Prostaglandin F2α Production Research Articles

Effects of Aspirin Eugenol Ester on Liver Oxidative Damage and Energy Metabolism in Immune-Stressed Broilers.

The aim of this study was to investigate the effects of aspirin eugenol ester (AEE) on liver oxidative damage and energy metabolism in immune-stressed broilers. In total, 312 broilers were divided into 4 groups (saline, LPS, SAEE, and LAEE). Broilers in the saline and LPS groups were fed a basal diet; the SAEE and LAEE groups had an added 0.01% AEE in their diet. Broilers in the LPS and LAEE groups were injected with lipopolysaccharides, while the saline and SAEE groups were injected with saline. Results showed that AEE increased the body weight, average daily gain, and average daily feed intake, as well as decreasing the feed conversion ratio of immune-stressed broilers. AEE protects against oxidative damage in immune-stressed broiler livers by elevating the total antioxidant capacity, superoxide dismutase activity, and glutathione S-transferase alpha 3 (GSTA3) and glutaredoxin 2 (GLRX2) expression, while decreasing malondialdehyde content. AEE lessened inflammation by reducing prostaglandin-F2α production and prostaglandin-endoperoxide synthase 2 (PTGS2) and interleukin-1beta (IL-1β) expression. AEE decreased oxidative phosphorylation rates by increasing succinic acid levels and lowering both adenosine diphosphate (ADP) levels and ceroid lipofuscinosis neuronal 5 (CLN5) expression. AEE modulated the metabolism of phenylalanine, tyrosine, lipids, and cholesterol by reducing the phenyllactate and L-arogenate levels, lowering dopachrome tautomerase (DCT) and apolipoprotein A4 (APOA4) expression, and increasing phenylpyruvic acid and dopa decarboxylase (DDC) expression. In summary, AEE can effectively alleviate liver oxidative damage and energy metabolism disorders in immune-stressed broilers.

Inducing cyclooxygenase-2 expression, prostaglandin E2 and prostaglandin F2α production of human dental pulp cells by activation of toll-like receptor-3, mitogen-activated protein kinase kinase/extracellular signal-regulated kinase and p38 signaling

Background/purposeBacterial infection was the major etiology for pulpal/root canal infection. This study aimed to investigate the activation of toll-like receptor-3 (TLR) on cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) and PGF2α production of human dental pulp cells (HDPCs) and associated signaling. Materials and methodsHDPCs were exposed to different concentrations of Poly (I:C) (a TLR3 activator). Cell viability was determined by 3- (4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) assay and alkaline phosphatase (ALP) activity was evaluated by ALP staining. Activation of extracellular signal-regulated kinase (ERK) and p38 by Poly (I:C) was determined by immunofluorescent staining. The COX-2 protein expression was analyzed by Western blot. PGE2 and PGF2α production was measured by enzyme-linked immunosorbent assay. The mRNA expression was studied by real-time polymerase-chain reaction. Moreover, HDPCs were exposed to Poly(I:C) with/without U0126 or SB203580 treatment and analysis of COX-2 expression and prostanoid production were conducted. ResultsPoly (I:C) showed little effect on ALP activity, but decreased viability of HDPCs. It stimulated COX-2 mRNA and protein expression. Poly (I:C) induced PGE2 and PGF2α production of HDPCs. Poly (I:C) activated p-ERK, and p-p38 protein expression. Treatment by U0126 (a mitogen-activated protein kinase kinase (MEK)/ERK inhibitor) and SB203580 (a p38 inhibitor) attenuated Poly (I:C)-induced COX-2 mRNA and protein expression as well as PGE2 and PGF2α production. ConclusionTLR3 activation is involved in the infection and inflammatory responses of pulp tissues, via MEK/ERK, and p38 signaling to mediate COX-2 expression as well as PGE2 and PGF2α production, contributing to the pathogenesis and progression of pulpal/periapical diseases.

Myeloid Bmal1 deletion suppresses the house dust mite-induced chronic lung allergy.

