Inflammation in the bovine female reproductive tract

Inflammation of the reproductive tract of a cow occurs when the physical and functional barriers to contamination are breached or specific infection occurs. Commonly, contamination occurs at parturition and to a lesser extent at estrus. Uterine contamination following calving is common, but most healthy cows are able to clear the uterus of bacteria in the first 2 to 3 wk after calving. Persistent infections are more likely to be caused by Actinomyces pyogenes. Specific venereal infections tend to be more host-adapted and produce a lower grade inflammation. Nonspecific bacterial contamination of the endometrium generally induces a neutrophilic influx into the stratum compactum and uterine lumen. Neutrophils phagocytize bacteria with the aid of opsonins in the uterine fluid. Mast cells and eosinophils may also contribute to the inflammatory reaction, which may damage the surface epithelium and release vasoactive substances that allow leakage of serum antibodies into the uterine secretions. Specific antibodies of immunoglobulin (Ig) isotype A, M, G1, and G2 in uterine secretions have been described. In model species, the immune capability of the uterus is influenced by steroid hormones, especially estradiol, which increases secretory component and both IgA and IgG content in uterine secretions and increases the activity of antigen-presenting cells in the uterus. Similar cyclic fluctuations in immune components have been described for cows, including changes in the population of subsurface cytotoxic and helper T cells and changes in the expression of major histocompatibility II antigen on surface cells.     

Immunity in the female sheep reproductive tract

Immune surveillance in the female reproductive tract is dependent on the interplay of many factors that include the expression of pattern recognition receptors on epithelial cells, resident leukocyte populations and hormones, none of which are uniform. The lower reproductive tract must accommodate the presence of commensal organisms whereas the upper reproductive tract is sterile. However, the upper female reproductive tract has its own immunological challenge in that it must tolerate the presence of a semi-allogeneic fetus if pregnancy is to succeed. So, immune activation and effector mechanisms to control pathogens may be qualitatively and quantitatively different along the reproductive tract. Our knowledge of innate and adaptive immunity in the sheep is less comprehensive than that of human or mouse. Nevertheless, comparative studies suggest that there are likely to be conserved innate immune sensory mechanisms (e.g. Toll-like receptors) and defence mechanisms (anti-proteases, defensins) that combine to limit infection in its early stages while shaping the adaptive response that leads to immunological memory and long-term protection. There are many pathogens that target the reproductive tract, and in particular the placenta, where specialised immunoregulatory mechanisms are operational. Among such pathogens are bacteria belonging to the genera Chlamydia/Chlamydophila that chronically infect the reproductive tracts of sheep and humans and ultimately cause disease through inflammation and tissue damage. An understanding of the immunological microenvironment of the reproductive tract is important for the design of novel control strategies to control chlamydial disease.     

Regulation by gonadal steroids of estrogen and progesterone receptors along the reproductive tract in female lambs.

The regulation of estrogen and progesterone receptor (ER, PR) expression by estradiol (E2) and progesterone (P4) in the oviduct, uterus and cervix of female lambs was studied. The animals received three intramuscular injections of E2, P4 or vehicle with an interval of 24 h and they were slaugthered 24 h after the third injection. Determinations of ER and PR were performed by binding assays and mRNAs of ER alpha and PR by solution hybridization. High levels of ER and PR in both cervix and oviduct were found in the female lamb, differing from other mammalian species. No significant effects by either E2 or P4 treatment on ER and PR levels in the cervix and oviduct could be observed. E2 treatment increased the mRNA levels of ERa and PR more than 3-fold in the cervix, while P4 treatment increased the mRNA levels of ERa and PR in the uterus. The results show differential effects of gonadal steroids on sex steroid receptor expression along the reproductive tract in female lambs, suggesting that steroid target tissues can modulate responses to the same circulating levels of steroid hormones.     

Sperm interaction with the female reproductive tract

Sperm transit in the female tract is a critical event for the success of fertilization. From their deposition in the vagina to final migration in the oviduct, sperm pass through the different compartments of the genital tract in which they encounter different environments. The cervix and the uterotubal junction (UTJ) are two barriers with different relative importance according to the species. The protein composition, the degree of glycosylation and the hydration of the cervical mucus change during the oestrous cycle. Several sperm surface proteins are associated with their migration through the cervical mucus and the UTJ. Data regarding the interaction of sperm with secretions of the epithelial tissue lining the different compartments of the female genital tract during the sperm transit are reviewed, with a particular emphasis on the migration of sperm through the cervix.     

