Preimplantation genetic diagnosis and screening.

Preimplantation genetic diagnosis (PGD) identifies genetic abnormalities in preimplantation embryos prior to embryo transfer. PGD is an exciting technology that may improve the likelihood of a successful pregnancy and birth for five distinct patient groups: (1) those with infertility related to recurrent miscarriages or unsuccessful in vitro fertilization (IVF) cycles, (2) those with unexplained infertility, (3) advanced maternal age, (4) severe male factor infertility, and (5) couples at risk for transmitting a hereditary disease to their offspring. PGD is always performed following an IVF cycle where multiple oocytes are retrieved and fertilized. Sophisticated techniques such as multiprobe, multicolor fluorescence in situ hybridization are used to test single cells for structural or numerical chromosome abnormalities, whereas the polymerase chain reaction, linkage analysis, and DNA sequencing are used to analyze single cells for disease-specific DNA mutations. PGD allows one to transfer only those embryos identified as being free of genetic abnormalities, thus potentially increasing the implantation rate and decreasing the miscarriage rate. These technologies identify embryos free of specific genetic abnormalities and may increase the likelihood of achieving the patient's goal: the birth of a healthy infant.

The adverse effect of hydrosalpinges on in vitro fertilization pregnancy rates and the benefit of surgical correction.

OBJECTIVE: To test the hypothesis that IVF-ET pregnancy rates (PRs) for patients with tubal factor infertility are decreased in patients with hydrosalpinges and that surgical correction reverses this effect. DESIGN: Retrospective chart review. SETTING: Private practice IVF-ET program. PATIENT(S): Patients (n = 160) undergoing 238 cycles of IVF-ET were stratified into groups based on the presence of hydrosalpinges and whether surgical correction had been performed. Patients >39 years old and patients with male factor infertility were excluded from the study. INTERVENTION(S): Patients with hydrosalpinges were offered surgical correction. MAIN OUTCOME MEASURE(S): Clinical pregnancy defined by an intrauterine gestational sac. RESULT(S): Patients with hydrosalpinges had significantly decreased implantation rates and PRs per transfer (2.8% and 8.5%, respectively) than patients with tubal factor infertility but without hydrosalpinges (15.7% and 38.6%). Surgical correction improved implantation rates and PRs in patients with prior failed cycles (16.1% and 37.5%) and in patients undergoing surgery before IVF-ET (21.8% and 51.7%). The type of surgery performed did not affect success rates in the small number of patients evaluated. CONCLUSION(S): The presence of a hydrosalpinx during an IVF-ET cycle results in significant decreases in implantation rates and PRs. Surgical treatment of hydrosalpinges before IVF-ET cycles improves implantation rates and PRs.

Modulation of implantation-associated integrin expression but not uteroglobin by steroid hormones in an endometrial cell line.

In order to test the hypothesis that integrin and uteroglobin (UG) expression in cultured endometrial cells are affected by hormone treatment, Ishikawa-CH endometrial cancer cells were cultured and exposed to oestradiol or oestradiol and progesterone regimens and assayed using immunohistochemistry. We evaluated the intensity of immunohistochemical staining for the integrin monomers alpha(v) and beta1, the dimers alpha(v)beta3 and alpha(v)beta6, and for the secretory protein uteroglobin under various experimental conditions. Cells grown in control media stained positively for the integrin monomers alpha(v) and beta1, the dimer alpha(v)beta3, and for UG. Oestradiol and sequential oestradiol/progesterone reversibly suppressed staining for the dimer alpha(v)beta3. Hormone treatment had no effect on the staining of the beta1 and alpha(v) monomers or UG. The alpha(v)beta6 dimer antibody did not stain under any experimental treatment conditions. These data indicate that expression of the integrin complex alpha(v)beta3 is reversibly suppressed by oestradiol in Ishikawa cells and that these cells may be a good model for studying hormone-driven molecular changes in endometrium.

Use of computerized morphometric analyses of endometrial hyperplasias in the prediction of coexistent cancer.

