Effect of endotoxin treatment on the expression and localization of spinal cyclooxygenase, prostaglandin synthases, and PGD2 receptors

Systemic inflammation leads to increased expression of spinal cyclooxygenase (COX)-2 and to a subsequent increase of prostaglandin (PG) biosynthesis, which contribute to the development of hyperalgesia and allodynia. In this study, endotoxin caused a sequential induction of membrane bound prostaglandin E synthase-1 and lipocalin-type PGD synthase (L-PGDS) in the mouse spinal cord. L-PGDS expression was detected in the leptomeninges, oligodendrocytes, and interestingly, in discrete perivascular cells. Endotoxin-caused increase was most prominent in oligodendrocytes. Endotoxin-induced COX-2 and membrane bound prostaglandin E synthase-1 were restricted to the leptomeninges and perivascular cells. COX-1 was not influenced by endotoxin. We found COX-1 expressed in microglia, some of them in close proximity to L-PGDS-positive oligodendrocytes and co-localization of COX-1 with L-PGDS in perivascular and leptomeningeal cells under control conditions. It can be assumed, that PGD₂ biosynthesis under control conditions is mediated via COX-1 and that during inflammation, increased PGD₂ is dependent on COX-2. We found the PGD₂ receptors DP1 and chemoattractant receptor homologous molecule expressed on T helper type 2 cells (CRTH2) localized in neurons of the dorsal, and motoneurons in the ventral horn. The localization of the PGD₂ receptors DP1 and CRTH in spinal cord neurons, particularly in neurons of lamina I and II involved in the processing of nociceptive stimuli, supports a role of PGD₂ under inflammatory conditions.     

Effects of short-term oilseed supplementation on plasma fatty acid composition,progesterone and prostaglandin F metabolite in lactating beef cows

Twenty-four 3-year-old Angus cows (512.2±21.6 kg) and six ruminally cannulated beef heifers (523.1±16.9 kg) were used to determine the impact of feeding oilseeds starting at the beginning of estrous synchronization until maternal recognition of pregnancy on plasma fatty acid composition. Starting ~60 days postpartum cows were synchronized with the Select Synch+controlled internal drug-release (CIDR) device and timed artificial insemination (AI) protocol. The day CIDR was inserted; cattle were randomly assigned to one of the three treatments being grazing only (CON) or a supplement containing whole soybeans (SOY); or whole flaxseed (FLX). Cattle continued to receive these diets for 28 days. Blood was collected every 3 days until 10 days after insemination and then every day until 18 days after insemination. All cattle grazed a common pasture and supplemented cattle were individually fed their respective supplements once daily. Ruminally cannulated heifers were used to evaluate the impact supplements had on forage intake, which was reduced (P=0.05) with oilseed supplementation. Feeding oilseeds increased total fatty acid intake (P<0.001) across treatments with SOY having greater (P<0.001) 18:2n-6 intake than either CON or FLX. Likewise, cattle fed FLX had greater (P<0.001) 18:3n-3 intake than either CON or SOY. There was a treatment×time interaction (P⩽0.05) for all fatty acids identified except for 20:5n-3 (P=0.99). Within 3 days after the start of supplementation, plasma concentrations of 18:2n-6 increased (P<0.001) for cattle fed SOY compared with CON or FLX, whereas flax-fed cattle did not exhibit an increase (P=0.02) until day 15 of supplementation over that of CON. Plasma concentrations for 18:3n-3 was greater (P<0.013) for FLX than both CON and SOY by day 12. Feeding flaxseed tended to (P=0.07) increase and increased (P=0.01) plasma concentrations of 20:4n-6 by day 18 over CON and SOY, respectively. Overall, treatment did not affect serum concentration of progesterone (P=0.18) or prostaglandin F metabolite (P=0.89). However, day after breeding had an effect on serum progesterone (P=0.01) with day 16 after timed AI being lower compared with other days. Feeding oilseeds during the time of estrous synchronization will not only increase the energy density of the diet but will provide key fatty acids around the time of maternal recognition of pregnancy.     

Rat osteogenic sarcoma cells: comparison of the effects of prostaglandins E1, E2, I2 (prostacyclin), 6-keto F1alpha and thromboxane B2 on cyclic AMP production and adenylate cyclase activity.

