Acetyl-L-carnitine (ALC) administration positively affects reproductive axis in hypogonadotropic women with functional hypothalamic amenorrhea.

BACKGROUND: Hypothalamic amenorrhea (HA) is characterized by neuroendocrine impairment that, in turn, negatively modulates endocrine function, mainly within the reproductive axis. HA presents with hypo-LH, hypoestrogenism and, until now, a definite therapeutic strategy has not yet been found. The aim of the following study was to test the efficacy of acetyl-L-carnitine (ALC) administration in HA-affected subjects. POPULATION: Twenty-four patients affected by stress-induced HA were divided into two groups according to LH plasma levels: group A, hypo-LH (LH≤3 mIU/ml; no.=16), and group B, normo-LH (LH>3 mIU/ml; no.=8), were treated with ALC (1 g/day, per os) for 16 weeks. DESIGN: Patients underwent baseline hormonal assessment, pulsatility test (for LH and FSH), naloxone test (for LH, FSH and cortisol) both before and after 16 weeks of treatment. RESULTS: Under ALC administration hypo-LH patients showed a significant increase in LH plasma levels (from 1.4±0.3 to 3.1±0.5 mIU/ml, p<0.01) and in LH pulse amplitude (p<0.001). No changes were observed in the normo-LH group. LH response to naloxone was restored under ALC therapy. Maximal LH response and area under the curve under naloxone were significantly increased (p<0.05 and p<0.01, respectively). No changes were observed in the normo-LH patients. CONCLUSIONS: Our data support the hypothesis of a specific role of ALC on counteracting the stress-induced abnormalities in hypo-LH patients affected by hypothalamic amenorrhea.

[Preoperative platelet count as index of the grading in endometrial carcinoma]. / Conta piastrinica preoperatoria come indice del grading nel carcinoma endometriale.

AIM: To investigate a possible relationship between preoperative platelet count and following clinicopathological variables of the endometrial carcinoma: age, stage, histological type, histological grading (G), myometrial invasion, lymphovascular space involvement, cervical involvement, lymph node metastasis. In particular the existence of a possible relationship between elevated preoperative platelet count (=or>300 000 microL) and negative prognostic factors. METHODS: The authors analyzed retrospectively 120 patients with endometrial carcinoma underwent to surgery as the initial treatment. All the patients were subjected to radical surgical procedure: peritoneal cytology, total abdominal hysterectomy, bilateral salpingo-oophorectomy, systematic pelvic lymphadenectomy and omentectomy. Blood platelet count was taken from the patients three days prior to the surgery. RESULTS: The patients with platelet count<300000/microL whom they had a G1, G2, G3 they were respectively the 23.1%, 44.2% and 32.7% versus the 0%, 12.5% and 87.5%, respectively for G1, G2, G3, of the patients with platelet count>300000/microL (P=0.024). Only considering the patients to the stage I of the Federazione Internazionale dei Ginecologi ed Ostetrici (FIGO). The patients with platelet count<300000/microL whom they had a G1, G2, G3 they were respectively the 27.3%, 43.2% and 29.5% versus the 0%, 0% and 100%, respectively for G1, G2, G3, of the patients with platelet count=or>300000/microL (P=0.008). There were no differences respect to age, stage, histological type, myometrial invasion, lymphovascular space involvement and cervical involvement. CONCLUSION: Elevated preoperative platelet count, in the patients with endometrial carcinoma, may reflect poor prognostic factor such as higher histological grade. This study allowed to observe: a significant correlation between elevated preoperative platelet count (=or>300000/microL) and tumoral grading (G3) of general population submitted to study; for the patients to the stage I FIGO a more significant correlation between elevated preoperative platelet count (=or>300000/microL) and tumoral grading: the 100% of the patients with platelet count=or>300 000/microL had a histological grading G3.

Randomized double-blind study evaluating the efficacy on uterine fibroids shrinkage and on intra-operative blood loss of different length of leuprolide acetate depot treatment before myomectomy.