Male reproductive traits and their relationship to reproductive traits in their female progeny: a systematic review

The overall objective of one of the major research programs in the Co-operative Research Centre (CRC) for Beef Genetic Technologies is to 'Improve female reproductive performance' in tropical, northern Australian beef cattle herds. To address this overall objective, a quantitative genetics project focused on investigation of male reproductive traits was designed and linked to three female reproduction-focussed projects, (i) discovery of genes associated with post-partum re-conception and age at puberty; (ii) expression of genes associated with post-partum re-conception; and (iii) early predictors of lifetime female reproductive performance. During the initial planning of this male reproductive traits project, the CRC Scientific Review Committee recommended that the research team investigate and evaluate potentially new, early-life (i.e able to be measured before 2 years of age) predictors of both male and female reproductive performance. To address this recommendation, the following was carried out: (i) criteria for selection of traditional and candidate traits were established; (ii) methodology for tabulation of potential traits/phenotypes that define male and female reproductive function was developed; and (iii) a systematic scientific review of early-life predictors of male and female fertility was prepared. This review concluded that although factors that might be useful in predicting male reproductive performance have been studied for many years, there was relatively little useful information available to meet the objectives of this review. It was also concluded that the direction of future research should be guided not only by previous research which was scarce, but also by speculative hypotheses arising from an understanding of the physiological, endocrinological and genetic processes active in reproduction. A small number of new traits were recommended in addition to traditional sperm morphology, sexual behaviour, anatomical structure and growth traits. Potential additional traits include measurement of gonadotrophin-releasing hormone-stimulated luteinizing hormone (GnRH-stimulated LH); inhibin; several seminal plasma proteins (osteopontin, spermadhesin and seminal plasma proteins BSP30 and phospholipase A(2) could be used in an index); 11β-hydroxysteriod dehydrogenase; and leptin. In addition, the potential also exists to screen animals for a number of genetic markers associated with age of puberty, follicular recruitment and ovulation rate and genes associated with bovine seminal plasma protein and testosterone production. Insulin-like growth factor-1 (IGF-1) measurements are included because of their association with growth parameters, and an additional analysis demonstrated associations with male and female reproductive traits. Some of these factors have been previously evaluated in small numbers of animals of various species under intensive management conditions. Therefore, there is a need to evaluate these factors in much larger numbers of beef cattle grazing semi-extensive tropical production systems in northern Australia to determine their value in improving beef cattle enterprise profitability through improved herd fertility.     

Mechanisms of sperm storage in the female reproductive tract: an interspecies comparison

Once semen has been collected for artificial insemination, it is diluted into extenders designed to prevent its deterioration over the period prior to insemination. If the semen is not frozen, the extenders provide protection for a period of a few hours to a few days, depending on species. Despite the efforts of biotechnologists to increase the duration of storage without compromising fertility, there has been relatively little progress for many years. However, comparative studies in diverse species have revealed that long-term sperm storage (up to months and years) within the female reproductive tract is relatively commonplace in reptiles, fishes, birds and amphibians. Even among mammals, some species of bat have evolved mechanisms for storing spermatozoa for several months in the uterus or oviduct so that they can mate in the autumn but postpone fertilization until the spring. We currently know little about the mechanisms that support such long-term sperm storage, mainly because evidence from such species is either absent or fragmentary. Nevertheless, parallels between mammalian and other systems, where spermatozoa are sequestered in sperm storage tubules, suggest that the enclosure of spermatozoa within pockets of epithelial cells may be sufficient to achieve long-term sperm storage. In addition, recent evidence from sperm-storing bats has suggested an alternative, or additional, hypothesis that the modulation of apoptosis within epithelial cells is important in controlling sperm survival. Despite a lack of direct experimental evidence from a wide variety of species, I propose that there is now enough evidence to warrant investigation of these hypotheses.     

Granular cell lesions in the distal female reproductive tract of aged Sprague-Dawley rats

