Identification of elevated concentrations of estradiol in bovine uterine endometrium

We have investigated the estradiol content of bovine endometrium and related this to circulating plasma estradiol content. In 9 heifers, mean ± S.E.M. plasma estradiol concentration was 0.64 ± 0.25 pg/ml while the mean ± S.E.M. endometrial estradiol content was 43.0 ± 14.7 pg/g tissue; there was a close relationship between plasma and tissue estradiol levels (R² = 0.81; P < 0.001). During culture of endometrial tissue there was a progressive transfer of estradiol from tissue to culture media but no change in total estradiol. Culture of endometrium from 4 heifers with 5 ng/ml testosterone for 72 h resulted in no increase in estradiol. Furthermore, immunohistochemistry revealed no aromatase protein in uterine endometrium. These results confirm high stored tissue concentrations of estradiol in bovine endometrium while providing no evidence for estradiol synthesis by this tissue. The mechanism(s) through which this sequestration of estradiol into uterine tissue occurs remains to be determined.     

子宫内膜炎患牛子宫内膜PGR和ESR-1 mRNA的表达特征

为探讨卵巢激素及其子宫内膜受体在奶牛子宫内膜炎中的作用,以产后6～10d健康及患急性化脓性子宫内膜炎的中国荷斯坦奶牛为研究对象,通过ELISA法检测子宫内膜中PGF2a,血清中孕酮和雌二醇浓度,SYBRGreenⅠ荧光定量PCR检测子宫内膜PGR和ESR-1mRNA的表达。结果显示,病牛子宫内膜PGF2α水平为（6.503±0.122）μg/L,血清孕酮为（0.816±0.281）μg/L,血清雌二醇为（35.462±2.523）μg/L,而对照组分别为（14.408±0.285）,（0.543±0.142）,（37.6±1.317）μg/L;子宫内膜炎患牛组PGF2a水平极显著低于对照组（P〈0.01）,孕酮含量显著高于对照组（P〈0.05）,PGF2a与孕酮水平呈显著负相关（r=0.893）,雌二醇含量组间差异不显著。PGR基因的mRNA水平极显著高于对照组（P〈0.01）,而ESR-1基因的mRNA水平低于对照组,且差异极显著（P〈0.01）。上述结果表明,孕酮浓度显著升高及PGR mRNA高表达,可能抑制子宫收缩;ESR-1mRNA低表达使子宫新陈代谢减慢,子宫内膜合成PGF2a功能下降,这些改变有可能促进了产后子宫内膜炎的发生发展。     

Effect of simulated transport stress on the rat small intestine: A morphological and gene expression study

The present study investigated the effects of simulated transport stress on morphology and gene expression in the small intestine of laboratory rats. Sprague Dawley rats were subjected to 35 °C and 0.1×g on a constant temperature shaker for physiological, biochemical, morphological and microarray analysis before and after treatment. The treatment induced obvious stress responses with significant decreases in body weight (P < 0.01), increases in rectal temperature, serum corticosterone (CORT), serum glucose (GLU), creatine kinase (CK) and lactate dehydrogenase (LDH) levels (P < 0.01), as well as expression of Hsp27/70/90 mRNA (P < 0.05; P < 0.01). The rat jejunum was severely damaged and apoptotic after mimicking transport stress, which may mainly be related to cell death, oxidation reduction and hormone imbalance determined by microarray analysis. The bioinformatics analysis from the present study would provide insight into the potential mechanisms underlying transport stress-induced injury in the rat small intestine.     

热应激对奶牛机体的影响及防控措施

随着全球气候变暖,以及奶牛产奶性能的提高,热应激成了影响我国奶牛生产的一大难题,尤其在南方高温高湿的夏季,奶牛极易发生热应激。本文就热应激对奶牛产奶量、乳品质和繁殖性能的影响等方面进行了综述,并提出了一些预防奶牛热应激的措施,以期指导生产实践,提高奶牛养殖的经济效益。     

Physiological and pathological expression of intermediate filaments in the equine endometrium

