Identification of mouse lines with HA-tagged prostaglandin receptors / 生理学报

Prostaglandins are a class of poly-unsaturated fatty acids-derived bioactive lipids with important physiological function by binding to specific receptors. Prostaglandin receptors lack specific antibodies, which greatly impedes the research on our understanding of the signaling of prostaglandins. The aim of this study was to identify nine mouse lines with amino terminal (-NH2, -N) HA-tagged prostaglandin receptors by using the combination of artificial sperm and CRISPR-Cas9 technology. The guide RNA expression plasmid and labeled targeting vector plasmids were transferred into "artificial sperm cells". The "artificial sperm cells" containing labeled proteins were selected and injected into mouse oocytes, and implanted into pseudopregnant mice to obtain labeled mice. The genomic DNA of the prostaglandin receptor tagged mice was extracted, and the genotypes of mice were detected by PCR method. We also isolated mouse peritoneal macrophages to verify the protein expression of HA-labeled prostaglandin receptor by Western blot. Specific DNA bands were amplified in prostaglandin receptor labeled mice, and specific HA protein bands were detected in macrophage proteins, which was not detected in wild type mice. In summary, we successfully constructed 9 mouse lines with HA-tagged prostaglandin receptors, providing a powerful tool for further study of the pathophysiological functions of prostaglandin signaling both in vivo and in vitro.

Cloprostenol administration in the first week postpartum reduces expression of oxytocin receptors in the endometrium in Holstein-Zebu cows / Administração de cloprostenol na primeira semana pós-parto pode reduzir a expressão de receptores de ocitocina no endométrio de vacas mestiças Holandês-Zebu

The present study investigated the hormonal profile and expression of prostaglandin F2α (PGF2α), oxytocin and estrogen receptors in uterine tissues of postpartum cows treated with cloprostenol. Twenty Holstein-Zebu crossbred cows were treated with saline solution (treatment CONT) or cloprostenol (treatment CLO), both administered two and five days postpartum. Blood samples were collected on days two, seven, 14, 21 and 28 postpartum for progesterone, PGF2α metabolite (PGFM) and estradiol determination, and endometrial biopsy was performed in order to quantify the expression of oxytocin receptor (OXTR), prostaglandin F receptor (PTGFR) and estrogen receptor 1 (ERS1) genes. In the CLO treatment, expression of OXTR was reduced (P<0.05) but no difference (P>0.05) between treatments was found for PTGFR and ERS1 expression. Estrogen concentrations increased progressively until day 14 (P<0.05) and the highest OXTR expression and lowest PTGFR expression were observed on day 14 (P<0.05) in both treatments. Serum PGFM concentrations were high throughout the experiment. In conclusion, cloprostenol administration at days two and five of postpartum seems to reduce OXTR expression in the endometrium in crossbred cows.(AU)

O presente estudo avaliou o perfil hormonal e a expressão gênica de receptores de prostaglandina F2α (PGF2α), ocitocina e estrógeno no endométrio de vacas pós-parto tratadas com cloprostenol. Vinte vacas mestiças Holandês-Zebu foram tratadas com solução salina (tratamento CONT, n = 10) ou cloprostenol (tratamento CLO, n = 10), ambos administrados dois e cinco dias após o parto. Amostras de sangue foram coletadas nos dias dois, sete, 14, 21 e 28 pós-parto para mensuração de progesterona, de metabólito de PGF2α (PGFM) e de estradiol, e foram obtidas biópsias endometriais para quantificar a expressão de PTGFR, OXTR e ESR1. No tratamento CLO, a expressão gênica de receptores de ocitocina foi menor (P<0,05). As concentrações de estrógeno aumentaram progressivamente até o dia 14 (P<0,05). A maior expressão de OXTR foi observada no dia 14 (P<0,05). A expressão de ESR1 foi semelhante entre os tratamentos (P>0,05). Os níveis de PGFM foram altos durante todo o estudo. Conclui-se que a administração de cloprostenol nos dias dois e cinco pós-parto parece diminuir a expressão de OXTR no endométrio em vacas mestiças.(AU)

Blockade of PGE2, PGD2 receptors confers protection against prepatent Schistosomiasis Mansoni in mice