OBJECTIVE: Our purpose was to determine whether computerized morphometric analysis is predictive of coexistent cancer in uteri that show endometrial hyperplasia in curettings or biopsy specimens. STUDY DESIGN: Forty-five patients with endometrial hyperplasia and 10 patients with well-differentiated cancers diagnosed from curettings or biopsy specimens and treated by hysterectomy at Thomas Jefferson University Hospital between 1989 and 1993 were identified from the pathology department archives. Curettings were analyzed by computerized morphometric analysis at the Free University Hospital in Amsterdam. Pathologists performing the morphometric analyses were blinded to the pathologic diagnoses obtained by examining the hysterectomy specimens. The histopathologic classification of the hysterectomy specimens were used as the end point. RESULTS: Twelve of 45 patients with endometrial hyperplasia (26.7%) by preoperative histopathologic classification showed coexistent carcinoma at hysterectomy. All instances of carcinoma occurred in patients with atypical hyperplasia. Sensitivity of morphometric analysis to predict carcinoma was 100%, with a specificity of 88.5%. The positive predictive value was 83.3%, and the negative predictive value was 100%. A blinded reanalysis of the quantitative analysis in 16 patients showed good reproducibility of this technique (r = 0.93). CONCLUSIONS: Morphometric analysis is useful for predicting which patients with endometrial hyperplasia have coexistent carcinomas. Computerized morphometric analysis may be useful in therapeutic decision making for complex atypical hyperplasia.

Immunocytochemical localization of growth factors and their receptors in human pre-embryos and Fallopian tubes.

We utilized indirect immunocytochemistry to demonstrate the presence of growth factors and their receptors in human pre-embryos and Fallopian tubes. In pre-embryos, only transforming growth factor-alpha (TGF-alpha) and the intracellular domain of epidermal growth factor receptor (EGFR) were found at the 4-cell stage. In 8- to 14-cell pre-embryos, TGF-alpha, the intracellular and extracellular domains of EGFR, and insulin-like growth factor-I and its receptor were found. Antibodies against TGF-alpha stained all Fallopian tube specimens, while the extracellular domains of EGFR was only found in specimens from patients with either blood type A or AB. These results suggest a cross-reactivity between the extracellular domain of the EGFR and blood group antigens. Our novel demonstration of growth factor receptor staining in human pre-embryos shows that growth factor receptor localization is dependent on the developmental stage of human pre-embryos. We have also established a potentially important link between the Fallopian tube which secretes growth factors and the localization of growth factor receptors in pre-embryos. These findings are compatible with the hypothesis that tubal secretions are embryotrophic for the early development of the pre-embryo.

Achieving multiple-order embryo transfer identifies women over 40 years of age with improved in vitro fertilization outcome.

OBJECTIVE: To define factors in patients > or = 40 years that may improve outcome and provide prognosis for success in IVF-ET. DESIGN: Retrospective. SETTING: University infertility center. PATIENTS: Patients (n = 501) undergoing IVF-ET from 1987 to 1994. INTERVENTIONS: IVF-ET (n = 713 cycles) with GnRH-analogue suppression and hMG stimulation. MAIN OUTCOME MEASURES: We evaluated data including age, diagnosis, prestimulation (day 3) FSH and E2, hMG ampules used, days of monitoring, follicle number and size, maximum E2, ova retrieved, cancellation rate, clinical pregnancy, nidation, and miscarriage rates. RESULTS: Overall, patients > or = 40 years had significantly decreased pregnancy rates (PRs), response to stimulation, and increased miscarriage rates. However, if these patients had four or more embryos transferred, their response and PRs (34.4% per ET) were not significantly different from younger women (47.7% per ET). The majority (77.8%) of pregnancies in women > or = 40 years occurred when four or more embryos were transferred. CONCLUSION: A subset (49%) of women > or = 40 years undergoing IVF-ET will respond to ovarian stimulation well enough to result in four or more embryos available for transfer with a resultant PR similar to that observed in younger patients. We recommend consideration of an attempt at IVF-ET before recommending oocyte donation.

Prognostic value of day 3 estradiol on in vitro fertilization outcome.

OBJECTIVE: To evaluate the prognostic value of day 3 E2 levels, independent of day 3 FSH levels, on responses to ovulation induction and subsequent pregnancy rates (PRs) in IVF-ET patients. DESIGN: Prospective, observational. SETTING: University-based tertiary care and private reproductive endocrine-infertility units. PATIENTS AND INTERVENTIONS: A total of 225 patients underwent 292 IVF cycles with luteal phase GnRH agonist suppression and hMG stimulation. MAIN OUTCOME MEASURES: We evaluated response and outcome data including age, day 3 FSH and E2 levels from a menstrual cycle before IVF, ampules of hMG used, maximum E2 level, cancellation rates, and clinical PR. RESULTS: Despite similar age, number of ampules of hMG, and peak E2 levels, patients with an elevated E2 level (E2 > or = 80 pg/mL) (conversion factor to SI unit, 3.671) on day 3 of a cycle before IVF-ET achieved a lower PR per initiated cycle (14.8% versus 37.0%) and had a higher cancellation rate (18.5% versus 0.4%) compared with those with E2 levels < 80 pg/mL. Even when patients with elevated FSH levels (FSH > or = 15 mIU/mL) (conversion factor to SI unit, 1.00) were excluded (leaving 279 cycles), those with an elevated day 3 E2 still had a lower PR per initiated cycle (14.8% versus 38.9%) and maintained a higher cancellation rate (18.5% versus 0.4%). When the day 3 E2 was > or = 100 pg/mL there was a 33.3% cancellation rate and no pregnancies were achieved. CONCLUSION: Patients who presented with an elevated day 3 E2 (> or = 80 pg/mL) in a cycle before IVF-ET had a higher cancellation rate and achieved a lower PR independent of FSH level. A day 3 E2 level, in addition to a day 3 FSH level, appears very helpful in prospectively counseling patients regarding cancellation risk and ultimate IVF-ET success.

