bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of Intercellular Adhesion Molecule‐1 (ICAM‐1) on Cultured Human Endometrial Stromal Cells and Its Role in the Interaction With Natural Killers
Authors: Viganó Paola , Pardi Ruggero , Magri Barbara , Busacca Mauro , Blasio Anna Maria Di , Vignali Mario
Published: May 2013

PROBLEM: Recent evidence emphasizes the role of natural killer cells (NKs) as potential effectors of peritoneal immune surveillance directed against the outgrowth of endometrial cells, refluxed with menstrual debris, in ectopic sites. This NK‐mediated cytotoxicity toward autologous endometrial antigens seems to be significantly decreased in endometriosis patients.

METHOD: We set up experiments to clarify which molecules are involved in NK‐endome‐trial cell interaction. In particular, we evaluated the surface expression and functional activity of intercellular adhesion molecule‐1 (ICAM‐1), a cell surface glycoprotein that has been identified as one of the ligands for lymphocyte function‐associated antigen‐1 (LFA‐1), present on almost all leucocyte cell types. Immunofluorescence flow cytometry was used to assess ICAM‐1 expression on resting and IL 1β‐activated endometrial stromal cells in culture. Dermal fibroblasts were used as control cells. Cytotoxicity and binding assays by 51Cr release in presence and absence of a specific monoclonal antibody (mAb) against ICAM‐1 were then performed in order to determine the effect of this molecule on NK‐mediated cytotoxic and binding activity toward endometrial stromal cells.

RESULTS: The results of this study indicated that ICAM‐1 expression on endometrial stromal cells seems to be constitutively higher than on dermal fibroblasts and can be up‐regulated upon exposure to IL 1β. Furthermore, a mAb against ICAM‐1 strongly inhibits the binding but not the cytotoxicity of NKs toward endometrial cells. No difference in the expression of this molecule was observed throughout the cycle.

CONCLUSIONS: The presence of ICAM‐1 on human endometrium might relate to the action of the immunocompetent cells in human specific reproductive events.
Immunosuppression by Hydatidiform Mole Trophoblast Is Neutralized by Monoclonal Antibodies to β‐Interferon
Authors: Bennett William A. , Brackin Martha N. , Long Cecil A. , Cowan Bryan D.
Published: May 2013

PROBLEM: In sheep and cattle, trophoblast‐derived interferons serve as signals for the maternal recognition of pregnancy and may regulate the immunologic relationship between the fetus and mother.

METHOD: In this study, soluble extracts prepared from human hydatidiform mole decidua (DE) and trophoblast (HME) were screened for immunosuppressive activity using an interleukin (IL)‐2‐dependent T‐cell line (CTLL2). Antibody neutralization studies were performed with monoclonal antibodies to α‐ and β‐interferon (IFN).

RESULTS: HME suppressed (P < 0.05) IL‐2‐stimulated (2 IU/well) CTLL2 proliferation at doses ranging from 500 (52 ± 2% of control) to 100 (74 ± 5%) μg/ml concentrations. DE also suppressed (P < 0.05) CTLL2 proliferation in a dose‐related fashion from 500 (20 ± 6% of control) to 100 (71 ± 8%) μg/ml doses. Preincubation with the α‐ and β‐IFN antibody preparations had no effect on CTLL2 suppression by the DE sample. In contrast, the β‐IFN antibody partially neutralized the suppressive activity of HME at each of the dilutions tested. The monoclonal antibody to α‐IFN failed to neutralize HME suppression at any of the doses tested.

CONCLUSIONS: These results suggest that hydatidiform mole trophoblast produces a β‐IFN‐like macromolecule that may abrogate maternal rejection responses that are harmful to the developing fetal allograft.
I. Organ Culture of Amniochorionic Membrane In Vitro
Authors: Fortunato Stephen J. , Menon Ramkumar , Swan Kenneth F. , Lyden Timothy W.
Published: May 2013

PROBLEM: The purpose of the study was to develop a novel method of amniochorionic membrane culture aimed at maintaining tissue integrity.

METHOD: Amniochorionic membranes were collected from women prior to labor, undergoing elective cesarean section, with no history of infection or pregnancy related complication. Fetal membranes were maintained in culture for up to ten days. Glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH), a “house keeping” gene and inflammatory cytokine mRNA was detected by reverse transcriptase‐polymerase chain reaction (RT‐PCR) at various periods of culture. mRNA localization was performed by in situ hybridization.

RESULTS: GAPDH gene expression was seen throughout the culture period. Inflammatory cytokine mRNA (IL‐1α, IL‐1β and IL‐6) were also detected during culture. Cellular and tissue morphology appeared normal.

CONCLUSIONS: The culture technique we propose is a simple organ explant system which maintains the morphology and autocrine/paracrine relationships within this tissue.
II. Expression of TNF‐α and TNFR p55 in Cultured Amniochorion
Authors: Fortunato Stephen J. , Menon Ramkumar , Swan Kenneth F.
Published: May 2013

PROBLEM: Preterm labor and PROM are major complications of pregnancy. We have reported the possible role of amniochorionic membrane as it relates to the production of cytokines and the early onset of labor. Amniochorion is capable of responding to an infectious process with the production of IL‐6 and IL‐1β. Here we examine the expression of TNF‐α and TNFR in amniochorion.

METHOD: Amniochorionic membranes were collected and maintained in an organ explant system. Samples were frozen and/or fixed for RT‐PCR, in situ hybridization, and immunocytochemistry.

RESULTS: RT‐PCR demonstrated mRNA for TNF‐α and in situ hybridization localized mRNA in chorion and amnion. Immunocytochemistry demonstrated TNF‐α peptide in amnion but not in chorion. Immunocytochemical localization of TNFR indicates presence of that peptide in both amnion and chorion.