Asthma is the chronic pulmonary inflammatory response that could lead to respiratory failure when allergic reactions exacerbate. It is featured by type 2 immunity with eosinophilic inflammation, mucus, and IgE production, and Th2 cytokine secretion upon repeated challenge of allergens. The symptom severity of asthma displays an apparent circadian rhythm with aggravated airway resistance in the early morning in patients. Bmal1 is the core regulator of the circadian clock, while the regulatory role of Bmal1 in asthma remains unclear. Here, we investigate whether the myeloid Bmal1 is involved in the pathogenesis of house dust mite (HDM)-induced lung allergy. We found that knockdown of Bmal1 in macrophages suppressed the time-of-day variance of the eosinophil infiltration in the alveolar spaces in chronic asthmatic mice. This was accompanied by decreased bronchial mucus production, collagen deposition, and HDM-specific IgE production. However, the suppression effects of myeloid Bmal1 deletion did not alter the allergic responses in short-term exposure to HDM. The transcriptome profile of alveolar macrophages (AMs) showed that Bmal1-deficient AMs have enhanced phagocytosis and reduced production of allergy-mediating prostanoids thromboxane A2 and prostaglandin F2α synthesis. The attenuated thromboxane A2 and prostaglandin F2α may lead to less induction of the eosinophil chemokine Ccl11 expression in bronchial epithelial cells. In summary, our study demonstrates that Bmal1 ablation in macrophages attenuates eosinophilic inflammation in HDM-induced chronic lung allergy, which involves enhanced phagocytosis and reduced prostanoid secretion.

Biochemical characterization of the cyclooxygenase enzyme in penaeid shrimp.

Cyclooxygenase (COX) is a two-step enzyme that converts arachidonic acid into prostaglandin H2, a labile intermediate used in the production of prostaglandin E2 (PGE2) and prostaglandin F2α (PGF2α). In vertebrates and corals, COX must be N-glycosylated on at least two asparagine residues in the N-(X)-S/T motif to be catalytically active. Although COX glycosylation requirement is well-characterized in many species, whether crustacean COXs require N-glycosylation for their enzymatic function have not been investigated. In this study, a 1,842-base pair cox gene was obtained from ovarian cDNA of the black tiger shrimp Penaeus monodon. Sequence analysis revealed that essential catalytic residues and putative catalytic domains of P. monodon COX (PmCOX) were well-conserved in relation to other vertebrate and crustacean COXs. Expression of PmCOX in 293T cells increased levels of secreted PGE2 and PGF2α up to 60- and 77-fold, respectively, compared to control cells. Incubation of purified PmCOX with endoglycosidase H, which cleaves oligosaccharides from N-linked glycoproteins, reduced the molecular mass of PmCOX. Similarly, addition of tunicamycin, which inhibits N-linked glycosylation, in PmCOX-expressing cells resulted in PmCOX protein with lower molecular mass than those obtained from untreated cells, suggesting that PmCOX was N-glycosylated. Three potential glycosylation sites of PmCOX were identified at N79, N170 and N424. Mutational analysis revealed that although all three residues were glycosylated, only mutations at N170 and N424 completely abolished catalytic function. Inhibition of COX activity by ibuprofen treatment also decreased the levels of PGE2 in shrimp haemolymph. This study not only establishes the presence of the COX enzyme in penaeid shrimp, but also reveals that N-glycosylation sites are highly conserved and required for COX function in crustaceans.

Prostaglandin F2α-PTGFR signaling promotes proliferation of endometrial epithelial cells of cattle through cell cycle regulation

There is production of prostaglandin F2α (PGF2α) and there is PGF2α receptor (PTGFR) mRNA transcript in endometrial epithelial cells of cattle. The aims of the present study were to (1) determine whether PGF2α-PTGFR signaling modulates the proliferation of endometrial epithelial cells and (2) increase knowledge of PGF2α-PTGFR signaling on the physiological and pharmacological processes in the endometrium of cattle. Amount of cellular proliferation was determined using real-time cell analysis and cell proliferation reagent WST-1 procedures. Abundance of cyclins, cyclin-dependent kinases (CDKs), cyclin-kinase inhibitors, proliferating cell nuclear antigen (PCNA), cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), PTGFR, epidermal growth factor (EGF) mRNA and protein abundances were evaluated using real-time RT-PCR and western blot analyses. The PGF2α-PTGFR signaling promoted the proliferation of endometrial epithelial cells by inducing changes in abundance of mRNA transcript and protein that resulted in an increase in the abundance for the cyclins (A, B1, D1, D3), CDKs (1, 2, 4, 6), and PCNA; decrease in abundance for p21; and increase in abundance for EGF, COX-1, COX-2, and PTGFR. There was a direct molecular association between PGF2α-PTGFR signaling and cell cycle regulation in endometrial epithelial cells of cattle. In addition, findings improve the understanding of PGF2α-PTGFR signaling in the physiological and pharmacological processes of the endometrium of cattle.

