Cells Of Reproductive Tract Research Articles

Aquaporin-1 expression in visceral smooth muscle cells of female rat reproductive tract.

The aquaporins (AQ-s) are a group of intrinsic membrane proteins which facilitate movement of water across cell membranes; their recent identification in the kidney has led to the reappraisal of the mechanisms and pathways of water movement across epithelia. Aquaporin-1, (CHIP-28) is reported distributed in cardiac myocytes and vascular smooth muscle cells of large arteries. A related protein, AQ-4, has been identified in the sarcolemma of skeletal muscle fibres. We report aquaporin expression in the cell membrane of smooth muscle cells of the rat genital tract; fluorescence immunohistochemistry of rat uterine (fallopian) tube and vagina demonstrated AQ-1 in visceral smooth muscle of these tissues. In the uterine tube, AQ-1 labelling is most pronounced in the innermost longitudinal and the inner cells of the circular muscle layer and is absent from the outer longitudinal muscle layer of the myosalpinx. The possibility of a specific role for AQ-1 in tubal transport by altering the tubal luminal diameter during the estrus cycle is suggested.

Human immunodeficiency virus type 1 infection of cells and tissues from the upper and lower human female reproductive tract.

Viable tissue sections and isolated cell cultures from the human fallopian tube, uterus, cervix, and vaginal mucosa were examined for susceptibility to infection with human immunodeficiency virus type 1 (HIV-1). We examined infectivity by using the monocytotropic strain HIV-1(JR-FL) and several primary isolates of HIV-1 obtained from infected neonates. HIV-1 infection was measured by p24 production in short-term culture and by immunofluorescence detection of HIV-1 Nef and p24 proteins by laser scanning confocal microscopy. Three-color immunofluorescence was used to phenotype HIV-infected cells within tissue sections from each site. Our findings indicate that epithelial, stromal, and dendritic cells and cells with CD14+ CD4+, CD14-CD4-, and CD4+ CD14- phenotypes from the female reproductive tract are infectable with HIV-1. Of importance is the finding that tissues from the upper reproductive tract are susceptible to infection with HIV-1. Moreover, tissue samples from women in all stages of the menstrual cycle, including postmenopausal women (inactive), could be infected with HIV-1. Female reproductive tract cells required a minimum of 60 min of exposure to HIV-1 in order for infection to occur, in contrast to peripheral blood lymphocytes, which became infected after being exposed to HIV-1 for only 1 min. These findings demonstrate that HIV-1 can infect cells and tissues from different sites within the female reproductive tract and suggest that multiple cell types, including epithelial cells, may be targets for the initial infection by HIV-1.

Mucosal immunity in the human female reproductive tract: cytotoxic T lymphocyte function in the cervix and vagina of premenopausal and postmenopausal women.

To investigate the mucosal immune system in the cervix and vagina of premenopausal women in terms of immune cells present and cytolytic capacity of mucosal CD3+ T cells in the lower reproductive tract. Fresh tissue fragments prepared by vibratome sectioning were analyzed for the presence of cells by confocal scanning laser microscopy (CSLM). Isolated reproductive tract cells prepared by enzymatic were analyzed for CD3+ T cell phenotype by FACS analysis and for cytolytic function by an anti-CD3 mAb mediated redirected lysis assay. As determined by CSLM, CD3+ cells as well as macrophages and dendritic cells are distributed throughout the lower female reproductive tract in both the epithelium and subepithelial mucosa. It was found that cervical and vaginal tissues from pre- and post-menopausal women contain CD3+ T cells (CTL) that have cytolytic activity, when measured in an antigen non-specific anti-CD3 mAb mediated redirected lysis assay. These results indicate that the lower reproductive tract of women is immuno-competent as judged by the presence of CD3, CD4, CD8, macrophage, and dendritic cells in the endocervix, ectocervix, and vagina of premenopausal and postmenopausal women. Further, these studies demonstrate that CD3+ T cells with cytolytic activity are present in the cervix and vagina during the proliferative and secretory phases of the menstrual cycle and following menopause.

Activation of androgen receptor in epidermal growth factor modulation of fetal mouse sexual differentiation

Previous studies from this laboratory indicated a role for epidermal growth factor (EGF) in androgen-dependent male sexual differentiation. The mechanism by which EGF modulates male sexual differentiation has not been determined and investigation has been made to assess the role for androgen receptor (AR) in mediating the EGF-induced effect. We report that EGF, like androgen, stabilized the Wolffian duct in the 13-day female specimen, grown in organ culture. Anti-AR, flutamide and cyproterone acetate blocked the Wolffian duct-stabilizing effect of EGF. EGF also induced cell proliferation of the fetal reproductive tract in a dose-dependent manner and a combination of physiological dosages of EGF and androgen-induced cell proliferation synergistically, suggesting an interactive effect of these two drugs. Cyproterone acetate blocked both EGF-induced normal cell proliferation and the synergistic cell proliferation induced by combination of EGF and androgen suggesting a role of AR in the effects of EGF. The role of AR was further assessed by determining the effect of EGF on AR binding directly. It was shown that EGF stimulated androgen binding activity of the male fetal reproductive tract cells significantly by increasing the number of binding sites by 3-fold with slight decrease in binding affinity. Thus, it appears that AR plays a role in mediating EGF-modulation of sexual differentiation.

