[Oestrus suppression in a dairy herd by means of anti-GnRH vaccination Improvac®: A prospective field study]. / Brunstunterdrückung mittels Anti-GnRH-Impfung Improvac® in einem Milchviehbestand: Eine prospektive Feldstudie.

INTRODUCTION: The aim of this study was to evaluate the duration of estrus suppression after a double administration of the anti-GnRH vaccine Improvac® (Zoetis Schweiz GmbH, 2800 Delémont) in cows. Furthermore, it should be investigated, if a third administration could prolong the effect of the cycle suppression. A total of 21 cows (more than four weeks post partum) were vaccinated twice, at least 35 days apart, with 2 ml Improvac® (0.4 mg of a GnRH-analogon) subcutaneously on one side of the neck. Over a period of 368 days and in the course of 18 farm visits these cows were examined gynecologically and re-vaccinated if they showed signs of estrus behaviour or ovarian activity. After the second vaccination the cycle of the cows was suppressed for an average period of 114 days (59-175 days) and the effect could be prolonged by a booster of the vaccine for another 127 days in three cows. Estrus behaviour was absent for a longer period than ovarian activity was. The vaccine was tolerated well: apart from slight swelling at the injection site, no side effects were observed. Our results demonstrate that two immunizations with Improvac® are an easily applicable method for the suppression of cyclic activity in cows for a mean period of 114 days. The duration of cycle suppression was prolonged by a booster of the vaccination.

INTRODUCTION: Le but de cette étude était d'évaluer la durée de la suppression des chaleurs chez les vaches après une double administration du vaccin anti-GnRH Improvac® (Zoetis Schweiz GmbH, 2800 Delémont). En outre, on a recherché si une troisième administration pourrait prolonger l'effet de la suppression du cycle. Au total, 21 vaches (plus de quatre semaines après vèlage) ont été vaccinées deux fois, à au moins 35 jours d'intervalle, avec 2 ml d'Improvac® (0,4 mg d'analogue de la GnRH) par voie sous-cutanée d'un côté du cou. Sur une période de 368 jours et au cours de 18 visites à la ferme, ces vaches ont été examinées gynécologiquement et revaccinées si elles présentaient des signes de comportement œstral ou d'activité ovarienne. Après la deuxième vaccination, le cycle des vaches a été supprimé pendant une période moyenne de 114 jours (59 - 175 jours) et l'effet a pu être prolongé par un rappel du vaccin pendant 127 jours supplémentaires chez trois vaches. Le comportement d'œstrus était absent pendant une période plus longue que l'inactivité ovarienne. Le vaccin a été bien toléré: à part une légère enflure au site d'injection, aucun effet secondaire n'a été observé. Nos résultats démontrent que deux immunisations avec Improvac® sont une méthode facilement applicable pour la suppression de l'activité cyclique chez les vaches pendant une période moyenne de 114 jours. La durée de la suppression du cycle a été prolongée par un rappel de vaccination.

Impact of tissue enzymatic digestion on analysis of immune cells in mouse reproductive mucosa with a focus on γδ T cells.

Mucosal tissues are enriched in γδ T lymphocytes, which maintain epithelial homeostasis, however, the homeostatic mechanisms are still incompletely understood. To elucidate their role in the tissue integrity governance within the female genital mucosa we employed flow cytometry, which is a powerful tool used for the characterization of tissue-resident immune cells, however, often requiring cell release upon tissue enzymatic disaggregation. Here, we analyzed the impact of various proteolytic enzymes in their ability to effectively isolate viable immune cells from the reproductive system of non-pregnant mice. Murine vaginas and uteri were digested using commercially available enzyme blends (liberases) and single enzymes (dispase II and collagenase IV). Among tested enzymes, liberases released the highest number of cells from digested tissues while dispase II and collagenase IV led to a significant decrease in the number of isolated live cells. Also, liberases had only minor detrimental effects on cell viability and detection of CD45, CD3ε, γδ TCR and CD11c positive cells. We found that a single liberase blend called Liberase TL was the most suited for the analysis of γδ T cells in the reproductive tract. By examining two distinct phases of the estrous cycle - estrus and diestrus, characterized by high and low epithelial stratification, respectively, we showed that higher numbers of γδ T lymphocytes were present in the latter cycle phase in vagina and uterus. Interestingly, the diestrus-associated increase in γδ T lymphocyte number was also observed in reproductive tract draining lumbar lymph nodes but not in more distant, inguinal lymph nodes. Our data indicate that enzymes used for reproductive mucosa digestion have profound effects on the cell viability and isolation efficiency, which consequently influence the phenotypic and quantitative analysis of immune cells.

