Paul Kaplan

Steroid regulation of oncofetal fibronectin expression in human cytotrophoblasts

Oncofetal fibronectin (onfFN) is a uniquely glycosylated form of FN suggested to play a critical role in uterine/placental adherence during pregnancy. In the present study we have examined steroid regulation of onfFN in highly purified preparations (> or = 95%) of cytotrophoblasts isolated from human term placentas. Based on immunoassays, relative to controls, treatment of cytotrophoblasts with 10(-6) M medroxyprogesterone acetate (MPA) down-regulated media levels of onfFN 25, 53, 59, and 62% on days 1, 2, 3 and 4, respectively. The pattern of steroid regulation and levels of total FN were nearly identical to that of onfFN suggesting that chronic steroid treatment regulates synthesis of FN and not its oncofetal glycosylation. MPA treatment induced a 2-fold stimulation in media levels of hCG indicating that increased placental function was associated with steroid-mediated changes in FN expression. Steroid specificity experiments demonstrated that MPA, cortisol, and dexamethasone were potent inhibitors of onfFN expression whereas estradiol (E2), deoxycorticosterone, testosterone, progesterone, and the synthetic progestin OD-14, were not. This suggested that glucocorticoids and not progestins may be the physiologic regulators of placental FN expression and that MPA may mediate its matrix-modifying activity through a glucocorticoid-like mechanism. Treatment of cells with dexamethasone (10(-7) M) did not affect the levels of total protein synthesis or the release of human placental lactogen to the culture medium. This indicated that steroid-mediated down-regulation of onfFN expression in cytotrophoblasts did not result from a general reduction of protein synthesis. Based on densitometric scanning of Western blots, MPA and dexamethasone treatments down-regulated media levels of onfFN 70% relative to control levels. Northern blotting revealed that MPA and dexamethasone mediated a 60-90% reduction in steady state levels of FN mRNA in the presence or absence of E2. Our in vitro model may provide a unique system to evaluate steroidal effects on extracellular matrix (ECM) protein expression. In addition, we suggest that steroids may critically regulate placental ECM protein synthesis, and thus affect trophoblast/uterine adherence throughout pregnancy and expulsion of the placenta and membranes following delivery of the fetus.

Relationship between circulating human chorionic gonadotropin levels and premature luteinization in cycles of controlled ovarian hyperstimulation.

OBJECTIVEnTo determine if premature luteinization (serum P levels > 1.1 ng/mL on or before the day of hCG administration) during controlled ovarian hyperstimulation (COH) is associated with elevated levels of serum hCG.nnnSETTINGnTertiary fertility center.nnnDESIGNnRetrospective evaluation of ovum donors undergoing COH.nnnPATIENTSnForty-four women underwent COH. Comparisons of serum hCG levels and hormonal and cycle characteristics were made between cycles with premature luteinization (group I) and without premature luteinization (group II).nnnRESULTSnGroup I (16 women) were similar to women in group II in age, amount of hMG, and the ratio of FSH:hMG received. Both groups received hCG on similar days, but women in group I had higher peak E2 levels. Serum hCG levels increased and correlated with serum P levels in group I only and were higher on the day of hCG administration (group I 1.8 +/- 0.9 mIU/mL versus group II 1.2 +/- 0.45 mIU/mL; conversion factor to SI unit, 1.00). Peak E2 and LH levels, ampules of hMG and the FSH:LH ratio, and day of hCG administration did not correlate with hCG levels. Human chorionic gonadotropin exposure, as measured by area under the curve, was significantly higher in group I compared with group II.nnnCONCLUSIONnHigher serum levels of hCG and integrated hCG exposure are found in COH cycles with premature luteinization compared with cycles without premature luteinization. Higher hCG levels may be due to decreased clearance of hCG from the circulation and/or the hCG content of hMG.