The aim of this study was to investigate the expression of the intermediate filaments cytokeratin, vimentin and desmin in the equine endometrium by immunohistological techniques. For this purpose, endometrial biopsies of 151 mares were examined to determine physiological cycle patterns and changes resulting from endometriosis. During the physiological cycle epithelial cells and mesenchymal cells express cytokeratin and vimentin, respectively, whilst desmin and vimentin were coexpressed by the smooth muscle cells. Epithelial coexpression of cytokeratin and vimentin was seen in numerous fibrotic glands and in the uterine glands of three mares with pathologically inactive endometria. Three different staining patterns (basal, perinuclear, diffuse) of vimentin were associated with typical morphological alterations of the affected epithelia. In addition, in 14 cases a stromal coexpression of vimentin and desmin was found, indicating an atypical stromal differentiation in inactive endometria of older mares, barren for several years.     

Expression of ADAMTS1 mRNA in bovine endometrium and placenta during gestation

A disintegrin and metalloproteinase with thrombospondin motifs 1 (ADAMTS1) is a secreted protease. Through the regulation of extracellular matrix remodeling or developmental processes or both, ADAMTS1 is involved in several biological functions, including ovulation and embryo receptivity. However, the expression and possible role of ADAMTS1 in bovine endometrium is unknown. In this study, we analyzed ADAMTS1 mRNA expression in bovine endometrium during the estrous cycle, peri-implantation period, and at different stages of gestation by using quantitative real-time RT-PCR (qPCR) and in situ hybridization. The qPCR results indicated that the expression of ADAMTS1 mRNA was not affected by the day of the estrous cycle and was similar to cyclic levels on day 35 of gestation; however, the expression was more abundant in cotyledonary tissues of the placenta during late gestation. The in situ hybridization study showed that ADAMTS1 mRNA was detected mainly in uterine luminal epithelia and stromal cells during the estrous cycle and peri-implantation period. A disintegrin and metalloproteinase with thrombospondin motifs 1 mRNA was also expressed in the peri-implantation conceptus as well as in trophoblast cells, which include binucleate cells, and increased during late gestation. Furthermore, treatment of stromal cell with progesterone (300 nM) stimulated the expression of ADAMTS1 mRNA. This study indicates that ADAMTS1 participates in bovine endometrial remodeling, which is required for implantation and placental development in coordination with ovarian steroids.     

Expression dynamics of bovine MX genes in the endometrium and placenta during early to mid pregnancy

MX belongs to a family of type I interferon (IFN)-stimulated genes, and the MX protein has antiviral activity. MX has at least two isoforms, known as MX1 and MX2, in mammals. Moreover, bovine MX1 has been found to have alternative splice variants—namely, MX1-a and MX1B. In ruminants, IFN-τ—a type I IFN—is temporarily produced from the conceptus before implantation and induces MX expression in the endometrium. However, the expression dynamics of MX after implantation are not clear. In the present study, we investigated the expression of MX1-a, MX1B and MX2 in the endometrium and placenta before and after implantation along with the expression of IFN-α, type I receptors (IFNAR1 and IFNAR2) and interferon regulatory factors (IRF3 and IRF9). Pregnant uterine samples were divided into five groups according to pregnancy days 14–18, 25–40, 50–70, 80–100, and 130–150. Tissue samples were collected from the intercaruncular endometrium (IC), caruncular endometrium (C) and fetal placenta (P). Although all the MX expressions were significantly higher in the IC and C at days 14–18, presumably caused by embryo-secreted IFN-τ stimulation, their expressions were also detectable in the IC, C and P after implantation. Furthermore, IFN-α expression was significantly higher in the IC. RT-PCR indicated IFNAR1, IFNAR2, IRF3 and IRF9 mRNA in all the tissues during pregnancy. These results suggest that all the MX genes are affected by the type I IFN pathway during pregnancy and are involved in an immune response to protect the mother and fetus.     

