Pär Hellberg

E-cadherin expression in human epithelial ovarian cancer and normal ovary

The ovarian surface epithelium (OSE) is the origin of the majority of human ovarian cancers. These adenocarcinomas are characterized by initial local growth followed by spreading into the peritoneal cavity at later stages of tumor progression. The cell‐adhesion molecule E‐cadherin (E‐cad) plays an important role in maintaining tissue integrity. Disappearance or impaired function of E‐cad have often been associated with tumor formation and invasion in vivo and in vitro. The cell‐specific expression of E‐cad was investigated in normal human ovaries (n = 12), in benign (n = 5) and borderline (n = 4) ovarian epithelial tumors and in adenocarcinomas of different stages and histological grades (n = 18), by immunohistochemistry and immunoblotting. An ovarian cancer cell line (NIH‐OVCAR3) was used as a reference. The epithelial origin of the cells was confirmed with cytokeratin (AE1/AE3) staining. In normal ovaries, the expression of E‐cad was limited to inclusion cysts or deep clefts lined with OSE, whereas no staining of the OSE could be demonstrated at the surface of the ovary. In contrast, benign and borderline tumors uniformly expressed E‐cad. This was observed in malignant tumors of all stages despite their degree of differentiation. E‐cad was also present in metastasis from such tumors. The cell‐specific expression of E‐cad in inclusion cysts of normal ovaries and in epithelial layers of borderline tumors indicates a role for E‐cad in the early events of the progression to a malignant phenotype. E‐cad was not downregulated in later stages of ovarian cancer progression. Int. J. Cancer 74:275‐280, 1997.

Wnt-signalling pathway in ovarian epithelial tumours: increased expression of β-catenin and GSK3β

Beta-catenin is involved in both cell–cell adhesion and in transcriptional regulation by the Wingless/Wnt signalling pathway. Alterations of components of this pathway have been suggested to play a central role in tumorigenesis. The present study investigated, by immunohistochemistry and immunoblotting, the protein expression and localisation of β-catenin, adenomatous polyposis coli (APC), glycogen synthase kinase 3β (GSK3β) and lymphocyte enhancer factor-1 (Lef-1) in normal human ovaries and in epithelial ovarian tumours in vivo and in vitro. Immortalised human ovarian surface epithelium and ovarian cancer cell cells (OVCAR-3) expressed β-catenin, APC, GSK3β and Lef-1. Nuclear staining of β-catenin and Lef-1 were demonstrated only in OVCAR-3 cells. There were significant increases of β-catenin and GSK3β, while APC was reduced in ovarian cancer compared to the normal ovary. Beta-catenin and Lef-1 were coimmunoprecipitated in ovarian tumours, but not in the normal ovary. Nuclear localisation of β-catenin or Lef-1 could not be demonstrated in the normal ovary or in the ovarian tumours. The absence of nuclear localisation of β-catenin could be due to an increased binding to the cadherin–α-catenin cell adhesion complex. In fact, we have earlier reported an increased expression of E-cadherin in ovarian adenocarcinomas. In summary, this study demonstrates an increase in the expression of components of the Wingless/Wnt pathway in malignant ovarian tumours. The increase suggests a role for this signalling pathway in cell transformation and in tumour progression. However, it remains to be demonstrated whether it is an increased participation of β-catenin in transcriptional regulation, or in the stabilisation of cellular integrity, or both, that is the crucial event in ovarian tumorigenesis.