Functionally Selective Inhibition of the Oxytocin Receptor by Retosiban in Human Myometrial Smooth Muscle.

Novel small molecule inhibitors of the oxytocin receptor (OTR) may have distinct pharmacology and mode of action when compared with first-generation oxytocin antagonists when used for the prevention of preterm birth. The aim was to determine the mechanism of action of small molecule OTR antagonists retosiban and epelsiban compared with the currently used peptide-based compound atosiban. Human myometrial samples were obtained at cesarean section and subjected to pharmacological manipulations to establish the effect of antagonist binding to OTR on downstream signaling. Retosiban antagonism of oxytocin action in human myometrium was potent, rapid, and reversible. Inhibition of inositol 1,4,5-trisphosphate (IP3) production followed single-site competitive binding kinetics for epelsiban, retosiban, and atosiban. Retosiban inhibited basal production of IP3 in the absence of oxytocin. Oxytocin and atosiban but not retosiban inhibited forskolin, and calcitonin stimulated 3',5'-cyclic adenosine 5'-mono-phosphate (cAMP) production. Inhibition of cAMP was reversed by pertussis toxin. Oxytocin and atosiban, but not retosiban and epelsiban, stimulated extracellular regulated kinase (ERK)1/2 activity in a time- and concentration-dependent manner. Oxytocin and atosiban stimulated cyclo-oxygenase 2 activity and subsequent production of prostaglandin E2 and F2α. Prostaglandin production was inhibited by rofecoxib, pertussin toxin, and ERK inhibitor U0126. Oxytocin but not retosiban or atosiban stimulated coupling of the OTR to Gα q G-proteins. Oxytocin and atosiban but not retosiban stimulated coupling of the OTR to Gα i G-proteins. Retosiban and epelsiban demonstrate distinct pharmacology when compared with atosiban in human myometrial smooth muscle. Atosiban displays agonist activity at micromolar concentrations leading to stimulation of prostaglandin production.

Non-Coding RNA Sequencing of Equine Endometrium During Maternal Recognition of Pregnancy.

Maternal recognition of pregnancy (MRP) in the mare is not well defined. In a non-pregnant mare, prostaglandin F2α (PGF) is released on day 14 post-ovulation (PO) to cause luteal regression, resulting in loss of progesterone production. Equine MRP occurs prior to day 14 to halt PGF production. Studies have failed to identify a gene candidate for MRP, so attention has turned to small, non-coding RNAs. The objective of this study was to evaluate small RNA (<200 nucleotides) content in endometrium during MRP. Mares were used in a cross-over design with each having a pregnant and non-mated cycle. Each mare was randomly assigned to collection day 11 or 13 PO (n = 3/day) and endometrial biopsies were obtained. Total RNA was isolated and sequencing libraries were prepared using a small RNA library preparation kit and sequenced on a HiSeq 2000. EquCab3 was used as the reference genome and DESeq2 was used for statistical analysis. On day 11, 419 ncRNAs, representing miRNA, snRNA, snoRNA, scaRNA, and vaultRNA, were different between pregnancy statuses, but none on day 13. Equine endometrial ncRNAs with unknown structure and function were also identified. This study is the first to describe ncRNA transcriptome in equine endometrium. Identifying targets of these ncRNAs could lead to determining MRP.

Neurobehavioral Effects of β-Escin

Background: Aescin or β-escin is the main and active constituent of horse chestnut seed (Aesculus hippocastanum) used for the treatment of inflammatory edema, venous insufficiency and ischemic ulcerations. Aescin has many actions due to induction of endothelial nitric oxide and prostaglandin F2-α production moreover; aescin antagonizes the effect of histamine and 5HT at receptor levels. &lt;/P&gt;&lt;P&gt; Objective: The aim of the present study was to evaluate the neurobehavioral effects of aescin on normal healthy volunteers. &lt;/P&gt;&lt;P&gt; Method: A total number of 65 healthy participants with mean age of 21+1.1 years were recruited to study the effects of aescin on the neurobehavioral effects of normal healthy volunteers compared to placebo. The neurobehavioral effects were assessed by psychomotor performances and sensorimotor reaction, cortical arousal and central integrity processes and assessment of memory capacity. Results: Placebo produced insignificant amelioration of TRT and RRT p&gt;0.05, with mild significant effect on MRT p=0.03. Aescin produced a significant effect in the amelioration of psychomotor performances and sensorimotor reaction p=0.0001. Regarding the differential effect of placebo and aescin on the cortical arousal and central integrity processes, placebo illustrated insignificant effect at p&gt;0.05 whereas; aescin showed mild significant effect on Critical Fusion Frequency (CFFA) p&lt;0.05 and highly significant effect on the other parameters p&lt;0.01 except for critical-fusion frequency threshold when aescin illustrated insignificant effect at p&gt;0.05. Aescin illustrated significant acceleration of ІІ-back WMA, ІІІ-back WMA and Second trial Short-Term Memory (STM) at p&lt;0.01 compared to the placebo effect. Conclusion: Short-term therapy with aescin improves the neurobehavioral effects on healthy volunteers.