CONCLUSIONS: We conclude that the fetal membranes are sources of TNF‐α and TNFR, supporting our previous work indicating that fetal membranes are active participants in the response to intraamniotic infection.
Decay‐Accelerating Factor Is Expressed in the Human Endometrium and May Serve as the Attachment Ligand for Dr Pili of Escherichia coli
Authors: Kaul Anil , Nowicki Bogdan J. , Martens Mark G. , Goluszko Pawel , Hart Audrey , Nagamani Manubai , Kumar Dhruv , Pham Tuan Q. , Nowicki Stella
Published: May 2013

PROBLEM: We evaluated the hypothesis that different tissue substructures in uteri may express decay accelerating factor (DAF), a complement regulatory protein that also may serve as ligand for bacterial attachment.

METHOD: Purified Dr pili, anti‐Dr pili IgG, anti‐DAF (SCR‐3) IgG, and fluoresceinisothiocyanate‐conjugated secondary IgG were used for binding and inhibition experiments. RESULT: We observed staining of endometrial glands, spiral arterioles, and myometrial arteries with Dr adhesin (pili) and anti‐DAF (SCR‐3) IgG, and found variation in distribution and amount of Dr ligands in different individuals. Anti‐DAF (SCR‐3) IgG blocked the binding of Dr pili to the endometrium.

CONCLUSION: Presence of DAF in endometrium may protect tissues from complement‐induced damage. Differences between individuals in DAF density in the endometrium may affect sensitivity to attachment of Dr‐bearing E. coli and/or complement activation.
The Presence of Cytomegalovirus Antigens in Karyotyped Abortions
Authors: Lijnschoten Gesina van , Stals Frans , Evers Johannes L.H. , Bruggeman Catharina A. , Havenith Michael H. , Geraedts Joseph P.M.
Published: May 2013

PROBLEM: About one out of six pregnancies ends in spontaneous abortion before the 17th week. In more than half of these a chromosome abnormality is present, which explains the abortion. The role of cytomegalovirus (CMV) infections in early pregnancy failure is unclear. If there is a role for CMV, a preponderance of infections would be expected in a chromosomally normal group.

METHOD: The significance of CMV in abortions has been studied by screening 80 spontaneous and nine induced abortions with known karyotype for the presence of phosphoprotein pp65, an early CMV antigen. Also, endometrial biopsies (n = 55) and menstruum (n = 10) were screened. In 11 patients more than one specimen was available for study. RESULT: The protein was present in the glandular epithelium of the decidua of spontaneous as well as induced abortions in 31 of 89 (35%) cases, irrespective of chromosomal constitution. Trophoblastic cells were pp65 positive in 6/89 cases. A total of 17 embryos could be studied, seven of which were positive. Positive staining of embryonic organs correlated strongly with the presence of the antigen in the decidua. The endometrial biopsies and menstrual discharges from women of comparable ages showed the same percentages of positive tests for pp65: 35 and 20%. Histologically positive and negative specimens could not be differentiated other than by the specific staining. In 10 of 11 patients with more than one specimen available, the results were consistent. One patient had two positive and one negative specimen.

CONCLUSION: The findings indicate that in early pregnancy cytomegalovirus is transmitted from the decidual glands to the foetus. However, we have not found indications that the presence of cytomegalovirus infection as shown by the presence of antigen is pathogenic for early pregnancy.
Decreased Beta‐Carotene Levels in Exfoliated Vaginal Epithelial Cells in Women With Vaginal Candidiasis
Authors: Mikhail Magdy S. , Palan Prabhudas R. , Basu Jayasri , Anyaegbunam Akolisa , Romney Seymour L.
Published: May 2013

PROBLEM: Women are more susceptible to vaginal candidiasis when the host immune response is suppressed. The antioxidant nutrient beta‐carotene is postulated to possess immunoenhancing properties. The purpose of this study was to investigate beta‐carotene concentrations in exfoliated vaginal epithelial (EVE) cells in women with vaginal candidiasis.

METHODS: Beta‐carotene levels in EVE cells, collected by a saline lavage technique from 22 women with vaginal candidiasis and 20 normal controls, were analyzed. The diagnosis of vaginal candidiasis was established by the presence of pruritus, white cheesy vaginal discharge, and a positive potassium hydroxide preparation. Beta‐carotene levels were assayed using high pressure liquid chromatography.

RESULTS: Vaginal cell concentrations of beta‐carotene were significantly decreased in women with vaginal candidiasis (P < 0.001).

CONCLUSIONS: Decreased beta‐carotene levels, and possibly other antioxidants, may alter the local immune response resulting in disturbances in the vaginal flora, overgrowth of Candida, and the development of vaginal candidiasis.
Expression of Complement Regulatory Proteins on Human Eggs and Preimplantation Embryos
Authors: FENICHEL PATRICK , DONZEAU MICHELE , CERVONI FLORENCE , MENEZO YVES , HSI BAE‐LI
Published: May 2013

PROBLEM: To investigate the relation between the complement system and reproduction, expression of complement regulatory proteins (C3b receptors and inhibitor of the membrane attack complex) were screened on unfixed human eggs and preimplantation embryos.

METHODS: Unfixed unfertilized oocytes and preimplantation embryos obtained from an in vitro fertilization program were stained by indirect immunofluorescence using monoclonal antibodies raised against membrane cofactor protein, (MCP or CD46), decay accelerating factor (DAF or CD55), protectin (CD59), human C3b/C4b receptor (CR1 or CD35), and major histocompatibility complex class I antigen (MHC class I).

RESULTS: CD55 and CD59 were both expressed by the plasma membrane of unfertilized oocytes and pre‐implantation embryos. CD46 was not expressed by unfertilized oocytes but appeared at the 6‐to‐8 cell stage embryo when human gene expression first occurs. CD35 and MHC class I antigens were not expressed at all on oocytes and preimplantation embryos.