Schistosomiasis is a chronic disease with considerable social impact. Despite the availability of affordable chemotherapy, drug treatment has not significantly reduced the overall number of disease cases. Among other mechanisms, the parasite produces PGE2 and PGD2 to evade host immune defenses. To investigate the role of PGE2 and PGD2 in schistosomiasis, we evaluated the effects of L-161,982, Ah6809 [PGE2 receptor antagonists alone or combined with each other] and MK-0524 [PGD2 receptor antagonist] during prepatent Schistosoma mansoni infection. Drugs were administered intraperitoneally an hour before and 24 hours after infection of C57BL/6 mice with 100 Schistosoma mansoni cercariae. L-161,982, Ah6809, their combination and MK-0524 caused partial protection against pre-patent S. mansoni infection which was mediated by biasing the immune response towards Th1 phenotype. These results showed that blockade of PGE2 and PGD2 receptors confers partial protection against pre-patent S. mansoni infection in mice and that they may be useful as adjunctive therapy to current anti-schistosomal drugs or vaccines

Prostaglandin F receptor expression in intrauterine tissues of pregnant rats

In this investigation, we studied the expression and localization of rat prostaglandin F (FP) receptor in uterine tissues of rats on gestational Days 10, 15, 18, 20, 21, 21.5 and postpartal Days 1 and 3 using Western blotting analysis, real-time PCR, and immunohistochemistry. A high level of immunoreactivity was observed on gestational Days 20, 21, and 21.5 with the most significant signals found on Day 20. FP receptor protein was expressed starting on gestational Day 15, and a fluctuating unsteady increase was observed until delivery. Uterine FP receptor mRNA levels were low between Days 10 and 18 of gestation (p < 0.05). The transcript level increased significantly on Day 20 and peaked on Day 21.5 just before labor (p < 0.05). There was a positive correlation between FP receptor mRNA expression and serum estradiol levels (rs = 0.78; p < 0.01) along with serum estradiol/progesterone ratios (rs = 0.79; p < 0.01). In summary, we observed an increase FP receptor expression in rat uterus with advancing gestation, a marked elevation of expression at term, and a concominant decrease during the postpartum period. These findings indicate a role for uterine FP receptors in the mediation of uterine contractility at term.

Inhibitory G proteins and their receptors: emerging therapeutic targets for obesity and diabetes

The worldwide prevalence of obesity is steadily increasing, nearly doubling between 1980 and 2008. Obesity is often associated with insulin resistance, a major risk factor for type 2 diabetes mellitus (T2DM): a costly chronic disease and serious public health problem. The underlying cause of T2DM is a failure of the beta cells of the pancreas to continue to produce enough insulin to counteract insulin resistance. Most current T2DM therapeutics do not prevent continued loss of insulin secretion capacity, and those that do have the potential to preserve beta cell mass and function are not effective in all patients. Therefore, developing new methods for preventing and treating obesity and T2DM is very timely and of great significance. There is now considerable literature demonstrating a link between inhibitory guanine nucleotide-binding protein (G protein) and G protein-coupled receptor (GPCR) signaling in insulin-responsive tissues and the pathogenesis of obesity and T2DM. These studies are suggesting new and emerging therapeutic targets for these conditions. In this review, we will discuss inhibitory G proteins and GPCRs that have primary actions in the beta cell and other peripheral sites as therapeutic targets for obesity and T2DM, improving satiety, insulin resistance and/or beta cell biology.

Prostaglandin D2 and TH2 Inflammation in the Pathogenesis of Bronchial Asthma

Prostaglandin D2 (PGD2) is a major prostanoid, produced mainly by mast cells, in allergic diseases, including bronchial asthma. PGD2-induced vasodilatation and increased permeability are well-known classical effects that may be involved in allergic inflammation. Recently, novel functions of PGD2 have been identified. To date, D prostanoid receptor (DP) and chemoattractant receptor homologous molecule expressed on TH2 cells (CRTH2) have been shown to be major PGD2-related receptors. These two receptors have pivotal roles mediating allergic diseases by regulating the functions of various cell types, such as TH2 cells, eosinophils, basophils, mast cells, dendritic cells, and epithelial cells. This review will focus on the current understanding of the roles of PGD2 and its metabolites in TH2 inflammation and the pathogenesis of bronchial asthma.