The effects of prostaglandins (PG's) E₁, E₂, I₂ (prostacyclin), 6-keto F_1α and thromboxane (Tx) B₂ were compared in freshly isolated cells from a rat osteogenic sarcoma and in membranes from cultured cells of the same tumour. Cyclic AMP production was measured in cells and adenylate cyclase activity was measured in cell membranes. In both systems PGI₂ was less potent than either of the PGE's, and both TxB₂ and 6-keto PGF_1α were only weak agonists. These effects on bone-derived cells suggest that PGI₂ is unlikely to be a potent bone resorbing agent. Resitance experiments suggested that all the PG's share the same receptor site which appears distinct from the site of action of parathyroid hormone.     

Abruquinone A,B,D,E,F and G from the Root of Abrus precatorius

Eight flavonoids have been isolated from the root of Abrus precatorius L. Among them, six isoflavanquinones, designated (3R)-abruquinone A, B, D and E, (3S)-abruquinone F and G, are characterized by chemical and spectral means including 1H-1H COSY, 1H-13C COSY and CD methods.     

肉毒A、B、E、F型抗毒素血浆的试生产与检定

按照《中国生物制品规程》(2000版)的规定,在肉毒A、B、E、F型抗毒素血浆试生产前,制定了肉毒A、B、E、F型类毒素马匹免疫计划及相应抗毒素血浆效价测定方法,免疫结果显示：肉毒A、B、E、F型类毒素免疫马匹成功率分别为75%、75%、60%、100%。     

Effect of prostaglandins A1, A2, B1, B2, E2 and F2 alpha on human umbilical cord vessels

Human umbilical blood vessels have the ability to close spontaneously following delivery at term. It has been suggested that prostaglandins may have a possible physiological role in its closure. This study investigates the effects of 6 naturally occurring prostaglandins (A1, A2, B1, B2, E2, F2a) on the umbilical blood vessels. Umbilical cords were collected from cases of normal spontaneous vaginal deliveries and cesarian section at term. A total of 41 strips of umbilical arteries and 26 strips of umbilical veins from 24 cords were used. A 4-point bioassay method was used to compare the potency of prostaglandins A1, A2, B1 and F2a with PGE2. The effect of Polyphloretin Phosphate (PPP) on prostaglandin-induced contractions was studied on umbilical artery strips from 12 cords. The 6 prostaglandins exerted a stimulant effect on the isolated strips of human umbilical arteries. Prostaglandin B2 was the most potent compound on the umbilical vein, followed by PGA2. PPP in the concentration range of 10 to 40 mcg/ml completely eliminated the responses of PGE2, F2a, A1, A2, and B1. Responses to PGB2 were considerably but not completely abolished. PPP (up to 40 mcg/ml) did not affect contractions induced by 5-hydroxytryptamine, suggesting the presence of discrete receptor sites in the blood vessels for different pharmacologically active compounds. This is the first report of the constrictor effect of PGA and PGB compounds. These naturally occuring prostaglandins with high potencies (compared with other prostaglandins and other vasoactive substances) may play a role in spontaneous closure of umbilical vessels. PGE1, E2, F1 and F2a are found in umbilical blood vessels obtained at term.     

Effects of thyrotropin and N6,O2′-dibutyryl cyclic 3′,5′-adenosine monophosphate on prostaglandin levels in thyroid

We have shown that TSH increases PG levels in isolated bovine thyroid cells. We now report that TSH also increases PG levels in rat and mouse thyroid, and that these effects may be mediated via cyclic AMP. PG and cyclic AMP levels in intact rat and mouse thyroid lobes were measured by radioimmunoassay. During 60-min incubations at 37°C, 25 mU/ml TSH effected a 75–83% increase in PGE₁ and PGF_2α ”equivalents“ in rat thyroid; parallel measurements of endogenous cyclic AMP in these intact thyroid lobes revealed that maximal TSH-induced increase in cyclic AMP also required 60-min incubations. In mouse thyroid, 5 mU/ml TSH increased PGE₁ and PGF_2α levels 38–82% above basal; this TSH effect was evident within 15 min of incubation, thus mimicking the time-course of TSH-induced increase in mouse thyroid cyclic AMP. Exogenous DBcAMP, 0.5 to 3 mM, effected a dose-related increase in mouse thyroid PG levels. The stimulatory effects of both TSH and DBcAMP on mouse thyroid PG levels were abolished by aspirin and indomethacin. These studies suggest that TSH-induced increase in endogenous PG levels in thyroid may be mediated by cyclic AMP.     