Immunization against GnRF in adult cattle: a prospective field study.

BACKGROUND: Suppression of cyclic activity in cattle is often desired in alpine farming and for feedlot cattle, not intended for breeding. A cattle specific anti-GnRF vaccine (Bopriva™) is registered for use in heifers and bulls in different countries. In adult cows vaccinated with Bopriva™, the median period until recurrence of class III follicles was 78 days from the day of the 2nd vaccination and reversibility could be proven, as out of 11 experimental cows 10 cows became pregnant at first, and one cow at second insemination. In the present study, 76 healthy, cyclic Eringer heifers and cows were vaccinated twice with Bopriva™ 3-7 weeks apart, to prevent estrus during alpine pasturing. Blood samples were taken for progesterone and GnRF antibody titer analysis on the day of inclusion (7-9 d before the first vaccination) and at the first vaccination. At the same time, gynaecological examinations were performed. When estrus occurred in the course of the alpine pasturing season, a gynaecological examination was done including analysis of a blood sample (progesterone, anti-GnRF antibody titer). Cows were followed for fertility out to 26 months post second vaccination. RESULTS: Median duration of estrus suppression was 191 days after the second vaccination (when the 2 vaccinations were given 28-35 days apart). From n = 13 cows showing signs of estrus on the alpine pasture, n = 7 could not be confirmed in estrus (serum progesterone value >2 ng/ml, no class III follicles seen using ultrasonography). Median duration between second vaccination and next calving was 496 days (25%/75% quartiles: 478/532 days). CONCLUSION: Bopriva™ induced a reliable and reversible suppression of estrus for more than 3 months in over 90% of the cows.

The reproductive cycle is a pathogenic determinant during gonococcal pelvic inflammatory disease in mice.

Women with asymptomatic Neisseria gonorrhoeae infection are at risk of developing pelvic inflammatory disease (PID) if the bacteria ascend from the endocervix into the uterus and oviducts. Factors that affect disease severity, ranging from mild discomfort to severe inflammation, pain, and infertility, remain elusive. Herein we perform direct transcervical inoculation of N. gonorrhoeae into the uterus of mice to establish an infection that leads to PID. Profoundly different disease outcomes were apparent at different stages of the reproductive cycle. Mice that were infected during the diestrus stage of the reproductive cycle displayed extensive gonococcal penetration into the submucosa, severe inflammation, and clinical signs reflecting discomfort. Meanwhile, infection during the intervening estrus stage showed only modest effects. Furthermore, a gonococcal-specific humoral response was only elicited following the penetrative upper genital tract (UGT) infection during diestrus but not estrus. Strikingly, the potential for antibodies to contribute to protection during re-infection also depends upon the reproductive stage, as antigonococcal antibodies within the genital tract were markedly higher when mice were in diestrus. Combined, this work establishes a robust new model reflecting gonococcal PID in humans and reveals how the reproductive cycle determines the pathogenic outcome of gonococcal infections of the UGT.

Development and application of an ELISA kit for the detection of milk progesterone in dairy cows.