CRTH2 antagonist ameliorates airway inflammation in rats with asthma / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the effect of prostaglandin D2 receptor antagonists on the airway inflammation in rats with asthma.</p><p><b>METHODS</b>Forty male SD rats were randomly divided into four groups: Group A (normal control), Group B (asthma group), Group C (CRTH2 antagonist BAYu3405 treatment group), Group D (DP1 antagonist BWA868C treatment group). Asthma was induced by ovalbumin (OVA) challenge. The rats in each group were sacrificed 24 h after the last challenge of OVA.DP1/CRTH2 receptors on eosinophils (EOS) were measured by radiological binding assay (RBA). The left lungs were used for histological examinations and bronchoalveolar lavage fluid (BALF) was collected from the right lungs. The total cell numbers, EOS absolute count and differential cell counts in BALF were performed. Serum concentrations of IL-4, 5 and IFN-gamma were measured by ELISA.</p><p><b>RESULTS</b>Rats in BAYu3405 treatment group showed profoundly decreased infiltrates of EOS and lymphocytes in the wall of bronchus when compared with those of asthma group and BWA868C treatment group. Serum concentrations of IFN-gamma in rats of BAYu3405 treatment group increased, but IL-4 and IL-5 decreased significantly when compared with those in rats of asthma group and BWA868C treatment group (P<0.01), and BALF EOS count was decreased significantly (P<0.01). Peripheral blood EOS count was higher than that in rats of normal control group, but was not significantly different from that in rats of asthma group and BWA868C treatment group. The combining capacity of CRTH2 and DP total combining capacity on EOS in asthma group, BAYu3405 treatment group and BWA868C treatment group were significantly higher than those in Group A (P<0.01). There was no significant difference in DP1 among all the groups (P>0.05).</p><p><b>CONCLUSION</b>CRTH2, but not DP1 antagonist can effectively ameliorate airway inflammation in rats with asthma.</p>

Changes of prostaglandin D2 receptor on T cells in peripheral blood of children with asthma / 中国当代儿科杂志

<p><b>OBJECTIVE</b>Chronic airway inflammation is associated with the polarization of TH2 cells in asthma. Prostaglandin D2 (PGD2) plays an important role in the polarization of TH2 cells. This study aimed to investigate the changes of PGD2 receptors (DP1/CRTH2) on T lymphocytes and their significance in asthma.</p><p><b>METHODS</b>Seventy-two children with asthma were assigned to two groups: acute attack (n=42) and remission (n=30). Thirty-five healthy children were used as the control group. Plasma levels of TH2 cytokines IL-4 and IL-5, and TH1 cytokine INF-gamma were detected using ELISA. Radiological binding assay (RBA) was used to measure the contents of DP1/CRTH2 receptors on T cells in peripheral blood (PPB).</p><p><b>RESULTS</b>The total combining contents of DP and CRTH2 on T cells in PPB in the acute attack and the remission groups were significantly higher than those in the control group (p<0.01). There was no significant difference in the DP1 content among the three groups. Serum levels of IL-4 and IL-5 significantly increased (p<0.01), in contrast, serum levels of TH1 cytokine IFN-gamma were significantly reduced in the acute attack and the remission groups compared with those in the control group (p<0.01).</p><p><b>CONCLUSIONS</b>The total combining contents of DP and CRTH2 on T cells increased, serum levels of TH2 cytokines also increased, but serum levels of TH1 cytokine decreased significantly in the acute attack and the remission phases in children with asthma. This showed that a polarization of TH2 cells occurred in children with asthma and suggested that CRTH2 antagonism may be a new target for the treatment of asthma.</p>

Inflamação: o certo e o errado / Inflammation: the right and tre wrong

A inflamação é caracterizada por uma complexa série de eventos que surgem como resposta a uma agressão orgânica. O presente artigo descreve os mecanismos através do qual esse processo ocorre e as principais substâncias envolvidas. Dor, calor, rubor e edema são os sintomas cardinais, mas além dessas alterações locais, manifestações sistêmicas, como febre, podem estar presentes

Effect of Prostaglandin E2 on Gel-forming Mucin Secretion in Normal Human Nasal Epithelial Cells / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: Recently, Prostaglandin E2 (PGE2) was found to induce MUC5AC production via an agonist of E-prostanoid (EP2/EP4), but not EP1/EP3, in normal human airway epithelium. However, the receptor that mediates MUC5AC has not been determined. This study was aimed to investigate the MUC5AC mucin gene and mucin secretion by PGE2 and its receptors in cultured normal human nasal epithelial cells. MATERIALS AND METHOD: After treatment with PGE2 and/or PGE2 antagonist, gel-forming mucin mRNA expression was determined by reverse transcription-polymerase chain reaction. Total mucin and MUC5AC mucin levels were measured using an immuno-dot blotting assay. RESULTS: PGE2 increased the MUC5AC gene expressions and MUC5AC mucin, but not the expressions of other gel-forming mucin genes. An EP2 receptor antagonist (AH6809) did not suppress the PGE2-induced MUC5AC gene expression or MUC5AC mucin. However, an EP4 receptor antagonist (AH23848) significantly suppressed the level of PGE2-induced MUC5AC gene expression and MUC5AC mucin. CONCLUSION: These findings indicate that PGE2 induces MUC5AC gene expression and mucin secretion via EP4 receptor in cultured normal human nasal epithelial cells.