Preovulatory Changes of Blood Flow in Different Regions of the Human Follicle

OBJECTIVE To assess regional changes in ultrasound-derived indices of blood flow in the dominant human follicle after the plasma LH surge. DESIGN A cross-sectional, prospective study. SETTING Reproductive medicine unit at a university. PATIENT(S) Women attending an assisted conception clinic to determine the appropriate time to transfer previously frozen embryos during a natural cycle. INTERVENTION(S) Transvaginal ultrasonography with color Doppler imaging and pulsed Doppler spectral analysis was used to obtain indices of blood flow and velocity from vessels in the base, lateral part, and apex of the dominant follicle on days 10-12 (from day 1 of menses) and after the LH surge, but before rupture. Immunoassays were used to measure the blood concentrations of LH twice daily (at 8-10 A.M. and 4-6 P.M.) from cycle day 10. MAIN OUTCOME MEASURE(S) The pulsatility index (PI), resistance index (RI), peak systolic velocity (PSV), and time-averaged maximum velocity (TAMXV) in the uterine arteries and three regions of the dominant follicle (apical, lateral, and basal parts); follicular volume; the day and time of the onset of the LH surge (defined as first concentration of LH > 22 U/L) and the times of each scan. RESULT(S) Twenty-two women (aged 28-39 years) were studied and seven were scanned on days 10-12. A retrospective examination of the data from the remainder showed that eight were scanned < 20 hours after onset of the LH surge and seven were scanned > 20 hours after the onset of the LH surge. There was a significant increase in follicular volume after the LH surge. The PI was similar in vessels from the base (0.86 +/- 0.11; mean +/- SEM), lateral part (0.72 +/- 0.51) and apex (0.67 +/- 0.09) at cycle days 10-12 and then gradually decreased in the apex. There were similar changes in the RI. The PSV (mean +/- SEM; cm/s) was similar in vessels from the base (10.1 +/- 1.64), lateral side (8.2 +/- 1.43), and apex (9.2 +/- 1.91) in follicles of days 10-12. Within 20 hours of the onset of the LH surge, the PSV had increased in basal vessels (23.4 +/- 4.10), remained similar in lateral vessels (11.64 +/- 3.18), and was undetectable in apex vessels from six of eight follicles. Twenty hours after the LH surge, there was no pulsatile blood flow observed in the apical part of the follicle, but there was a sustained high PSV in the base (15.73 +/- 3.42) and lateral side (9.02 +/- 1.5). There were corresponding changes in the TAMXV. CONCLUSION(S) During the ovulatory process there are prominent changes in the regional blood flow of the follicle with a marked increase of the flow to the base of the follicle and a concomitant decrease of blood flow to the apex. These changes may be essential for the release of a mature oocyte.

The transcription factor C/EBP-beta and its role in ovarian function; evidence for direct involvement in the ovulatory process.

Gonadotropins are responsible for maturation of the ovarian follicle and the oocyte. Ovulation is the ultimate step in this process and involves disintegration of the follicular wall and subsequent release of an oocyte into the oviduct. These events are triggered by a surge of luteinizing hormone (LH). Genes expressed in the ovary, that respond to LH, are likely to be involved in the biochemical pathways that regulate ovulation. The transcription factor C/EBP‐β is induced promptly in the ovary, as a response to an ovulatory dose of gonadotropins. We used an ex vivo perfusion system to demonstrate that a specific reduction in ovarian C/EBP‐β expression inhibits ovulation. In such ovaries the oocytes appeared to be entrapped within the follicle. We have found a correlation between the expression level of the activating isoform of C/EBP‐β and the number of oocytes ovulated in response to gonadotropins. Since a reduction in C/EBP‐β expression does not affect the level of the ovulatory mediator prostaglandin endoperoxide synthase‐2 (PGS‐2), these findings support the view of C/EBP‐β as an important factor in the ovulatory process and highlight a C/EBP‐β‐dependent and PGS‐2‐independent pathway that takes part in regulation of ovulation.

Ultrasound studies of vascular and morphological changes in the human uterus after a positive self-test for the urinary luteinizing hormone surge

