Oncosuppressor proteins of fragile sites are reduced in cervical cancer.

FHIT and WWOX are tumor suppressor genes that span the common fragile sites FRA3B and FRA16D, respectively. To analyze possible synergisms among these genes in cervical cancer progression, we considered 159 cervical intraepithelial neoplasias, and 58 invasive squamous cell carcinomas of the uterine cervix. All cases were previously selected as high risk HPV. FHIT and WWOX proteins were examined by immunohistochemistry and their expression was inversely correlated with precancerous vs. invasive lesions. Statistics among biological markers indicated an association between FHIT and WWOX. Protein expression of these two genes was also absent or reduced in cancer cell lines. Thus, WWOX may be considered as a novel important genetic marker in cervical cancer and the association between the altered expression of FHIT and WWOX may be a critical event in the progression of this neoplasia.

Tissue inhibitor of metalloproteinase 2 (TIMP-2) expression in adenocarcinoma pleural effusions.

Serous effusions are frequently a clinical manifestation of metastatic disease, with lung, breast and ovarian carcinoma and mesothelioma leading the list. The diagnosis of malignant effusion signifies disease progression and is associated with a worsening patient prognosis. The ability to grow in a dense exudative fluid suggests that the malignant cells are capable of acquiring nutrients, surviving and proliferating, despite the lack of a solid-phase scaffold. During proliferation, neoplastic cells release ligands and matrix metalloproteinases (MMPs) into their environment, which dissolve the extracellular matrix (ECM). Tissue inhibitors of metalloproteinase (TIMPs) are endogenous regulators of MMPs, the principal enzymes responsible for the degradation of ECM in metastasis, and reduce their proteolytic activity. TIMP-2 has demonstrated an association between high tumor tissue expression levels and poor prognosis. The purpose of this preliminary study is to investigate, by immunocytochemistry, TIMP-2 expression in non-neoplastic and metastatic adenocarcinoma pleural effusions. We selected 16 cases of reactive mesothelio, 7 of normal mesothelio, 14 of lung adenocarcinoma, 9 from the ovary, 4 from the gastrointestinal tract and 3 from the breast. In 23/30 cases (76%), we detected adenocarcinoma cells with strong TIMP-2 expression. Positive TIMP-2 expression was found in 2/7 cases (28%) of normal and 2/16 (12%) of reactive mesothelio. A statistical association was detected between TIMP-2 expression and metastatic adenocarcinoma cells compared to reactive and normal mesothelial cells (p<0.00003). The calculated sensitivities for TIMP-2 compared to CEA and Ber-EP4 were, respectively, 76.7, 80.0 and 93.3%, and the specificities 82.6, 95.7 and 87.0%. In conclusion, immunocytochemical detection of TIMP-2 could be considered an interesting marker in metastatic adenocarcinoma pleural effusions, and could possibly be used as a component of an antibody panel in diagnostic cytopathology.

Interlaboratory quality control in gynecologic cytopathology using the novel CONQUISTADOR software. Interobserver reproducibility in the Latin American screening study.

OBJECTIVE: The recently developed software (CONQUISTADOR), capable of computing all intralaboratory and interlaboratory quality control (QC) indicators, was used to evaluate the diagnostic agreement among 4 cytology laboratories participating in the LAMS Study. STUDY DESIGN: The study was an interlaboratory exchange of specially designed 5 slide sets, each comprising 20 (conventional cytology) slides. At the first step, 80 slides (with "clear-cut" cases) were divided into four sets (A, B, C, D) of 20 specimens, each including inadequate and negative cases as well as in different proportions of all diagnostic TBS 2001 categories. In the second round, a fifth set (E) of 20 slides ("difficult cases") was designed, with all diagnostic categories, ASC and AGC included. Common measures of reproducibility (kappa and weighted kappa), accuracy (SE, SP, PPV, NPV) and 3 indices of diagnostic variability were calculated for sets A-D and set E, separately. RESULTS: For the 5 slide sets together, the weighted kappa was 0.8 (95% CI 0.76-0.85), which is the lower limit of the "almost perfect" ranking of kappa statistics, indicating an excellent interlaboratory agreement. The interlaboratory reproducibility was lower only for the difficult set (E). Similarly, the sensitivity for set E (70.0%) was lower than that (92.1%) for sets A-D. The diagnostic variability indices were not substantially different between the difficult (set E) and clearcut (sets A-D) cases. CONCLUSION: High interlaboratory reproducibility was obtained for sets A-D ("clear-cut" cases), while more interlaboratory variation was evident in the difficult samples. The new CONQUISTADOR software is a valuable tool in calculating the indicators needed in this intralaboratory and interlaboratory.

Prevalence of hepatitis C virus (HCV) genotypes and increase of type 4 in central Italy: an update and report of a new method of HCV genotyping.

