MicroRNA-551b expression profile in low and high-grade cervical intraepithelial neoplasia.

OBJECTIVE: To evaluate the expression of microRNA (miR)-551b in patients with low and high grade cervical intraepithelial neoplasia (CIN) and to find an association with high-risk Human Papillomavirus (HR-HPV) infection-related prognostic biomarkers. PATIENTS AND METHODS: The expression level of miR-551b was determined in 50 paraffin-embedded cervical specimens (10 normal squamous epithelium, 18 condylomas, 8 CIN1, and 14 CIN2-3) using quantitative Real-time polymerase chain reaction (qRT-PCR). χ2-test compared miR-551b expression in different diagnosis groups. An Ordered Logistic Regression and a Probit correlation were made to correlate miR-551b expression levels with the cervical tissue histological findings. The immunohistochemical distribution of p16 and Ki-67 according to histopathological findings was also assessed. RESULTS: The distribution of the miR-551b expression profile was significantly lower in CIN1-3 samples compared to other histological diagnosis groups (condyloma and negative). The expression levels were inversely correlated to the cervical pathological grade, from negative to CIN2-3. A 1% increase in miR-551b expression level produced an increase of 19% to the probability of a minor histological grade diagnosis in a range from negative to CIN2-3 and an increase of 13% to the probability of a negative histological grade diagnosis. Among the cases with miR-551b expression < 0.02 (considered as cut-off value) a significant statistical correlation was found between p16 and Ki-67 expression and the diagnosis of CIN2-3. CONCLUSIONS: Our data showed a significant inverse correlation between miR-551b expression and the histological grading of the lesions, suggesting a tumor suppressive function in the different stages of cervical dysplasia.

Microcolposcopy in the diagnostic evaluation of abnormal cervical cytology: when and why to do it.

Microcolposcopy is an in vivo cytological examination of the uterine cervix allowing the localization of exoendocervical precancerous lesions. The aim of this study was to assess the diagnostic reliability of microcolposcopy by means of correlation with histology, colposcopy and Pap test results. For the study, 256 patients with abnormal Pap test results were selected and subjected to colposcopy and microcolposcopy with the aim of evaluating the presence of any intraepithelial lesions. One hundred and nine of these patients were subjected to a biopsy. Colposcopy, histology and cytology results were compared with those obtained by microcolposcopy. In low-grade squamous intraepithelial lesion (LSIL) cytology cases, the percentage agreement on lesion grade between Pap test and microcolposcopy results was 74%, while in high-grade squamous intraepithelial lesion (HSIL) cytology cases, it was equal to 80%. The comparison between colposcopy and microcolposcopy showed a level of agreement of 72% for lower grades and 68% for higher grades. Finally, histology was in agreement with microcolposcopy in 73% of cervical intraepithelial grade 1 neoplasia (CIN 1) cases and reached 71% for CIN 2-3. Microcolposcopy proved to be accurate with regard to the diagnosis of lesion grade, and showed to be definitive in patients where cytology was positive for HPV infection and colposcopy was not able to identify any lesions.

Myc down-regulation affects cyclin D1/cdk4 activity and induces apoptosis via Smac/Diablo pathway in an astrocytoma cell line.

OBJECTIVES: We investigated the antiproliferative effect of Myc down-regulation via cell proliferation inhibition, cell cycle perturbation and apoptosis in two human astrocytoma models (T98G and ADF) steadily expressing an inducible c-myc Anti-sense RNA. MATERIALS AND METHODS: Cell growth experiments were performed using the trypan blue dye exclusion test and cell cycle analysis was evaluated by flow cytometry. Cell cycle molecules were detected by Western blot analysis, co-immunoprecipitation and reverse transcription-polymerase chain reaction assays. RESULTS: We showed that Myc down-regulation in astrocytoma cells led to G1 accumulation and an inhibition of cell proliferation characterized by S phase delay. Co-immunoprecipitation experiments detected formation of inactive cyclin D1/cdk4 complexes as evaluated by presence of an active unphosphorylated form of retinoblastoma protein, the best characterized target substrate for cyclin D1/cdk4 complex, in ADF pINDc-myc anti-sense 7 cells. We also found that either p57Kip2 "apice" or p27Kip1 "apice" inhibitors bound to cyclin D1/cdk4 complex, thus, suggesting that they cooperated to inhibit the activity of cyclin D1/cdk4. Moreover, c-Myc down-regulation led to activation of the apoptotic mitochondrial pathway, characterized by release of cytochrome c and Smac/Diablo proteins and by reduction of c-IAP levels through activation of proteasome-mediated protein degradation system. CONCLUSIONS: Our results suggest that c-Myc could be considered as a good target for the study of new approaches in anticancer astrocytoma treatment.