CONCLUSIONS: Selective expression of complement regulatory proteins (DAF and protectin) associated with the lack of MHC class I antigens may represent an immune protective mechanism by which human oocytes and preimplantation embryos escape complement‐mediated damage during their travel through the female genital tract. Furthermore, participation of these complement regulatory proteins including MCP in cell to cell interaction during fertilization and/or implantation cannot be excluded.
Validation of an Embryotoxicity Assay
Authors: ROUSSEV ROUMEN G. , STERN J. JAROSLAV , THORSELL LILLI P. , THOMASON EDWARD J. , COULAM CAROLYN B.
Published: May 2013

PROBLEM: Culture of mouse blastocysts has served as a tool for identifying various embryotoxic factors in human serum. While inactivated, sera from recurrently aborting women inhibit mouse blastocyst development in vitro. Variation in results from individual serum samples has limited the usefulness of this assay in establishing a new classification of idiopathic recurrent spontaneous abortion (RSA).

METHOD: Two‐cell embryos were collected from superovulated mated CB6F1/J mice and cultured in Ham's F‐10 media supplemented with 10% fetal bovine serum (FBS) or tested human serum at 37°C with 5% CO2 and high humidity. Each sample was assayed in triplicate using three mice with at least five embryos from the same mouse per dish. Development was evaluated at 72 h and the frequency of atretic embryos was recorded.

RESULTS: Intrasample (interassay) variation yielded a coefficient of variation of 9%. When repeated, samples from a given individual were evaluated and the coefficient of variation was 8.7%. Interoperator variability was 4% interassay and 2% intraassay. Atresia of embryos was 23% when incubated with FBS (N=122), 21% in FC (N=122), and in the sera of patients with RSA 34.6% (N=95). Results of percentage of atresia from the fertile control group had a nonparametric distribution. Using 2.2 multiples of the median to determine the 95% confidence interval, a threshold at 44.0% of atresia was established.

CONCLUSIONS: The critical step in maintaining low variability in this bioassay is to control mouse variability by averaging the percentage atresia from different mice as embryo donors for each tested serum. A subgroup of 24% (23/95) RSA patients who displayed embryotoxic activity was identified with a specificity of 95% and positive predictive value of 83%, P=0.001.
Isolation of Decidual Lymphocytes From Chorionic Villus Samples: Phenotypic Analysis and Growth in Vitro
Authors: HAYNES MARK K. , FLANAGAN MAUREEN T. , PERUSSIA BICE , JACKSON LAIRD G. , SMITH J. BRUCE
Published: May 2013

PROBLEM: Giemsa stained cell isolates prepared from chorionic villus samples (CVS) contain granulated cells morphologically similar to large granular lymphocytes.

METHOD: Phenotypic characterization of these cellular isolates by two‐color immunofluorescence and subsequent in vitro culture in the presence of recombinant interleukin‐2 (rIL‐2) were done in order to determine whether CVS could serve as a source of decidual lymphocytes.

RESULTS: A major fraction of the CVS‐derived lymphocytes were characterized as decidual NK cells, exhibiting high levels of CD56 expression (CD56+bright), without concomitant expression of CD16. The T cell population present in CVS‐derived lymphocytes contained both CD4+ and CD8+ cells in a ratio somewhat reduced compared to that found in peripheral blood. While both T cells and CD56+bright cells from CVS proliferate in vitro in response to rIL‐2 alone, preferential growth of CD56+bnghtcells was accomplished using a selective culture technique wherein co‐culture with an irradiated, B lymphoblastoid cell line promoted the growth of CD56+ cells.

CONCLUSION: CVS contains decidual NK cells and T cells that proliferate in response to rIL‐2 and/or third party stimulator cells. These culture techniques will allow investigations into the maturation and/or activation of decidual NK cells and T cells.
An 80‐kDa Syncytiotrophoblast Alloantigen Bound to Maternal Alloantibody in Term Placenta
Authors: JALALI G.R. , REZAI A. , UNDERWOOD J.L. , MOWBRAY J.F. , SURRIDGE S.H. , ALLEN W.R. , MATTHIAS S.
Published: May 2013

PROBLEM: We have shown that most of the IgG present on term syncytiotrophoblast, membrane, microvesicles is bound to an 80 kDa protein antigen (R80K).

METHODS: Microvesicles were prepared from term human placenta, and the IgG eluted at pH3.

RESULTS: When IgG antibody was eluted at pH3 and reacted with acid‐treated vesicles of other placentae, the alloantibody always bound to the preparation from which it was obtained, but only to about 10% of acid‐treated preparations from other placentae. A similar polymorphic protein found in association with IgG antibody was found in term horse placentae. Cross‐reactivity of the antibodies between species was not found. Using binding of labelled antibody, complement dependent cytotoxicity and FACS two‐color analysis, the human polymorphic antigen was present on peripheral blood monocytes and B‐lympho‐cytes. The R80k antigen on intact microvesicles was resistant to trypsin, but after acid elution of IgG, trypsin released a soluble 50 kDa fragment which reacted with the acid‐eluted IgG antibody.

CONCLUSION: The presence of antibodies to R80K in all term placentae studied, including first pregnancies, suggests that development of this alloantibody may be a normal requirement for successful pregnancy.
Clinical Significance of Anti‐GM3 Antibodies in Recurrent Pregnancy Loss With Elevated Level of Antiphospholipid Antibodies
Authors: OZAKI YASUHIKO , AOKI KOJI , AOYAMA TOMOMI , KUNIMATSU MITOSHI
Published: May 2013

PROBLEM: The ganglioside‐GM3 neutralizes the reactivity of antiphospholipid antibodies (APLs) to phospholipids in vitro. The question of whether anti‐GM3 antibodies might exert influence in APLs‐positive recurrent pregnancy loss patients who are undergoing prednisolone and aspirin (PSL/ASA) treatment was investigated.

METHOD: The anti‐GM3 antibody assay of sera was accomplished by ELISA. Sera of 56 patients with recurrent pregnancy loss, including 30 APLs‐positive cases given PSL/ASA treatment, were examined.

RESULTS: Patients positive for IgG or IgM type anti‐GM3 antibodies constituted 13/30 (43%) of the APLs‐positive group as compared with only 2/26 (8%) of those who were negative (P<0.01). In pregnant women with APLs‐positive treated with PSL/ASA, live births occurred in only 6/13 (46%) patients with detectable anti‐GM3 antibodies, while in 16/17 (94%) who tested negative for anti‐GM3 antibodies (P<0.01).