Hepatitis C virus (HCV) genotyping is very useful for identifying the patients (type 1 and 4) that need more aggressive management. In recent years, genotype 4 has shown spread in different parts of Europe. The aim of this study was to update on the prevalence of HCV genotypes of 288 patients in Central Italy, to analyze the possible increase of genotype 4 and to evaluate a new simple genotyping method. A line-probe assay (LiPA, Bayer) was used based on the reverse hybridization of HCV genome fragments previously amplified and biotinylated by a polymerase chain reaction (PCR) assay, COBAS System Amplicor HCV monitor version 2.0 (Roche) or previously amplified by COBAS Ampliprep/TaqMan HCV test (Roche). This last method uses non-biotin-labeled primers, therefore we added for each sample 10 microl of amplified HCV products, 10 microl of denaturation solution and 10 microl of biotinylated-nested primers (Bayer) to utilize the genotyping procedure previously used. The results showed that the prevalence of type 1, 2 and 3 (482, 34.6 and 10.5%, respectively) as well as the prevalent subtypes, 1b and 2a/2c (30.7 and 27.2%, respectively) were similar to previous data. Type 1 and 2 were statistically associated with an older group of patients when compared with type 3 and 4 (p < 0.001). Type 3 and 4 showed a significant prevalence of male patients compared to type 1 and in particular to type 2 (p < 0.014). The prevalence of type 4 was 5.6% in 2004, 6.1% in 2005 and 9.9% from January to July 2006. Type 4 showed an increase of male prevalence over a 3-year period (p < 0.001). In conclusion, subtype 1b and 2a/2c showed a very similar prevalence, age and gender distribution in Central Italy. The type 4 patient group was analyzed because an increase of this genotype (1.8 times) was detected.

Cytological value of sputum in workers daily exposed to air pollution.

In this study, quantitative modifications of dust cells, siderocytes, Curschmann's spirals and asbestos bodies and qualitative modifications (cellular changes and inflammatory infiltrate) in the sputum of 164 traffic police officers and 218 railway workers, occupationally exposed to environmental pollution, and the sputum of 119 inhabitants of a rural area, were evaluated. The results were correlated with time of exposure and smoking habits. Seventy-three (45%) traffic police officers (TPO), 76 (35%) railway workers (RW) and 29 (24%) of the rural population (RP) were smokers. The sputum, collected over a 3-day period, was smeared on glass slides and stained according to the Papanicolaou, Perl and yellow eosin methods. The results of the qualitative cytological diagnosis revealed a statistically significant difference between the TPO, RW and the RP (p < 0.001). The results of the qualitative and quantitative cytological examinations were not significantly correlated to time of occupational exposure, which was considered to be a continuous variable. The qualitative cytological examination of sputa was not statistically significant for the smoking habits of the TPO and the RP, but was significant for the RW (p < 0.0067). In the TPO, the number of dust cells was higher in smokers, and the relative risk (RR) was 3.95. In the RW, the RR was 2.84. The results of our study revealed that for the RW, the qualitative-quantitative cytological alterations in sputum were due much more to smoking habits than to occupational exposure, while the presence of asbestos bodies correlated with work activity. The qualitative-quantitative cytological examinations of the TPO differed significantly from that of the other two populations.

PRb2/p130, p107 and p53 expression in precancerous lesions and squamous cell carcinoma of the uterine cervix.

The aim of this study was to investigate pRb2/p130, p107 and p53 expressions in precancerous lesions and squamous cell carcinoma (SCC) of the uterine cervix. We evaluated Human Papillomavirus (HPV) testing and typing and pRb2/p130, p107 and p53 expressions (antibody D07) of 48 patients showing low-grade cervical intraepithelial neoplasia (LCIN, 18 cases), high-grade CIN (HCIN, 13 cases) and SCC (17 cases). Paraffin-embedded tissue sections were analyzed for the study. High-risk HPV types were present in 67%, 89% and in 100% of HPV-positive LCIN, HCIN and SCC, respectively (Spearman's correlation coefficient: 0.393, p=0.035). Positive pRb2/p130 expression was detected in 89% of LCIN, 77% of HCIN and in 35% of SCC (p=0.001), whereas diffuse p107 expression was 72%, 62% and 100%, respectively (p=0.024). The results of p53 expression in CINs and SCCs showed values (not statistically significant) comparable with the literature data concerning the antibody D07. For the first time, we tested pRb2/p130 and p107 expressions in CINs and SCCs. We found a progressive decrease in pRb2 expression from CINs to SCCs that suggests an important role of pRb2 in cervical carcinogenesis. Indeed, p107 expression does not seem to be a useful factor. In our opinion, confirmed by the literature data, p53 immunostaining helps to biologically characterize CIN (in particular LCIN) when each case is evaluated separately considering HPV testing/typing.

