglandin profile: TxB2>6-keto PGF1alpha and PGF2alpha>PGE2, despite the comparable release of total prostaglandins. No significant change in expression pattern of key prostaglandin-synthesizing enzymes was detected between the genotypes. We then further profiled genotype-related differences in the eicosanoid profile using macrophages pre-stimulated with lipopolysaccharide followed by a 10-min incubation with 10 microm [3H]arachidonic acid. Eicosanoid products were subsequently identified by reverse phase high pressure liquid chromatography. The dramatic reduction in [3H]PGE2 formation from mPGES-1-/- macrophages compared with controls resulted in TxB2 and 6-keto PGF1alpha becoming the two most abundant prostaglandins in these samples. Our results also suggest a 5-fold increase in 12-[3H]hydroxyheptadecatrienoic acid release in mPGES-1-/- samples. Our data support the hypothesis that mPGES-1 induction in response to an inflammatory stimulus is essential for PGE2 synthesis. The redirection of prostaglandin production in mPGES-1-/- cells provides novel insights into how a cell processes the unstable endoperoxide PGH2 during the inactivation of a major metabolic outlet.     

Effects of fluticasone propionate on arachidonic acid metabolites in BAL-fluid and methacholine dose-response curves in non-smoking atopic asthmatics

Hyperresponsiveness of the airways to nonspecific stimuli is a characteristic feature of asthma. Airway responsiveness is usually characterized in terms of the position and shape of the dose-response curve to methacholine (MDR). In the study we have investigated the influence of fluticasone propionate (FP), a topically active glucocorticoid, on arachidonic acid (AA) metabolites in broncho-alveolar lavage (BAL) fluid (i.e. TxB(2), PGE(2), PGD(2), 6kPGF(1alpha) and LTC(4)) on the one hand and MDR curves on the other hand. The effect of FP was studied in a randomized, double-blind, placebo-controlled design in 33 stable nonsmoking asthmatics; 16 patients received FP (500 mug b.i.d.) whereas 17 patients were treated with placebo. We found that the forced expiratory volume in 1s (FEV(1) % predicted) increased, the log(2)PC(20) methacholine increased and the plateau value (% fall in FEV(1)) decreased after a 12 week treatment period. No changes in AA-metabolites could be determined after treatment except for PGD(2) which decreased nearly significantly (p = 0.058) within the FP treated group, whereas the change of PGD(2) differed significantly (p = 0.05) in the FP treated group from placebo. The levels of the other AA metabolites (i.e. TxB(2), PGE(2), 6kPGF(1alpha) and LTC(4)) remained unchanged after treatment and were not significantly different from the placebo group. Our results support the hypothesis that although FP strongly influences the position, the shape and also the maximum response plateau of the MDR curve, this effect is not mainly achieved by influence on the level of AA metabolites. Other pro-inflammatory factors may be of more importance for the shape of the MDR curve. It is suggested that these pro-inflammatory factors are downregulated by FP.     

Effect of alterations in the cyclopentane ring on bone resorptive activity of prostaglandin

Previous studies have shown that the natural prostanoids, PGE2, PGE1 and PGF2 alpha are potent stimulators of bone resorption. In this study, we have examined the effects of alterations in the cyclopentane ring of these prostanoids for their effect on the resorptive response of cultured long bones from 19-day fetal rats as measured by the release of previously incorporated 45Ca. Indomethacin (10(-6)M) was added to minimize endogenous prostaglandin production. In this system PGE2 and PGE1, the 9 keto, 11 alpha hydroxy compounds, were approximately equally effective at concentrations of 10(-8) to 10(-6) M. The 9 alpha hydroxy, 11 alpha hydroxy compound, PGF2 alpha, was active at 10(-7) to 10(-5) M. In contrast, the 9 alpha hydroxy, 11-keto compound, PGD2, showed only a minimal stimulation of bone resorption at 10(-5) M. While these data suggested that the 11 alpha hydroxy group was important for bone resorbing activity, 11 beta PGE2 and 11-deoxy PGE1 were only slightly less potent than their physiologic counterparts. Both 9 beta, 11 alpha PGF2 and 9 alpha, 11 beta PGF2 were less potent than PGF2 alpha but did cause substantial stimulation of bone resorption and were equally effective at 10(-6) to 10(-5) M. 9 alpha, 11 beta PGF2 alpha is of particular interest since it is major metabolite of PGD2. These results suggest that the binding of prostanoids to the receptor which mediates bone resorption is affected by changes at the 9 and 11 positions of the pentane ring but do not support the hypothesis that the 11 alpha OH function is essential for this biological activity.     