CONCLUSIONS: This observation suggests the possibility that presence of anti‐GM3 antibodies may be an indicator for determining the prognosis in recurrent pregnancy loss with elevated level of APLs.
Early Pregnancy Factor (EPF) as a Marker for Detecting Subclinical Embryonic Loss in Clomiphene Citrate‐Treated Women
Authors: SHAHANI SAVITRI K. , MONIZ CLARA L. , GOKRAL JYOSTNA S. , MEHERJI PARVEEN K.
Published: May 2013

PROBLEM: A discrepancy exists between the apparently normal ovulation and the pregnancy rates in women treated with clomiphene citrate (CC). Our previous studies have indicated that immuno‐suppressive “early pregnancy factor” (EPF) is a novel marker to detect subclinical embryonic loss in infertile women.

METHOD: In the present study EPF was used as a marker to detect subclinical embryonic loss in women treated with CC with/without gonadotropins. In some of the women treated with CC, conception was assisted by artificial insemination with husband's semen (AIH).

RESULTS: Our results have indicated that fertilization occurred (EPF + ve) in 47.7% (52/ 109) of women treated with CC with/without gonadotropins; 13.46% (7/52) retained the fetus and continued pregnancy till full term, whereas 78.9% (41/52) did not retain the fetuses. In the group where after stimulation, conception was assisted by AIH, fertilization was observed in 38.24% (26/68), retention in 11.54% (3/26) but subclinical embryonic loss was observed in 80.8% (21/26) cases.

CONCLUSION: Thus, our results have indicated that subclinical embryonic loss may account for some of the discrepancy observed between the apparently normal ovulation and the pregnancy rates in women treated with clomiphene citrate.
Seminal Fluid and the Expression of MHC Class I Antigens in the Placenta of the Rat
Authors: KANBOUR‐SHAKIR AMAL , ARMSTRONG DAVID T. , ROULEAU ANNE , KUNZ HEINZ W. , GILL THOMAS J.
Published: May 2013

PROBLEM: To determine whether seminal fluid influences the expression of MHC class I antigens on the surface of basal trophoblast cells in the placenta of the rat.

METHODS: Transfer of DA × DA embryos into a WF (allogeneic) or DA (syngeneic) recipient made pseudopregnant by hormonal treatment followed by mating with a vasectomized male (seminal fluid) or by mechanical stimulation (no seminal fluid). Antigen expression was determined by electron microscopic immunocytochemistry using the appropriate gold‐labeled monoclonal antibodies.

RESULTS: Seminal fluid did not affect the expression of MHC class I antigens on the surface of the basal trophoblast in either allogeneic or syngeneic matings.

CONCLUSIONS: The suppression of the expression of paternal class I antigens on the surface of the basal trophoblast cells in allogeneic pregnancies most likely occurs at the genome level shortly after fertilization.
Qualitative and Quantitative Analysis of T Lymphocytes During Normal Human Pregnancy
Authors: SABAHI FARZANEH , ROLA‐PLESCZCYNSKI MAREK , O'CONNELL SEAN , FRENKEL LAWRENCE D.
Published: May 2013

PROBLEM: Human reproduction involves contact between cells which are allogeneic to one another, however the fetus not only survives but thrives.

METHODS: Aspects of T‐cell‐mediated immunity during normal human pregnancy were studied. PBMNCs of pregnant and nonpregnant women were stimulated with PHA and cytomegalovirus antigens (CMV). The capacity of stimulated cells to proliferate, to produce IL‐2 and IFN‐γ, to express IL‐2 receptor (IL2R1) and the effect of rIL2 on the proliferation rate of lymphocytes were examined. FACS was utilized for T‐cell subset comparisons.

RESULTS: The proliferation rate, IL‐2, and IFN‐γ synthesis were all significantly impaired at suboptimal concentration of PHA throughout pregnancy. Exogenous rIL‐2 corrected this depression of cell‐mediated immunity (CMI). At optimal concentration of PHA, proliferation rate and production of IFN‐γ and IL‐2 were all decreased. Exogenous rIL‐2 corrected these deficits only in the third trimester. Third trimester pregnant women demonstrated a significant depression of proliferation as well as IL‐2 and IFN‐γ production after CMV stimulation, which was partially corrected by exogenous rIL‐2. FACS analysis suggested that after stimulation by CMV and optimal concentration of PHA, T cells were activated and both CD4+ and CD8+ lymphoblasts expressed normal density of IL‐2R1. With suboptimal PHA, the number of activated CD4+ and CD4+IL2R1+ cells were diminished and CD4+ and CD8+ T lymphoblasts expressed lower number of IL2R1.

CONCLUSIONS: CD4 T helper (Thl) cell function is down regulated progressively during the three trimesters of pregnancy without changes in the quantity of T cell subsets.
Characteristics of Perforin Expressing Lymphocytes Within the First Trimester Decidua of Human Pregnancy
Authors: RUKAVINA DANIEL , RUBEŠA GORDANA , GUDELJ LEA , HALLER HERMAN , PODACK ECKHARD R.
Published: May 2013

PROBLEM: The number of perforin (P)‐positive cells in decidua of pregnancy is larger than that observed in any other pathological condition. The aim was to investigate the distribution and the phenotype of P+ cells.

METHOD: Decidual tissue was obtained from the first trimester vaginal termination of pregnancy. Tissue distribution of P+ cells was analyzed by immunohistochemistry. The method for simultaneous measurement of P and cell surface is presented.

RESULTS: There is no difference in number and distribution of P+ cells between decidua basalis (DB) and decidua parietalis (DP). The percentage of P+ decidual lymphocytes (DL) is two times higher than in peripheral blood lymphocytes (PBL) (55% vs. 27%), and the prevalent phenotype is CD3− CD4− CD8− CD2+ (95%) CD11c+ (68%) and CD56+ (82%). CD56bright+ DL are also Pbright+ and this is the largest DL subpopulation (42.4% DL). Two different subpopulations of CD8+ DL exist: 1) CD8bright+, which are CD3+ CD56− P− and 2) CD8dim+, which are CD3− CD56+ P+.