Endothelial cell transient receptor potential channel C5 (TRPC5) is essential for endothelium-dependent contraction in mouse carotid arteries

Augmented endothelium-dependent contractions (EDC) contributes to endothelial dysfunction and vascular disease progression. An early signal in EDC is cytosolic [Ca2+]i rise in endothelial cells, which stimulates the production of contractile prostanoids, leading to vascular contraction. In this study, the molecular identity of Ca2+-permeable channels in endothelial cells and its function were investigated. Vascular tension was measured by wire myograph. EDCs were elicited by acetylcholine (ACH) in the presence of NG-nitro-l-arginine methyl ester (L-NAME). [Ca2+]i was measured using a Ca2+-sensitive fluorescence dye. Enzyme Immunoassay (EIA) was used for prostaglandin measurement. Immunohistochemical staining found the expression of transient receptor potential channel C5 (TRPC5) in endothelial and smooth muscle cells of mouse carotid arteries. ACH-induced EDC in male mouse carotid arteries was found to be substantially reduced in TRPC5 knockout (KO) mice than in wild-type (WT) mice. TRPC5 inhibitors clemizole and ML204 also reduced the EDC. Furthermore, ACH-induced Ca2+ entry in endothelial cells was lower in TRPC5 KO mice than in WT mice. Moreover, the EDC was abolished by a cyclooxygenase-2 (COX-2) inhibitor NS-398, but not affected by a COX-1 inhibitor valeryl salicylate (VAS). Enzyme immunoassay results showed that TRPC5 stimulated the COX-2-linked production of prostaglandin F2α (PGF2α), prostaglandin E2 (PGE2), and prostaglandin D2 (PGD2). Exogeneous PGF2α, PGE2, and PGD2 could induce contractions in carotid arteries. Our present study demonstrated that TRPC5 in endothelial cells contributes to EDC by stimulating the production of COX-2-linked prostanoids. The finding extends our knowledge about EDC.

86 Difficulty of Transfer of In Vivo-Derived Bovine Embryos and Route of Administration of Flunixin Meglumine at the Time of Transfer may Affect Pregnancy Rate

Recipient handling during embryo transfer (ET) induces prostaglandin F2α (PGF2α) production in 2 periods: an early transient and rapid increase around the time of ET, followed by another 2 to 4 h later. This PGF2α is associated with embryonic loss during early gestation by affecting both the embryo and the corpus luteum. To control this, antiprostaglandins such as flunixin meglumine (FM) have been applied IM at the time of ET with varying results. In such studies, the interaction of IM administration of FM and difficulty of transfer has not always been evaluated, possibly confusing the interpretation of the results. Furthermore, IV FM injection at ET and its relationship with pregnancy rates (PR) has not been determined. The objectives were (1) to determine the relationship between difficulty of ET and PR; and (2) to evaluate the efficacy of IM v. IV FM on pregnancy outcomes. One hundred and ten crossbred (Bos taurus × Bos indicus) heifers (18-24 months old) from 3 farms were used as recipients. Two evaluation systems of ET difficulty were used: (1) duration of transfer (objective determination of the elapsed time measured in seconds between the introduction of the catheter and embryo release), and (2) level of difficulty experienced by the practitioner (subjective determination; 1 = minimum and 2 = medium to extreme manipulation). Quality 1 and 2 fresh embryos from superovulated cows were transferred by the same practitioner. At ET, recipients were randomly divided into 3 groups: (1) Control (no treatment, n = 31); (2) FM-IM (n = 39): injected IM with 2.2 mg kg−1 FM at ET; and (3) FM-IV (N = 40): injected with 2.2 mg kg−1 FM IV at ET. Pregnancy was diagnosed at 30 to 40 and 60 to 90 days after ET. Spearman’s test was performed to determine the correlation between duration and difficulty at ET and Chi-square test was used to compare PR. The mean duration of transfer for all heifers was 62.3 ± 57.5 s (11 to 357 s; median: 44.5 s). There was a high correlation (0.8; P &lt; 0.001) between the ET difficulty evaluation systems. Overall, ET difficulty 1 had higher PR than ET difficulty 2 (64.2 v. 40.7; P = 0.013). The PR was significantly improved (P &lt; 0.01) in the FM-IV group (75 and 70% at 30 and 60 days after ET) compared with control (45.2 and 32.3%) and FM-IM (33.3 and 30.7%). In conclusion, results indicate that the difficulty of transfer affects PR achieved following the transfer of in vivo-derived bovine embryos. Treatment with FM-IV following transfer resulted in significantly higher PR compared with control and FM-IM recipients. The IV injection of FM may antagonize the very early and transient increase of PGF2α caused by genital tract manipulation (even gently performed) at embryo transfer. Further research is necessary to confirm the results of the present study.

