Laura Fazzuoli

Estrogen and Progesterone Receptors in Uterine Leiomyomas

The purpose of our study was to evaluate the contrasting data between estrogen receptors (ER) and progesterone receptors (PgR) in uterine leiomyomas, endo‐metrium and myometrium. Uteri were obtained from 30 women undergoing hysterectomy for uterine leiomyomas (24 premenopausal and 6 postmenopausal patients). Cytosolic and nuclear ER and PgR were analysed in uterine tissues by the method of single saturation point assay at a final concentration 5 nM of [3H‐17P]estradiol and [3H]ORG‐2058. Our results confirm that leiomyomas undergo changes similar to those observed in other uterine tissues during the menstrual cycle. The ER levels show decreased values during the postovulatory phase. After subdividing leiomyomas according to their topographic criteria, as submucous or subserosal, we observed that ER and PgR are significantly more numerous in submucous than in subserosal myomas, both in proliferative and in the secretive phase of the cycle. These results agree with the hypothesis that leiomyomas have a different etiology and may explain why some myomas decrease after progestin therapy (submucous), while others (subserosal) remain unchanged.

Normal age-dependent values of serum insulin growth factor-I: Results from a healthy Italian population

Serum IGF-I levels were measured in 547 non-hypopituitaric, non-acromegalic healthy subjects of both sexes in Italy to develop reference values in relation to age and sex. Participant subjects were stratified in three age classes (25–39, 40–59 and ≥60 yr) and IGF-I assay was carried out by double-antibody radio immunoassay. Pearson’s correlation coefficient between age and IGF-I values was calculated by sex and predefined age ranges. IGF-I levels significantly decreased with age (p<0.001, Kruskal-Wallis test) while sex was not a significant factor. The median IGF-I levels were 206 ng/ml in the 25–39 yr range, 147 ng/ml in the 40–59 yr range and 103 ng/ml in the ≥60 yr range. Pearson’s correlation coefficient confirmed the negative correlation between age and IGF-I levels in the total sample of subjects (r=−0.529). The r coefficient between age and IGF-I levels did not differ between sexes (r=−0.570 in males and r=−0.529 in females), thus reflecting no sex-effect on IGF-I levels decline over years. No correlations were found in the 25–39 yr range (r=−0.036) or in the 40–59 yr range (r=−0.080) either, while in subjects aged >60 yr, IGF-I levels tended to further decrease with increased age (r=0.389). Ranges of normal values set at the 2.5th–97.5th percentile in the three age ranges were 95.6–366.7 ng/ml between 25 and 39 yr, 60.8–297.7 ng/ml between 40 and 59 yr and 34.5–219.8 ng/ml in subjects aged ≥60 yr. This study may contribute to the development of age-specific reference ranges for IGF-I determination in serum of normal subjects of both sexes in Italy.

Institutional experience of PTH evaluation on fine-needle washing after aspiration biopsy to locate hyperfunctioning parathyroid tissue.

Assaying parathyroid hormone (PTH) in the washing liquid after fine-needle aspiration biopsy (FNAB) seems to be a valid approach to locate parathyroid tissue. PTH-FNAB was evaluated in 47 patients with a clinical picture of primary hyperparathyroidism (PHP) and ultrasonography (US) suggestive of parathyroid lesion. The patients were subdivided into two groups on the basis of the absence or presence of US thyroid alterations. The result of PTH-FNAB was compared with those of cytology, scintigraphy and, in 24 patients, surgical outcome. PTH-FNAB samples with a value higher than that recorded in the serum and higher than our institutional cut-off were deemed to be probable samples of parathyroid tissue. Cytology proved diagnostic for benign thyroid lesions, non-diagnostic for thyroid lesions, hyperplastic parathyroid tissue, undetermined or malignant thyroid lesions and other lesions in 45%, 30%, 17%, 4%, and 4% of cases, respectively. In 47% of cases, PTH-FNAB indicated that the sample had been taken in parathyroid tissue. In patients without US alterations, the diagnostic accuracy of PTH-FNAB was greater than that of scintigraphy. After surgery, comparison between the results of PTH-FNAB and scintigraphy, in terms of positive predictive value (PPV), revealed the superiority of PTH-FNAB; PPV was 94% for FNAB and 71% for scintigraphy, while sensitivity was 83% and 69%, respectively. PTH-FNAB evaluation after FNAB appears to be more diagnostic than cytology and scintigraphy. Of all the procedures used, PTH-FNAB appears to be the method of choice when the target is US suggestive and reachable. PTH-FNAB appears to be a useful method of guiding surgical intervention.