Excretion of primary prostanoids and their metabolites during acute volume expansion

Simultaneous determination of urinary excretion rates of primary unmetabolized prostanoids and their enzymatic metabolites were performed by gas chromatography-mass spectrometry (GC/MS) or tandem mass spectrometry (GC/MS/MS). Changes in kidney function were induced by acute (4 h) volume expansion. Despite marked changes in urine flow, GFR, urinary pH, osmolality, sodium and potassium excretion, only a insignificant or transient rise in the enzymatic prostanoid metabolites (2,3-dinor-6-keto-PGF1 alpha, PGE-M, 2,3-dinor-TxB2 and 11-dehydro-TxB2) was observed. The excretion rates of the primary prostanoids were elevated in parallel with the rise in urine flow: PGE2 rose (p less than 0.05) from 14.2 +/- 4.0 to 86.2 +/- 20.7, PGF2 alpha from 60.0 +/- 4.9 to 119.8 +/- 24.0, 6-keto-PGF2 alpha from 7.2 +/- 1.3 to 51.5 +/- 17.0, and TxB2 from 11.2 +/- 3.3 to 13.6 +/- 3.6 ng/h/1.73 m2 (means +/- SEM) at the maximal urine flow. Except for 6-keto-PGF1 alpha and TxB2, this rise in urinary prostanoid levels was only transient despite a sustained fourfold elevated urine flow. We conclude that urine flow rate acutely affect urine prostanoid excretion rates, however, over a prolonged period of time these effects are not maintained. The present data support the concept that urinary levels of primary prostanoids mainly reflect renal concentrations whereas those of enzymatic metabolites reflect systemic prostanoid activity. From the excretion pattern of TxB2 one can assume that this prostanoid represents renal as well as systemic TxA2 activity.     

Neonatal urinary prostanoid excretion

Urinary excretion of prostanoids prostaglandin E2 (PGE2), PGE-M (7alpha-hydroxy-5,11-diketo-2,3,4,5,20-penta-19-carboxyprostano ic acid), 6-keto-PGF1alpha, 2,3-dinor-6-keto-PGF1alpha, thromboxane B2 (TxB2) 2,3-dinor-TxB2 and 11-dehydro-TxB2 was determined by gas chromatography/mass spectrometry in preterm and term infants to show that there is an age-dependent excretion rate of the above prostanoids in infants this young. Group I included premature children with normal postnatal development, Groups II and III included term children who were admitted in the neonatal period for observation because of feeding problems but who were subsequently found to be completely healthy. We present normal data of three primary prostanoids and four prostanoid metabolites. In Group I, excretion rates of 2,3-dinor-TxB2 were significantly lower than in Group II (P = 0.04) and in Group III (P = 0.05). Furthermore, the excretion rate of 11-dehydro-TxB2 in group I was significantly lower than in Group II (P = 0.05). We found no significant age-dependent differences between the three groups in excretion rates of PGE2, PGE-M, 6-keto-PGF1alpha, 2,3-dinor-6-keto-PGF1alpha, and TxB2.     

Synthetic pathways of gallbladder mucosal prostanoids: the role of cyclooxygenase-1 and 2.