Progesterone (P4) is a steroid gonadal hormone that is mainly produced from the corpus luteum and placenta and has various biological functions, especially reproductive regulation. It is important to establish a specific and sensitive P4 enzyme-linked immunosorbent assay (ELISA) for the study of ovary activity and reproductive disorders. Therefore, we prepared a monoclonal antibody (MAb) in a completed antigen (11α-OH-P4-HS-OVA). Based on the MAb and our previously prepared completed antigen, a highly specific and sensitive ELISA was developed. In the present study, a competitive ELISA for the determination of P4 was described in dairy cow milk. It was found that P4 concentration in milk samples from five pregnant cows was significantly higher than that from five estrus cows. The diagnosis rate for pregnancy and non-pregnancy in 54 dairy cows were 93.3% and 95.8%, respectively, at 19 to 23 days after pregnancy by detecting milk P4 concentration. In summary, the developed ELISA is a potential tool for P4 research and offers an alternative, simple, rapid technique for detecting P4, especially in future large clinical investigations on pregnancy identification and reproductive disorders in dairy farms in China.

Effects of a commercial canine gonadotropin releasing hormone vaccine on estrus suppression and estrous behavior in mares.

We investigated the effect of immunization against gonadotropin releasing hormone (GnRH) using a commercial canine GnRH vaccine on estrus suppression and unwanted estrous behavior in mares. In experiment 1, mares were immunized (n = 6) twice with vaccine (5 mL) given intramuscularly 4 weeks apart or received a control diluent (n = 5). Transrectal ultrasonographic examination of the reproductive tracts was performed three days a week for 40 weeks after initial vaccination. Blood samples were collected weekly for GnRH antibody titer and progesterone concentration determination. In experiment 2, privately-owned mares (n = 12) were immunized twice with vaccine (1 mL) given intramuscularly 4 weeks apart. Blood samples were collected prior to each vaccination as well as 12 and 20 weeks after initial treatment, and transrectal ultrasonographic examinations of the reproductive tracts were performed 12 weeks after the first vaccination. Vaccinated mares in experiment 1 responded with a GnRH antibody titer, progesterone concentrations significantly lower than controls, and cessation of ovarian activity. Vaccinated mares in experiment 2 also responded with a GnRH antibody titer, progesterone concentrations that remained basal for the duration of the study, and cessation of ovarian activity. Owners of vaccinated mares in experiment 2 reported that the number of unwanted estrous behaviors present before vaccination significantly decreased following vaccination. In conclusion, GnRH immunization using a canine GnRH vaccine is an effective method for suppressing estrus and unwanted estrous behavior.

In vivo multiphoton microscopy technique to reveal the physiology of the mouse uterus.

PROBLEM: Pregnancy is a challenge to the maternal immune system as it allows the growing of a semiallogeneic fetus within the uterus. Such tolerance suggests a set of complex cellular distributions and interactions inside the organ. Until now, direct observation of such processes was absent because proper intravital imaging techniques were not available. METHOD: We developed a new two-photon microscope stage together with a set of surgical procedures to provide direct observation of immune cell within the mouse uterus. RESULTS: Using our technique, we observed an accumulation of dendritic cells (DCs) in the uterus during the estrus phase of the estrus cycle. Some of the observed DC clusters were located near the lumen of the uterus or small blood vessels, each situated on the antimesometrium side. CONCLUSION: While two-photon microscopy has become a widely used technology for intravital imaging, new advances in the development of staging and experimental protocols can still push the limits of this technique for exploring new biology. As proof of this, we demonstrated that with specially designed staging and surgical protocols, we observed the formation of DC clusters in the uterus; structures that may play a role in the complex immunology of the uterus-fetal interface.

Coxsackievirus B3-induced myocarditis: infection of females during the estrus phase of the ovarian cycle leads to activation of T regulatory cells.

Transgenic female mice expressing the TNFalpha gene under the cardiac myosin promoter (TNF1.6) develop substantially increased myocarditis and increased numbers of CD4+Th1 (interferon gamma+) cells when infected with coxsackievirus B3 (CVB3) during the diestrus and proestrus phases of the estrus cycle compared to females infected during the estrus and metestrus phases. Cardiac virus titers were increased in females infected in estrus compared to females infected during the other phases. T regulatory cells (CD4+CD25+FoxP3+) were increased in both peripheral blood and inflammatory cells in the heart in females infected during estrus. Exogenous administration of 200 ng/mouse 17-beta-estradiol to females protected against CVB3 induced myocarditis and increased CD4+CD25+FoxP3+ cells. These results demonstrate that hormonal fluctuations occurring in normally cycling females can determine T regulatory cell response and control virus-induced pathogenesis.