CONCLUSION: P expressing DL are prevalently nonclassical NK cells (CD16−) with low cytolytic activity but fully equipped with potent cytolytic machinery (pbright+). There are no classical cytotoxic lymphocytes (CTL) (CD3+ CD8+ P+) in the decidua, and all CD8+ P+ cells are CD3− CD56+. The number of P+ cells is even higher in DP in the vicinity of noninvasvie trophoblast, than in DB.
Lack of Magainin‐Like Activity in Human Cervical Tissue
Authors: HANSEN KEITH A. , KILIANSKI JOSEPH , NEWHALL KATHY , WRIGHT GEORGE , STOCK RICHARD J. , ARCHER DAVID
Published: May 2013

PROBLEM: The cervix plays an integral role in innate immunity of the human reproductive tract. Magainins are antimicrobial and spermicidal peptides recently described in human submandibular glands. We investigated the human cervix for magainin‐like peptides.

METHOD: Histologic sections of frozen and paraffin embedded cervical tissue from nine subjects were separately incubated with two rabbit, polyclonal, anti‐magainin antibodies (CB‐2 and CB‐7) to investigate for magainin‐like activity in the human cervix.

RESULTS: There was no specific staining of cervical columnar cells within the endocervical canal or in the endocervical crypts. Magainin‐like immunoreactivity in the human submandibular gland confirmed previous observations.

CONCLUSION: Antigen related to magainin‐like peptides were not discovered in the human cervix.
Histochemical Demonstration of Interleukin‐2 in Decidua Cells of Patients With Preeclampsia
Authors: HARA NAONORI , FUJII TOMOYUKI , OKAI TAKASHI , TAKETANI YUJI
Published: May 2013

PROBLEM: The objective of this study was to gain insight into the immunological aspects of preeclampsia.

METHOD: The presence of interleukin‐2 (IL‐2) protein in the decidua was examined in five preeclamptic patients and seven normal pregnant women employing the immunohistochemistry.

RESULTS: The decidua cells were strongly stained for IL‐2 in four out of five preeclamptic patients, while only very weak, if any, staining was observed in all the uncomplicated pregnant women.

CONCLUSIONS: IL‐2 was clearly present in decidua cells in the preeclamptic setting, suggesting the possible involvement of immune‐activation in the phathophysiology of preeclampsia.
A Subset of Patients With Recurrent Spontaneous Abortion Is Deficient in Transforming Growth Factor β‐2‐Producing “Suppressor Cells” in Uterine Tissue Near the Placental Attachment Site
Authors: LEA RICHARD G. , UNDERWOOD JENNY , FLANDERS KATHY C. , HIRTE HAL , BANWATT DALJEET , FINOTTO SUZETTA , OHNO ISAO , DAYA SALIM , HARLEY CALVIN , MICHEL MAGDY , MOWBRAY JAMES F. , CLARK DAVID A.
Published: May 2013

PROBLEM: To determine if patients with unexplained recurrent miscarriage have a deficiency of decidual immunosuppressor cells that produce transforming growth factor β type 2, as has been found in mice with abortion due to rejection and/or trophoblast failure.

METHODS: Decidual biopsy specimens were taken as near to the placental attachment site as possible under ultrasound guidance from first trimester legal termination (control) patients with recurrent miscarriage and non‐viable pregnancy, and from patients with sporadic missed abortion. The tissue was tested for TGFβ‐2+ suppressor cells by in situ hybridization, immunohistochemistry, and analysis of supernatants.

RESULTS: TGFβ‐2‐related suppressor molecules similar but not identical to those identified in pregnant mice were released by decidual lymphoid cells. Fifty percent of 14 recurrent miscarriage patients showed a lack of suppressor cells and 59% were subnormal in comparison to 20 controls and 5 sporadic miscarriage patients, where 80–85% of the patients had detectable suppressor cells.

CONCLUSIONS: Suppressor cell deficiency is compatible with a role for rejection and/or trophoblast failure in some patients with recurrent miscarriage. Presence of suppressor cells in most patients with missed abortion (4/5) is compatible with an alternative cause of fetal death, similar to findings reported in genetic fetal death mice.
Interleukin‐1β Induces Cyclooxygenase‐2 in Cultured Human Decidual Cells
Authors: KENNARD ELIZABETH A. , ZIMMERMAN PETER D. , FRIEDMAN CHAD I. , KNISS DOUGLAS A.
Published: May 2013

PROBLEM: The purpose of this study was to examine the hypothesis that interleukin‐1β (IL‐1β)‐elicited increases in decidual prostaglandin E2and F2α (PGE2 and PGF2α) biosynthesis are due to the de novo expression of the inducible isoform of cyclooxygenase (i.e., COX‐2).

METHOD: Primary human decidual cell cultures were established from term placentas delivered by cesarean section. After 8 days in vitro, when the cultures secreted immunoreactive prolactin, the cells were incubated for 24 h in serum‐free medium, and then challenged with IL‐1β from 1 to 48 h. PGE2 and PGF2α content in the media were measured by specific radioimmunoassays.

RESULTS: IL‐1β stimulated a time‐dependent enhancement in PGE2 and PGF2α production, with PGF2α synthesis predominating over PGE2. IL‐1β also induced a dose‐dependent increase in the output of both arachidonic acid metabolites. When Northern blots of IL‐1β‐treated and control cells were probed with cDNAs encoding either COX‐1 or COX‐2 isoforms or an oligonucleotide probe encoding a portion of the human β‐actin, we detected a time‐and dose‐dependent increase in the steady‐state levels of COX‐2, but not COX‐1 or β‐actin mRNA transcripts. Moreover, the expression of COX‐2 mRNA in IL‐1β‐stimulated cells was superinduced by preincubation with cycloheximide, but completely abolished by actinomycin D.