Endometrial hyperplasia and the risk of carcinoma.

Recent reports suggest that atypical endometrial hyperplasia diagnosed by biopsy or curettage is accompanied by a higher than expected risk of coexistent invasive cancer. In order to test this hypothesis we reviewed the pathology and clinical history of all patients at our institution who underwent hysterectomy for endometrial hyperplasia with or without cytologic atypia. We found 24 patients of 45 with a preoperative diagnosis of hyperplasia with cytologic atypia, and 21 with simple or complex hyperplasia without atypia. No cancers were found at surgery in the latter group nor were any significant historical differences found between the two groups. Of the patients with atypia, 12/24 (50%) had an endometrial carcinoma and nine patients (37.5%) were stage IB or greater. This is a significantly greater risk than previously reported in the literature. Endometrial hyperplasia with cytologic atypia may carry a higher risk of coexistent invasive endometrial carcinoma than previously believed. Methods to identify those patients at highest risk should be determined.

The predictive value of serum progesterone and 17-OH progesterone levels on in vitro fertilization outcome.

PURPOSE: In order to identify parameters which predict prognosis for success with in vitro fertilization, 17-hydroxyprogesterone and progesterone levels were evaluated in 254 patients undergoing 296 in vitro fertilization cycles. Selected response and outcome data were recorded. RESULTS: Patients with intermediate values of serum progesterone (0.7-0.8 ng/ml) at the time of human chorionic gonadotropin administration achieved significantly higher pregnancy rates than patients with lower (< 0.7 ng/ml) or higher (> 0.8 ng/ml) levels. The clinical pregnancy rates were 46%, 31%, and 27% respectively (P = 0.02). There was no change in 17-hydroxyprogesterone concentration which predicted a higher pregnancy rate. CONCLUSION: Excellent clinical pregnancy rates were noted in cycles with a progesterone level of 0.7-0.8 ng/ml, as well as good results in cycles above 0.8 ng/ml. There is therefore no reason to administer human chorionic gonadotropin at a smaller follicle size to prevent a rise in serum progesterone.

Evaluation of the atypical cytologic smear. Validity of the 1991 Bethesda System.

The 1991 Bethesda System states that atypical squamous or glandular cells of undetermined significance should be further classified as reactive or premalignant/malignant. The validity of this qualification for identification of patients with cervical intraepithelial neoplasia (CIN) was tested. One hundred twenty-four cytologic smears with squamous atypia were reviewed retrospectively by two cytopathologists blind to the colposcopy results. The smears were classified as favoring either reactive or premalignant/malignant processes. Subjective criteria used in the classification were based on the pathologists' experience. All patients underwent colposcopy and selected biopsy under the direction of a gynecologic oncologist. Of the 124 atypical smears, 69 were classified as favoring reactive processes and 55 as favoring premalignant/malignant processes by cytopathologist 1. Cytopathologist 2 classified 68 as reactive and 56 as premalignant/malignant. Colposcopy and selected biopsy revealed the following lesions: 34 cases of human papillomavirus (27.4%), 17 of CIN 1 (13.7%), 4 of CIN 2 (3.2%), 2 of CIN 3 (1.6%) and 67 without pathology (54.0%). All six patients with squamous atypia and underlying CIN 2 and 3 lesions had their cytology classified as premalignant/malignant by the cytopathologists. In these patients this qualification had high sensitivity (100%) and negative predictive value (100%). The 1991 Bethesda System classification above, when applied to patients with squamous atypia, was effective in identifying patients with serious pathologic cervical lesions. If used as a triage method, colposcopy should be reserved for atypical lesions classified as premalignant/malignant, potentially decreasing the cost of health care without decreasing the quality of that care.