Jeffrey A. Keenan

Regression of endometrial explants in a rat model of endometriosis treated with the immune modulators loxoribine and levamisole.

OBJECTIVE To investigate the effects of the immune modulators levamisole and loxoribine in a rat model of endometriosis. DESIGN Prospective, placebo-controlled study. SETTING Hospital-based research facility. ANIMAL(S) Nineteen rats with experimentally induced endometriosis. INTERVENTION(S) Rats were treated with three weekly intraperitoneal injections of levamisole (2 mg per rat; n = 6), loxoribine (1 mg per rat; n = 6), or saline (control; n = 7) and killed 8 weeks after treatment. MAIN OUTCOME MEASURE(S) Histologic and immunohistochemical analysis of endometriotic explants. RESULT(S) The loxoribine-treated group showed marked regression of both epithelial and stromal components. Epithelial regression was noted in the control group, but the epithelium was strikingly preserved in the levamisole group. There were significantly greater numbers of dendritic cells in the explants of animals treated with loxoribine and levamisole. The number of natural killer cells was significantly reduced in loxoribine-treated explants. CONCLUSION(S) Loxoribine, a potent immunomodulatory drug, appeared to cause regression in both stromal and epithelium components in a rat model of endometriosis. Further, specific cell-mediated immune responses in this model of endometriosis were elucidated.

Immunohistochemical Studies of the Adult Human Ovary: Possible Contribution of Immune and Epithelial Factors to Folliculogenesis

PROBLEM: Formation of primordial follicles in adult ovaries could be a cryptic process limited to relatively small areas of the ovarian cortex and occurring during a certain stage of the menstrual cycle. Such an event may require a specific milieu provided by factors involved in developmental processes, i.e., morphoregulatory molecules and macrophages.

Association of mesenchymal cells and immunoglobulins with differentiating epithelial cells

BackgroundMesenchymal-epithelial interactions play an important role in the physiology and pathology of epithelial tissues. Mesenchymal cells either associate with epithelium basement membrane [pericytes and perivascular monocyte-derived cells (MDC)] or reside within epithelium (MDC and T cells). Although intraepithelial mesenchymal cells were suggested to contribute to the epithelium physiology, their association with particular steps in differentiation of epithelial cells, interactions among themselves, and their fate remain unclear. We studied epitopes of mesenchymal cells and their products (immunoglobulins) in stratified epithelium of uterine ectocervix, which is one of the prototypes of complete cellular differentiation from stem into the aged cells.ResultsPerivascular CD14 primitive MDC associated with basal (stem) epithelial cells. Thy-1 pericytes of microvasculature secreted intercellular vesicles, which associated with Ki67 postmitotic epithelial cells expressing MHC class I. Intraepithelial T cells showed an association with veiled type MDC [dendritic cell (DC) precursors] among parabasal cells, and exhibited fragmentation after entering intermediate (mature) epithelial layers. Mature DC secreted CD68 and exhibited fragmentation after reaching mid intermediate layers. Binding of IgM was detected at the top of each layer: in the upper parabasal, upper intermediate, and most surface epithelial cells. IgG was confined to the entire superficial layer.ConclusionsThese data suggest that the phylogenetically and ontogenetically developed hierarchy of mesenchymal cells (MDC, pericytes, T cells) and immunoglobulins (IgM, IgG) accompanies differentiation of epithelial cells from immature into the mature and aged phenotype. Further studies of an involvement of mesenchymal cells in the regulation of tissue homeostasis may bring novel approaches to the prevention and therapy of tissue dysfunctions characterized by permanent tissue immaturity (muscular dystrophy) or accelerated aging (degenerative diseases).

Changes of ovarian interstitial cell hormone receptors and behavior of resident mesenchymal cells in developing and adult rats with steroid-induced sterility.

