Gloria Giovannini

Characterization of laboratory working standard for quality control of immunometric and radiometric estrogen receptor assays. Clinical evaluation on breast cancer biopsies. Italian Committee for Hormone Receptor Assays Standardization.

The objective of the study was to characterize a low-cost and reliable working standard material for quality control of estrogen receptor (ER) determination with dextran-coated charcoal (DCC) and enzyme immunoassay (EIA) methods. Human fibromatous uterine lyophilized cytosol demonstrated good characteristics of stability and applicability for this purpose. Eleven laboratories participated in the intralaboratory and interlaboratory quality control study, and they achieved slightly higher coefficients of variation for ER-EIA (interlaboratory, 37.7 %; intralaboratory, 22.9 %) than for ER-DCC (Interlaboratory, 24.2 %; intralaboratory, 15.7 %). There was an excellent correlation between ER results with ER-EIA and ER-DCC for 268 breast cancer biopsies. Quality assurance for ER assays using DCC techniques and immunometric methods with monoclonal antibodies (ER-EIA) can be set up with this available material of human origin to satisfy the characteristics of both techniques and the species specificity of monoclonal antibodies.

Testis cytological structure, plasma sex steroids, and gonad cytosol free steroid receptors of heterologous gonadotropin (hCG)-stimulated silver eel, Anguilla anguilla L

Complete testicular maturation was induced in silver eels, kept at 24 degrees in fresh water, by a single injection of 1000 I.U. of heterologous gonadotropin (hCG). Each week, for 4 weeks, some eels were examined for testis structural pattern, plasma sex steroids, and gonad cytosol steroid receptors. The first effect of the hCG was on the tubular organization of the testis, followed by spermatogenesis. Plasma androgens were not detectable in the untreated eels, whereas a peak was detected a week after in those treated with the injection and afterward a decline. Plasma progesterone and estradiol showed a peak 2 weeks after treatment. Untreated eel gonads showed a high content of cytosolic free estradiol receptors which disappeared in the hCG-treated ones, a peak of free progesterone receptors was found 1 week after injection. The results are discussed in relation to the differentiation and maturation of eels testes.

Interleukin-1β levels in gingival crevicular fluid and serum under naturally occurring and experimentally induced gingivitis

AIMS To evaluate the interleukin-1 beta (IL-1 beta) levels in gingival crevicular fluid (GCF) and serum in either naturally occurring (N-O) or experimentally induced (E-I) plaque-associated gingivitis. MATERIAL AND METHODS Thirty-seven periodontally healthy subjects were evaluated in real life conditions (N-O gingivitis) as well as after 21 days of experimental gingivitis trial (E-I gingivitis). During the experimental gingivitis trial, in one maxillary quadrant (test quadrant), gingival inflammation was induced by oral hygiene abstention, while in the contralateral (control) quadrant, oral hygiene was routinely continued. IL-1 beta concentrations in N-O and E-I gingivitis were investigated for IL-1B(+3954) and IL-1B(-511) gene polymorphisms. RESULTS (i) GCF IL-1 beta concentrations in E-I gingivitis were significantly higher compared with N-O gingivitis; (ii) an intra-individual correlation between GCF concentrations of IL-1 beta detected in N-O and E-I gingivitis was observed in control quadrants, but not in test quadrants; (iii) IL-1 beta concentration in GCF was associated with IL-1B(+3954) genotype only at test quadrants; (iv) IL-1 beta was detectable in serum only at low levels in a limited number of subjects, without difference between gingivitis conditions. CONCLUSIONS Aspects of the bacterial challenge to the gingival tissues, such as the amount of plaque deposits and plaque accumulation rate, appear to affect the IL-1 beta levels in GCF in subjects with a specific IL-1B genotype.

Inhibition of amniotic interleukin-6 and prostaglandin E2 release by ampicillin.

OBJECTIVE: To test the effect of ampicillin on amniotic interleukin-6 (IL-6) and prostaglandin E2 (PGE2) release. METHODS: In an in vitro study, IL-6 and PGE2 release from amnion-like Wistar Institute Susan Hayflick cells was assayed under basal conditions, as well as after incubation with ampicillin. In an in vivo study, amniotic fluid IL-6 was assayed in a total of 212 patients submitted to genetic amniocentesis during the 17th week of their singleton physiological pregnancy. The study population was subdivided as follows: 92 patients sampled before ampicillin administration, 70 patients sampled 4 hours after administration of 1 g ampicillin, and 50 patients sampled 12 hours after administration of 1 g ampicillin. RESULTS: At doses ranging from 10−7 to 10−4 M, ampicillin decreased IL-6 release from Wistar Institute Susan Hayflick cells. The drug effect was already statistically significant (−30%; P < .05) at the lowest concentration tested (10−7 M), reaching the maximum (−50%) at 10−6 M after 4 hours of incubation. Moreover, ampicillin concentrations ranging from 10−7 to 10−4 M decreased PGE2 release from Wistar Institute Susan Hayflick cells; maximal inhibition was reached at 10−6 M after 4 hours (−40%; P < .05). Finally, IL-6 levels measured in amniotic fluid of patients sampled 4 hours after ampicillin administration proved strongly and significantly reduced when compared with those sampled either before or 12 hours after treatment (P < .001). CONCLUSION: The capacity of ampicillin to directly decrease amniotic IL-6 and PGE2 release should be considered in the management of bacterial and nonbacterial inflammatory complications of pregnancy mediated by the cytokine and prostanoid interaction. LEVEL OF EVIDENCE: III

A High Cytosol Value of Urokinase-Type Plasminogen Activator (uPA) May Be Predictive of Early Relapse in Primary Breast Cancer