Acute cholecystitis is associated with increased gallbladder prostanoid formation and the inflammatory changes and prostanoid increases can be inhibited by nonsteroidal anti-inflammatory agents. Recent information indicates that prostanoids are produced by two cyclooxygenase (COX) enzymes, COX-1 and COX-2. The purpose of this study was to determine the COX enzymatic pathway in gallbladder mucosal cells involved in the production of prostanoids stimulated by inflammatory agents. Human gallbladder mucosal cells were isolated from cholecystectomy specimens and maintained in cell culture and studied in comparison with cells from a well differentiated gallbladder mucosal carcinoma cell line. COX enzymes were evaluated by Western immunoblotting and prostanoids were measured by ELISA. Unstimulated and stimulated cells were exposed to specific COX-1 and COX-2 inhibitors. In both normal and transformed cells constitutive COX-1 was evident and in gallbladder cancer cells lysophosphatidyl choline (LPC) induced the formation of constitutive COX-1 enzyme. While not detected in unstimulated normal mucosal cells and cancer cells, COX-2 protein was induced by both lipopolysaccharide (LPS) and LPC. Unstimulated gallbladder mucosal cells and cancer cells produced prostaglandin E2 (PGE2) and prostacyclin (6-keto prostaglandin F1alpha, 6-keto PGF1alpha) continuously. In freshly isolated normal gallbladder mucosal cells, continuously produced 6 keto PGF1alpha was inhibited by both COX-1 and COX-2 inhibitors while PGE2 levels were not affected. Both LPS and LPC stimulated PGE2 and 6 keto PGF1alpha formation were blocked by COX-2 inhibitors in freshly isolated, normal human gallbladder mucosal cells and in the gallbladder cancer cells. The prostanoid response of gallbladder cells stimulated by proinflammatory agents is inhibited by COX-2 inhibitors suggesting that these agents may be effective in treating the pain and inflammation of gallbladder disease.     

Effects of ozone on cyclooxygenase metabolites in the baboon tracheobronchial tree

Short-term exposure to 0.5 parts per million (ppm) ozone has been shown to cause an increase in respiratory resistance in primates that can be diminished by 50% with pretreatment with cromolyn sodium. Because of the known membrane-stabilizing effects of cromolyn and the resultant inhibition of mediator production, we hypothesized a role for the products of arachidonic acid (AA) metabolism in these events. We exposed five adult male baboons to 0.5 ppm ozone on two occasions, once with cromolyn pretreatment and once without. Pulmonary resistance (RL) was monitored and bronchoalveolar lavage (BAL) was performed before and after each exposure. The BAL was analyzed for a stable hydrolysis product of prostacyclin, 6-keto-prostaglandin (PG) F1 alpha, PGE2, a stable hydrolysis product of thromboxane (Tx) A2, TxB2, and PGF2 alpha. RL increased after ozone exposure (1.62 +/- 0.23 to 3.77 +/- 0.51 cmH2O.l-1.s, difference 2.15; P less than 0.02), and this effect was partially blocked by cromolyn (1.93 +/- 0.09 to 3.18 +/- 0.40 cmH2O.l-1.s, difference 1.25; P less than 0.02). The base-line levels of the metabolites of AA in the BAL were as follows (in pg/ml): 6-keto-PGF1 alpha 72.78 +/- 12.6, PGE2 145.92 +/- 30.52, TxB2 52.52 +/- 9.56, and PGF2 alpha 22.28 +/- 5.42. Ozone exposure had no effect on the level of any of these prostanoids (P = NS). These studies quantify the magnitude of cyclooxygenase products of AA metabolism in BAL from baboon lungs and demonstrate that changes in the levels of these mediators in BAL are not prerequisites for ozone-induced increases in respiratory resistance.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prostaglandin production by phagocytic cells of the mouse thymic reticulum in culture and its modulation by indomethacin and corticosteroids