The effect of female antisperm antibodies on in vitro fertilization, early embryonic development, and pregnancy outcome *

STUDY OBJECTIVEnTo evaluate the extent to which human in vitro fertilization-embryo transfer (IVF-ET) alleviates immunological infertility.nnnDESIGNnRetrospective.nnnSETTINGnIn vitro fertilization program.nnnPATIENTSnThirty-three patients with positive antisperm antibodies undergoing 50 cycles of IVF-ET in which maternal serum was replaced by 5 mg/mL of bovine serum albumin (BSA) comprised the study group. Seventy-one patients with tubal infertility served as controls. In 50 of these, medium was supplemented with 7.5% maternal serum, and 21 were assigned to BSA substitution.nnnRESULTSnPercentage of fertilization in the study group was significantly lower (41 +/- 31; mean +/- SD) than that of controls with maternal serum (77 +/- 15) and BSA (76 +/- 22). Early embryonic quality, as assessed by percentage of cleavage and morphological grading, was found to be inferior in patients with antisperm antibodies. The percentage of advanced embryos (greater than or equal to 4 blastomeres) at the time of transfer was 42 +/- 39 in the study group, compared with 65 +/- 23 and 75 +/- 35 for maternal serum and BSA controls, respectively. Percentage of morphologically favorable embryos (grades 1 and 2 in a 1 to 5 grading system) was 49 +/- 31 in the study group, compared with 78 +/- 35 and 74 +/- 23 for the controls. Percentage of clinical pregnancy was somewhat lower in the study group (12.5%) than in controls with either maternal serum (18%) or BSA (19%).nnnCONCLUSIONSnAntisperm antibodies may have an adverse effect on fertilization and early embryonic development. Female immunological infertility may not be completely alleviated by IVF-ET.

Intrauterine insemination-ready versus conventional semen cryopreservation for donor insemination: a comparison of retrospective results and a prospective, randomized trial

OBJECTIVEnTo compare fecundity rates following intrauterine insemination (IUI) with donor sperm frozen conventionally versus an IUI-ready preparation.nnnDESIGNnBoth retrospective results and a prospective, randomized study where recipients were assigned to one of two sperm cryopreservation methods in each cycle of intrauterine insemination are reported.nnnSETTINGnUniversity-based infertility practice, affiliated private practices, and andrology laboratory.nnnPATIENT(S)nWomen desiring therapeutic insemination in an effort to establish pregnancy.nnnINTERVENTION(S)nIntrauterine insemination with donor sperm frozen conventionally or by an IUI-ready protocol.nnnMAIN OUTCOME MEASURE(S)nCycle fecundity in donor IUI recipients.nnnRESULT(S)nIn a retrospective analysis involving 642 inseminations in 209 recipients, 79 pregnancies were recorded for an overall pregnancy rate of 12.3% per insemination (or cycle): 11.3% with IUI-ready sperm and 13.9% with conventionally preserved sperm. In a follow-up prospective, randomized study, the pregnancy rate for IUI-ready sperm preparations was 36% per cycle (14 of 39) whereas that for conventionally preserved sperm was 19.6% per cycle (9 of 46). Thirteen of the 23 pregnancies occurred in the first study cycle of insemination; only two pregnancies were observed in patients undergoing more than four cycles of insemination.nnnCONCLUSION(S)nCycle fecundity for IUI-ready donor sperm is equivalent to conventional cryopreserved sperm based on both prospective and retrospective assessments.

Developmental Outcomes of Children Born After Assisted Reproductive Technologies

Since the birth of Louise Brown in England more than 25 years ago, an estimated 2 million children have been born worldwide through in vitro fertilization and related assisted reproduction technology (ART) procedures. Most children born after ART are healthy and develop without complications. Largely because of multiple births (twins, triplets, etc), however, some ART offspring experience developmental problems. In addition, recent studies suggest increased risk of prematurity and smaller birth weight after ART. Regular periodic screening and careful developmental follow-up of ART offspring are recommended as well as rigorous follow-up studies of children conceived after these procedures.