Профілактика ембріональної смертності у корів

The article analyzes the scientific data and own research on study and formulation the preventive efficacy of ketoprofen during embryonic mortality in cows for estrus synchronization, depending on the duration of the period after birth. It is known that in cows at spontaneous recovery of sexual cyclicity after calving for its next synchronization form smaller corpus luteum, although their fertilization is higher than in animals without sexual cyclical recovery after giving birth to synchronize estrus. The application for cows with small yellow bodies of NSAIDs inhibit the production of prostaglandin F2α by uterine lining through blocking Cyclogenesis of type II. Stimulation and sexual cyclical synchronization was performed using surfagon and estrofan and at the 10th day cows were inseminated. Animals of the experimental subgroups at the 11th day after insemination were injected in therapeutic dose NSAIDs ainil. The prophylactic use of ainil towards embryonic mortality after insemination of cows for previous estrus synchronization promotes animal insemination by average of 7.3%. Such effect of NSAIDs on cows fertilization depending on the period after calving can point to the prevention of fetal death in animals with restored sex cycles after calving regardless the cause of the formation of the corpus luteum at spontaneous manifestation of the sexual cycle or estrus synchronization.

Pseudomonas aeruginosa produces aspirin insensitive eicosanoids and contributes to the eicosanoid profile of polymicrobial biofilms with Candida albicans

The interaction of clinically relevant microorganisms is the focus of various studies, e.g. the interaction between the pathogenic yeast, Candida albicans, and the bacterium, Pseudomonas aeruginosa. During infection both release arachidonic acid, which they can transform into eicosanoids. This study evaluated the production of prostaglandin E2, prostaglandin F2α and 15-hydroxyeicosatetraenoic acid by biofilms of P. aeruginosa and C. albicans. The influence of co-incubation, acetylsalicylic acid and nordihydroguaiaretic acid on biofilm formation and eicosanoid production was evaluated. Acetylsalicylic acid decreased colony forming units of P. aeruginosa, but increased metabolic activity and eicosanoid production of the cells. In contrast to prostaglandin E2, prostaglandin F2a production by C. albicans was insensitive to acetylsalicylic acid, indicating that different enzymes are responsible for their production in this yeast. Nordihydroguaiaretic acid inhibited biofilm formation by P. aeruginosa, however co-incubation provided protection against this inhibitor. Production of these eicosanoids could affect pathogen-clearance and infection dynamics and this previously uncharacterized facet of interaction could facilitate novel therapeutic intervention against polymicrobial infection.

A role for the endocannabinoid system in premature luteal regression and progesterone withdrawal in lipopolysaccharide-induced early pregnancy loss model.

