George A. Hill

Development of a method to isolate and culture highly purified populations of stromal and epithelial cells from human endometrial biopsy specimens

Appropriate endometrial maturation is of paramount importance to achieve reproductive success. Practical and ethical considerations require that in vitro methods be available to evaluate regulation of human endometrial function. Additionally, tissue complexity requires separation of individual cell populations. This report describes an improved method for isolation of endometrial epithelial and stromal cells, using biopsy specimens as a tissue source. Separated cells were obtained using selective enzymatic digestion in conjunction with physical separation procedures. Isolated populations exhibited over 95% homogeneity, ascertained immunocytochemically. Using this system, isolated cells from normal endometrium can readily be obtained for in vitro studies. Within the defined conditions of a culture system, important areas of current concern in the endometrium such as ectopic endometrial growth and implantation can be addressed.

Efficacy of a Modified Oxidized Cellulose Fabric in the Prevention of Adhesion Formation

A new fabric of regenerated oxidized cellulose (TC-7) was evaluated for the prevention of postoperative adhesions. Standardized injuries were performed on the uterine fundal serosa, the unilateral uterine cornu, and the contralateral ovary in 24 New Zealand rabbits. TC-7 was applied to the injured surfaces in 12 rabbits utilizing a randomized study design. At second-look laparotomy 3 weeks later, adhesions were graded without knowledge of the treatment category in the 22 rabbits surviving the study. There were no statistically significant differences in adhesion formation between treated and control animals at any of the injury sites. No gross or histologic residua of the TC-7 were evident 3 weeks after intraperitoneal application. Further studies will be needed to clarify the clinical role of this new fabric.

Comparison of human menopausal gonadotropin, clomiphene citrate, and combined human menopausal gonadotropin-clomiphene citrate stimulation protocols for in vitro fertilization

Human menopausal gonadotropins (hMG) and clomiphene citrate (CC), either alone or in combination, are frequently used for in vitro fertilization (IVF) in an attempt to maximize the number of oocytes recovered and the number of embryos transferred. However, direct comparison of the relative efficacy of these protocols in the same institution has been limited. To evaluate this question, the authors examined the outcome of 304 consecutive women attempting IVF. One hundred eighty-one women received hMG, 42 received CC, and 81 received combination hMG/CC. The percentages of women undergoing laparoscopy were not different among the groups (69%, 71%, and 74%, respectively), nor were the rates of oocyte recovery (94%, 100%, and 100%). However, the percentage of women achieving oocyte fertilization (77%, 83%, and 93%) and embryo transfer (73%, 83%, and 90%) were significantly greater among those who had received hMG/CC stimulation. A comparison of hMG/CC with hMG and CC cycles revealed a statistically significant increase in the total number of developing follicles (4.5 +/- 0.3, 3.3 +/- 0.2, and 3.1 +/- 0.3, respectively; P = 0.0137), total oocytes recovered (4.1 +/- 0.3, 3.2 +/- 0.2, and 2.5 +/- 0.2; P = 0.0011), and embryos transferred (2.2 +/- 0.2, 1.4 +/- 0.2, and 1.4 +/- 0.2; P = 0.0013). However, there was no significant difference in the occurrence of ongoing pregnancies. Thus, in terms of the per-patient number of follicles, oocytes, and embryo transfers, combined hMG/CC stimulation appears to be superior to either hMG or CC alone. However, to date the combined regimen has not improved pregnancy rates.

Male partner screening before in vitro fertilization: preselecting patients who require intracytoplasmic sperm injection with the sperm penetration assay

OBJECTIVES To determine the diagnostic accuracy of the sperm penetration assay (SPA) and standard semen parameters for subsequent fertilization in in vitro fertilization-embryo transfer (IVF-ET). DESIGN Prospective study. SETTING Andrology Laboratory, and university research laboratory. PATIENTS Two hundred sixteen couples undergoing male-partner screening before IVF-ET (265 cycles). INTERVENTION(S) Male-partner screening (semen analyses [SA] and SPA), standard IVF-ET procedures, follow-up of fertilization in IVF-ET. MAIN OUTCOME MEASURE(S) Diagnostic accuracy of SA and SPA for prediction of fertilization in IVF-ET. RESULT(S) The SPA predicted IVF fertilization with high negative (84%) and positive (98%) predictive rates, and correct prediction in 88% of cycles. In contrast, sperm concentration, motility, morphology, and complete SA showed poor diagnostic accuracy, with correct prediction of IVF fertilization in 64%, 65%, 45%, and 68% of cycles, respectively. CONCLUSION(S) Very low sperm concentration and/or motility were good predictors of poor IVF fertilization, however, low to normal semen parameters were not predictive of successful IVF fertilization. The SPA is a useful screening tool that predicts IVF fertilization with high diagnostic accuracy. The SPA may be useful to discriminate between those couples with a high probability of normal fertilization in IVF and those with a low probability of normal fertilization that may benefit from assisted fertilization by intracytoplasmic sperm injection (ICSI).

Therapeutic donor insemination: the impact of insemination timing with the aid of a urinary luteinizing hormone immunoassay *

The records of 120 patients undergoing therapeutic donor insemination were reviewed to determine if the use of the enzyme immunoassay of urinary luteinizing hormone (LH) to plan inseminations decreased the number of cycles required to achieve conception. All inseminations were performed with fresh semen. Patients in group 1 (n = 26) utilized urinary LH testing in addition to basal body temperature (BBT) and cervical mucus examinations to time their inseminations, while inseminations in group 2 (n = 94) were timed with only BBT and cervical mucus examinations. The monthly fecundability of patients in group 1 was 0.13, whereas the monthly fecundability of patients in group 2 was 0.11. The mean number of inseminations was 1.4 and 1.6 per cycle for groups 1 and 2, respectively. There were no significant differences between groups 1 and 2 in regard to the number of cycles required to achieve conception. The use of a urinary LH immunoassay for insemination timing offers no benefit over conventional methods of timing (BBT, cervical mucus) when fresh donor semen is used.