During the review of a rat carcinogenicity study, a spectrum of granular cell lesions was recognized in the distal female reproductive tract. To verify the diagnoses, cell populations of nine granular cell alterations of the cervix or vagina were characterized immunohistochemically and four were evaluated ultrastructurally. Immunoreactivity was demonstrated in 8/9 cases with S100 protein, 6/9 cases with neuron-specific enolase, and 7/9 cases with Leu-7. Granular cells were negative for smooth muscle-specific actin and calretinin. The immunohistochemical profile of these lesions was similar to that previously reported in other species, including humans. Ultrastructurally, as expected many lysosomal bodies were present in the cytoplasm of granular cells in all specimens evaluated. Based on the detailed evaluation of a series of lesions, we adopted the following diagnostic criteria and nomenclature for the granular cell changes of the female reproductive tract of rats. Granular cell aggregates were non-space-occupying lesions composed of clusters of typical granular cells. Benign granular cell tumors were space occupying lesions that typically contained prominent interstitial collagen and were either discrete masses or were difficult to discern from the surrounding tissues. Some benign tumors also contained foci of spindle cells with decreased granularity. Malignant tumors exhibited pleomorphism and an increased nucleus: cytoplasm ratio morphologically but had the same biologic behavior as the benign tumors. We applied these diagnostic criteria during the review of controls from 9 carcinogenicity studies. Up to 23% of control females in those carcinogenicity studies had granular cell lesions that could be classified into one of the three categories. Granular cell lesions appear to be common in the cervix/vagina of the Sprague-Dawley rat, and tumors may develop from granular cell aggregates.     

Distribution of immune cells in the female reproductive tract in uninfected and FIV infected cats

Cell-free and cell-associated FIV effectively cross the mucosa of the feline female reproductive tract. To identify possible cellular targets of FIV and to characterize changes in mucosal immunity after infection, we examined the types and numbers of immune cells residing in the reproductive tracts of control and intravaginally FIV-infected cats. Sections of the vestibule, vagina, cervix, uterus, and ovaries, were examined by immunohistochemistry for CD4+ and CD8+ T lymphocytes, CD22+ B lymphocytes, CD1a+ dendritic cells, and CD14+ macrophages. The reproductive tract of uninfected cats contained substantial numbers of CD8+ T lymphocytes, CD4+ T lymphocytes and macrophages, as well as moderate numbers of CD1a+ dendritic cells, and few B lymphocytes. The most prominent change between FIV- and FIV+ cats was a marked decrease in the concentration of CD4+ T lymphocytes resulting in inverted CD4+:CD8+ ratios throughout the reproductive tract of infected cats. There was also a trend towards increasing numbers of CD1a+ dendritic cells in the intravaginally-infected FIV+ cats, and decreasing numbers of macrophages and CD22+ B lymphocytes. This study indicates that similar to the peripheral immune system, FIV infection is associated with CD4+ cell loss and reduced CD4+:CD8+ ratios in the female reproductive mucosal tissue.     

Distribution of eosinophil granulocytes and mast cells in the reproductive tract of female goats in the preimplantation phase

Changes in eosinophil granulocytes and mast cells post-insemination may affect conceptus implantation, but information regarding the numbers of such cells in the mammalian reproductive tract is limited. This study investigated the preimplantation distribution of eosinophil granulocytes and mast cells (MCs) in the reproductive tract organs of female goats. Uterus, uterine cervix and uterine tubes samples were obtained at slaughter. Cornu uteri were washed in phosphate buffer solution (each animal contained at least one embryo). Tissues were fixed in 10% neutral buffered formol, Carnoy solution and Mota’s fixative (basic lead acetate) for 48 h and embedded in paraffin. Six-micrometre-thick sections were stained with Congo red (for eosinophil granulocytes) and toluidine blue in 1% aqueous solution at pH 1.0 for 5 min (for MCs). In the uterus, MCs occurred in highest numbers in the myometrium. Higher MC numbers were observed in uterus, uterine and uterine tubes in the preimplantation (experimental) group (cycle synchronised through 7 days intravaginal sponge with 0.3 g P₄) compared with the control group (P < 0.05). Eosinophil granulocyte numbers were significantly higher in the experimental group than in the control group (P < 0.05). These results indicate preimplantation-related changes in numbers of eosinophil granulocytes and MCs in goat reproductive tract organs.     

Expression of progesterone receptor membrane component 1, serpine mRNA binding protein 1 and nuclear progesterone receptor isoforms A and B in the bovine myometrium during the estrous cycle and early pregnancy