Bioactive apocarotenoids annuionones F and G: structural revision of annuionones A, B and E

The polar bioactive fractions of Helianthus annuus cv. Stella and SH-222 have yielded eight apocarotenoids, two of them isolated for the first time as natural products (annuionones F and G). The isolation of higher amounts of annuionones A and E allowed us to realize a more comprehensive spectroscopical study. We propose a revised structure for annuionone A, B and E based on careful re-analyses of new spectroscopical data.     

The use of Endopep-MS for the detection of botulinum toxins A, B, E, and F in serum and stool samples

Botulinum neurotoxin (BoNT) causes the disease botulism, which can be lethal if untreated. Previous work in our laboratory focused on developing Endopep-MS, a mass spectrometric-based endopeptidase method for the detection and differentiation of BoNT serotypes. We have expanded this effort to include an antibody capture method to partially purify and concentrate BoNT from serum and stool extract samples for the Endopep-MS assay. Because complex matrices such as serum and stool contain abundant endogenous proteases, this technique was needed to remove most proteases from the sample while concentrating BoNT from a sample size of 100 to 500 microl to 20 microl. When this antibody capture method is combined with the Endopep-MS reaction, limits of detection in 500mul of spiked human serum are 10 mouse LD50 (20 mouse LD50/ml) for BoNT A, 0.5 mouse LD50 (1 mouse LD50/ml) for BoNT B, 0.1 mouse LD50 (0.2 mouse LD50/ml) for BoNT E, and 0.5 mouse LD50 (1 mouse LD50/ml) for BoNT F. The limits of detection in spiked stool extracts are somewhat higher due to the high-protease environment of stool extract that also requires use of protease inhibitors. The entire method can be performed in as short a time as 4 h.     

Gamahonolides A,B, and Gamahorin,Novel Antifungal Compounds from Stromata of Epichloe typhina on Phleum pratense

Four novel antifungal compounds, gamahonolides A and B, gamahorin, and 5-hydroxyl-4-phenyl-2(5H)-furanone, were isolated from stromata of Epichloe typhina on Phleum pratense. Their structures were determined by spectroscopic methods. The absolute configuration of gamahonolide A was determined by its ORD spectrum and ¹H-NMR shift difference between the diastereomeric pair of its O-methylmandelates. The stereochemistry of gamahorin was determined by NOE difference spectra and its CD spectrum.     

Effect of leukotriene B4, prostaglandin E2 and arachidonic acid on cytosolic-free calcium in human neutrophils

Changes in cytosolic free calcium [Ca2+]i and release of beta-glucuronidase in response to leukotriene B4 (LTB4) were measured in intact neutrophils loaded with the fluorescent Ca2+ indicator, quin 2. LTB4 (10(-10) M or higher) caused a rapid rise in [Ca2+]i due to influx from the extracellular medium and release from intracellular pools as well as enzyme release. PGE2 (3 microM) did not alter [Ca2+]i whereas arachidonic acid (10 microM) raised [Ca2+]i. Pretreatment of cells with the chemotactic peptide FMLP inhibited the subsequent rise of [Ca2+]i induced by LTB4. Since chemotactic peptides activate the lipoxygenase pathway of arachidonic acid metabolism, it may be speculated that endogenous LTB4 generation is involved in neutrophil activation.     

Effects of nitric oxide synthase inhibitors and a substrate,L-arginine,on the cardiac function of juvenile salmonid fish

Control of cardiac function was investigated juvenile brown trout (Salmo trutta L.) and rainbow trout (Oncorhynchus mykiss Walbaum) using inhibitors of nitric oxide synthase (NOS), (L-NAME, NG-nitro-L-arginine and L-NMMA, NG-monomethyl-L-arginine) and a substrate of NOS (L-arginine). Salmonid alevins are excellent models for such studies since they are transparent, the beating heart is easily observed, diffusing distances are small, and they respond within a few seconds to exogenously administered chemicals. The response to inhibitors of NOS (L-NAME or L-NMMA) was tachycardia interpreted as vasoconstriction through lowered capacity for synthesis of NO. This could be reversed by addition of L-arginine and the subsequent bradycardia was explained as a vasodilation resulting from increased synthesis of NO. Blood flow into the heart is mainly via the vitelline vein and changes of flow resulting from constriction or dilation of this vessel may be probably major determinants of heart rate. The results provide evidence for the presence NOS in juvenile fish and indicate a physiological role for NO in cardiovascular control.     