In the present paper, we report that injection of testosterone propionate (500 microg) during the critical window of rat development (postnatal day 5) induces temporary appearance of aged interstitial cells in developing ovaries (days 7 and 10). Aged interstitial cells showed large size (> or = 12 microm), enhanced androgen receptor (AR) and low estrogen (ER) and luteinizing hormone receptor (LHR) expression. Although normal mature interstitial cells (large size and strong ER and LHR expression) appeared later (day 14), and ovaries of androgenized rats were similar to normal ovaries between days 14 and 35, ovaries of adult androgenized females showed only aged and no mature interstitial cells. Androgenization on day 10 caused the development of aged interstitial cells on day 14, but adult ovaries were normal. Long lasting postnatal estrogenization (estradiol dipropionate for four postnatal weeks) caused in developing and adult ovaries a lack of interstitial cell development beyond the immature state. Immature interstitial cells were characterized by a small size (< or = 7 microm) and a lack of AR, ER and LHR expression. Because the critical window for steroid-induced sterility coincides with the termination of immune adaptation, we also investigated distribution of mesenchymal cells (Thy-1 mast cells and pericytes, ED1 monocyte-derived cells, CD8 T cells, and cells expressing OX-62 of dendritic cells) in developing and adult ovaries. Developing ovaries of normal, androgenized and estrogenized females were populated by similar mesenchymal cells, regardless of differences in the state of differentiation of interstitial cells. However, mesenchymal cells in adult ovaries showed distinct behavior. In normal adult ovaries, differentiation of mature interstitial cells was accompanied by differentiation of mesenchymal cells. Aged interstitial cells in ovaries of androgenized rats showed precipitous degeneration of resident mesenchymal cells. Immature interstitial cells in ovaries of estrogenized rats showed a lack of differentiation of resident mesenchymal cells. These observations indicate that an alteration of interstitial cell differentiation during immune adaptation toward the aged phenotype results in precipitous degeneration of resident mesenchymal cells and premature aging of ovaries in adult rats, and alteration toward immature phenotype results in a lack of differentiation of mesenchymal cells and permanent immaturity of ovaries in adult females.

Thy-1 Differentiation Protein and Monocyte-Derived Cells During Regeneration and Aging of Human Placental Villi*

PROBLEM: The classification of placental villi was reviewed, and regeneration of villous trees in mature human placentae was examined.

Assisted reproduction using donated embryos: outcomes from surveillance systems in six countries

BACKGROUND Embryo donation, though less often performed than other assisted reproductive technology (ART), can represent an attractive option for couples who do not wish to discard their embryos remaining after IVF, and for those who cannot or should not conceive naturally. Clinicians and potential participants could benefit from information comparing outcomes of embryo donation with those of other ARTs, in various countries. METHODS We analyzed outcome information from ART treatment cycles using 2001-2008 data from national surveillance systems in the USA, Canada, the UK, Australia, New Zealand and Finland. We calculated the live birth rate (LBR) with relative risks, the average number of embryos transferred per cycle and the ratio between them (LBR per embryo transferred). We compared outcomes of embryo donation cycles with those for autologous IVF, frozen embryo transfer (FET) and oocyte donation (OD). RESULTS LBRs for embryo donation cycles were 14-33%, compared with 16-28% for autologous FET, 22-35% for autologous IVF and 15-52% for OD. In every country except Australia/New Zealand, and in all countries combined, the LBR for embryo donation approximated that for IVF, with no statistically significant differences in Finland and Canada. The average number of embryos transferred per cycle was 1.5-2.8. The LBR per embryo transferred was 11-12% for donor embryo cycles, compared with 8-11% for autologous FET, 12-15% for autologous IVF and 9-21% for OD. CONCLUSIONS We found that transfer of donated embryos in these countries yields pregnancy outcomes comparable to those of autologous ART procedures. The variation in outcome rates among countries is not entirely explained by the number of embryos transferred. The relatively high success rates and low costs make embryo donation an attractive family building alternative.

Variability of placental expression of cyclin E low molecular weight variants.