OBJECT: To determine whether length of pre-operative treatment with gonadotrophin-releasing hormone agonists (GnRHa) may have different effects on uterine shrinkage and intra-operative blood loss, 36 patients with symptomatic uterine fibroids awaiting myomectomy were randomly divided into two groups. METHOD: Twenty patients received long-term GnRHa administration, six monthly depot injections of leuprolide acetate (LA), while 16 patients were treated with two monthly LA injections before surgery. The hemoglobin concentration and estradiol, follicle-stimulating hormone and luteinizing hormone concentrations were measured before and after treatment in both groups. RESULTS: Uterine volume decreased in the long-term treated group from 680+/-276 cm3 to 486+/-195 cm3 (36%) after two and to 388+/-172 cm3 (51%) after six LA injections. In the short-term treated group the basal uterine volume decreased from 745+/-320 cm3 to 456+/-177 cm3 (39%) after two LA injections. The uterine volume decrease was statistically significant (p<0.05) after two LA injections in both groups while the decrease observed between two and six LA injections was not significant (p>0.05). The intra-operative blood-loss was not significantly different between the two groups studied, 315+/-93 cm3 and 336+/-88 cm3. CONCLUSION: Two pre-operative GnRHa depot injections offer similar results, in terms of uterine shrinkage and intra-operative blood loss, and a longer treatment seems to be justified in cases of anemia.

B cell leukaemia/lymphoma-2 protein concentrations during normal pregnancy.

We evaluated bcl-2 protein levels in whole blood, and intraerythrocytic products of lipoperoxidation, in 65 healthy pregnant primiparous women at the end of the first, second and third trimesters of pregnancy. During pregnancy we found a progressive increase in concentrations of bcl-2 and intraerythrocytic products of lipoperoxidation. Moreover, a positive correlation was found between bcl-2 concentrations and lipoperoxidation products, and between bcl-2 protein, E2 and progesterone. The results show that modifications occurring during pregnancy are accompanied by variations of bcl-2 protein.

Anticardiolipin antibodies in pre-eclampsia and intrauterine growth retardation.

OBJECTIVE: To establish an association of anticardiolipin antibody (ACA) levels and pre-eclampsia or intrauterine growth retardation (IUGR). METHODS: Twenty-eight patients with pre-eclampsia, 28 with IUGR and 28 normotensive control group were matched for maternal age, race, weight, cigarette smoking, and parity. All had plasma anticardiolipin antibodies (GPL and MPL) detected by the modified enzyme-linked immuno-absorbent assay (ELISA) technique. RESULTS: No statistical significant difference in ACA values, both GPL and MPL, was found among the three groups studied Furthermore, none reached a value of ACA that could be considered clinically relevant (> 15). CONCLUSION: No association was found in anticardiolipin antibody levels between pre-eclamptic and IUGR versus the control group.

Lack of endogenous estrogens effects on circulating adhesion molecule ICAM-1.

The serum vascular adhesion molecule ICAM-1 that is involved in atherogenesis was determined in fertile, postmenopausal and at term pregnant women. The aim of the study was to ascertain if the antiatherogenic estrogens action may involve this adhesion molecule. The serum ICAM-1 concentrations resulted similar in the three groups studied: 331 +/- 35 ng/ml, 333 +/- 28 ng/ml and 302 +/- 53 ng/ml in fertile, postmenopausal and pregnant subjects respectively, despite the very different estrogen plasma levels. These data, the first on the ICAM-1 serum levels in women with different natural hormonal milieu, demonstrate that estrogens antiatherogenic action is not involving ICAM-1.

Salmon calcitonin plus intravaginal estriol: an effective treatment for the menopause.