The fertilization antigen-1 does not have proteolytic/acrosin activity, but its monoclonal antibody inhibits sperm capacitation and acrosome reaction**Supported in part by grant HD 24425 from the National Institutes of Health (to R.K.N.), Bethesda, Maryland.

OBJECTIVEnTo determine if human sperm surface fertilization antigen exhibits proteolytic or acrosin activity and to investigate the mechanism(s) whereby monoclonal antibody (mAb) to fertilization antigen inhibits human sperm penetration of zona-free hamster ova.nnnDESIGNnProteolytic and acrosin activities of human fertilization antigen were determined. Acrosomal status, acrosin activity, and motion characteristics were evaluated after incubation of human sperm with immunoaffinity-purified mAb to fertilization antigen.nnnSETTINGnAcademic research environment.nnnPARTICIPANTSnFertile donors used as controls for infertile patients for fertility evaluation.nnnINTERVENTIONSnHuman spermatozoa were treated with mAb to fertilization antigen and induced to undergo acrosome reaction using calcium ionophore A23187.nnnMAIN OUTCOME MEASURESnProteolytic and acrosin activities of fertilization antigen. Sperm penetration assay, acrosomal status, and motion parameters.nnnRESULTSnFertilization antigen does not exhibit proteolytic or acrosin activity; however, its mAb completely blocks human sperm penetration of zona-free hamster ova. The mAb to fertilization antigen inhibits ionophore-induced acrosome reaction and blocks development of the hyperactivated state of human sperm cells.nnnCONCLUSIONSnMonoclonal antibody to fertilization antigen blocks fertilization by inhibiting capacitation and acrosome reaction.

The Predictive Value of Zona-Free Hamster Egg Sperm Penetration Assay for Failure of Human in Vitro Fertilization and Subsequent Successful Zona Drilling

The value of various sperm parameters and the zona-free hamster egg sperm penetration assay (SPA) in predicting human in vitro fertilization (IVF) failure and subsequent successful fertilization with zona drilling was assessed. In 19 couples, throughout 31 IVF cycles, a total of 153 oocytes failed to be fertilized. In subsequent 12 cycles with zona drilling, 33 of 131 (25%) were fertilized. The incidence of teratospermia and asthenospermia was significantly higher in the study group than in the control, 74% versus 32% and 42% versus 5%, respectively. Although the mean values for the performance of sperm in SPA and fertilization of human eggs after zona drilling were remarkably similar (28 +/- 6 versus 28 +/- 4), there was no correlation between individual parameters (r = 0.15). Thus, whereas male factor infertility is more likely to be associated with teratoasthenospermia, neither the SPA nor other sperm parameters have any predictive value for failure in IVF. In addition, no criterion of sperm function has yet been identified that would eliminate oligoteratoasthenozoospermic males from consideration of IVF with zona drilling.

The predictive value of zona-free hamster egg sperm penetration assay for failure of human in vitro fertilization and subsequent successful zona drilling**Presented at the 45th Annual Meeting of The American Fertility Society, San Francisco, California, November 13 to 16, 1989.

The value of various sperm parameters and the zona-free hamster egg sperm penetration assay (SPA) in predicting human in vitro fertilization (IVF) failure and subsequent successful fertilization with zona drilling was assessed. In 19 couples, throughout 31 IVF cycles, a total of 153 oocytes failed to be fertilized. In subsequent 12 cycles with zona drilling, 33 of 131 (25%) were fertilized. The incidence of teratospermia and asthenospermia was significantly higher in the study group than in the control, 74% versus 32% and 42% versus 5%, respectively. Although the mean values for the performance of sperm in SPA and fertilization of human eggs after zona drilling were remarkably similar (28±6 versus 28±4), there was no correlation between individual parameters ( r =0.15). Thus, whereas male factor infertility is more likely to be associated with teratoasthenospermia, neither the SPA nor other sperm parameters have any predictive value for failure in IVF. In addition, no criterion of sperm function has yet been identified that would eliminate oligoteratoasthenozoospermic males from consideration of IVF with zona drilling.