The effect of fibroids without cavity involvement on ART outcomes independent of ovarian age.

BACKGROUND: The effect of fibroids that do not distort the endometrial cavity on pregnancy rate (PR) and implantation rate (IR) is controversial. Use of oocyte donor-derived embryos offers an ideal patient population to study the effect of fibroids in patients utilizing assisted reproductive technologies (ARTs). METHODS: We conducted a retrospective cohort study of patients undergoing oocyte donor recipient (ODR) IVF cycles at two tertiary care fertility centres. We examined medical records for the presence of non-cavity-distorting fibroids and evaluated subsequent PR and IR. RESULTS: Three hundred and sixty-nine patients, 94 with fibroids, underwent oocyte donor recipient transfer cycles with fresh embryos. There was no statistical difference in IR (36 versus 38%) or clinical PR (47 versus 54%) between patients with or without fibroids. Neither the location (subserosal versus intramural) and the presence of multiple myomas nor the size of the myomas affected outcomes. Fibroids were more likely to be present in patients with increasing recipient age. CONCLUSIONS: Clinical PR and IR are not affected by the presence of non-cavity-distorting leiomyomata. This evidence does not support myomectomy before ART in patients with asymptomatic fibroids that do not significantly distort the endometrial cavity or cause abnormal uterine bleeding.

Birth after intracytoplasmic sperm injection with use of testicular sperm from men with Kartagener/immotile cilia syndrome.

OBJECTIVE: To describe two cases of in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) with testicular sperm in men with immotile cilia syndromes. DESIGN: Case report. SETTING: A university-based male infertility clinic and assisted reproduction unit. PATIENT(S): Two couples with male factor infertility due to Kartagener/immotile cilia syndrome. INTERVENTION(S): IVF/ICSI with testicular sperm. MAIN OUTCOME MEASURE(S): Semen characteristics, sperm viability, fertilization rate, and pregnancy. RESULT(S): With testicular sperm, the two pronuclear fertilization rates were 63% and 60% in two cases. One case resulted in the birth of normal healthy girl. CONCLUSION(S): With testicular sperm, successful oocyte fertilization after ICSI in couples with male Kartagener/immotile cilia syndrome is possible despite the lack of sperm motility.

A comparison of ICSI outcomes with fresh and cryopreserved epididymal spermatozoa from the same couples.

The published experience with frozen-thawed epididymal spermatozoa and intracytoplasmic sperm injection (ICSI) suggests that fertilization and pregnancy success rates are comparable to those achieved with freshly retrieved spermatozoa. However, no study has exactly compared clinical outcomes between the two IVF/ICSI cycles in the same couples. To formally address this issue, we assessed ICSI outcomes in couples each of whom had had two IVF/ICSI cycles: one using fresh and the second using frozen-thawed epididymal spermatozoa obtained from a single aspiration procedure. From a pool of 101 consecutive patients undergoing IVF/ICSI with epididymal spermatozoa, 19 couples initially used fresh epididymal spermatozoa and subsequently underwent a second IVF/ICSI procedure with frozen-thawed spermatozoa from the same aspiration. Normal (2PN) oocyte fertilization rates, embryo quality and pregnancy rates were compared between the two IVF/ICSI cycles for each couple. In the fresh epididymal sperm group, 58.4% of the injected oocytes fertilized normally compared with 62.0% of the injected oocytes in the frozen-thawed epididymal sperm group, revealing no statistically significant difference. Graded embryo quality also did not differ significantly between the paired IVF/ICSI cycles. The clinical pregnancy rates were 31.6% (6/19) and 36.8% (7/19) in the first and second cycles respectively. All but one pregnancy were singletons. In summary, this study provides strong evidence to support the notion that motile, cryopreserved and thawed epididymal spermatozoa are equal to freshly retrieved spermatozoa for ICSI in couples with obstructive azoospermia.

Ovarian steroid and cytokine modulation of human endometrial angiogenesis.