The aim of this study was to investigate the (1) expression of progesterone membrane component 1 (PGRMC1), serpine mRNA binding protein 1 (SERBP1) and progesterone receptor (PR) mRNA and (2) protein expression levels of PGRMC1, SERBP1 and PR isoforms A and B in the bovine myometrium during the estrous cycle and early pregnancy. Uteri from cows on days 1-5, 6-10, 11-16 and 17-21 of the estrous cycle and weeks 3-5, 6-8 and 9-12 of pregnancy were used (n=5-6 per period). There were no changes (P>0.05) in PGRMC1 mRNA expression during the estrous cycle, while expression of SERBP1 and PR mRNA was the lowest (P<0.05) on days 11-16 relative to other days of the cycle. The highest mRNA expression of PGRMC1, SERBP1 and PR was found during pregnancy. There were no changes (P>0.05) in SERBP1 protein expression in cycling and pregnant cows, while the highest (P<0.05) PGRMC1 protein expression was found during weeks 3-5 of pregnancy. Similar protein expression profiles for PRA and PRB were found, and protein levels were highest on days 1-5 of the estrous cycle. From day 6 of the cycle, PRA and PRB protein expression decreased and were maintained at this lower level during pregnancy. In conclusion, our study assessed mRNA and protein expression levels of PGRMC1, SERBP1 and PR in the bovine myometrium during the estrous cycle and the first trimester of pregnancy. It is possible that progesterone (P4) affects myometrial function in a genomic and nongenomic manner.     

Isolation of pathogenic strains of Haemophilus somnus from the female bovine reproductive tract.

The prevalence of Haemophilus somnus in the genital tract of slaughtered and live cows in southern Ontario was investigated. The vagina and uterus of slaughtered cows were swabbed separately. Live cows were examined and sampled in two field surveys: Centre A and Centre B. In the former, aspirated mucus secretions and in the latter, specimens obtained by guarded swabbing were examined bacteriologically. Haemophilus somnus was isolated from 28 genital tracts of 461 slaughtered (6.1%), and seven of 199 live (3.5%) cows during the centre B survey. The isolates were recovered from both normal and diseased reproductive tracts. Fourteen strains isolated from genital organs were examined for pathogenicity in vivo to test the occurrence of pathogenic isolates. In the initial stage of the in vivo study on pathogenicity, each of the fourteen isolates was examined on one calf using an intracisternal inoculation. Subsequently, one pathogenic and one nonpathogenic strain were inoculated into five calves each to statistically confirm their pathogenic potential. Of 14 genital isolates of H. somnus examined in an intracisternal calf assay, six (43%) caused a fatal peracute neurological disease, while eight were nonpathogenic. A comparative pathological study of pathogenic and nonpathogenic isolates showed that the former caused a severe fatal suppurative meningoencephalitis whereas the latter caused no lesions whatsoever or a mild leukocytic leptomeningitis. The salient data obtained in this study indicate that there are pathogenic strains of H. somnus in the genital tract of apparently normal cows as well as of those with inflammatory disease.     

蒙古绵羊雌性生殖道β-防御素(sBD-1)的序列分析及组织表达

从蒙古绵羊输卵管、子宫、子宫颈、阴道组织中提取总RNA,根据GenBank中羊的β-防御素的cDNA序列设计合成引物,通过RT-PCR进行cDNA扩增,获得了301bp的片段。将该片段克隆于pMD-18T载体后进行序列分析,确认该PCR产物为β-防御素。通过相同的RT-PCR反应体系和条件后,对PCR产物进行琼脂糖凝胶电泳,以β-actin为参照,根据PCR产物的亮度推断不同组织中β-防御素的表达量。结果,蒙古绵羊雌性生殖道上皮各组织均有β-防御素表达,说明β-防御素在蒙古绵羊雌性生殖道的先天免疫中起重要作用。β-防御素的表达量在雌性生殖道各组织内不同,提示雌性生殖道各组织的防御功能强度不同。     

植物甾醇对去卵巢KM小鼠血清生殖激素及乳腺组织孕激素受体的影响

本试验旨在研究植物甾醇对去卵巢小鼠血清激素及乳腺组织中孕激素受体的影响。50只雌性小鼠,随机选40只进行去卵巢手术,恢复7 d。分组如下：正常组、去卵巢对照组、植物甾醇低、中、高3个剂量组（每日灌胃20、80和320 mg/kg植物甾醇）。连续灌胃3周,第22天采集血样和乳腺组织,对小鼠血清中的雌二醇、孕酮和催乳素水平进行检测,对乳腺组织的孕激素受体基因表达进行荧光定量检测。结果表明,去卵巢对照组小鼠雌二醇水平显著低于正常组（P〈0.05）。植物甾醇组小鼠血清雌二醇水平持续升高,其中,高剂量组小鼠血清雌二醇水平与正常组最接近;与去卵巢对照组相比,去卵巢植物甾醇组小鼠血清孕酮水平均有提高,但差异不显著（P〉0.05）;与去卵巢对照组相比,去卵巢植物甾醇组小鼠催乳素水平先下降后上升,但他们之间无显著差异（P〉0.05）。与去卵巢对照组相比,植物甾醇各剂量组小鼠孕激素受体基因相对表达量呈上升趋势,低、中、高剂量组小鼠孕激素受体基因相对表达量分别是去卵巢对照组的1.05、1.13和2.78倍。植物甾醇可改善去卵巢小鼠内源激素不平衡的状况,促进乳腺组织PR基因的表达。     