Comparison of enzyme activities and gene expression profiling between yak and bovine skeletal muscles

The objective of the present study was to reveal the differences between yak (Bos grunniens) and cattle in energy metabolic characteristics and gene expressions in skeletal muscles. Activities of lactate dehydrogenase (LDH), malate dehydrogenase (MDH) and β-hydroxyacyl-CoA dehydrogenase (HOAD), which are involved in metabolism of carbohydrate and fatty acid respectively, were measured in longissimus dorsi and biceps femoris. Yak contained higher total LDH activity and higher proportion of LDH5 in longissimus dorsi than cattle (P < 0.05), indicating its more anaerobic potential characteristics in carbohydrate metabolism. However, yak contained higher activities of MDH and HOAD (P < 0.05) in biceps femoris than cattle, exhibiting high oxidative capacity under hypoxic environment. Semi-quantitative RT-PCR analysis of mRNA abundance showed that yak and cattle contained a similar level of adipocyte fatty acid-binding protein (A-FABP) in both longissimus dorsi and biceps femoris, while myosin heavy chain I (MyHC I) in longissimus dorsi and biceps femoris, and peroxisome proliferator-activated receptor coactivator 1α (PGC-1α) level in biceps femoris were significant lower in yak compared with cattle (P < 0.01 and P < 0.05 respectively), indicating the anaerobic characteristic of yak skeletal muscles. In conclusion, our experiment showed that yak skeletal muscles exhibit different energy metabolic property and gene expressions compared to cattle.     

γ-氨基丁酸在缓解奶牛热应激上的应用前景

γ-氨基丁酸作为机体一种重要的神经抑制剂,因具有提高食欲、增强免疫力、缓解热应激等作用,已被广泛应用于食品、药品等领域。在我国,奶牛夏季面临严峻的热应激问题,高温严重影响了奶牛正常的生理活动,对此开发具有缓解热应激功能的新型饲料添加剂成为研究热点。目前,γ-氨基丁酸已应用于畜禽生产,但对反刍动物的相关研究少有报道。本文对γ-氨基丁酸的生理功能及其在奶牛热应激中的应用进行了综述,以供同行参考。     

Interleukin-1 receptor antagonist expression in the equine endometrium during the peri-implantation period

To identify factors involved in the establishment of pregnancy in the mare, endometrium was collected from day 13 (day 0 = day of ovulation) cyclic and day 13, 19, and 25 pregnant animals. From initial cDNA subtraction studies, interleukin-1 receptor antagonist (IL-1RN) mRNA was found as a candidate molecule expressed uniquely in the pregnant endometrium. Expression of IL-1RN mRNA was markedly increased in day 19 and 25 gravid endometrium. In situ hybridization analysis revealed that IL-1RN mRNA was localized to the glandular epithelium. Interleukin-1 receptor antagonist (IL-1RN) protein was found in the extracts of day 25 gravid endometrium and was immunochemically localized to the glandular epithelium/luminal cavity of the pregnant uterus. High concentrations of estradiol-17β (E₂) were detected in day 25 conceptuses. Concentrations of E₂ were higher in the gravid endometrial portion than in other endometrial regions. On the other hand, progesterone concentrations did not differ among endometrial samples analyzed. Furthermore, the expression of IL-1RN mRNA was up-regulated in endometrium culture samples treated with 10 ng/mL E₂ and 10 ng/mL progesterone. In the analysis of related gene expression, increased amounts of IL-1α and IL-6 mRNA were also found in the day 25 gravid endometrium; however, these expressions in endometrial culture samples were not up-regulated by the steroid treatment. These results indicate that expression of IL-1RN in the endometrium is likely regulated by E₂ and progesterone and suggest that IL-1RN regulates the degree of IL-1 signal transduction and thereby plays an important role in the establishment of equine pregnancy.     

The dynamic expression of extracellular matrix in the bovine endometrium at implantation

Remodeling of uterine endometrial extracellular matrix (ECM) is pivotal to successful implantation and placentation, and has been well described in the rodents and humans. However, bovine endometrial ECM remodeling is still vaguely defined, especially at the time of implantation. Therefore, this study investigated the distribution of four ECMs namely, types I and IV collagen, laminin and fibronectin, from days 0 to 30 of gestation in bovine endometrium by immunofluorescence microscopy. A change in the distribution pattern of ECMs was evident by day 14 of gestation as features at this stage were clearly different from those of day 14 of the estrous cycle. The immunoreactivity of type I collagen, fibronectin and laminin decreased from day 14 of gestation and was obscured by day 24 of gestation. The type I collagen fibers formed were of thinner consistency than those of the estrous cycle and showed a coarser meshwork within the epithelium sites during the implantation period. In addition, the type IV collagen and laminin immunoreactivities of epithelial basement membrane also remarkably declined at exactly the same time. By day 30 of gestation, the four ECMs had regenerated with the formation of the placentome. In conclusion, this study reveals that remodeling of ECM is essential for the successful establishment of pregnancy in the bovine.     