Effect of mifepristone on pregnancy,pregnancy-specific protein B (PSPB), progesterone,estradiol-17beta,prostaglandin F2alpha (PGF2alpha) and prostaglandin E (PGE) in ovariectomized 90-day pregnant ewes

The objective of this experiment was to determine the effect of mifepristone, a progesterone receptor antagonist, on pregnancy and secretion of steroids, pregnancy-specific protein B (PSPB) and prostaglandins at mid-pregnancy in ewes. Ninety-day pregnant ewes were ovariectomized (OVX) and treatments were initiated 72 h post-OVX. Ewes received (1) vehicle, (2) prostaglandin F2alpha (PGF2alpha, 8 mg/58 kg/bw, i.m.) 84 h post-OVX, (3) mifepristone (50 mg intrajugular at 72, 84, 96, and 108 h post-OVX), (4) mifepristone (50mg) + PGF2alpha, (5) mifepristone (100 mg intrajugular at 72, 84, 96, and 108 h), and (6) mifepristone (100 mg) + PGF2alpha. Ewes treated with vehicle or PGF2alpha alone did not abort (P > or = 0.05). But, 60, 80, 60, and 100% of ewes treated with mifepristone (50 mg), mifepristone (50 mg) + PGF2alpha, mifepristone (100 mg), and mifepristone (100 mg) + PGF2alpha, respectively, aborted (P < or = 0.05). Profiles of progesterone, estradiol-17beta, prostaglandin E (PGE), or PSPB did not differ (P > or = 0.05) among treatment groups. Profiles of PGF2alpha of treatment groups receiving mifepristone with or without PGF2alpha differed (P < 0.05) from vehicle or PGF2alpha alone-treated ewes. It is concluded that progesterone actions are necessary to suppress uterine/placental secretion of PGF2alpha and that maintenance of critical progesterone: estradiol-17beta and PGE:PGF2alpha ratios are necessary for maintenance of pregnancy.     

The effects of prostaglandin A1 and prostaglandin B1 on the differentiation of cartilage in the chick embryo

Summary In organ cultures of chick embryonic limb rudiments the mean length of explants treated with 25g/ml prostaglandin B₁ (PGB₁) was significantly smaller than that of paired controls (P<0.001) after 4, 6 and 8 days in vitro. The deceleration of linear growth was constant during 8 days in vitro. Growth inhibition was confirmed by a statistically significant decrease in explant dry weight after 8 days of culture. However, PGB₁ caused no observable alteration in the histological structure of the explants. The possible role of PGB₁ in the physiological control of cartilage growth is postulated. Explants similarly treated with prostaglandin A₁ (PGA₁) at concentrations of 15 g/ml for 8 days or 20g/ml for 4 and 8 days exhibited comma and inverted commas phenomena, caused by the intermingling of chondroblasts from the epiphyseal and flattened-cell zones, which thus ceased to be distinct entities. Adenylate cyclase in the plasma membrane may be involved in this disturbance of cartilage differentiation.     

A study of prostaglandin F2alpha as the luteolysin in swine: III effects of estradiol valerate on prostaglandin F, progestins, estrone and estradiol concentrations in the utero-ovarian vein of nonpregnant gilts

Polyvinyl catheters were placed into the right and left utero-ovarian veins and saphenous vein and artery of three control (C) and four estradiol valerate (EV) treated gilts on Day 9 after onset of estrus. The EV treated gilts received 5mg EV/day on Days 11 through 15 after onset of estrus. On Days 12 through 17 utero-ovarian vein blood samples were collected at 15 min intervals from 0700 to 1000 hr and 1900 to 2200 hr and single samples were taken at 1100 and 2300 hr. Peripheral blood samples (saphenous vein or artery) were taken at 0700, 1100, 1900 and 2300 hr from Day 12 until the control gilts returned to estrus or until Day 25 for EV treated gilts and used to measure plasma steroid hormone concentrations. Utero-ovarian vein prostaglandin F (gf) concentrations (ng/ml, n-1,177) were measured by RIA. Status (control vs EV treated gilts) by day interactions were detected (P=.10). Curvilinear day trends were detected for plasma PGF concentrations in control (P less than .01) but not EV treated gilts. PGF concentrations (X +/- S.D.) for control and EV treated gilts were 1.20 +/- 2.08 and .26 +/- .84 ng/ml, respectively. PGF peaks (concentrations greater than X + 2 S.D.) occurred with greater frequency in control gilts (X2 =4.87; P less than .05). The interestrus interval (X +/- S.E.) for control and treated gilts was 19.0 +/- .6 and 146.5 +/- 74.8 days, respectively. Data indicate tht t estradiol valerate may exert its luteotrophic effect by preventing PGF release from the uterus.     