A key mechanism underlying the cyclical growth of the endometrium is its ability to regenerate a vascular capillary network. In normal cycling human endometrium, angiogenesis is influenced by both endocrine and paracrine factors. Hormonal manipulation of the endometrium, such as that occurring during the use of steroidal contraception, appears to result in capillary proliferation and fragility. As a consequence of these vascular changes, contraceptive users may be predisposed to unpredictable uterine bleeding, which is responsible for the high frequency of contraceptive discontinuation. In this paper we address mechanisms responsible for vascular endothelial cell proliferation in normal and contraceptive steroid-exposed endometria. We propose that regulation of endometrial angiogenesis is mediated indirectly, via steroid and cytokine actions on vascular endothelial growth factor (VEGF), and we present data indicating that VEGF expression in normal endometrial stromal cells is increased by oestrogens and progestins. Three proinflammatory cytokines with angiogenic effects in other systems (i.e. interleukin-1beta, tumour necrosis factor-alpha and interferon-gamma) do not appear to up-regulate VEGF expression in normal endometrial stromal cells. Well-characterized in-vitro models in conjunction with immunohistochemistry provide useful experimental systems to study endometrial neovascularization under physiological conditions and in those potentially perturbed via the use of contraceptive steroids.

Oestrogen receptor (ER)-alpha and ER-beta isoforms in normal endometrial and endometriosis-derived stromal cells.

Several investigators have noted that hormone-dependent development of endometriosis implants lags behind that of simultaneously analysed eutopic endometrium. With the recent discovery of the oestrogen receptor-beta (ER-beta) isoform, the aim of this study was to investigate whether differences in the expression of ER-alpha and ER-beta might explain this observation. mRNA transcripts from endometrial stromal cells isolated from normal endometrium (NE) and from endometriomas (EI) were analysed using a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) technique. RT-PCR and Southern blot analyses of the two ER isoforms indicated that NE and EI stromal cells predominantly express ER-alpha mRNA, however the relative concentrations of ER isoform mRNA transcripts differed between the two cell types. Steady-state ER-alpha:ER-beta mRNA ratios were 15.5 +/- 2.8 and 5.2 +/- 0.9 respectively for NE and EI cells (P = 0.02). NE and EI stromal cells expressed ER proteins with similar Kd ( approximately 0.9 nM) and densities ( approximately 24 500 binding sites/cell) respectively. Functional ER expression was indicated by an increase in progesterone receptor concentrations of approximately 60% (P = 0.03) after incubation with 10 nM oestradiol. We postulate that differential transcript processing, ligand specificity and biological actions of the ER-alpha and -beta isoforms may influence differential growth responses in normal and ectopic endometrium.

Angiogenesis and macrophage activation in endometriosis.

Recent studies suggest that the symptoms associated with endometriosis are the result of local peritoneal inflammation. Increased concentrations of activated pelvic macrophages and lymphocytes and the elevated levels of specific cytokines and growth factors reviewed above support this hypothesis. The precise roles of these soluble factors are currently unknown, but we propose that a complex network of endometrial cytokines are normally regulated by hormones produced during the ovulatory cycle. Ectopic endometrial implants also are subject to these same endocrine cues. The secretion of these proinflammatory proteins by endometriosis lesions into the peritoneal microenvironment appears to cause a recruitment of capillaries and activated inflammatory cells to the implant. Future therapeutic strategies directed to ameliorate the inflammatory reaction associated with endometriosis should not ignore the likely physiological actions of many of the same bioactive molecules in normal eutopic endometrial function.

Endometriosis and infertility: new concepts.