ANDROGEN RECEPTORS IN NORMAL AND PATHOLOGICAL THYROIDS

The purpose of this study was to determine the presence of cytosolic and nuclear androgen receptors (AR) in both normal and nodular thyroid tissues from twelve women and six men. Samples of benign thyroid nodules and corresponding surrounding normal tissue were processed by the single saturation point assay, using [3H] R1881 ([3H] Methyltrienolone) at final concentration of 5 nM. The results show the presence of AR (cytosolic and/or nuclear) in all examined tissue samples. The nuclear AR content was higher (p < 0.01) in normal rather than in nodular thyroid tissues. The same pattern was observed when nuclear AR were analyzed according to the sex. In addition, nuclear AR content was significantly (p < 0.05) higher in normal thyroid tissue from men than from women. Our data suggest an androgen influence on thyroid tissue. If androgens are supposed to exert an antagonist role on estrogen actions also in thyroid tissue, the presence of higher nuclear AR concentration in the male rather than in the female normal thyroid may justify the lower incidence of thyroid diseases in men. Moreover, the lower AR levels found in male as well as in female nodular and goitrous tissues support the hypothesis that androgens may act with an antagonist mechanism on thyroid growth.

Thyroid and steroid receptors

Estradiol and progesterone receptor proteins (ER, PgR) have been demonstrated in neoplastic and non-neoplastic human thyroid. The aim of this study was to determine the sexual steroid receptor content of pathological non-malignant thyroid (solitary adenoma, simple nontoxic goiter), and the adjacent normal tissue of the thyroid. The results show the presence of ER and PgR (cytosolic and/or nuclear) in most of examined tissues both pathological and normal. Low levels of steroid receptors are found in the cytosol fraction, whereas the receptor content is higher in the nuclear fraction. No correlation could be found between receptor levels and patients’ age, menstrual state in females, and sexual steroid circulating hormones. A significant difference has been observed between adenomas and simple goiters (p < 0.001) in the nuclear ER, and between normal tissue surrounding adenoma and normal tissue in goiters (p < 0.05). This result suggests that estradiol and progesterone receptors are present in thyroid tissue and may have a physiological function.

Estradiol and progesterone receptors in normal and pathologic colonic mucosa in humans

The presence of estradiol (E) and progesterone (Pg) receptors (R) has been demonstrated also in normal and neoplastic tissues known to be hormone-independent and in particular in primitive colonic cancer, and, possibly, in healthy colonic mucosa. In this study endoscopic and surgical colonic mucosa specimens from 55 subjects were analyzed and divided as follows: 21 samples from healthy subjects, 12 normal mucosa samples from subjects affected with colonic cancer, 8 adenomatous polyps specimens, 5 samples from ulcerative colitis drawn on areas showing macroscopic lesions and 9 colonic cancer specimens. In the control group we have observed 6 cases positive for ER (28.6%) and 2 positive for PgR (14, 3%). Six normal mucosa specimens from subjects affected with colonic cancer were found to be positive for ER (50%) and 2 for PgR (16.7%). Five colonic cancers resulted ER positive (55.5%) and 4 PgR positive (44.4%). Four polyps were ER positive (50%) and 3 PgR positive (37.5%); in this group only one subject showed positive binding in the surrounding normal tissue. These data confirm the presence of ER and PgR in colonic cancer and colonic adenomas (so-called precancerous disease); in these subjects the finding of steroid receptors also in normal mucosa suggests that the presence of steroid binding could be considered as a marker of a precancerous condition.

Cytosolic and nuclear androgen receptor activity in the cancer of the larynx

It is well known that the larynx is a target organ for androgens and the cancer of larynx is more frequent in male subjects. We have evaluated the androgen receptors (AR) in the cytosol (ACR) and in salt extractable (ANR) and salt resistant nuclear fraction (AMR) in a group of 24 male patients with cancer of the larynx surgically removed. In addition specimens obtained from the normal mucosa of the same subjects were analyzed. In 5 patients estrogen (ER) and progesterone (PgR) receptors were also assayed. In all subjects blood samples were taken before surgery for the assay of the following hormones: LH, FSH, estradiol, testosterone, dihydrotestosterone, Δ 4-androstenedione, dehydroepiandrosterone sulfate, Cortisol. The results observed showed that 18 out of 24 normal larynx mucosa specimens and 17 out of 24 larynx cancer specimens were positive for ACR or ANR or AMR. The 5 samples of normal and cancer tissues analyzed for ER and PgR were negative. In conclusion there is no significant correlation between AR positivity from one size, histology, degree of differentiation and invasivity of the cancer, age of patients and hormonal blood levels from the other. The high ANR and AMR positivity (normal hormonal translocation and binding on DNA acceptors) confirm that the normal and cancer larynx are target tissue for androgens and establish the hormone dependence of this cancer. Hormonal therapy could be envisaged as an alternative or a complementary therapy for this type of cancer at least in the cases in which the analysis of hormone receptors will prove to be positive.