Comparative analysis of proteomic profiles between endometrial caruncular and intercaruncular areas in ewes during the peri-implantation period

The endometrium of sheep consists of plenty of raised intercaruncular areas （IC）. In order to better understand aglandular areas called caruncular （C）, and intensely glandular the endometrium involved mechanisms of implantation, we used LC-MS/MS technique to profile the proteome of ovine endometrial C areas and IC areas separately during the peri-implantation period, and then compared the proteomic profiles between these two areas. We successfully detected 1740 and 1813 proteins in C areas and IC areas respectively. By comparing the proteome of these two areas, we found 170 differentially expressed proteins （DEPs） （P 〈 0.05）, functional bioinformatics analysis showed these DEPs were mainly involved in growth and remodeling of endometrial tissue, cell adhesion and protein transport, and so on Our study, for the first time, provided a proteomic reference for elucidating the differences between C and IC areas, as an integrated function unit respectively, during the peri-implantation period. The results could help us to better understand the implantation in the ewes. In addition, we established a relatively detailed protein database of ovine endometrium, which provide a unique reference for further studies.     

Comparative analysis of proteomic profiles between endometrial caruncular and intercaruncular areas in ewes during the peri-implantation period

The endometrium of sheep consists of plenty of raised aglandular areas called caruncular (C), and intensely glandular intercaruncular areas (IC). In order to better understand the endometrium involved mechanisms of implantation, we used LC-MS/MS technique to profile the proteome of ovine endometrial C areas and IC areas separately during the peri-implantation period, and then compared the proteomic profiles between these two areas. We successfully detected 1740 and 1813 proteins in C areas and IC areas respectively. By comparing the proteome of these two areas, we found 170 differentially expressed proteins (DEPs) (P < 0.05), functional bioinformatics analysis showed these DEPs were mainly involved in growth and remodeling of endometrial tissue, cell adhesion and protein transport, and so on. Our study, for the first time, provided a proteomic reference for elucidating the differences between C and IC areas, as an integrated function unit respectively, during the peri-implantation period. The results could help us to better understand the implantation in the ewes. In addition, we established a relatively detailed protein database of ovine endometrium, which provide a unique reference for further studies.     

Effect of retinoic acid on gene expression profiles of bovine intramuscular preadipocytes during adipogenesis

To investigate genes involved in intramuscular adipogenesis in ruminants, 16 genes with dramatic variable expression were selected. These were selected from the differentiation‐ and proliferation‐phase libraries of our previous serial analysis of gene expression (SAGE) studies of a clonal bovine intramuscular preadipocyte (BIP) cell line. We harvested the BIP cells over 12 days after adipogenic stimulation with all‐trans retinoic acid (ATRA). Quantitative real‐time PCR confirmed the earlier SAGE study results of the expression patterns of 15 of the genes. On day 6, TG accumulation increased significantly in the BIP cells but was completely inhibited in the 3T3‐L1 cells (the monogastric reference). ATRA enhanced expression levels of six genes whereas it suppressed expression of eight genes on day 3 of adipogenesis in the BIP cells. Forty‐eight hours after transfection, the messenger RNA expression level of the adipose differentiation‐related protein (ADFP), encoded by one of the upregulated genes, in the ADFP small interference RNA (siRNA)‐transfected cells was 3.5% of that in negative control‐transfected cells. Also, 6 days after induction the TG level in the ADFP siRNA‐transfected cells was 21.8% lower than that in negative control‐transfected cells. This analysis of gene expression profiles after ATRA treatment will contribute to our understanding of the molecular mechanisms involved in bovine intramuscular adipogenesis.