Distribution and heterogeneity of mast cells in female reproductive tract and ovary on different days of the oestrus cycle in Angora goats.

The physiological distribution of mast cells (MCs) in the reproductive tract and ovary of 12 Angora goats was determined using light microscopic histochemical techniques. Uterus (corpus uteri and cornu uteri), uterine cervix, uterine tubes (isthmus and ampulla) and ovary samples were obtained by laparatomy from groups of animals during metoestrus, dioestrus and proestrus (days 5, 10 and 16 of the oestrous cycle). Tissues were fixed in Mota's fixative (basic lead acetate) for 48 h and embedded in paraffin. Six-micrometre-thick sections were stained with toluidine blue in 1% aqueous solution at pH 1.0 for 5 min and alcian blue-Safranin at pH 1.0 for 30 min. MCs were generally associated with blood vessels in all reproductive organs. In the uterus, they were concentrated mainly in the close of the uterine gland and deep stroma in the endometrium. Higher MC numbers were observed by toluidine blue staining in the uterus, uterine cervix and uterine tubes on days 10 (corpus uterine: 4.7 +/- 3.8 and cornu uterine: 4.9 +/- 3.5) and 16 (corpus uterine: 5.9 +/- 4.5 and cornu uterine: 5.4 +/- 2.4) of the oestrous cycle compared with day 5 (p < 0.05). Mast cells were not observed in the follicles, the corpus luteum and the underside of the surface epithelium of the ovarian cortex, but were observed in the interstitial cortical stroma and the ovarian medulla. In the ovary, MC numbers were significantly higher on day 16 of the oestrous cycle (cortex: 3.4 +/- 2.4 and medulla: 5.7 +/- 4.5, p < 0.05). Safranin-positive connective tissue MCs were not observed in the uterine tube on any occasion. These results indicate oestrous cycle-related changes in the number and location of MCs in goat reproductive organs.

Impact of sex and age on bone marrow immune responses in a murine model of trauma-hemorrhage.

Although studies have demonstrated that trauma markedly alters the bone marrow immune responses, sex and age are crucial determinants under such conditions and have not been extensively examined. To study this, 21- to 27-day-old (premature), 6- to 8-wk-old (mature), and 20- to 24-mo-old (aged) male and female (proestrus) C3H/HeN mice were sham operated or subjected to trauma (i.e., midline laparotomy) and hemorrhagic shock (30 +/- 5 mmHg for 90 min) followed by fluid resuscitation. Twenty-four hours after resuscitation, bone marrow cells were harvested. Trauma-hemorrhage induced an increased number of the early pluripotent stem cell-associated bone marrow cell subsets (Sca1(+)CD34(-)CD117(+/-)lin(+/-)) in young mice. The CD117(+) proportion of these cell subsets increased in mature proestrus females, but not in males. Aged males displayed significant lower numbers of Sca1(+)CD34(-)CD117(+/-)lin(+/-) cells compared with young male mice. Trauma-hemorrhage also increased development of granulocyte/macrophage progenitor cells (CD11b(+)Gr-1(+)). Proliferative responses to granulocyte macrophage colony-stimulating factor were maintained in mature and aged proestrus females, but decreased in young mice and mature males. Augmented differentiation into monocyte/macrophage lineage in mature and aged proestrus females was observed and associated with the maintained release of TNF-alpha and IL-6. Conversely, increased IL-10 and PGE(2) production was observed in the male trauma-hemorrhage groups. Thus, sex- and age-specific effects in bone marrow differentiation and immune responses after trauma-hemorrhage occur, which are likely to contribute to the sex- and age-related differences in the systemic immune responses under such conditions.

Characterisation of lymphocyte subsets in the equine oviduct.