Coculture of human spermatozoa with reproductive tract cell monolayers can enhance sperm functions better than coculture with Vero cell monolayers.

In order to develop a better system for support of human sperm function in vitro, we conducted studies to evaluate whether reproductive tract cells are better than non-reproductive tract cells as an adjunt in that regard. Human spermatozoa were cocultured with Vero cells, with human oviduct cells and endometrial cells, and without cells (control) for either 1, 4, or 24 hr. Sperm motility was then analyzed with a computer-aided sperm analyzer (CASA-Hamiliton Thron, HTM IVOS Motility Analyzer). Aliquots of spermatozoa incubated for 24 hr were also stained with Hoechst 33258 and FITC-PNA to evaluate the status of acrosome in live cells. Significant differences (P < 0.05) between the oviduct cell and the control groups after 24 hr were evident in the curvilinear velocity (VCL) (81.4 +/- 13.4 vs 60.0 +/- 14.1 microns/sec) and amplitude of lateral head displacement (ALH) (5.2 +/- 0.6 vs 4.1 +/- 0.5 microns). The incidence of acrosome reaction of live sperm was significantly higher in the endometrial cell group than in the controls (25.4 +/- 9.9 vs 6.6 +/- 2.4%; P < 0.001). Coculture with human reproductive tract cells seems to improve some functional parameters of human spermatozoa. Coincubation with such cell lines, especially oviduct cells, might be a feasible approach to optimization of human spermatozoa for assisted fertilization using subfertile or frozen-thawed samples. We think coincubating human spermatozoa with a human reproductive tract cell line, especially oviduct cells, might be a feasible approach in preparing human spermatozoa for assisted fertilizatioin in subfertile and frozen-thawed semen samples.

Insulin-like growth factor binding proteins are transcribed by preimplantation mouse embryos.

Preimplantation embryos have been reported to synthesize insulin-like growth factors (IGF) and their receptors, and reproductive tract fluid has been found to contain insulin and IGF I. In this communication, we report that all stages of preimplantation mouse embryos transcribe IGF binding proteins (IGF-BP) 2, 3 and 4, and that blastocysts also transcribe IGF-BP6. IGF-BP5 was not detected at any preimplantation stage. Reproductive tract cells in proximity to preimplantation mouse embryos transcribe all of IGF-BP2 through 6. Thus studies of the mechanisms of IGF action on preimplantation mouse development must consider the IGF-BP.

Insulin-like growth factor binding proteins are transcribed by preimplantation mouse embryos

Preimplantation embryos have been reported to synthesize insulin-like growth factors (IGF) and their receptors, and reproductive tract fluid has been found to contain insulin and IGF I. In this communication, we report that all stages of preimplantation mouse embryos transcribe IGF binding proteins (IGF-BP) 2, 3 and 4, and that blastocysts also transcribe IGF-BP6. IGF-BP5 was not detected at any preimplantation stage. Reproductive tract cells in proximity to preimplantation mouse embryos transcribe all of IGF-BP2 through 6. Thus studies of the mechanisms of IGF action on preimplantation mouse development must consider the IGF-BP.

Effect of Prolactin Isolated from Human Amniotic Fluid on Cultured Bovine Cells of the Reproductive Tract and Splenocytes

Contents: Prolactin (PRL) was isolated from human amniotic fluid and than fractionated on Concanavalin A‐Sepharose (ConA‐S). Two fractions were tested for the 3H‐thymidine incorporation in the DNA of bovine granulosa, oviductal and stromal cells as well as splenocytes. All cell types reacted to PRL I or PRL II in a dose dependent manner. In general the effect of PRL I was greater than PRL II, and low concentrations of both enhanced the cell proliferation more than higher doses. The incorporating effect of 3H‐thymidine under the influence of PRL I was higher in stromal cells and splenocytes than in granulosa and oviductal cells. PRL II did not have such a stimulating effect, even the presence of 14 ng/ml resulted in a decrease of 3H‐thymidine incorporation in granulosa cells. The reason of this different intensity of tested PRL variants may depend on the molecular heterogeneity. The results indicate that PRL forms possess a cel type‐specific effect on the proliferation of cells of the bovine female reproductive tract and splenocytes.