OBJECTIVE: Intravaginal estriol (E3) effectively improves postmenopausal genito-urinary disturbances, without stimulating endometrial proliferation. The aim of the present study was to evaluate the effect of intravaginal estriol (E3) plus nasal spray salmon calcitonin (sCT), to improve neurovegetative symptoms and to prevent the decline of bone mineral density (BMD) of postmenopausal women. METHODS: Two hundred and fourteen (214) healthy postmenopausal women were treated for 12 months with: (1) E3 (0.5 mg every other day) + Ca (0.5 g/day); (2) E3 + Ca + sCT (50 IU x 2/day); (3) sCT + Ca; (4) Ca. Climacteric complaints, such as hot flushes and sweating, BMD at the distal 1/10 of the radius, analyzed by dual photon absorptiometry, urinary excretion of hydroxyproline and serum alkaline phosphatase were evaluated at baseline and every 6 months. At the same time, patient compliance and drug tolerability were evaluated. RESULTS: E3 but not sCT, improved hot flushes and sweating. E3 blunted but not completely counteracted the BMD decline observed in women treated with only Ca, and reduced urinary hydroxyproline excretion. sCT markedly increased BMD values and reduced both urinary hydroxyproline excretion and serum alkaline phosphatase. These effects were not potentiated by E3 coadministration. All treatments were well tolerated. CONCLUSIONS: Present data indicate that the combined administration of intravaginal E3 and sCT may represent an alternative therapeutic regimen for those postmenopausal women who do not accept or have contraindications to classical hormone replacement therapy.

Epithelial cells are the major source of biologically active granulocyte macrophage colony-stimulating factor in human endometrium.

Granulocyte macrophage colony-stimulating factor (GM-CSF) has emerged as an important growth factor for trophoblast and other placental cells, leading to improved placental functioning and fetal survival. Recent observations have indicated that GM-CSF is synthesized by epithelial cells in the murine pregnant and non-pregnant uterus. In this study, the production of GM-CSF by cells derived from human endometrium is assessed using a sensitive bioassay and specific neutralization of the cytokine bioactivity with a monoclonal antibody to GM-CSF. Originally, GM-CSF was assayed in the culture supernatants of explant cultures of human endometria. Concentrations of GM-CSF up to 4440 pg/ml were detected. Subsequently, enriched epithelial cell cultures were prepared from glands isolated from human endometrium. The purity of epithelial cultures was demonstrated by the expression of cytokeratin, a weak immunoreactivity for vimentin and a lack of immunoreactivity for leukocyte common antigen, CD68, a macrophage-specific protein and endothelial marker (factor VIII-related antigens). Detected concentrations of GM-CSF were as high as 18,800 pg/ml. Furthermore, pure epithelial cells of a neoplastic endometrial cell line ECC1 secreted GM-CSF, confirming the ability of endometrial epithelial cells to secrete this cytokine. The immunostaining of dated endometria from proliferative and secretory phases showed primarily that epithelial cells, and to a lesser extent stromal cells, exhibited immunoreactivity for GM-CSF. A Western blot analysis, performed to validate the immunohistochemical data, confirmed the presence of an immunoreactive gene product for GM-CSF in human endometrium throughout the menstrual cycle. These findings indicate that human endometrium synthesizes GM-CSF and that epithelial cells are a major contributor to its production.

Epidermal growth factor receptor (EGF-R) and transforming growth factor alpha (TGFA) expression in different endometrial cancers.

The role of epidermal growth factor receptor (EGF-R) and transforming growth factor alpha (TGFA) in endometrial cancer is still controversial. In this study the immunostaining of EGF-R and TGFA, investigated in 44 cases, proved to be related to endometrial cancer histotype regardless of the tumor grade or the extent of myometrial invasion. In fact, EGF-R expression was observed in 12.5% (1/8) of cases with classical endometrioid adenocarcinoma, in 91% (10/11) with endometrial adenocarcinoma and benign squamous metaplasia, and in 35% (6/17) with adenosquamous carcinoma. TGFA expression was concomitant with EGF-R in the first two histotypes, and was present in about 50% (9/17) of the latter. Conversely, in mucinous and serous papillary adenocarcinoma, EGF-R and TGFA immunostaining was not observed. In conclusion, EGF-R positivity was observed in endometrial cancer with squamous differentiation, especially in those cases with benign squamous metaplasia. EGF-R and TGFA coexpression appeared as a constant feature of benign squamous metaplasia.