What is the role of the endocannabinoid system (eCS) in the alterations of the endocrine system in a murine model of lipopolysaccharide (LPS)-induced miscarriage? In 7-days pregnant wild type, but not cannabinoid receptor type 1 knockout (CB1-KO) mice, LPS increased COX-2 expression and prostaglandin F2α (PGF2α) production in the uterus leading to lower expression of prolactin receptor in the ovary and a marked regression of corpora lutea (CL), suggesting that the eCS mediates the deleterious effects of LPS on reproductive events. Appropriate systemic progesterone levels are critical for a successful pregnancy outcome. Precocious loss of luteal progesterone (P4) secretion leads to miscarriage in rodents. We have previously shown that LPS administration to pregnant mice induces embryonic resorption accompanied by a dramatic decrease in systemic progesterone levels in a murine model of inflammatory miscarriage, with the eCS mediating these LPS-induced deleterious effects. CD1 wild-type (WT) and CB1-KO mice were randomly allocated to Vehicle (saline; i.p.) or LPS (0.5 μg/g body weight; i.p.) treated groups: (WT-Vehicle; WT-LPS; CB1-KO-Vehicle and CB1-KO-LPS). A single injection was given on day 7 of pregnancy and tissues (blood, ovary, uterus) were collected 6, 12, 24 and 48 h later. P4 and PGF2α plasma levels were determined by radioimmunoassay. Cyclooxygenase-2 (COX-2) mRNA (RT-PCR) and protein (Western blot) content in uterus was assayed. COX-2 and prolactin receptor (PrlR) mRNA levels in the ovary were assayed by RT-PCR. Tissue morphology of the CL was assessed by haematoxylin-eosin staining. Treatment of 7-day pregnant WT mice with LPS induced a P4 withdrawal (p<0.05), increased in uterine COX-2 mRNA and protein expression (p<0.05) as well as an increase in uterine PGF2α production (p<0.05). These changes were absent in LPS-treated 7-day pregnant CB1-KO mice. In ovarian tissues, LPS treatment to 7-day pregnant WT mice induced a downregulation of PrlR mRNA expression (p<0.05) together with an increase in COX-2 mRNA expression (p<0.05) and PGF2α content (p<0.05). These effects were absent in the CB1-KO mice. Collectively, our results suggest a role for the eCS mediating LPS-induced deleterious effects on reproductive tissues. An important caveat of this study is the endocrine differences between mice and humans during pregnancy (e.g. P4 is produced by the CL throughout pregnancy in mice, whereas this is not the case in humans), which limits the extrapolation of the results presented here. Our findings provide new insights in the role of the endocannabinoid system in the physiopathology of reproduction as well as the role of this endogenous system as a mediator of LPS deleterious effects on reproductive tissues. None. Dr Ana María Franchi was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2010/0813 and PICT 2013/0097) and by Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2012/0061). The authors have no competing interests.

PPAR ligand association with prostaglandin F2α and E2 synthesis in the pig corpus luteum—An in vitro study

The present study evaluated the involvement of PPARs in prostaglandin (PG) E2 and F2α production in the corpus luteum (CL) of pigs on days 10–12 and 14–16 of the estrous cycle or pregnancy. The tissue explants were incubated for 6h in the presence of PPARα, PPARβ, PPARγ ligands. The concentration of PGs in the incubation media was determined by radioimmunoassay, while mRNA abundance of PG synthetases (PGES and PGFS) was analyzed by quantitative real-time PCR. It was found that L-165,045 and rosiglitazone stimulated PGES synthesis on days 10–12 of the estrous cycle, whereas all factors that were assessed did not affect PGE2 release. The PGFS mRNA abundance in the CL did not change in the presence of PPAR ligands during the assessment periods. However, PPARβ agonist inhibited PGF2α secretion on days 10–12 of the estrous cycle and on days 14–16 of pregnancy. Interestingly, PPAR antagonists, MK 886, GW 9662 or T0070907 decreased PGF2α release by the slices on days 10–12 of the estrous cycle. It is concluded that the CL has a different susceptibility (greatest during mid-luteal phase of the estrous cycle) to the PPAR ligands, which is related to the physiological status of animal. The inhibition of PGF2α release and augmentation of PGES mRNA concentration during mid-luteal phase of the estrous cycle might suggest luteotropic properties of PPAR ligands.

Possibility of Diagnosing Uterine Function in Cows

Declining in reproductive efficiency is a worldwide problem, although remarkable progress has been made in reproductive techniques including artificial insemination programs, estrous cycle synchronization, and effective embryo production. One means of solving this problem is to select breeding cows based on highly precise diagnosis for high reproductive ability, including ovarian and uterine function. Many recent studies have documented the regulation of uterine function, with the results suggesting that said functions might be an effective diagnostic index for reproductive performance. Therefore, the production of oxytocin-induced prostaglandin F2α and epidermal growth factor in bovine endometrium, such as the physiological reaction in vivo, may be useful in detecting repeat breeder cows. Although the diagnosis is possible using relatively simple techniques such as biopsy, culture, and hormone assay, it is difficult to apply such diagnosis to the field conditions. However, further improvements in the exhaustive gene expression analysis of uterine function in repeat breeder or low fertility cows, so as to determine the genes associated with physiological reactions, could lead to new diagnostic indexes resulting in improved reproductive efficiency. Discipline: Animal Industry Additional key words: endometrial biopsy, epidermal growth factor, oxytocin, prostaglandin F2α

Dietary Compound Kaempferol Inhibits Airway Thickening Induced by Allergic Reaction in a Bovine Serum Albumin-Induced Model of Asthma.