REASONS FOR PERFORMING STUDY: The equine oviduct is the site of fertilisation and location of embryonic development during the first 5 or 6 days. It therefore has an important influence on mare fertility. Although histopathological changes have been described previously, there is limited information regarding lymphocyte subtypes present in the mucosa of the normal equine oviduct. OBJECTIVES: To characterise the distribution of CD3+, CD4+, CD8+ and B lymphocytes in the equine oviduct from inseminated mares during oestrus and dioestrus, and from noninseminated mares during the immediate post ovulatory period. METHODS: Oviductal tissues were collected from noninseminated mares at oestrus (> 30 mm follicle, n = 4), at Day 1 post ovulation (n = 3) and at dioestrus (Day 7 post ovulation; n = 4). Oviducts were also collected from inseminated mares at Days 1, 2, and 3 post ovulation (n = 4 for each period). Cross-sections of tissues from the ampullar-isthmic junction from each oviduct were snap frozen and cryostat sections stained by the immunoperoxidase technique with monoclonal antibodies directed against equine lymphocyte surface markers for B cells as well as CD3+, CD4+ and CD8+ cells. RESULTS: In all oviductal sections examined, B cells were rare whereas T cells were relatively abundant. The predominant cell type found was the CD8+ phenotype, with a lesser number of CD4+ cells. Among mares, individual variation was large; therefore, although breeding status and stage of oestrous cycle appeared to alter lymphocyte populations, these differences were not significant. CONCLUSIONS AND POTENTIAL RELEVANCE: A population of CD3+, CD4+ and CD8+ cells exists within the mucosal region of the equine oviduct. The density of these cells is similar to that described in the human oviduct. Their function is not currently known, but they may be involved with modulation of the maternal response to the presence of spermatozoa or the early conceptus within the equine oviduct. As our capacity to differentiate these cell types improves, along with the ability to identify the specific cytokines they produce, their functional significance will become more apparent.

Cytokine profile of mouse vaginal and uterus lymphocytes at estrus and diestrus.

It is known that sex hormones regulate IgA and IgG levels in the female reproductive tract. Moreover, antigen presentation by uterine and vaginal epithelial cells is also under strict hormonal control. The effect of the estrous cycle on cytokine secretion by vaginal and uterine lymphoid cells has been examined in mice using simultaneous staining for cytoplasmic cytokines and surface markers after ex vivo culture with PMA/ionomycin in the presence of Brefeldin A, and flow cytometry analysis. Two different mice strains, BALB/c and C57BL/6 mice, were used. The most relevant finding was the increase in the proportion of vaginal cells secreting IFN-gamma at diestrus in both strains of mice. Other cytokines (IL-2 and IL-4) as well as some T cell subsets seemed to be modified in a strain dependent fashion. Data also suggest that NK cells are at least partially responsible for IFN-gamma secretion. Our data indicate that vaginal and uterus lymphoid cells isolated at diestrus were in vivo activated to secrete cytokines after ex vivo culture. IFN-gamma seems to be the key cytokine, since it increases in both strains of mice.

Presence of the P2X(7) purinergic receptor on immune cells that invade the rat endometrium during oestrus.

Eosinophils, macrophages and other leucocytes invade the uterine endometrium during oestrus and play a role in the tissue remodeling and immune responses that occur prior to implantation of the fertilized ovum. Adenosine 5'-triphosphate (ATP) and its metabolites influence uterine function via ATP receptors. In this study, we investigated the presence and localisation of the P2X(7) nucleotide receptor in the cells that infiltrate the uterine endometrium of adult female rats during oestrus at the electron microscope level, using gold-silver pre-embedding immunocytochemical techniques. P2X(7) receptor expression was found in the cytoplasm and the cell membrane of eosinophils, macrophages and fibroblasts in the endometrium during oestrus. These results suggest that ATP-mediated responses may be important in uterine preparation and remodeling before implantation and that this may involve several types of cells. In particular, the presence of P2X(7) receptors on endometrial stromal cells may indicate their involvement in apoptosis and immune and inflammatory responses.

The sow endosalpinx at different stages of the oestrous cycle and at anoestrus: studies on morphological changes and infiltration by cells of the immune system.