Ovarian mesothelial and extramesothelial cells in interactive culture.

The ovarian mesothelium (OM) represents the tissue of origin of ovarian epithelial cancer. To gain insight into the regulation of this tissue, OM organoids and submesothelial ovarian stromal cells (SC) were isolated from New Zealand White rabbits by a stepwise tissue dispersal technique, while granulosa cells (GC) were aspirated from mature follicles (14 +/- 4 groups/animal). OM and SC dispersal were sequentially accomplished by: a) 1-h incubation in collagenase type I (300 U/ml), gentle scraping of the ovarian surface, and 1 g sedimentation of OM organoids (equivalent to 0.93 +/- 0.40 x 10(6) cells/animal) on 5% bovine serum albumin (BSA); b) 2-h incubation in pronase-collagenase (0.5%-300 U/ml) under periodical resuspension and gentle scraping of SC (1.40 +/- 0.25 x 10(6)/animal) from OM-denuded ovaries. After a week-long in vitro expansion, OM cells (OMC) were cultured alone and with SC or GC within monocameral vessels or bicameral transfilter vessels in serumless, fibronectinrich (4 micrograms/ml) HL-1 medium. After 7 d of contact cell-cell interaction, cytokeratin-positive OMC became surrounded by fibroblastoid, vimentin-positive SC or by cytokeratin and vimentin-weakly positive GC. Filter-bound OMC humorally interacting with underlying SC or GC displayed a biphasic, epithelioid and spindle, morphology with universal cytokeratin expression. Bromo-2'-deoxyuridine (BrdU) immunoperoxidase revealed mean cell proliferation indices of 14.88% for OMC cultured alone, 11.21% and 19.39% for OMC cultured with GC or SC in monocameral dishes, and 15.25% or 22.47% for OMC cultured in bicameral vessels over GC or SC, respectively. This model provides an experimental tool for investigating the unexplored role of stromal-mesothelial interaction in OM pathobiology.

Epidermal growth factor receptor expression and endometrial cancer histotypes.

Epidermal growth factor receptor (EGF-R) has been implicated in the growth of endometrial cancer. In the present study, immunostaining of EGF-R was investigated in different endometrial cancer histotypes. For this purpose 34 surgical samples of primary endometrial carcinoma were studied using monoclonal anti-EGF-R antibody. The endometrial carcinomas were classified according to the International Society of Gynaecological Pathology (I.S.G.P.). EGF-R expression was observed in: 1 of 5 classical endometrioid adenocarcinomas (AE), in 8 of 9 endometrial adenocarcinomas with benign squamous metaplasia (AA), and in 3 of 14 adenosquamous adenocarcinomas (AS). In mucinous (M) and serous papillary adenocarcinoma (SP) EGF-R immunostaining was not observed. The EGF-R positivity was observed in endometrial cancer with squamous (SQ) differentiation, especially in those with benign SQ metaplasia. These results suggest that EGF-R immunostaining is related to the endometrial cancer histotype regardless of the tumor grade or extent of miometrial invasion.

A feasibility study of ovarian cancer screening: does fine-needle aspiration improve ultrasound specificity?

AIMS AND BACKGROUND: The prognosis for ovarian cancer patients depends on the stage at diagnosis. As a prerequisite for any proposed procedure for ovarian cancer screening, high levels of specificity should be obtained using combinations of tests. Based on preliminary data from an ongoing feasibility study, this report is focused on the possible gain in specificity provided by fine-needle aspiration of occult ovarian masses detected by ultrasound. METHODS: Ultrasonography was used as a basic test, and fine-needle aspiration was obtained from selected patients. Those with (a) positive aspiration cytology or histology, (b) complex or solid masses showing volume increase, (c) complex or irregular masses and inadequate samples, and (d) recurrent cystic lesions were operated on. RESULTS: A total of 3541 asymptomatic patients 50-69 years of age underwent ultrasonography, and 98 were selected for fine-needle aspiration. Of these, 19 (positivity rate 19/3541 or 0.5%) were operated on. Two ovarian cancer cases (FIGO Stage II and III) were detected (detection rate 2/3541 or 0.6 x 1,000; positive predictive value 2/19 or 10.5%). Specificity was 3443/3539 or 97.3% for ultrasound alone and 3522/3539 or 99.5% for the procedure as a whole. Surgical exploration for relapses of benign cysts reduced by some 50% the potential gain in positive predictive value provided by aspiration cytology compared with ultrasound alone. CONCLUSIONS: Although fine-needle aspiration has improved the performance of ultrasound, this 2-level procedure does not appear to achieve acceptable levels of specificity.