Activation of the ERK/MAP kinase pathway in cervical intraepithelial neoplasia is related to grade of the lesion but not to high-risk human papillomavirus, virus clearance, or prognosis in cervical cancer.

We subjected 302 archival samples (150 squamous cell carcinomas [SCCs] and 152 cervical intraepithelial neoplasia [CIN] lesions) to immunohistochemical staining with extracellular signal-regulated kinase-1 (ERK1) antibody and human papillomavirus (HPV) testing with 3 primer sets. Follow-up data were available for all SCC cases and 67 CIN cases. High-risk (HR) HPV types were associated with CIN (odds ratio [OR], 19.12; 95% confidence interval [CI], 2.31-157.81) and SCC (OR, 27.25; 95% CI, 3.28226.09). There was a significant linear relationship between lesion grade and ERK1 staining intensity (P = .0001). ERK1 staining was a 100% specific indicator of CIN, with a 100% positive predictive value, but a poor predictor of HR HPV. ERK1 expression did not predict clearance or persistence of HR HPV after CIN treatment. ERK1 staining did not significantly predict survival in cervical cancer in univariate (P = .915) or multivariate analysis. After adjustment for HR HPV, stage, age, and tumor grade in the Cox regression model, only stage (P = .0001) and age (P = .002) remained independent prognostic factors. ERK1 expression seems to be an early marker of cervical carcinogenesis. ERK1 overexpression is not a specific marker of HR-HPV in CIN and cervical cancer, nor does it predict virus clearance after CIN treatment or disease outcome in cervical cancer.

p53 polymorphism at codon 72 is not a risk factor for cervical carcinogenesis in central Italy.

Recently Storey et al. showed that p53 polymorphism at codon 72 was related to cervical cancer. This polymorphism encodes either arginine (p53Arg) or proline (p53Pro). p53Arg was found to be more susceptible than p53Pro to E6-mediated degradation. Many studies were performed but conclusions are controversial. In this paper, we report our results from 80 women of central Italy, 30 patients showing High-grade Cervical Intraepithelial Neoplasia or squamous cell carcinoma (SCC) and 50 healthy women of the same age. The polymorphism was examined using the Storey's procedure, a molecular method, from formalin-fixed, paraffin-embedded biopsies (patients) and from cytological oral-samples (controls). The distribution of the genotypes among the cases was 27% heterozygous, 27% p53Pro-homozygous and 46% p53Arg-homozygous, while among the controls it was 52%, 2% and 46%, respectively. There was not an increased risk of SCC associated with p53Arg-homozygous; indeed there is a tendency for the contrary (odds ratio, 0.06; 99.4% confidence interval, 0.006-0.63; p = 0.019). Considering: 1) the biological characteristics of p53Pro in vitro; 2) the DNA quality, from formalin-fixed tissue, of our patients; and 3) the small number of samples performed in our study, we can only confirm that p53Pro is not a risk factor in vivo for cervical carcinogenesis in central Italy.

Aberrant bcl-2 and bax protein expression related to chemotherapy response in neuroblastoma.

Neuroblastoma (NB) is a tumor of the sympathetic nervous system which develops in young children. It derives from cells of the embryonal neural crest that form many different tissues, including the adrenal medulla and the sympathetic ganglia. Survival rates for patients with neuroblastoma have remained unchanged despite intensive efforts to develop more effective treatment strategies. The intrinsic resistance of many tumor types to antineoplastic therapies and the appearance of resistant cell populations upon relapse of an originally responsive tumor represent major impediments to successful treatment. The possibility exists that these neoplasms, particularly those that have become refractory to chemotherapeutic drugs, may occur in part because of failed apoptosis. In this study we investigated the immunocytochemical expression, before and after a trial of chemotherapy of the bcl-2 and bax proteins in 15 cases of NB, including 8 stroma-poor and 7 stroma-rich NBs. Patients with strong expression of bcl-2 before the treatment had shorter survival than those with weak or moderate expression of the protein (p = 0.004). Moreover, bcl-2 protein expression was correlated to the stroma-poor histotype only after the treatment (p = 0.031). An association between bax protein expression before treatment and longer survival (p = 0.014) was also found. We conclude that bcl-2 and bax expression, before the treatment, could be of prognostic value in neuroblastomas.

Human papillomavirus in virgins and behaviour at risk.

Human papillomavirus (HPV) infection is one of the most common sexual transmitted diseases (STDs). We compared two groups of virgins with genital HPV lesions to evaluate the behaviour at risk in the transmission of HPV infection. Partners were also examined. HPV lesions were detected in 88 virgins, who have never had sexual intercourse. This can be due to vertical transmission, fomities and skin-to-skin contact. Many other hypothesis can be proposed to explain HPV genital infection, however, further studies are required.