Detection of EP1 and FP receptor mRNAs in the iris-ciliary body using in situ hybridization / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To determine the expression of E-prostanoid1 (EP(1)) and F-prostanoid (FP) receptor mRNAs in iris-ciliary bodies of the human eye using in situ hybridization.</p><p><b>METHODS</b>EP(1) and FP receptor mRNAs were detected by riboprobes labeled with digoxigenin on paraffin sections of the iris-ciliary body tissue of the human eye using in situ hybridization.</p><p><b>RESULTS</b>EP(1) and FP receptor mRNAs were highly expressed in blood vessels, muscles and the endothelia of the iris. EP(1) receptor hybridization signals were present in all muscle fibers of the ciliary body. Hybridization signal corresponding to FP receptor mRNA transcript was predominantly expressed in the circular muscle and in the collagenous connective tissues of the ciliary body. FP receptor mRNA was not detected in radial and longitudinal muscles.</p><p><b>CONCLUSIONS</b>EP(1) and FP receptor mRNAs in human ocular tissues appear to be widely localized in the functional sites of the respective receptor agonists. Selective localization of EP(1) and FP receptor mRNAs in the circular muscles and collagenous connective tissues of the ciliary body suggests that EP(1) and FP receptors play an important role in enhancing uveoscleral outflow of aqueous humor.</p>

Edad y cáncer mamario / Age and breast cancer

Antecedente: En México la incidencia de cáncer mamario es de 18.3 por 100 mil habitantes, con edad promedio de presentación de 48 años. En los Estados Unidos de Norteamérica, 12.5 por ciento de las mujeres tiene cáncer de mama de las que 3.5 por ciento fallece anualmente. Se consideran factores pronósticos el tamaño tumoral, involucro de ganglios axilares, estado de receptores hormonales, grado histológico nuclear y los factores inmunohistoquímicos. La edad se considera como factor pronóstico importante, la relación entre edad y supervivencia aún no está bien definida. Objetivo. Evaluar las características y el comportamiento tumoral entre mujeres menores de 35 y mayores de 65 años con cáncer de mama. Pacientes. Grupo A:28 mujeres mayores de 65 años y grupo b:20 enfermas menores de 35 años; todas fueron atendidas entre enero de 1986 y febrero de 1997. Métodos. Se determinó edad al momento del diagnóstico, historia familiar de cáncer de mama y ginecobstétrica, estirpe neoplásica, lado afectado, estadio clínico, tamaño tumoral, participación de ganglios axilares, estado de receptores hormonales, tratamiento utilizado, tiempos de seguimiento y tiempo libre de recurrencia. Resultados. Edad promedio: 72.6 ñ 1.1 años en el grupo A y 31.3 ñ 3 en el B. Tiempo de seguimiento: 63.8 meses en el grupo A y 38 en el B. No se detectó diferencia con relación a menarca, paridad, edad al primer embarazo, uso de fármacos hormonales, historia familiar y sitio afectado. El 67.8 por ciento del grupo A y el 40 por ciento del grupo B mostraron estado ganglionar axilar negativo. Tuvieron más de 10 ganglios afectados el 26.6 por ciento del grupo B; en el grupo A ningún caso se presentó esta característica. La estirpe ductal infiltrante fue común para ambos grupos. Se detectó estadio IIA en el 67.8 por ciento del grupo A y en el 70.5 por ciento del B. Grados nucleares más altos predominaron en el B. Se detectó comorbilidad en 50 por ciento de las pacientes del grupo A. El tiempo libre de recurrencia fue de 80 meses en el grupo A y de 25.4 en el B. El grupo A se observaron tres casos de recurrencias (local, pulmonar y ósea); mientras que en el B se detectaron seis casos (mama contralateral en dos; local en otro; ganglio cervical en uno más; y pulmón, hueso y encéfalo en los otros dos)...