Biochemical diagnosis and assessment of disease activity in acromegaly: a two-decade experience

The objective of this study is to assess the secretory pattern of GH after Oral Glucose Tolerance Test (OGTT) or day-curve (DC), in relation with IGF-I and to evaluate the influence of therapy on OGTT. A retrospective analysis in 279 OGTTs performed in 93 acromegalic patients in our unit from January 1988 to December 2005, in 77 patients also DC data were retrived. GH concentration was evaluated by 3 different systems (RIA, IRMA and chemiluminescence assays), and IGF-I by two RIAs. About 12% of OGTT samples were discordant with the baseline, while discordance between nadir and 120th minute was much lower (5%), with all discordant values, except one, near the cut-off lines. Correlation between DC and OGTT data was around 0.99 among all values, discordance rate between nadir and minimum DC was much lower than that with mean DC. In almost 80% of cases there was a complete concordance between OGTT and DC results, and in about 30% IGF-I was discordant with GH. Correlation analysis between IGF-I and GH was highest with DC data and lowest with OGTT baseline (T0). Considering different treatments discrepancy rates between GH and IGF-I were comparable. The best GH parameter is the minimum GH DC, although in the clinical practice the evaluation of OGTT GH in association with IGF-I is the most practical approach. In this case, the basal and T120 GH values can replace multiple sampling. Different treatment modalities do not influence the discordance rate between GH and IGF-I.

Circulating nitric oxide levels increase after anti-androgen treatment in male-to-female transsexuals.

Circulating nitric oxide is produced by the vascular endothelium under the influence of the sex steroid milieu and shows gender difference. Since data on hormonal manipulation in males are scant, the present study was designed to evaluate nitric oxide levels before and after anti-androgen treatment in young male-to-female (MF) transsexuals. Fifteen MF transsexuals aged 23.7±1.3 yr, with normal testicular volume and normal body mass index were studied. Twenty adult males aged 28.2±2.4 yr served as controls. A low nitrate diet was administered to all subjects throughout the study, starting 15 days before the beginning. Blood samples were drawn from all subjects on day 0; flutamide 750 mg/day was then administered to transsexuals for 30 days, and another sample was taken on day 30. In all subjects the concentration of nitrite plus nitrate (NOx), two stable compounds into which nitric oxide spontaneously decomposes, was determined; also total testosterone (T) and free testosterone (fT), 17βestradiol (E2), SHBG, δ4-androstenedione (A), DHEAS, 17-hydroxy-progesterone (OHP), LH, FSH and PRL were assayed. All hormones determined in controls and transsexuals were comparable at the beginning of the study. NOx was also comparable in controls (11.0±1.0 μM/l) and transsexuals (11.1±1.2 μM/l) and did not significantly correlate with any of the hormones assayed. After 30 days of flutamide administration, LH, T, fT, A and E2 increased; DHEAS decreased, while FSH, SHBG and PRL were unchanged; NOx rose significantly (18.7±1.7 μM/l; p<0.05), and its percentage increase with respect to pre-treatment levels correlated with that of E2 (R=0.77; p<0.01). Healthy males and MF transsexuals do not differ in terms of sex hormones and NOx levels. In neither group is NOx significantly correlated to any sex hormone assayed. Treatment with flutamide in MF transsexuals elicits an increase in androgens, which are not biologically active because of the androgen receptor blockade, and an increase in the estrogenic milieu, which correlates with the increase in NOx.

Responsiveness to hormone, growth factor and drug treatment of a human breast cancer cell line: Comparison between early and late cultures

SummaryGrowth rate, morphology, and responsiveness to mitogenic stimuli and pharmacological treatments were evaluated in early and late cell passages derived from the same clone of the widely used MCF-7 human breast adenocarcinoma cell line. Our results indicate dissimilarities between early (E) and late (L) passages for some of the parameters analyzed. The cells that underwent many subcultivations grew faster than the others; both appeared homogeneous in size and shape. The E cells, subcultured for almost 1 yr, displayed higher sensitivity to the mitogenic action of both estradiol, according to the level of estrogen receptor, and insulin-like growth factor-I than did the L cells, kept in culture for more than 10 yr. Cell responsiveness to two drugs, a novel steroid antiestrogen and a polysulfonated distamycin A derivative, was more pronounced in the early cultures only at the longer time of exposure to the higher concentration of the estrogen antagonist. In addition, a drug-induced inhibition of insulin-like growth factor-I binding to its receptor was shown in both E and L cells, the latter being less sensitive than the former when exposed to the antiestrogen. Finally, MCF-7 E and L cells showed similar behavior when drug-induced apoptosis was tested.