Blazeispirols B, C, E and F, des-A-ergostane-type compounds, from the cultured mycelia of the fungus Agaricus blazei

Four new des-A-ergostane derivatives including blazeispirols B, C, E and F were isolated from the cultured mycelia of fungus Agaricus blazei Murill and were established to be (20S, 22R, 23R, 24S)-14beta,22: 22,25-diepoxy-5-methoxy-des-A-ergosta-5,7,9,11-tetraen-23-ol; (20S, 22S, 23R, 24S)-14beta,22: 22,25-diepoxy-5-methoxy-des-A-ergosta-5,7,9-trien-23-ol; (20S, 22S, 23R, 24S)-14beta, 22: 22, 25-diepoxy-5-methoxy-des-A-ergosta-5,7,9,11-tetraene-19,23-diol and (20S, 22S, 23R, 24S)-14beta,22: 22,25-diepoxy-des-A-ergosta-5,7,9-triene-5,23-diol by comparison of extensive 1D and 2D NMR spectral data with that of blazeispirol A.     

Effects of collagen, ionophore A23187 and prostaglandin E1 on the phosphorylation of specific proteins in blood platelets

Human platelets that had been preincubated with 5-hydroxy[(3)H]tryptamine and [(32)P]P(i) were stirred with various agents; the secretion of 5-hydroxy[(3)H]tryptamine from platelet granules and the radioactivity of platelet [(32)P]phosphopolypeptides separated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis were then measured. Exposure of the platelets to collagen fibres or ionophore A23187 selectively increased the phosphorylation of polypeptides with apparent mol.wts. of 47000 (P47) and 20000 (P20) by approx. 3-fold, in association with the release of 5-hydroxy[(3)H]tryptamine. The 47000-mol.wt. phosphopolypeptide (P47) was clearly separated from platelet actin by the electrophoresis system used. Prostaglandin E(1), which inhibits platelet function by increasing platelet cyclic AMP, decreased the phosphorylation of polypeptides caused by collagen as well as the release of 5-hydroxy[(3)H]tryptamine. Prostaglandin E(1) also selectively increased the phosphorylation of distinct polypeptides with apparent mol.wts. of 24000 (P24) and 22000 (P22) by approx. 2-fold. As the phosphorylation reactions caused by collagen are probably mediated by an increase in Ca(2+) concentration in the platelet cytosol and may have a role in the release reaction [Haslam & Lynham (1977) Biochem. Biophys. Res. Commun.77, 714-722; (1978) Thromb. Res.12, 619-628], we suggest that a cyclic AMP-dependent phosphorylation of the 24000- and/or 22000-mol.wt. polypeptides caused by prostaglandin E(1) may initiate processes that decrease the Ca(2+) concentration in the cytosol, so inhibiting both the Ca(2+)-dependent phosphorylation reactions and the release reaction. Treatment of platelets with prostaglandin E(1) did not inhibit the increased phosphorylation of polypeptides with apparent mol.wts. of 47000 and 20000 (P47 and P20) caused by ionophore A23187, which may therefore short-circuit cyclic AMP-dependent mechanisms that decrease the Ca(2+) concentration in the platelet cytosol. As prostaglandin E(1) did inhibit the release of 5-hydroxy[(3)H]tryptamine by ionophore A23187, cyclic AMP may also inhibit the release reaction by additional mechanisms.     

Total synthesis of (+)-macrosphelides A,C, E,F, and G based on enzymatic function

The total synthesis of (+)-macrosphelide A (1) (18.5% overall yield in 11 steps), (+)-macrosphelide C (2) (25% overall yield in 9 steps), (+)-macrosphelide E (3) (23.9% overall yield in 11 steps), (+)-macrosphelide F (4) (20% overall yield in 9 steps), and (+)-macrosphelide G (5) (22% overall yield in 9 steps) was achieved from a chemoenzymatic reaction product (4R,5S)-4-benzyloxy-5-hydroxy-2(E)-hexenoate 10.