The aim of this study was to investigate the morphological changes of the sow endosalpinx and the distribution of leukocytes throughout the oestrous cycle and at anoestrus. Nineteen crossbred sows (Swedish Landrace x Swedish Yorkshire) at late dioestrus (three), prooestrus (three), oestrus (three), early dioestrus (three), dioestrus (three) and anoestrus (four) were used. Oviductal samples from three different parts (isthmus, ampulla and infundibulum), taken immediately after slaughter, were fixed, embedded in plastic resin and stained with toluidine blue or stored in a freezer at -70 degrees C until analysed by immunohistochemistry (prooestrus and anoestrus) with an avidin-biotin peroxidase method. Quantitative and qualitative examinations of oviductal epithelium and subepithelial connective tissue were performed by light microscopy. During all stages, a lower degree of morphological changes (pseudostratification, mitosis and secretory granules) was found in the isthmus compared with ampulla and infundibulum. In ampulla and infundibulum, pseudostratification, mitotic activity and secretory granules of the epithelium were high at prooestrus/oestrus. Cytoplasmic protrusions of epithelial cells with some extruded nuclei were prominent in ampulla and infundibulum at all stages except for oestrus and early dioestrus. Lymphocytes as well as CD2- and CD3-positive cells were the predominant immune cells in the epithelial layer. The numbers of lymphocytes and CD3-positive cells did not differ among segments and stages. Numbers of CD2-positive cells did not differ between prooestrus and anoestrus while the numbers were significantly higher in the infundibulum than in ampulla and isthmus. Neutrophils were only occasionally found and mainly in the infundibulum. In the subepithelial connective tissue layer, the two most commonly observed immune cell types were lymphocytes and plasma cells. The numbers of lymphocytes as well as CD2- and CD3-positive cells was lower in isthmus than in the other segments (p < or = 0.001). Higher numbers of plasma cells (p < or = 0.001) were found in infundibulum than in ampulla and isthmus. The numbers of lymphocytes and plasma cells were not significantly different between stages of the oestrous cycle. However, the number of neutrophils differed and were highest at prooestrus in ampulla and infundibulum. The numbers of CD2-, CD3- and CD79-positive cells did not differ between prooestrus and anoestrus whereas for CD14- and SWC3-positive cells, the numbers were higher at prooestrus (p < or = 0.05) than at anoestrus. In the oviduct, the morphology differed in ampulla and infundibulum with oestrous cycle stages, which indicates an effect by ovarian steroid hormones. The immune cell infiltration was less influenced by cyclic changes. However, the immune cell infiltration (in the connective tissue) in the upper part, especially infundibulum, differed significantly from the one in the lower part, isthmus, indicating different immune functions within various parts of the oviduct.

Myocarditis susceptibility in female mice depends upon ovarian cycle phase at infection.

Female BALB/c mice were infected with coxsackievirus B3 in the diestrus, proestrus, estrus, or metestrus phases of the ovarian cycle. Cycle stage was determined by vaginal smear. All mice were killed 7 days after infection. Females infected in the diestrus and especially the proestrus phases developed myocarditis. CD4+ T cells expressing interferon-gamma (IFNgamma) infiltrate the myocardium in these two phases, whereas CD4+ T cells expressing IL-4 are more frequent during estrus. Cardiac virus titers were determined 15 h and 7 days after infection. No differences in virus titer were seen at 7 days. These studies show that natural hormone variations can have substantial effects on viral pathogenicity presumably due to hormone effects on the immune system.

Effect of immunization with bovine luteinizing hormone receptor on ovarian function in cats.