[Detection, characterization and clinical significance of circulating cancer cells in patients surgically treated for breast cancer]. / Ricerca, caratterizzazione e significato clinico di cellule tumorali circolanti in pazienti operate per cancro della mammella.

Recent technological advances have led to an increasing ability to detect isolated or groups of tumour cells in blood, lymph nodes or bone marrow in patients with different tumour types. However, the clinical evidence of these advances is unclear. The detection and the characterisation of circulating breast cancer cells and the eventually micrometastasis represent an important prognostic factor with therapeutic implications. The number of neoplastic cells being very small, these are not easily detected by using only cytomorphology, possibly associated to immunocytochemistry. In the last decade many studies have been directed in order to identify new assays. In the present review the Authors summarize advantages and disadvantages about two different technical approaches: molecular and immunomagnetic selection with cellular enrichment and immunocytochemistry.

[The role of some micronutrients in the evolution of HPV infection]. / Ruolo di alcuni micronutrienti (Vit. A, Vit. E, GSH-Px) nell'evoluzione dell'infezione da HPV.

BACKGROUND: Over the past few years a series of research projects has shown that the scant or deficient immune response in HIV infection may be secondary to reduced cell resistance and/or the uncontrolled formation of free radicals. In line with these findings, subjects with HIV infection present a deficit of polyunsaturated fatty acids (the principal components of cell membranes) and many antioxidating substances, like Vitamin E and glutathione peroxidase. The high incidence of heterosexual transmission of HIV has now shown the close correlation between HIV infection and HPV infection. By analogy, we wanted to ascertain whether these deficits were also present in subjects with HPV infection and dysplastic and neoplastic lesions of the uterine cervix. Published data confirm that a HPV-positive subject has an increased risk, ranging from 40 to 200%, of contracting HIV infection. METHODS: Eighty women with HPV infection of the genital tract, at various stages. Blood levels of vitamin E and polyunsaturated fatty acids were measured using gas-chromatography; glutathione was assayed using the spectrophotometric technique. RESULTS: The alternation of the aforesaid parameters is correlated to the progress of infection and increases with the severity of lesions; Statistically significant data were recorded by comparing the group with condylomatosis with patients diagnosed with cervical carcinoma (p<0.001). CONCLUSIONS: The increased possibility that some patients are affected by an association of HPV and HIV depends on the anomalous or scarce function of many immunocompetent cells, as well the quantitative immune deficiency induced by the initial virus and the presence of various mechanisms that facilitate the development of the infection.

Biological indicators of aggressiveness in T1 ductal invasive breast cancer.

BACKGROUND: Apoptosis plays an important role in the maintenance of tissue homeostasis. When defective, this process could contribute to the pathogenesis and the progression of tumors. On this basis, we investigated the combined effect of Bcl-2 and Bax expression, known regulators of apoptotic processes, in the activation of apoptosis in breast cancer. Their relationship with DNA content and proliferative activity was also studied in order to more accurately define breast cancer patients' prognosis and treatment. MATERIALS AND METHODS: In this study we investigated 76 T1 ductal invasive breast cancers and 76 normal epithelium samples for Bcl-2 and Bax expression by immunohistochemistry, for apoptosis by tunel assay and for DNA content and proliferative activity by flow cytometry. RESULTS: High levels of Bcl-2 were associated with prevention of apoptosis. Conversely high Bax expression was found to be related to apoptosis. DNA ploidy was strictly related to the proliferative activity. In addition most of the tumors showing high Bcl-2 expression were aneuploid. CONCLUSION: This report suggests that Bax over-expression could accelerate apoptotic cell death by counteracting the ability of Bcl-2 to inhibit apoptosis. These data also suggest that the ratio Bcl-2/Bax and their relationship with the activation of apoptosis could be used as predictive indicators of breast cancer patients' prognosis and response to conventional therapy.

E-cadherin and alpha-catenin expression during tumor progression of cervical carcinoma.