CONCLUSIONS: Taken together, the data suggest that COX‐2 mRNA expression is largely responsible for the robust increase in PG formation seen in IL‐1β‐treated decidual cells.
Detection of Antiendometrial Antibodies in Sera of Patients With Endometriosis by Dual‐Colored, Double‐Labeling Immunohistochemical Method and Western Blot
Authors: KIM JUNG GU , KIM CHUL WOO , MOON SHIN YONG , CHANG YOON SEOK , LEE JIN YONG
Published: May 2013

PROBLEM: This study was undertaken to determine whether specific binding activities against endometrial proteins in sera of patients with endometriosis are detectable and, if so, to identify endometrial antigens involved in autoimmunity in endometriosis.

METHOD: Sera from 33 patients with endometriosis and 20 cord sera (controls) were tested against endometria of patients and their protein extracts by dual‐colored, double‐labeling immunohistochemical method, and Western blotting.

RESULTS: Antiendometrial binding activities were detected in sera of 2 (10.0%) control patients and 13 (48.2%) patients with endometriosis by the immunohistochemical method. Endogenous immunoglobulin G (IgG) binding to endometrial proteins had molecular weights (MW) of 26, 28, 54, 85, 107 and 116 kDa. Most sera of both control and patients showed reactivity against endometrial proteins with MW of 34, 36, 56 and 77 kDa. However, there were specific IgG autoantibodies reactive against the endometrial proteins of 71, 92, and 103 kDa in sera of 55.2% (16/29) of patients but not in the control sera. Over 80% (10/12) of patients' sera with binding activities detectable by the immunohistochemical method also tested positive by Western blot analysis.

CONCLUSIONS: These data show that specific IgG antibodies reactive against endometrial antigens are detectable in sera from some patients with endometriosis.
Co‐Expression of Mannose‐Ligand and Non‐Nuclear Progesterone Receptors on Motile Human Sperm Identifies an Acrosome‐Reaction Inducible Subpopulation
Authors: BENOFF SUSAN , RUSHBROOK JULIE I. , HURLEY IAN R. , MANDEL FRANCINE S. , BARCIA MICHELE , COOPER GEORGE W. , HERSHLAG AVNER
Published: May 2013

PROBLEM: To determine whether surface expression of receptors for progesterone and mannose can be used to identify spermatozoa likely to undergo an acrosome reaction after zona binding and to compare the reactivity of these receptors with naturally occurring sperm head‐directed anti‐sperm antibodies (ASAs).

METHOD: Progesterone binding sites on the surface of fresh and capacitated motile human sperm in relation to acrosome status were visualized using a cell‐impermeant progesterone. Free progesterone and/or mannose ligands were compared for percent sperm binding and ability to induce an acrosome reaction. Western blots of sperm proteins localized to the plasma membrane and surface proteins precipitated following passive transfer of serum ASAs were probed with progesterone‐horseradish peroxidase. The effects of the same ASAs on ligand binding and on the induced acrosome reaction were examined.

RESULTS: The two receptors are located in close proximity on a subset of capacitated motile sperm and are coordinately cleared from the plasma membrane overlying the acrosomal cap prior to exocytosis. The surface appearance of functional binding sites for each ligand, however, is regulated by different mechanisms and the progesterone receptor alone is specifically precipitated by ASAs. Passive transfer of ASAs to capacitated sperm selectively inhibits the progesterone‐stimulated acrosome reaction but not the ionomycin‐induced acrosome reaction or the ability of sperm to bind mannose ligands.

CONCLUSIONS: Sperm from fertile donors incubated under capacitating conditions in vitro can be subdivided into acrosome reaction inducible and noninducible subpopulations on the basis of the co‐expression or total absence of these receptors. The combined data indicate that reaction of sperm surface progesterone receptors with ASAs contributes to the acrosome reaction insufficiency observed in anti‐sperm immune infertility.
Generation and Characterization of Monoclonal Antibodies Specific to Surface Antigens of Human Trophoblast Cells
Authors: YOSHIKI TATSUHIRO , YANG YUNG‐YU , LEE YITA , LEE CHI‐YU GREGORY
Published: May 2013

PROBLEM: To generate and utilize specific monoclonal antibodies for routine fetal cell isolation from the maternal circulation.

METHODS: Monoclonal antibodies specific to human trophoblast cell surface antigens were generated and characterized. After cell fusion, antibodies secreted by hybridomas were screened by enzyme‐linked immunosorbent assay and immunohistochemical assays.

RESULTS: By using cultured BeWo choriocarcinoma cells or the membrane fraction of human placenta as the immunogen, seven (BW‐108, 110, 123, 124, HP‐15, 16 and 17) antibodies specific to the surface antigens of trophoblast were produced. They were shown to have little cross‐reactivity to other human tissues. Among the antibodies raised against human sperm, HSA‐10 was also found to cross‐react with human trophoblast, but not detected in other tissues. When immobilized to magnetic beads, these antibodies were shown to react only with BeWo cells in suspension, but not blood cells and ovarian carcinoma cell line, OC‐3‐VGH.

CONCLUSION: Therefore, these antibodies may have potential application in fetal trophoblast cell isolation from the maternal circulation for prenatal genetic diagnosis.
Soluble Tumor Necrosis Factor Receptor in Serum and Urine Throughout Normal Pregnancy and at Delivery
Authors: ARNTZEN KNUT JØRGEN , LIABAKK NINA‐BEATE , JACOBSEN GEIR , ESPEVIK TERJE , AUSTGULEN RIGMOR
Published: May 2013

PROBLEM: To determine the concentration of the two soluble tumor necrosis factor receptors (sTNFR), sp55 and sp75, in healthy pregnant women.

METHOD: Serum and urine samples were longitudinally collected from a group of pregnant women (N=53) five times throughout pregnancy. Maternal and umbilical sera were obtained from some of the deliveries (N=31). The samples were analysed using ELISA based on two monoclonal antibodies (IV4E and 3H5) against the soluble tumor necrosis factor receptors (sp55 and sp75).