OBJECTIVE: To determine the effect of immunization with bovine luteinizing hormone receptor (LH-R) on ovarian function of cats. ANIMALS: 9 adult female domestic cats. PROCEDURE: 7 cats were immunized with 0.5 mg of LH-R encapsulated in a silastic subdermal implant (3 x 10 mm); 2 served as control cats. Receptors had 80% specific binding to 125I-human chorionic gonadotropin with a binding capacity of 2,682 pM/mg. Cats received booster injections of LH-R. Cats were induced to ovulate with luteinizing hormone (LH) releasing hormone on day 345. Samples of venous blood and vaginal cells were collected through day 395. Observation of estrus behavior continued until day 516. Serum concentrations of estradiol, progesterone, thyroid gland hormones, LH, and LH-R antibody were determined. RESULTS: LH-R antibody was detected in the sera of immunized cats within 21 days after implantation. Detection of LH-R antibody was associated with suppression of serum progesterone to < or = 0.5 ng/mL during the study period, compared with concentrations of 5 to 10 ng/mL in control cats. Immunized cats did not display signs of estrus. Release of LH after administration of LH-releasing hormone indicated an intact hypothalamic-pituitary axis but poor corpus luteum function. Serum estradiol concentrations remained between 30 to 40 pg/mL in immunized and control cats. With the decrease antibody titers, hormone concentrations returned to a pattern consistent with that during fertility. CONCLUSIONS AND CLINICAL RELEVANCE: Active immunization with LH-R suppressed corpus luteum function in cats. The effect was reversible. An LH-R-based antifertility vaccine may have clinical application in other vertebrates.

Resistance to glomerulosclerosis in B6 mice disappears after menopause.

The frequency of chronic renal failure increases with age, especially in women after menopause. Glomerulosclerosis is a common cause of chronic renal failure in aging. We reported that pre-menopausal female C57BL6 (B6) mice are resistant to glomerulosclerosis, irrespective of the type of injury. However, we now show that B6 mice develop progressive glomerulosclerosis after menopause. Glomerular lesions, first recognized in 18-month-old mice, consisted of hypertrophy, vascular pole sclerosis, and mesangial cell proliferation. Diffuse but moderate mesangial sclerosis and more marked hypertrophy were present at 22 months. At 28 to 30 months the glomerulosclerosis was diffuse and increased levels of type I and type IV collagen and transforming growth factor-beta 1 mRNA were present. Urine albumin excretion was significantly increased in 30-month-old mice. Mesangial cells isolated from 28-month-old mice retained their sclerotic phenotype in vitro. Comparison of the effects of uninephrectomy (Nx) in 20-month-old and 2.5-month-old mice revealed a 1.7-fold increase in urine albumin excretion, accelerated glomerulosclerosis, and renal function insufficiency in 20-month-old Nx mice, but not in 2.5-month-old Nx mice. Glycemic levels, glucose, insulin tolerance, and blood pressure were normal at all ages. Thus, B6 mice model the increased frequency of chronic renal failure in postmenopausal women and provide a model for studying the mechanism(s) of glomerulosclerosis in aging women.

Interleukin-5 transgene expression and eosinophilia are associated with retarded mammary gland development in mice.

Eosinophils are prevalent in the female reproductive tract, where they may contribute to regulation of development and maintenance of epithelial integrity. The present study examined the effects of constitutive interleukin-5 (IL-5) expression and overabundance of eosinophils on the development and function of the mammary gland, uterus, and ovary in mice. Eosinophils were up to 13-fold and 4-fold more abundant in the uterus and mammary gland, respectively, in female IL-5 transgenic (IL-5Tg) mice than in wild-type (Wt) animals. Eosinophils were present in large numbers in regressing corpora lutea in IL-5Tg mice but not in ovaries from Wt mice. Postpubertal mammary gland development was retarded in IL-5Tg mice, with impaired terminal end bud formation and an altered pattern of epithelial cell proliferation across the mammary fat pad coincident with disrupted ductal branching and extension. By 10 wk of age, the ductal tree was complete in both genotypes. Onset of first estrus was also delayed in IL-5Tg mice, but once IL-5Tg mice reached puberty, serum estrogen content across the cycle and estrous cycle duration were normal. The histology of uterine tissue and epithelial cell turnover were unchanged. Capacity to mate and achieve pregnancy was not affected by maternal IL-5 transgene expression, although at Day 18 of gestation, a modest decrease in the fetal:placental weight ratio was observed. Furthermore, parturition and ability to lactate and nurture postnatal pup development were not compromised. These data demonstrate an effect of IL-5 overexpression on ductal morphogenesis during postpubertal mammary gland development that is consistent with a direct regulatory role for eosinophils in these events, but these data also show that eosinophil excess does not have long-term consequences for adult reproductive function.