The production of prostaglandins by phagocytic cells of the thymic reticulum in culture (P-TR) was studied by using high pressure liquid chromatography and radioimmunoassay. Radioimmunologic determinations showed that thromboxane B2 (TXB2), prostaglandin E2 (PGE2), and 6-keto-prostaglandin F1 alpha (6 keto-PGF1 alpha) were the major compounds released into the culture medium, whereas prostaglandin F2 alpha (PGF2 alpha) was only a minor component. Indomethacin and dexamethasone exerted a similar pattern of differential inhibition of the secretion of prostanoids. PGE2 and 6-keto PGF1 alpha productions were markedly decreased by these anti-inflammatory drugs, whereas those of TXB2 and PGF2 alpha were not or were only slightly affected. Experiments performed with an antiglucocorticoid compound (RU 38486) showed that the steroid-induced inhibition of prostanoid secretion is a classical receptor-mediated action. These results demonstrated that phagocytic cells of the thymic reticulum, which resemble the thymic interdigitating cells, produce several types of prostaglandins. Because it has been described that P-TR regulate thymocyte proliferation in vitro via the secretion of both interleukin 1 and PGE2, these results suggest that anti-inflammatory agents may be able to modulate the thymic microenvironment and, consequently, thymocyte proliferation.     

Prostaglandin D2 induces programmed cell death in Trypanosoma brucei bloodstream form

African trypanosomes produce some prostanoids, especially PGD2, PGE2 and PGF2alpha (Kubata et al. 2000, J. Exp. Med. 192: 1327-1338), probably to interfere with the host's physiological response. However, addition of prostaglandin D2 (but not PGE2 or PGF2alpha) to cultured bloodstream form trypanosomes led also to a significant inhibition of cell growth. Based on morphological alterations and specific staining methods using vital dyes, necrosis and autophagy were excluded. Here, we report that in bloodstream form trypanosomes PGD2 induces an apoptosis-like programmed cell death, which includes maintenance of plasma membrane integrity, phosphatidylserine exposure, loss of mitochondrial membrane potential, nuclear chromatin condensation and DNA degradation. The use of caspase inhibitors cannot prevent the cell death, indicating that the process is caspase-independent. Based on these results, we suggest that PGD2-induced programmed cell death is part of the population density regulation as observed in infected animals.     

Prostaglandin E(2) and stem cell factor can deliver opposing signals to B lymphocyte precursors

The synthetic prostanoid, 16,16-dimethyl PGE(2), suppressed B lymphopoiesis in mice and proliferation of normal B cell precursors or the F10 pro-B cell line to interleukin 7 in culture. This was not the case with two other prostanoids, PGD(2) and PGF(2alpha), or agonists for PGI(2) agonist and thromboxane A(2) agonist receptors. PGE(2), but not the related prostanoids or agonists, induced apoptosis in F10 cells. The apoptotic response was mediated by the EP2 class of PGE(2) receptors and required an increase in intracellular cyclic adenosine 3',5'-monophosphate, activation of protein kinase A, and protein synthesis. The influence of PGE(2) on F10 cells was diminished in the presence of a cloned stromal cell line or stem cell factor. These findings describe another potential regulatory circuit in bone marrow which might influence B lymphopoiesis under disease or steady-state conditions.     

Evidence for separate PGD2 and PGF2 alpha receptors in the canine mesenteric vascular bed.

Potential interactions between PGD2 and PGF2 alpha in the mesenteric and renal vascular beds were investigated in the anesthetized dog. Regional blood flows were measured with electromagnetic flow probes. PGD2, PGF2 alpha and Norepinephrine (NE) were injected as a bolus directly into the appropriate artery, and responses to these agents were obtained before, during and after infusion of either PGD2 or PGF2 alpha into the left ventricle. In each case, the infused prostaglandin caused vascular effects of its own. Left ventricular infusion of PGD2 reduced responses to local injections of PGD2 in the intestine, and a similar effect was observed for PGF2 alpha, suggesting significant receptor or receptor-like interactions for each of the prostanoids. However, systemic infusion of prostaglandin F2 alpha (20--100 ng/kg/min) had no effect on renal or mesenteric vascular responses to local injection of prostaglandin D2. Similarly, PGD2 administration (100 ng/kg/min) did not affect responses to PGF2 alpha in the intestine. The present results therefore suggest that these prostaglandins, i.e., D2 and F2 alpha, act through separate receptors in the mesenteric and renal vascular beds. In addition, increased prostaglandin F2 alpha levels produced by infusion of F2 alpha reduced mesenteric but not renal blood flow, suggesting that redistribution of cardiac output might participate in side effects often observed with clinical use of this prostaglandin, such as nausea and abdominal pain.