Abstract Cyclin E, a G1 cyclin serving to activate cyclin-dependent kinase 2, is the only cyclin gene for which alternative splicing leading to structurally different proteins has been described. Different cyclin E proteins are present in tumor tissues but absent from normal (steady) tissues. Cyclin E contributes to the regulation of cell proliferation and ongoing differentiation and aging. Because trophoblast has invasive properties and differentiates into syncytium and placental aging may develop at term, we examined cyclin E protein variants in human placenta. Placental samples were collected from 27 deliveries between 33 and 41 wk and were compared with ovarian cancer (positive control). Both placental and tumor tissues showed seven cyclin E low molecular weight (LMW) bands migrating between 50 and 36 kDa. Placental expression of cyclin E showed certain variability among cases. Lowest cyclin E expression was detected in normal placentas (strong expression of Thy-1 differentiation protein in villous core and low dilatation of villous blood sinusoids). Abnormal placentas (significant depletion of Thy-1 and more or less pronounced dilatation of sinusoids) showed significant increase either of all (early stages of placental aging) or only certain cyclin E proteins (advanced aging). Our studies indicate that a similar spectrum of cyclin E protein variants is expressed in the placental and tumor tissues. Low cyclin E expression in normal placentas suggests a steady state. Overexpression of all cyclin E proteins may indicate an activation of cellular proliferation and differentiation to compensate for developing placental insufficiency. However, an enhanced expression of some cyclin E LMW proteins only might reflect an association of cyclin E isoforms with placental aging or an inefficient placental adaptation.

Birth of twins unrelated to each other or to their gestational mother

This report records the first documented instance of a birth in which twins, genetically unrelated to each other, were born to a mother genetically unrelated to either of them. After an extensive history of infertility with multiple unsuccessful treatments, a 42-year-old woman gave birth to healthy twins following transfer of three embryos from two different donor sources. DNA testing confirmed that the twins represented both sources. This report adds to the growing body of literature documenting the successes of embryo donation to other couples, and supports the option of embryo mixing for those couples who wish to consider it.

Experiences of Families Inquiring About Donating or Adopting Embryos: Results of an Online Survey

Four hundred thirty-four individuals who had previously inquired with the National Embryo Donation Center (NEDC) or Nightlight Christian Adoptions about donating or adopting embryos responded to an online survey regarding which issues had been raised with them during their inquiry and how well they perceived NEDC and/or Nightlight had answered their questions regarding those issues. In general, respondents perceived that the operational issues surrounding donating or adopting embryos received more attention than the longer-term issues involving family dynamics and relationships. Overall, the inquirers were pleased with the responses to their questions (average >8 on a satisfaction scale of 1–10).

Vascular endothelial growth factor expression in cycling human endometrium**Supported in part by the Physicians’ Medical Education and Research Foundation, University of Tennessee Medical Center, Knoxville, Tennessee and a grant-in-aid (R.J.T.) from The American Heart Association, Indiana Affiliate.††Presented in part at the Sixth International Congress of Reproductive Immunology, Washington, DC, July 19 to 23, 1995.

OBJECTIVE To determine the spatial distribution of vascular endothelial growth factor protein in human endometrium and to assess temporal fluctuations in vascular endothelial growth factor gene expression and variant isoform production by stromal and epithelial cells during the menstrual cycle. DESIGN Prospective study design. PATIENTS Early proliferative endometrial biopsies were obtained from women undergoing gynecologic surgery for benign conditions; secretory stage biopsies were obtained from patients undergoing routine infertility investigations without evidence of luteal insufficiency. MAIN OUTCOME MEASURE Immunohistochemical detection of vascular endothelial growth factor protein in endometrial biopsies, analyses of vascular endothelial growth factor RNA expression, and isoform production in intact endometrium and isolated endometrial stromal and epithelial cells. RESULTS Strong vascular endothelial growth factor immunoreactivity was detected in the glandular epithelial cells of the secretory endometrium with no discernible immunoreactivity in stroma cells. The proliferative endometrium demonstrated prominent glandular immunoreactivity and faint, inconsistent stromal cell immunoreactivity. Preincubation of the antibody with excess cognate peptide abolished all immunoreactivity. A threefold to sixfold increase in vascular endothelial growth factor messenger RNA expression occurs in secretory versus proliferative endometrial samples. Endometrial stromal and epithelial cell isolates from both phases of the menstrual cycle express VEGF121, VEGF165, and VEGF189 isoforms, however, vascular endothelial growth factor variant 206 was not detected. CONCLUSIONS Expression of vascular endothelial growth factor in the endometrium throughout the menstrual cycle suggests that vascular endothelial growth factor may promote the vascular growth, maintenance, and hyperpermeability required for adequate receptivity in the cycling human endometrium.