Endometriosis is a common gynecological disorder with varied symptomatology including chronic pelvic pain, dysmenorrhea, and infertility. The association of endometriosis and infertility has been recognized for years, although definite evidence of causality still eludes us. In this review, we will explore three general concepts that enhance our understanding of the cellular and molecular interactions contributing to the pathophysiology of this disorder and that have steered current research in endometriosis. First, we review evidence of a local peritoneal inflammatory process, supported by the findings of elevated cytokine and growth factor concentrations in peritoneal fluid of affected patients. Second, we propose a role for angiogenic factors in the establishment of ectopic implants. Third, we review evidence for biochemical differences of eutopic and ectopic endometrium in endometriosis patients, which may contribute to both the pathogenesis and sequelae of this important disorder. Through information derived from these research efforts, we hope to develop better therapeutic interventions as adjunctive or alternative therapies to our current medical and surgical armamentarium.

Immunolocalization and regulation of the chemokine RANTES in human endometrial and endometriosis tissues and cells.

Retrograde menstruation is postulated as the initiating event in the histogenesis of endometriosis; however, subsequent steps in the pathogenesis of this common disorder remain poorly characterized. The ip accumulation of activated leukocytes and the infiltration of endometriosis lesions by macrophages and T cells are cytological markers of the inflammatory nature of this syndrome. The apparent recruitment of these leukocytes prompted us to search for chemokine expression by endometriosis cells. We reported previously that pelvic fluid RANTES (regulated upon activation, normal T cell expressed and secreted) concentrations correlated with the stage of endometriosis. In the current study, RANTES messenger ribonucleic acid (mRNA) was identified in normal endometrium and endometriosis lesions, and techniques were developed to localize RANTES protein within these tissues. Using isolated endometrial and endometriosis cell cultures, we demonstrated that RANTES mRNA and protein can be induced by the proinflammatory cytokines tumor necrosis factor-alpha and interferon-gamma in endometrial stromal, but not in epithelial or adenocarcinoma cells. Immunocytochemical studies confirmed the biochemical findings. Metabolic labeling experiments verified that nascent RANTES secreted by cytokine-stimulated endometriosis stromal cells was the mature, 8-kDa protein predicted by the mRNA encoding this chemokine. The results indicate that RANTES is a normal constituent of the eutopic endometrium. We propose that secretion of RANTES by ectopic endometriosis implants provides a mechanism for peritoneal leukocyte recruitment.

Microglandular adenocarcinoma of the endometrium: a form of mucinous adenocarcinoma that may be confused with microglandular hyperplasia of the cervix.

Microglandular adenocarcinoma of the endometrium may cause diagnostic problems because of its bland cytologic appearance and its histologic similarity to benign microglandular hyperplasia of the cervix. We present two cases of microglandular adenocarcinoma and discuss the clinical, pathologic, and immunohistochemical findings. Both patients were postmenopausal women, one of whom was taking exogenous hormones. Endometrial biopsy specimens contained polypoid tissue fragments, within which were microcystic spaces lined by flattened, cuboidal, or columnar cells. Solid nests or sheets of tumor cells surrounded glands in some tissue fragments. The nuclei were uniform and bland, and mitotic figures, although readily identifiable, were infrequent (1 per 10 high-power fields). A majority of tumor cells contained intracytoplasmic mucin. Numerous neutrophils were present in gland lumens and tissues. Immunohistochemical stains for carcinoembryonic antigen and TAG72 (B72.3) revealed focal moderate to intense apical and cytoplasmic staining; immunostains for p53 protein were negative. One carcinoma was confined to the endometrium, whereas the other invaded into the inner one-third of the myometrium. Both patients were well after a limited follow-up of 1 year. Microglandular adenocarcinoma is a distinctive variant of endometrial carcinoma that is most likely a form of mucinous adenocarcinoma.

Ovarian steroid regulation of vascular endothelial growth factor in the human endometrium: implications for angiogenesis during the menstrual cycle and in the pathogenesis of endometriosis.