The aim of the study reported here was to establish complementary data for changes in uterine size, echogenicity and vascularity during the menstrual cycle relative to a positive self-test for urinary luteinizing hormone (LH) and day 1 of next menses. Thirteen volunteers (aged 23-32 years) with apparently regular menstrual cycles were recruited from the nursing staff. The plan was to examine all women by transvaginal ultrasonography with colour Doppler imaging on day 11 of the menstrual cycle. A urinary LH self-test was to be used daily until a positive result was obtained and the women were to be re-scanned daily until the dominant follicle had ruptured. All women were then to be scanned at least every 48 h (within +/- 2 h of the same time of day) until day 6 of the next menstrual cycle. Matched samples of peripheral blood were taken at the time of each scan for hormone analysis. The main outcome measures were the times of follicular rupture, a positive test result for urinary LH and the start of menses, uterine volume, cavity length, endometrial thickness and grade, pulsatility index (PI), and time-averaged and peak systolic maximum velocities in uterine and radial arteries and in subendometrial vessels. Nine women fulfilled the criteria for an ovulatory cycle, and seven provided data over the complete study. The principal changes relative to a positive urinary LH test were (i) a continued rise in endometrial thickness to days 3 and 4 (this index then remained relatively constant, but the layered appearance was lost) and (ii) a gradual decrease in the uterine arterial PI. There was a significant rise in uterine volume, cavity length and uterine arterial PI around the time of the next menses, and a fall in endometrial thickness and blood velocity in the uterine and radial arteries and subendometrial vessels. The data may have implications for the assessment of reproductive status and the design of future studies on disorders of implantation or menstruation.

The endometrium in breast cancer patients on tamoxifen.

Abstract We restudied histologically and immunohistochemically 17 endometrial carcinomas, 2 malignant mixed tumors and 180 endometria with benign changes during or after tamoxifen therapy. The carcinomas were subtyped according to the 1994 WHO-classification. Endometrial biopsies were taken only if the endometrial thickness was > 8 mm sonographically, when a polyp was seen, or for postmenopausal bleeding. About half of the endometrial specimens showed simple or cystic atrophy, 55–76% had cystic-atrophic polyps or regressive hyperplasia. Depending upon the dose of tamoxifen, 7–19% (30 mg) to 27– 36% (20 mg) showed moderate glandular proliferation. 20–33% had foci of mucinous, clear cell or serous-papillary metaplasia. 68–70% revealed diffuse extensive fibrosis of the endometrial stroma. None of 11 patients biopsied before starting tamoxifen therapy had advanced endometrial glandular proliferation in the second endometrial biopsy after tamoxifen treatment. None of the 19 endometrial neoplasms after tamoxifen therapy was of the endometrioid type: 11 were mucinous adenocarcinomas, 4 clear cell carcinomas, 2 serous-papillary carcinomas, one carcinosarcoma and one malignant Müllerian mixed tumor. The reasons for discrepancies between suspicious sonograms and endometrial atrophy are discussed.

Saralasin-induced inhibition of ovulation in the in vitro perfused rat ovary is not replicated by the angiotensin II type-2 receptor antagonist PD123319

OBJECTIVE Our aim was to explain the effect of the nonspecific angiotensin II antagonist saralasin and the specific angiotensin II type-2 receptor antagonist PD123319 on ovulation. STUDY DESIGN Saralasin, 1 micromol/L (n = 5), and PD123319 10 micromol/L (n = 6), were administered to in vitro perfused rat ovary. Prostaglandin (prostaglandin E2, prostaglandin F2alpha, 6-keto-prostaglandin F1alpha), hydroxy-eicosatetraenoic acid (12-hydroxy-eicosatetraenoic acid, 15-hydroxy-eicosatetraenoic acid), estradiol, and progesterone levels in the perfusate and the ovulation rate were compared (Mann-Whitney U test) with controls. RESULTS Saralasin significantly (P < .01) inhibited the ovulation rate (3.0 +/- 1.4) versus control (13.1 +/- 1.0) and reduced prostaglandin E2 (at 3 hours P < .01 and 20 hours P < .05) and 6-keto-prostaglandin F1alpha (at 20 hours P < .05) levels. Saralasin did not alter prostaglandin F2alpha, hydroxy-eicosatetraenoic acids, or steroid levels. PD123319 decreased 15-hydroxy-eicosatetraenoic acid levels at 3 hours (P < .05) but had no effects on other eicosanoids, steroid levels, or the ovulation rate. CONCLUSION Angiotensin II plays an important role in ovulation in the rat and is associated with ovarian prostaglandin synthesis. This effect is not selectively regulated via the angiotensin II type-2 receptor.