Asthma is characterized by aberrant airways including epithelial thickening, goblet cell hyperplasia, and smooth muscle hypertrophy within the airway wall. The current study examined whether kaempferol inhibited mast cell degranulation and prostaglandin (PG) release leading to the development of aberrant airways, using an in vitro model of dinitrophenylated bovine serum albumin (DNP-BSA)-sensitized rat basophilic leukemia (RBL-2H3) mast cells and an in vivo model of BSA-challenged asthmatic mice. Nontoxic kaempferol at 10–20 μM suppressed β-hexosaminidase release and cyclooxygenase 2 (COX2)-mediated production of prostaglandin D2 (PGD2) and prostaglandin F2α (PGF2α) in sensitized mast cells. Oral administration of ≤20 mg/kg kaempferol blocked bovine serum albumin (BSA) inhalation-induced epithelial cell excrescence and smooth muscle hypertrophy by attenuating the induction of COX2 and the formation of PGD2 and PGF2α, together with reducing the anti-α-smooth muscle actin (α-SMA) expression in mouse airways. Kaempferol deterred the antigen-induced mast cell activation of cytosolic phospholipase A2 (cPLA2) responsive to protein kinase Cμ (PKCμ) and extracellular signal-regulated kinase (ERK). Furthermore, the antigen-challenged activation of Syk-phospholipase Cγ (PLCγ) pathway was dampened in kaempferol-supplemented mast cells. These results demonstrated that kaempferol inhibited airway wall thickening through disturbing Syk-PLCγ signaling and PKCμ-ERK-cPLA2-COX2 signaling in antigen-exposed mast cells. Thus, kaempferol may be a potent anti-allergic compound targeting allergic asthma typical of airway hyperplasia and hypertrophy.

Genomic and non-genomic effects of progesterone on prostaglandin (PG) F2? and PGE2 production in the bovine endometrium.

Progesterone (P4) acts through different actuating pathways called genomic and non-genomic pathways. Here we investigated whether P4 regulates prostaglandin (PG) F2? (PGF) and PGE2 production in bovine endometrium through different pathways. Cultured endometrial cells were exposed to P4 for a short time (5-20min) or bovine serum albumin (BSA)-conjugated P4 (P4-BSA) for 24h. Progesterone treatment for 24h stimulated PGE2 production in epithelial cells, but suppressed both PGF and PGE2 production and the expression of PG-metabolising enzymes including phospholipase A2 (PLA2) and cyclooxygenase-2 (COX2) in stromal cells. Short-term (5-20min) P4 treatment did not affect PLA2 or COX2 transcript levels in either cell type. P4-BSA increased PGF and PGE2 production only in epithelial cells. Nuclear P4 receptor mRNA expression in endometrium was higher at the follicular phase than at the early- to mid-luteal stages, whereas membrane P4 receptor mRNA expression did not change throughout the oestrous cycle. The overall results suggest that P4 controls PG production by inhibiting enzymes via a genomic pathway and by stimulating signal transduction via a non-genomic pathway. Consequently, P4 may protect the corpus luteum by attenuating PGF production in stromal cells and by increasing PGE2 secretion from epithelial cells.

107 PRODUCTION OF EPIDERMAL GROWTH FACTOR BY BOVINE ENDOMETRIAL CELLS AND ITS EFFECTS ON EMBRYONIC INTERFERONT AND PROSTAGLANDIN