The human endometrium undergoes a complex process of vascular and glandular proliferation, differentiation, and regeneration with each menstrual cycle in preparation for implantation. Vascular endothelial growth factor (VEGF) is an endothelial cell-specific angiogenic protein that appears to play an important role in both physiological and pathological neovascularization. To investigate whether VEGF may regulate human endometrial angiogenesis, we examined VEGF messenger ribonucleic acid (mRNA) and protein throughout the menstrual cycle and studied the regulation of VEGF by reproductive steroids in isolated human endometrial cells. By ribonuclease protection analysis, VEGF mRNA increased relative to early proliferative phase expression by 1.6-,2.0-, and 3.6-fold in midproliferative, late proliferative, and secretory endometrium, respectively. In histological sections, VEGF mRNA and protein were localized focally in glandular epithelial cells and more diffusely in surrounding stroma, with greatest VEGF expression in secretory endometrium. Consistent with these in vivo results, the treatment of isolated human endometrial cells with estradiol (E2), medroxyprogesterone acetate (MPA), or E2 plus MPA significantly increased VEGF mRNA expression over the control value by 3.1-, 2.8-, and 4.7-fold, respectively. The VEGF response to E2 was rapid, with steady state levels of VEGF mRNA reaching 85% maximum 1 h after the addition of steroid. E2 also caused a 46% increase in secreted VEGF protein, and the combination of E2 and MPA caused an 18% increase. VEGF expression in endometriosis, an angiogenesis-dependent, estrogen-sensitive disease was similar to that seen in eutopic endometrium. Peritoneal fluid concentrations of VEGF were significantly higher in women with moderate to severe endometriosis than in women with minimal to mild endometriosis or no disease. VEGF, therefore, may be important in both physiological and pathological angiogenesis of human endometrium, as it is an estrogen-responsive angiogenic factor that varies throughout the menstrual cycle and is elevated in women with endometriosis.

Interleukin-6 secretion in vitro is up-regulated in ectopic and eutopic endometrial stromal cells from women with endometriosis.

An in vitro model developed to compare human endometrial and endometriosis stromal cells was used to examine basal and stimulated expression of interleukin (IL-6). Stromal cells isolated from normal endometrium (NE) exhibited the lowest level of IL-6 secretion (84 pg/10(6) cells-48 h), whereas those cells isolated from endometriosis implants (EI) secreted the highest concentration of this inflammatory cytokine (46,284 pg/10(5) cells-48 h; P < 0.01). Eutopic endometrial stromal cells from women with endometriosis (EE) expressed an intermediate concentration of IL-6 (831 pg/10(6) cells-48 h). Stimulation of the various cultures with IL-1 beta dramatically augmented stromal cell production of IL-6. The mean concentrations of stimulated IL-6 secretion were 16,257, 37,800, and 264,290 pg/10(5) cells-48 h for NE, EE, and EI cells, respectively (P < 0.03). Exposure of the cell cultures to 10 nmol/L estradiol had little direct effect on IL-6 production. The results indicate that endometrial stromal cells isolated from tissues of women with and without endometriosis express IL-6 under basal and cytokine-stimulated conditions. Differential responsiveness among the three cell sources indicates that NE, EE, and EI cells have intrinsic quantitative differences in cytokine regulation.

Interleukin-8 concentrations are elevated in peritoneal fluid of women with endometriosis.

OBJECTIVE: To investigate the presence of interleukin-8 (IL-8), a macrophage-derived angiogenic factor, in peritoneal fluid (PF) of women with and without endometriosis. DESIGN: Case-control study. SETTING: University hospital. PATIENTS: Eighteen women with laparoscopic findings of mild to severe endometriosis, and nine women with no visual evidence of pelvic pathology. MAIN OUTCOME MEASURES: Peritoneal fluid IL-8 levels were determined using an ELISA. Interleukin-8 concentrations were compared among women with and without endometriosis. Correlation between PF IL-8 concentration and endometriosis stage was investigated. RESULTS: Interleukin-8 was detectable in the PF of a majority of women (67%). Interleukin-8 concentrations were higher in the PF of women with endometriosis than in matched normal controls. A significant correlation between PF IL-8 concentration and endometriosis stage was noted. CONCLUSIONS: We hypothesize that IL-8 is an important angiogenic factor that contributes to the pathogenesis of endometriosis by promoting the neovascularization of ectopic endometrial implants.