Background: There is now much data that suggest a relationship between angiogenesis and breast cancer prognosis. Angiogenesis is a multistep process resulting from an ordered set of events and regulated by positive and negative modulators of microvessels growth and by the expression of various proteolytic enzymes. Materials and methods: We prospectively evaluated VEGF and microvessels density on tumor specimen and cytosolic levels of uPA and PAI-1. Results: We enrolled 81 primary breast cancer patients. The median follow-up was 38 months. Using the median value as cutoff for the statistical analysis, we found significant correlation between cytosolic levels of uPA and PAI-1(r=0.61;p<.0001), between VEGF and steroid hormone receptor status(p=.01), between PAI-1 and tumor grading(p=.009), and between uPA and tumor size greater than 1 cm(p=.04). With respect to the prognosis, we observed a significant correlation between low uPA levels and RFS and an unforeseen, direct correlation between high VEGF values and better RFS. Conclusions: Our preliminary results indicate that the cytosolic level of uPA at diagnosis may be predictive of early relapse in primary breast cancer.

Plasminogen activator system in serum and amniotic fluid of euploid and aneuploid pregnancies.

Objective To compare euploid and aneuploid pregnancies with respect to maternal serum and amniotic fluid (AF) levels of the components of the plasminogen system. Methods The study population consisted of 123 single pregnancies at the 17th gestational week, 16 with minor chromosomal abnormalities, 15 aneuploid, and 92 euploid. Results Both groups with chromosomal abnormalities had significantly higher serum levels of urokinase plasminogen activator and its complexed form with its type-1 inhibitor compared with euploid pregnancies. In AF, tissue plasminogen activator was significantly lower in the aneuploid than the euploid group, whereas type-1 inhibitor of plasminogen activator was significantly higher in the cases with minor chromosomal abnormalities compared with euploid. At cutoff levels set at 100% sensitivity, the complexed form of urokinase plasminogen activator with its type-1 inhibitor had the strongest specificity (66.3%); after logarithmic transformation, its serum level was 7.53 times higher in aneuploidies than euploidies. Conclusion Aneuploid pregnancies appear to be accompanied by abnormalities of the plasminogen activation system, which could lead to impaired placental perfusion and thus to abortion, fetal death, and fetal growth restriction.

Comparison between single saturating dose ligand binding assay and enzyme immunoassay for low-salt extractable oestrogen and progesterone receptors in breast cancer: A multicentre study

An excellent correlation between ligand binding assay (LBA) and enzyme immunoassay (EIA) for both oestrogen (ER) and progesterone (PR) receptors has been reported. Nevertheless, considering that the clinical value of any discrepancy between LBA and EIA probably varies with the receptor level, we undertook a collaborative study in which a single saturating dose (SSD) LBA and EIA were compared in different ER and PR dose ranges. The values of ER measured by EIA were higher in tumours with low or intermediate receptor content, causing a misclassification of ER status in 9% of cases (ER+: 77.5%, EIA, 68.8% SSD). In the case of ER, EIA values tended to be higher than SSD in all centres. For PR, EIA and SSD were generally more comparable (PR+: 66.0% EIA, 72.0% SSD, discordance rate 6%), with EIA showing, however, different trends in different centres. PR concentration was not significantly different in ER SSD-/EIA+ and in ER SSD+/EIA+ cases, suggesting that EIA detects at least in part integer ER. We conclude that although EIA may be a reliable methodological alternative to SSD, the two methods are not interchangeable until effective cut-off levels for clinical decisions are assessed for EIA.

Hormone receptors and breast cancer: Correlations with clinical and histologic features

The possible relationships between hormone receptor status and several clinical (age, gynecologic history, clinical stage) and morphologic aspects (histologic grade, vascular invasion, lymphocytic infiltration, necrosis, fibrosis, elastosis and lymph node metastasis) were evaluated. A highly significant correlation between estrogen receptor levels, patient age, menses regularity and postmenopausal status was found. The histologic features most significantly related to tumor receptor status were histologic grade, lymphocytic infiltrate, necrosis and elastosis. Since these same histologic aspects appear to influence prognosis in breast cancer, the prognostic significance attributed to tumor receptor levels is substantiated. Therefore the importance of this assay is confirmed, not only for its diagnostic and therapeutic purposes, but also for its prognostic value.

Neuropeptide release from slices of rat and guinea pig trigeminal ganglia: modulation by dihydroergotamine and sumatriptan

Abstract The trigeminovascular system is considered to play a role in the mechanism of migraine headache. Novel in vitro animal models that investigate the release of neuropeptides may be of help to understand the pathophysiology and pharmacology of trigeminal neurons. Here, we examined the release of the immunoreactivity (LI) of the sensory neuropeptides calcitonin gene-related peptide (CGRP) and substance P (SP) from slices of rat and guinea pig trigeminal ganglia with proximal nerve trunks attached. Electrical field stimulation (EFS, 10 Hz), high K+ medium (50 mM) and capsaicin (1 μM) caused a significant increase in CGRP-LI outflow. SP-LI was also released after exposure to EFS, high K+ and capsaicin. The increase in CGRP-LI outflow induced by EFS was markedly reduced in a Ca2+-free medium and by pretreatement with a high capsaicin concentration, tetrodotoxin, ω-conotoxin, dihydroergotamine and sumatriptan. Sensory neuropeptide release from slices of rat trigeminal ganglia with nerve trunks attached fulfills the criteria required to define it as a neurosecretory event. This is a novel method for studying trigeminal neuron pathophysiology and the action of antimigraine drugs.