Selective fallopian tube cannulation: Initial experience in an infertile population

We have studied a group of 19 patients presenting with proximal tubal occlusion. Seventy-three percent (22/30) of tubes attempted were successfully cannulated. Sixty-two percent (5/8) of tubes remained patent in limited follow-up at 3 to 6 months. We conclude that selective fallopian tube cannulation may provide an alternative to microsurgical tubal anastomosis in selected patients.

Response to repetitive cycles of ovulation induction in the same women

It has been theorized that the administration of human menopausal gonadotropin (hMG) in consecutive menstrual cycles will result in a poor follicular response in the second cycle. To examine this, 50 women undergoing ovulation induction in two consecutive cycles were assessed, using in each the same induction regimen during the initial 5 days. The remainder of each cycle was individualized according to their response. Nine women were anovulatory, 19 were oligoovulatory, and 22 ovulated regularly in unstimulated cycles. In repeat cycles only 3 of 50 had poor follicular development and did not receive human chorionic gonadotropin (hCG); all were anovulatory. Forty-two of 50 of the first cycles had continually rising estradiol (E2), while 43 of 47 of the second cycles had rising E2 patterns. Grouping the peak E2 prior to hCG in the ranges <300, 300–699, 700–1099, and ⩾1100 pg/ml, peaks in the second cycle were similar in 25 of 50, lower in 16, and higher in 9. Only 3 of 9 anovulatory women had similar peaks, as compared to 22 of 41 of the oligoovulatory and regularly ovulating women. Comparing the second to the first cycle, the day of hCG was within 1 day in 28 of 50 women, 2 or more days less than the first cycle in 6, and 2 or more days greater than the first cycle in 11. We conclude that in a successive cycle of ovulation induction (i) the follicular response is impaired in anovulatory women, but (ii) in oligoovulatory or regularly ovulating women, clinically significant differences in the estradiol response do not occur.

EMBRYO DISPOSITION: CHOICES MADE BY PATIENTS AND DONOR OOCYTE RECIPIENTS

OBJECTIVE To compare final embryo disposition between patients and donor oocyte recipients. DESIGN Retrospective study. SETTING Private infertility practice. PATIENT(S) Patients undergoing IVF with embryo cryopreservation. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Final cryopreserved embryo disposition. RESULT(S) A total of 1,262 patients using autologous oocytes had 5,417 embryos cryopreserved. A majority either used their embryos (39%) or continued storage (35%). Of 364 patients, who did not use their remaining 1,406 embryos, 77 (21%) donated 290 embryos to other infertile couples, 41 (11%) donated 160 embryos for research, and 246 (68%) discarded 956 embryos. Concurrently, 272 donor oocyte recipients had 1,233 embryos cryopreserved. A majority either used their embryos (40%) or continued storage (23%). Of 110 recipients that did not use their remaining 455 embryos, 62 (56%) donated 280 embryos to other infertile couples, 6 (6%) donated 31 embryos for research, and 42 (38%) discarded 144 embryos. CONCLUSION(S) In our patient population, a higher proportion of patients with infertility ultimately used or stored their cryopreserved embryos for future reproduction compared with donor oocyte recipients. However, recipients were much more likely to donate to other infertile couples and less likely to discard their remaining embryos compared with patients.

Epithelial cells from normal human endometrium express a tumor-associated glycoprotein (TAG-72) epitope in vitro

Monoclonal antibody B72.3 identifies a tumor-associated glycoprotein (TAG-72) epitope derived from a human breast carcinoma metastasis. Recently, expression of this epitope was noted in normal endometrium during the secretory, but not proliferative, menstrual interval. In light of known hormonal control of normal endometrial growth and differentiation, we investigated in vitro expression of TAG-72 epitope in purified endometrial epithelium cultured under serum-free conditions on Matrigel biomatrix. Cells from secretory endometrium exhibited homogeneous tumor-associated glycoprotein 72 epitope expression. Unexpectedly, epithelium from the proliferative interval developed expression after 5 to 6 days of culture. Epithelial cells from both intervals maintained expression over 12 days of culture without exogenous estradiol and progesterone. Spontaneous, uniform expression of tumor-associated glycoprotein 72 epitope by normal endometrial epithelial cells in vitro is in marked contrast to the cyclic, heterogeneous expression observed in vivo. Such expression also differs from published in vitro observations of cancer cell lines that express this epitope.

Endometrial inadequacy after treatment with human menopausal gonadotropin/human chorionic gonadotropin

The incidence of abnormal endometrial histology in patients undergoing human menopausal gonadotropin/human chorionic gonadotropin (hMG/hCG) treatment during cycles of in vitro fertilization (IVF) or gamete intrafallopian transfer has been reported to be 52% to 91%. The incidence of endometrial inadequacy, as judged by a single late luteal endometrial biopsy, has not been studied in hMG/hCG non-IVF cycles. In the current study, 30 patients (30 cycles) undergoing hMG/hCG treatment were evaluated by an endometrial biopsy. The incidence of endometrial inadequacy was found to be 27%. No preovulatory predictors for endometrial inadequacy could be identified. Therefore, luteal support for patients undergoing non-IVF hMG/hCG cycles should be considered.