Demonstration of cytokeratin intermediate filaments in oocytes of the developing and adult human ovary.

The intermediate filaments (IF) present in the various cells of human ovaries were studied by immunolocalization using antibodies to cytokeratins (CKs), vimentin, desmin and alpha-smooth muscle (alpha-SM) actin. Oocytes revealed a single paranuclear aggregate, which reacted with antibodies to CKs 8, 18 and 19 both in adult and fetal ovaries. The existence of this aggregate was also documented by electron microscopy. Ovarian surface epithelium and granulosa cells consistently coexpressed CKs 8, 18, 19 and vimentin. During follicle maturation vimentin remained unchanged in the granulosa layer while CKs content decreased, showing variation in the amount and distribution of the different CK-types. Thecal cells of secondary and mature follicles showed alpha-SM actin positivity. These contractile fibres increased in mature follicles. Ordinary fibrous stromal cells showed isolated cells which were desmin and alpha-SM actin positive. A similar pattern of IF expression and distribution existed in all stages of development in fetal and embryonic ovaries. These results indicate that CKs are present in human oocytes and that the coexpression of vimentin and CKs can be regarded as a peculiar feature of all ovarian cell types except oocytes and ordinary stromal cells. Contractile properties have been documented associated with a modification in expression of IF proteins. This is likely to represent an integral part of folliculogenesis along with the functional hormone-dependent changes.

Treatment of hirsutism by an association of oral cyproterone acetate and transdermal 17 beta-estradiol.

Twenty-four hirsute women were treated with an inversal sequential scheme of cyproterone acetate, 50 mg/d by oral route from the 1st to the 15th day of the menstrual cycle, along with 100 micrograms/24 h of 17 beta-estradiol transdermally administered from days 1 to 21, for nine cycles at weekly intervals. The acne and seborrhea as well as hirsutism showed a significant improvement in all subjects studied. The plasma testosterone and dehydroepiandrosterone sulfate decrease from 1.5 +/- 1.3 ng/mL and 6.9 +/- 1.3 micrograms/mL to 0.5 +/- 0.03 ng/mL and 2.7 +/- 1.7 micrograms/mL, respectively. Similar values were observed in subjects with idiopathic hirsutism during the treatment. The metabolic parameters, as well as the plasma levels of sex hormone-binding globulin, appeared unaffected by the therapy. Furthermore, the luteinizing hormone and follicle-stimulating hormone secretion was strongly inhibited from the first cycle of treatment. In conclusion, considering the good clinical results and the avoidance of any hepatic effect, this association should be taken into account in the treatment of hirsutism, especially in case of oral estrogen intolerance.

Tissue factors influencing growth and maintenance of endometriosis.

The unpredictable response of endometriosis to steroids and its recurrence after therapy, led us to hypothesize a possible further control of this pathology by factors other than steroids. The presence of estrogen, progesterone and epidermal growth factor receptors (ER, PR, EGFr) was evaluated using immunohistochemistry before and after therapy with Danazol or a gonadotropin-releasing hormone analogue (GnRHa), Buserelin. EGFr, ER and PR were present in 100% of endometrial specimens, and in 71%, 29% and 49% of endometriotic implants, respectively. Danazol and GnRHa reduced immunohistochemical staining for EGFr antisera in the endometrial and endometriotic specimens. About 21% of endometriosis were EGFr positive and ER negative, suggesting a potential role of epidermal growth factor in growth and maintenance of endometrial ectopia.