Coculture of mouse embryos with cells isolated from the human ovarian follicle, oviduct, and uterine endometrium

To examine the specificity of somatic cell support by comparing embryonic development during long-term in vitro coculture with feeder cells derived from the human ovarian follicle, oviduct, and endometrium. Comparative study of murine embryo development and degeneration during 6 days of in vitro coculture. All feeder-cell cultures were beneficial to embryonic development and viability. Few differences were observed between feeder cell types (epithelial or fibroblastic) or cell origin (ovarian follicle, oviductal, or endometrial). Embryos developed to the eight-cell stage in 24 hours whether in coculture (83.6% to 100%) or in media alone (85.2%); however, further development in media alone decreased compared with coculture (15.6% versus 63.4% to 87.7%, plating) and embryo degeneration increased (67.9% versus 5.5% to 19.4%) after 6 days. [1] Coculture of embryos with human reproductive tract cells is beneficial to embryonic development and viability. [2] Human somatic cell support of murine embryos during long-term in vitro coculture is not tissue specific nor dependent on cell type.

Formulation of a complex serum-free medium (CSM) for use in the co-culture of mouse embryos with cells of the female reproductive tract

Co-culture of pre-implantation embryos with cells of the reproductive tract requires a medium that is beneficial to both embryos and cells. However, many studies in this area utilize media originally formulated for specific cell lines. In the present study, a complex serum-free medium (CSM) was formulated on the basis of the ionic compositions of existing embryo culture media and mouse oviductal fluid as well as the concentrations of growth factors that appear to benefit mouse embryo development. The study began by investigating the effect of altering the concentrations of K+ ions (0-40 mM) and sulfate ions (0-10 mM) in embryo culture media on the development of 2-cell mouse embryos. Mouse embryos showed improved cell numbers at the blastocyst stage when cultured in 10 mM K+ compared with Whittingham's T6 medium. Embryos were also cultured in T6 supplemented with bovine serum albumin (BSA) containing various concentrations of insulin, insulin-like growth factors I and II, fibroblast growth factor, and epidermal growth factor. Insulin concentrations of 100 ng mL-1 significantly (P less than 0.05) improved the cell numbers of 2-cell embryos cultured to the morulae and blastocyst stages compared with those cultured in T6 + BSA alone. CSM was formulated on the basis of the results of these experiments and was found to support both improved development of 2-cell mouse embryos and the culture of mouse fibroblast and mouse oviduct cells.

Co-culture of domestic animal embryos

Early bovine and ovine embryos co-cultured on monolayers of oviductal cells and uterine fibroblasts had improved viability and cleaved more frequently than embryos cultured in medium alone or co-cultured on monolayers of non-reproductive tract cells. Co-culture of early bovine and ovine embryos resulted in the development beyond the 8- to 16-cell “blocks” to development in vitro but which lagged behind in vivo development of contemporaneous embryos from superovulated females. Transfer of co-cultured embryos to recipients resulted in successful pregnancies although early embryonic wastage appeared to be high. Co-culture of porcine embryos with uterine fibroblasts improved their developmental scores and their ability to attach to the culture substrate. In several studies, co-culture of embryos on monolayers of cells from the reproductive tract improved the development of embryos more than co-culture with non-reproductive tracts cells. Co-culture may be a valuable tool for identification of interactions between the embryo and reproductive tract cells that play a specific role in the early development of embryos. In addition, co-culture may become a tool for evaluation of viability of ova that have been manipulated by procedures such as splitting, microinjection and nuclear transfer.

Immunohistologic characterization and quantitation of leukocyte subpopulations in human semen

Various major leukocyte subpopulations (granulocytes, monocytes/macrophages, B lymphocytes, helper and suppressor/cytotoxic T lymphocytes) were detectable in human semen by applying monoclonal antibodies (MAbs) specific for pan-leukocyte and leukocyte subpopulation markers. MAb-labeled cells in smears of washed human semen were visualized by a standard streptavidin-biotin immunoperoxidase method. This approach was validated by testing the MAb panel against sections of human testis and epididymis to rule out cross-reactivity with immature germ cells or other reproductive tract cells, and against peripheral blood leukocytes that had been incubated in human semen to ascertain leukocyte marker stability in seminal plasma. When the immunoperoxidase technique was applied to semen smears from 17 proven fertile men and 51 randomly selected infertility patients, the authors found highly varying total leukocyte numbers, ranging from 8970 to 20,520,000 per ejaculate among fertile men (median, 170,000), and from 43,120 to 104,580,000 among infertility patients (median, 1,035,000). Higher means and medians of granulocytes, monocytes/macrophages, and lymphocyte subsets were seen in the infertility group. These results warrant further investigation on the diagnostic utility of semen leukocyte profiles and the possible role of seminal leukocytes in male infertility.

Carbonic anhydrase isoenzymes in certain tissues of the male anf female reproductive tract and red blood cells.

Carbonic anhydrase isoenzymes in certain tissues of the male anf female reproductive tract and red blood cells.