Macroscopic anatomy of the reproductive tract of the reproductively quiescent female emu (Dromaius novaehollandiae)

Three reproductively quiescent female emus (Dromaius novaehollandiae) were embalmed with 10% formalin solution. The reproductive tract was dissected and described. The reproductive tract consists of an ovary and oviduct situated on the left side of the abdominal cavity. The left ovary is dark brown to black in colour with follicles covering the ventral surface. The ovary is located medial to the spleen and closely associated with the ventral surface of the cranial and middle lobes of the left kidney. The oviduct is a relatively straight tube that extends from the level of the cranial extent of the left ilium to the caudal border of the left pubic bone. The oviduct is grossly divided into the infundibulum, magnum, isthmus, uterus and vagina using variations in the mucosal fold pattern.     

Characterization of the reproductive cycle in female bharals (Pseudois nayaur) based on the changes in serum progesterone concentrations and parturition season

The purpose of present study was to determine annual changes in serum progesterone (P(4)) concentrations and to clarify basic reproductive characteristics, such as breeding season, estrous cycle, and puberty in female bharals (Pseudois nayaur). Blood was collected from 9 female bharals once or twice weekly for approximately one year. Serum P(4) concentrations were determined by radioimmunoassay. Serum P(4) concentrations showed remarkable and cyclic changes between November/December (winter) and May/June (late spring). The mean estrous cycle was 24.9 +/- 0.5 days. Chasing insistently to other females and discharge of mucus from the vulva were observed around the time when the serum P(4) concentrations began to increase. The chasing behavior and discharge of mucus were considered to be external indicators of estrus in female bharals. Serum P(4) concentrations of a pregnant female had non-cyclic changes, and the values remained high. In this study, all 37 deliveries were between April and September, and about 70% of these were concentrated in May and June. The conception month determined on the day of birth was between October and April for all animals, and the most common month was in December (54%). This month corresponded to an early stage of the period when the serum P(4) concentrations changed cyclically. These results indicate that many female bharals become pregnant at the beginning of the breeding seasons and, if they do not become pregnant, the estrous cycle, about 25 days in length, is repeated.     

Air‐liquid interface cell culture: From airway epithelium to the female reproductive tract

The air‐liquid interface (ALI) approach is primarily used to mimic respiratory tract epithelia in vitro. It is also known to support excellent differentiation of 3D multilayered skin models. To establish an ALI culture, epithelial cells are seeded into compartmentalized culture systems on porous filter supports or gel substrata. After an initial propagation period, the culture medium is removed from the apical side of the epithelium, exposing the cells to the surrounding air. Therefore, nutritive supply to the cells is warranted only by the basolateral cell pole. Under these conditions, the epithelial cells differentiate and regain full baso‐apical polarity. Some types of epithelia even generate in vivo‐like apical fluid or mucus. Interestingly, the ALI culture approach has also been shown to support morphological and functional differentiation of epithelial cells that are not normally exposed to ambient air in vivo. This review aims at giving a brief overview on the characteristics of ALI cultures in general and ALI models of female reproductive tract epithelia in particular. We discuss the applicability of ALI models for the investigation of the early embryonic microenvironment and for its implications in assisted reproductive technologies.     

Nongonadal LH receptors, their involvement in female reproductive function and a new applicable approach

Luteinising hormone (LH) and human chorionic gonadotropin (hCG) share a common receptor in gonadal cells. The receptors have also been detected in several nongonadal but reproduction-associated tissues of pigs, cattle, and other species including the uterus (myometrium, endometrium), oviduct, cervix, blood vessels, mammary gland and other tissues. The main role of LH/hCG receptors in the myometrium is stimulation of growth and hyperplasia, and relaxation of uterine motility; hCG also boosts blood flow in the uterine artery. LH and hCG can increase production of prostaglandins in the endometrium, oviduct, and blood vessels. We suggest that the preovulatory surge of LH plays an important role in controlling oviductal contractions. Awareness of LH binding to many tissues of the female reproductive tract and integration with embryonic factors may lead to the elaboration of new strategies for improved reproductive efficiency in domestic species. Mammary glands also possess LH/hCG receptors through which gonadotropins can affect the metabolism of steroid hormones and could play an inhibitory role in mammary carcinogenesis and in the growth of breast tumours. A novel approach to target and ablate carcinoma cells through LH receptors is described.