RESULTS: Serum concentration of sp55 and sp75 were increased in pregnant women compared to that of nonpregnant controls. Concentration of both sTNFRs increased towards term. Labor was associated with further increase of sp55. Concentrations of sp55 and sp75 in umbilical serum were significantly higher than those of maternal serum. Significant correlations were observed between maternal and umbilical sTNFR concentrations.

CONCLUSIONS: The current study suggests that pregnancy is associated with an activation of mechanisms regulating the biological activities of TNF.
Human Peritoneal Macrophage and T Lymphocyte Populations in Mild and Severe Endometriosis
Authors: BECKER JEANNE L. , WIDEN RAYMOND H. , MAHAN C. SCOTT , YEKO TIMOTHY R. , PARSONS ANNA K. , SPELLACY WILLIAM N.
Published: May 2013

PROBLEM: The aim of this study was to characterize the phenotype of peritoneal lymphocyte and macrophage populations in mild versus severe endometriosis.

METHOD: Using dual staining, antigen expression on peritoneal leukocytes from 24 women with endometriosis and 21 control patients was analyzed by flow cytometry.

RESULTS: All groups had CD4:CD8 ratios of 0.6, with subpopulations of CD8+ cells expressing cytotoxic marker S6F1. Mild and severe endometriosis patients had increased CD3/ DR+ cells, relative to controls. Two populations of macrophages were identified by size in all groups. Mild endometriosis patients had increased percentages of small macrophages expressing CD14 and HLA DQ, compared to controls and severe disease patients. In severe disease patients, antigen expression on small macrophages did not differ from controls, but decreased percentages of large macrophages expressed CD14 relative to controls and mild disease patients.

CONCLUSION: All women with endometriosis exhibit activated peritoneal lymphocytes, whereas macrophage expression of CD 14 is differentially expressed as a function of disease stage. Alterations in the functional capacity of these cells may contribute to the pathophysiology of this disease.
The Ovarian Dysgenesis Normally Induced by Neonatal Thymectomy is Prevented by the Prior Administration of Estrogen
Authors: CHAPMAN JOHN C. , GRIFFIN WILLIAM J. , VASSALO MILO F. , MICHAEL SANDRA D.
Published: May 2013

PROBLEM: Neonatal thymectomy (Tx) and estrogen (E2) administration disrupt the reproductive and immune systems of female mice. The current experiment examined the combined effects of the two procedures on ovarian function, performed in sequence, and in reverse sequence.

METHOD: Groups of (C57BL/6J × A/J)F, (B6A) female mice were given four daily injections of 20 μg estradiol‐17β, either from 0 days to 3 days, or from 3 days to 6 days postpartum. In some groups this regimen was combined with thymectomy performed either prior to steroid injection (TX‐3), or after steroid treatment (TX‐4). Animals were sacrificed between 100 and 110 days of age then ovaries evaluated via light microscopy for dysgenesis and follicular cysts.

RESULTS: When E2 treatment followed Tx, the incidence of ovarian dysgenesis was unchanged (study 1, Tx + E2 = 60% ovarian dysgenesis; Tx = 63% ovarian dysgenesis) (study 2, Tx + E2 = 46% ovarian dysgenesis; Tx = 45% ovarian dysgenesis). In contrast, when E2was given before Tx, ovarian dysgenesis did not occur (study 2, E2 + Tx = 0% ovarian dysgenesis; Tx = 46% ovarian dysgenesis). Ovaries from E2 + Tx animals were characteristic of ovaries from E2‐injected animals without Tx.

CONCLUSION: The results indicate that E2 injection prevents Tx‐induced ovarian dysgenesis, suggesting E2‐activation of an extrathymic pathway for thymus function.
Active Immunization Against LHRH Alone or Combined With LHRH‐Analogue Treatment Impedes Growth of Androgen‐Dependent Prostatic Carcinoma
Authors: LADD ANNA , WALFIELD ALAN , TSONG Y‐Y. , THAU ROSEMARY
Published: May 2013

PROBLEM: To determine whether active immunization against LHRH can serve as treatment for androgen‐dependent prostatic carcinoma.

METHOD: Male rats of Copenhagen X Fisher strain, implanted with Dunning R‐3327 prostatic carcinoma cells were either immunized against LHRH, treated with LHRH‐antago‐nist, or received a combined treatment of active immunization against LHRH and LHRH‐antagonist.

RESULTS: Testicular histology was consistent with infertility in all treatment groups. The rate of tumor growth was inhibited by all three treatment regimens. Tumor size increased by 3.8 ± 1.4 cm2 in the LHRH‐antagonist group, 3.2 ± 1.1 cm2 in the immunized group, and 1.0 ± 0.4 cm2 in the combined treatment group, as compared to 8.2 + 2.6 cm2 in non‐treated control group.

CONCLUSION: LHRH‐antagonist administration combined with immunization against LHRH appeared to exert a synergistic effect. This may be due to the blockade of prostatic LHRH‐like receptors by the antagonist, while androgen depletion was rapidly achieved by LHRH‐antagonist, and maintained by continued gonadotropin suppression caused by active immunization against LHRH once antagonist treatment had been discontinued.
Human Leukocyte Antigen DQ α Sharing Is Not Increased in Couples With Recurrent Miscarriage
Authors: DIZON‐TOWNSON DONNA , NELSON LESA , SCOTT J.R. , BRANCH D. WARE , WARD KENNETH
Published: May 2013

PROBLEM: The results regarding human leukocyte antigen (HLA) DQ a allele sharing in recurrent miscarriage couples are conflicting. The purpose of this study was to determine the frequency of HLA DQ α allele sharing in our unexplained recurrent spontaneous abortion (RSA) patients using modern DNA analytical techniques.