A dynamic interaction between bioactive products of the embryo (blastocyst) and the endometrium is crucial for the successful establishment of pregnancy. In ruminants, the principal signal for maternal recognition of pregnancy is interferon-τ (IFNT) secreted by the trophoectoderm between Days 8 and 20 post-fertilization. Epidermal growth factor (EGF) produced by the endometrium acting through EGF receptors (EGFR) present in the blastocyst seems to regulate embryonic production of IFNT. Epidermal growth factor and IFNT have been shown to play crucial roles in controlling luteolytic prostaglandin (PG) F2α (PGF) and luteotropic PGE2 production by bovine endometrium. However, it is unknown how these bioactive molecules regulate uterine function during maternal recognition of pregnancy. To clarify the main source of EGF in bovine endometrium and the mechanisms regulating the interaction between the hatched blastocyst and maternal uterine environment, the production of EGF by cultured endometrial epithelial and stromal cells and the effects of EGF on embryonic IFNT and PG were investigated. In addition, the effects of EGF on PGE2 and PGF production by cultured epithelial or stromal cells were examined. Endometrial epithelial and stromal cells were enzymatically isolated on the day of ovulation, seeded at a density of 100 000 viable cells mL–1, and cultured at 38°C in a humidified atmosphere of 5% CO2 in air. After the cells reached 90% confluence, they were cultured in the presence or absence of EGF (0.1, 1.0, 10, and 100 ng mL–1) for 24 h. Cells cultured in the absence of EGF and their cultured media were collected separately for protein analysis. Hatched bovine blastocysts (Days 8–10) were also cultured and exposed to EGF (1, 10, and 100 ng mL–1) for 24 h. Protein concentrations of EGF and IFNT in the cultured media were determined by commercial enzyme immunoassay kit. Hormonal concentrations were analysed by ANOVA followed by Fisher's protected least-significant difference procedure (PLSD) as a multiple comparison test by StatView (Abacus Concepts Inc., Berkeley, CA, USA). The concentration of EGF in the culture media of epithelial cells cultured in the absence of EGF was significantly (P &lt; 0.05) higher than in the cultured media of endometrial stromal cells. Epidermal growth factor (10 and 100 ng mL–1) increased embryonic production of IFNT and luteotropic PGE2 production but not luteolytic PGF by hatched blastocyst. EGF (100 ng mL–1) increased both PGE2 and PGF production (P &lt; 0.05) by cultured endometrial epithelial and stromal cells. The overall results suggest that endometrial epithelial cells rather than stromal cells are the main source of EGF. Epidermal growth factor produced by epithelial cells stimulates the production of IFNT by bovine trophoblasts. The capacity of conceptus to increase IFNT and luteotropic PGE2 production rather than luteolytic PGF in response to EGF stimulation may be essential for the establishment of pregnancy in cattle.

Molecular Evolution, Characterization and Expression Profiling of Uterine Aldoketoreductase 1B5 Gene in Endometrium of Goat (Capra hircus)

Aldoketoreductase 1B5 (AKR1B5), a member of the Aldoketoreductase family, is involved in the production of Prostaglandin F2α (PGF2α) as one of vital prostaglandin F synthase (PGFS). PGs (Prostaglandins) play a crucial role in female reproductive system. In the present study, we cloned and characterized the full-length open reading frame of AKR1B5 gene in Black Bengal (BB) goat. The complete coding sequence of AKR1B5 comprises an entire open reading frame of 951 bp, encoding 316 amino acid (AA) residues. BB AKR1B5 showed >82.9% identity with that of cattle, rabbit, human, and rat at nucleotide and amino acid levels, respectively. Further, a systematic study of AKR1B5 sequence evolution was also conducted using Phylogenetic Analysis by Maximum Likelihood (PAML), entropy plot, and Blossum 62 in a phylogenetic context. Analysis of nonsynonymous to synonymous nucleotide substitution rate ratios (Ka/Ks) revealed that negative selection may have been operating on this gene during evolution in goat, cattle, rabbit, human, and rat, which showed its conservation across species. Further, expression of AKR1B5 was determined by quantitative real-time PCR in goat endometrial tissues at different stages of the estrous cycle and early pregnancy. Our results indicated its high expression at luteolytic phase (stage III; day 16–21) during the estrous cycle. However, during early (day ∼30–40) pregnancy the expression was highest as compared to estrous cycle.

Role of prostaglandin F2α production in lipid bodies from Leishmania infantum chagasi: insights on virulence.

Lipid bodies (LB; lipid droplets) are cytoplasmic organelles involved in lipid metabolism. Mammalian LBs display an important role in host-pathogen interactions, but the role of parasite LBs in biosynthesis of prostaglandin F2α (PGF2α) has not been investigated. We report herein that LBs increased in abundance during development of Leishmania infantum chagasi to a virulent metacyclic stage, as did the expression of PGF2α synthase (PGFS). The amount of parasite LBs and PGF2α were modulated by exogenous arachidonic acid. During macrophage infection, LBs were restricted to parasites inside the parasitophorous vacuoles (PV). We detected PGF2α receptor (FP) on the Leishmania PV surface. The blockage of FP with AL8810, a selective antagonist, hampered Leishmania infection, whereas the irreversible inhibition of cyclooxygenase with aspirin increased the parasite burden. These data demonstrate novel functions for parasite-derived LBs and PGF2α in the cellular metabolism of Leishmania and its evasion of the host immune response.