Three consecutive pregnancies in a patient with glycogen storage disease type IA (von Gierke's disease).

Glycogen storage disease type IA is associated with metabolic abnormalities that can compromise fetal outcome. Normal outcome can be achieved by maintaining euglycemia throughout gestation. We report three consecutive pregnancies in a patient with glycogen storage disease type IA. The patient, a 35-year-old woman, has been maintained on a regimen of nightly nasogastric or cornstarch feedings for the past 12 years with improving metabolic control, reduced liver size, and no progression of multiple hepatic adenomas. On confirmation of each pregnancy, early in the first trimester nightly feeding was changed from cornstarch ingestion to Polycose by nasogastric intubation, with good metabolic control. During the last trimester of each pregnancy metabolic control showed further improvement, with lowering of lactate, urate, and triglyceride levels. During the first pregnancy unexpected fetal death occurred at 33 weeks. During the last two pregnancies, the patient was admitted at 33 and 34 weeks, respectively, for closer supervision of metabolic status and fetal monitoring. She underwent a cesarean section at 35 weeks 4 days of gestation and was delivered of a girl. She underwent a repeat cesarean section at 35 weeks 2 days for the subsequent gestation and was delivered of a boy. Both infants are healthy and appear to be unaffected by von Gierke's disease. Hepatic adenomas did not enlarge during the pregnancies. Meticulous management resulted in normal pregnancy outcomes in two consecutive gestations. Rapid fetal growth late in the third trimester may require particularly careful supervision to maintain euglycemia.

Isolation, characterization, and comparison of human endometrial and endometriosis cells in vitro.

Endometriosis is a common gynecological disorder of unclear pathogenesis. We have established an in vitro model to investigate phenotypic similarities and differences between normal endometrial and endometriosis cells. Highly purified cultures of epithelial and stromal cells were isolated from normal endometrium and endometriosis implants. Morphological features as well as immunocytochemical markers confirm these isolates as epithelial and stromal cells. Potential hormone responsiveness was established by the documentation of estrogen receptor mRNA in epithelial and stromal cells isolated from both tissue types. Expression of this receptor protein was verified in stromal cells by competitive radioligand binding, revealing comparable receptor numbers and dissociation constants. CA-125 is selectively secreted in similar concentrations by epithelial cells isolated from both tissue types. PRL secretion is selectively exhibited by progestin-stimulated stromal cells from both tissue types. Our findings demonstrate that highly purified epithelial and stromal cells cultured from normal endometrial and endometriosis tissues express the same phenotypic and functional markers as their in vivo counterparts. These cultures provide useful models to identify endometriosis-specific cell products that contribute to the pathogenesis of this disorder.

Peritoneal fluid concentrations of the cytokine RANTES correlate with the severity of endometriosis.

OBJECTIVE: Endometriosis is a common gynecologic disorder in which the concentration and activation of peritoneal macrophages are increased. The goal of this study was to quantify pelvic fluid concentrations of two cytokines involved in macrophage recruitment and activation. STUDY DESIGN: A case-control study of women undergoing pelvic surgery was conducted by collecting peritoneal fluid from 12 women without evidence of endometriosis (controls), 12 with mild, and 12 with moderate to severe endometriosis. Concentrations of RANTES and interferon gamma, soluble cytokines known to recruit and activate macrophages, were quantified by enzyme-linked immunosorbent assays. RESULTS: Pelvic fluid concentrations of RANTES are elevated in women with endometriosis and the levels correlate with the severity of disease. By contrast, concentrations of interferon gamma appear unaffected by the presence of or severity of endometriosis. CONCLUSION: The findings indicate that RANTES, a cytokine with potent chemotactic activity for human monocytes, may play an important role in the recruitment of peritoneal macrophages in endometriosis.