METHODS: DNA was extracted from whole blood samples of 1) 51 couples with at least three miscarriages, and 2) 43 fertile couples (with at least seven children and no known history of recurrent miscarriage). The polymerase chain reaction (PCR) was used to amplify the second exon of the HLA DQ α locus on chromosome 6. Genotypes were identified by allele specific hybridization with 12 sequence‐specific oligonucleotide probes.

RESULTS: 47% of recurrent miscarriage couples and 35% of fertile couples shared no alleles. 47% of recurrent miscarriage couples compared to 58% of fertile couples shared one allele, and 6% of recurrent miscarriage couples and 7% of fertile couples shared two alleles.

CONCLUSIONS: Reproductive partners with unexplained recurrent pregnancy loss have no increased frequency of HLA DQ α allele sharing. It is unlikely that HLA DQ α genotyping will be helpful in the management of patients with RSA.
Reciprocal Alteration in Circulating TJ6+ CD19+ and TJ6+ CD56+ Leukocytes in Early Pregnancy Predicts Success or Miscarriage
Authors: COULAM C.B. , BEAMAN K.D.
Published: May 2013

PROBLEM: TJ6 is a cytokine whose membrane form is regularly expressed on the B lymphocytes of women during pregnancy. In a previous study we have shown that pregnancies that end in a spontaneous abortion are characterized by an increase in natural killer (NK) cells and that NK cells in these pregnancies also expressed TJ6, whereas NK cells from pregnancies that terminate normally neither increase in number nor do they express significant amounts of the cytokine.

METHODS: To evaluate the ability of TJ6 to predict pregnancy outcome, 61 blood samples from pregnant women were studied. Blood samples were drawn between 5 and 12 weeks of gestation and analyzed for lymphocyte expression of TJ6 using Cytoron Absolute flow cytometry and two‐color fluorescence. The percentage of CD19+ (B) cells and CD56+ (NK) cells that expressed TJ6 was calculated, and this percentage was correlated with subsequent pregnancy outcome classified as successful (viable birth) or unsuccessful (abortion, stillbirth). TJ6 CD19 and TJ6 CD56 was determined in 32 blood samples from women with successful pregnancy outcomes and 29 samples from women with unsuccessful pregnancy outcome.

RESULTS: The mean TJ6 CD19 expression for successful pregnancies was 10.6% and for unsuccessful pregnancies was 5.1% (P < 0.03). The mean TJ6 CD56+ percentage of circulating cells that express TJ6 expression for successful pregnancies was 3.3% and for unsuccessful pregnancies was 10.4% (P = 0.02). All unsuccessful pregnancies had less than 90% of circulating CD19 cells express TJ6 and/or greater than 50% of circulating CD56 cells express TJ6. Use of the expression of TJ6 on CD19 and CD56 cells during the first trimester predicts viable pregnancy with a sensitivity of 100%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 100%.

CONCLUSIONS: The work now being reported validates the hypothesis that expression of TJ6 on NK cells obtained from pregnant women predicts the outcome of pregnancy. In addition we show that this prognosticator can be reliably demonstrated early in the first trimester.
Effect of Leukemia Inhibitory Factor on Human Cytotrophoblast Differentiation Along the Invasive Pathway
Authors: BISCHOF P. , HAENGGELI L. , CAMPANA A.
Published: May 2013

PROBLEM: Leukemia inhibitory factor (LIF) is a pleiotropic secreted cytokine that was shown to be essential for blastocyst implantation in mice. Since it is well documented that LIF is produced by the human endometrium, we wondered if this cytokine was capable of modulating the invasive behaviour of human cytotrophoblastic cells (CTB).

METHODS: CTB were isolated and purified from first trimester abortions, separated or not into cells bearing a laminin or a fibronectin receptor (α6β4 or α5β1 respectively) using specific monoclonal antibodies and magnetic particles.

RESULTS: We observed that rhLIF inhibited the secretion of gelatinases and of hCG by CTB but remained without effects on the secretion of fetal fibronectin (fFN). These effects were exerted on different CTB subsets: although rhLIF inhibited the secretion of gelatinases by α6 positive cells, it stimulated the fFN secretion by α5 positive cells. The inhibitory effect of rhLIF on the secretion of hCG was mainly due to its effect on the hCG secretion of α5 positive CTB.

CONCLUSIONS: Taken together these results suggest that in vitro LIF inhibits the differentiation of CTB towards an invasive phenotype by inhibiting the secretion of metalloproteinases, by increasing the deposition of fFN into the extracellular matrix and by inhibiting the differentiation of CTB into syncytium.
Regulation of Human Decidual Cell Macrophage Inflammatory Protein‐1α (MIP‐1α) Production by Inflammatory Cytokines
Authors: DUDLEY DONALD J. , SPENCER SLADE , EDWIN SAM , MITCHELL MURRAY D.
Published: May 2013

PROBLEM: Inflammation of human gestational tissues is a key pathophysiologic event in the genesis of infection‐associated preterm labor. Human gestational tissues produce several inflammatory cytokines after stimulation with bacterial products. These include interleukin‐1β (IL‐1β), tumor necrosis factor‐α (TNFα), and IL‐6. Another class of cytokines includes chemokines of the “C‐C” subclassification such as macrophage inflammatory protein‐1α (MlP‐1α). The purpose of this study was to determine whether cultured human decidual cells produce MIP‐1α in response to other inflammatory cytokines.

METHODS: Various concentrations of IL‐1β, TNFα, IL‐6, and IL‐4 were incubated with confluent monolayer cultures of decidual cells isolated from normal term placentae for 16 h at 37°C, and MlP‐1α concentrations in culture supernatants were measured by ELISA.

RESULTS: We found that incubation of decidual cells with IL‐1β, TNFα, and IL‐4 resulted in significant concentration‐dependent increases in M1P‐1α production. IL‐6 had no effect on MlP‐1α production.

CONCLUSIONS: Our data are the first to show that human decidual cells in culture produce MlP‐1α in response to other inflammatory cytokines. We suggest that decidual cell production of MIP‐1α is an important early event in the pathophysiology of infection‐associated preterm labor.