Corrigendum to "The sow endometrium at different stages of the oestrus cycle: studies on morphological changes and infiltration by cells of the immune system." [Anim. Reprod. Sci. 65 (2001) 95-114].

The aim of this study was to investigate the distribution of leukocytes and the morphological changes of the sow endometrium throughout the oestrous cycle. Fifteen crossbred multiparous sows (Swedish Landrace x Swedish Yorkshire), with an average parity number of 3.4+/-0.7 (mean+/-S.D.) were used. Blood samples were collected from the jugular vein 1 h before slaughter for analyses of oestradiol-17beta and progesterone levels. Uterine samples from the mesometrial side of both horns, taken immediately after slaughter at late dioestrus, prooestrus, oestrus, early dioestrus and dioestrus, were fixed, embedded in plastic resin and stained with toluidine blue. The surface and glandular epithelium as well as subepithelial and glandular connective tissue layers were examined by light microscopy (LM). The significantly highest surface and the glandular epithelium were observed at oestrus and dioestrus, respectively. The largest number of capillaries (underneath the surface epithelium) was found at oestrus. In the surface epithelium, the largest number of intraepithelial lymphocytes (IELs, round nucleus) was found at early dioestrus. The largest number of lymphocytes and macrophages within the glandular epithelium were found at early dioestrus and oestrus, respectively. In the subepithelial connective tissue layer, the most common type of leukocytes during all stages was the lymphocyte. The largest numbers of lymphocytes and neutrophils were found at oestrus while the largest number of eosinophils was found at dioestrus. The dominating cells of the immune system in the connective tissue of the glandular layer were lymphocytes and macrophages. The significantly largest numbers of lymphocytes and plasma cells were found at early dioestrus and dioestrus, respectively. The number of lymphocytes in the connective tissue of the glandular layer and the number of plasma cells in the subepithelial layer were positively correlated with the plasma level of progesterone (P < or = 0.05). The numbers of capillaries and neutrophils in the subepithelial layer underneath the surface epithelium as well as the number of macrophages in both surface and glandular epithelium were positively correlated with the plasma level of oestradiol-17beta (P < or = 0.05). In conclusion, the present study showed a variation om the infiltration and distrobution of lymphocytes, neutrophils, eosinophils, macrophages, mast cells, and plasma cells in the sow endometrium during different stages of the oestrous cycle. Also morphological parameters (e.g. height of surface and glandular epithelium, capillaries density and degree of oedema) varied throughout the oestrous cycle.

Interleukin 2 receptor gamma chain (gamma(c)) knockout mice show less regularity in estrous cycle but achieve normal pregnancy without fetal compromise.

PROBLEM: Implication of cytokines in pregnancy suggested but remains to be established. We studied the effect of cytokines on reproductive functions such as ovulation and pregnancy, with mutant mice lacking interleukin 2 receptor gamma (IL-2Rgamma), the so-called common gamma chain (gamma(c)), which is shared among receptors for multiple cytokines such as IL-2, IL-4, IL-7, IL-9 and IL-15. METHOD OF STUDY: Regularities of estrous cycles were observed by vaginal smear. Ovaries stimulated with postmenopausal serum gonadotropin (PMSG) were examined for the ovarian capacities. The uteri at 13 days of gestation were used for histological analysis of the maternal-fetal interface. RESULTS: The estrous cycles in gamma(c) knockout (gamma(c) KO) mice were irregular compared with wild-type mice, although, the mutants could become pregnant. No uterine natural killer (uNK) cell was found in the uterus at 13 days of pregnancy, and poor decidual formation and thickness of blood vessel walls were observed. Apparent differences were not seen in the numbers and weight of the fetuses between wild type and mutant animals, and fetuses were not compromised throughout pregnancy. CONCLUSIONS: The gamma(c) KO mice showed irregular estrous cycle but they could carry out normal pregnancy despite lacking uNK and cytokines actions of IL-2, 4, 7, 9 and 15.