OBJECTIVE: The objective of this study was to determine whether a relationship between the E-caherin molecule and the E-cadherin-associated cytoplasmic molecule, alpha-catenin, could provide additional information in the neoplastic progression of cervical cancer. METHODS: In this study we investigated by immunohistochemistry E-cadherin (E-cad) and alpha-catenin (alpha-cat) expression in 57 cervical biopsy samples representative of normal exocervical epithelium, viral (human papillomavirus (HPV) infection) and dysplastic lesions (low- and high-grade lesions), and invasive squamous cell carcinoma. The presence of immunostaining and the pattern of distribution of these molecules were correlated with the histological diagnosis (normal cervical epithelium vs HPV, cervical intraepithelial neoplasia (CIN), and invasive carcinoma). RESULTS: The correlation between alpha-cat expression and the histological diagnosis was statistically significant (chi2 test, P < 0.0077); moreover, E-cad and alpha-cat distributions were significantly correlated with the histological classification (P < 0.0001 and P < 0.0043, respectively). CONCLUSION: These results suggest that not only E-cad but also alpha-cat are associated with the loss of differentiation in CIN and squamous cell carcinomas; thus the coexpression of E-cad and alpha-cat may represent a discriminant of malignant potential and could provide an additional criterion to define high-grade CIN.

Expression of keratinocyte growth factor receptor compared with that of epidermal growth factor receptor and erbB-2 in endometrial adenocarcinoma.

Immunohistochemical analysis of the expression of keratinocyte growth factor receptor (KGFR) was performed in human endometrial carcinomas from 18 patients and in normal proliferative and secretory endometrium. The level of immunostaining was correlated with the clinico-pathological characteristics of the endometrial carcinoma patients and with the parallel expression of the epidermal growth factor receptor (EGFR) and erbB-2. The results showed that KGFR expression increased with the stage of the tumor and that the simultaneous overexpression of the three growth factor receptors appeared to be related to the depth of myometrial invasion. Taken together, these observations suggest that KGFR may represent an additional prognostic indicator in endometrial cancer.

[Peritoneal pseudomyxoma]. / Pseudomixoma del peritoneo.

The authors report a case of Pseudomyxoma Peritonei (PP) which was referred to their attention. PP is a rare pathology which is manifested by an autonomous growth of amorphous mucous material in the peritoneal cavity and serosa. PP may be caused by the perforation of a "mucinous cystoadenoma" with secretion of cellular material capable of implanting itself and spreading throughout the peritoneal cavity, or the presence of a mucocele of the appendix, or lastly the possibility of the intrabdominal transformation of the peritoneal mesothelium into mucin-secreting tissue.

[Clinical use of leuprolide acetate depot in a group of gynecologic patients in the preoperative period]. / Impiego clinico del leuprolide acetato depot in un gruppo di pazienti ginecologiche in attesa di intervento chirurgico.

BACKGROUND: The authors examined 33 patients with symptomatic uterine leiomyomas due to undergo total hysterectomy in order to evaluate the effects of treatment with GnRH analogues on leiomyoma and estrogen and progesterone receptors. METHODS: The patients were divided into two groups: one group was treated with leuprolide acetate (Group A) and the other did not receive treatment (Group B). RESULTS: A significant reduction in the volume of leiomyomas and estrogen and progesterone receptors was noted in patients in Group A. CONCLUSIONS: Treatment with GnRH analogues therefore represents a valid aid for patients with uterine leiomyomas and sideropenic anemia awaiting surgery.

EGF-R expression in ductal breast cancer: proliferation and prognostic implications.

We studied epidermal growth factor receptor (EGF-R) expression in relation to steroid receptor status, flow cytometric DNA content and S-phase fraction (%S) in a selected case series of 129 ductal primary operable breast cancer to determine the possible role of EGF-R in prognosis assessment. EGF-R expression was positively related with proliferation activity, suggesting that EGF-R could be involved in the regulation of breast cancer cell growth. We found about 80% of highly proliferating DNA aneuploid tumors in the EGF-R positive category, while the EGF-R negative tumors showed a lower frequency of highly proliferating DNA aneuploid tumors (57%), confirming the important role of EGF-R in breast cancer aggressiveness and progression. No relationship between EGF-R expression and steroid receptor status was observed. To better understand how EGF-R and estrogen receptor (ER) operate together to stimulate breast cancer cell growth, we analyzed the %S in the two groups of ER negative (ER-) and ER positive (ER+) tumors, stratifying the patients on the basis of EGF-R tumor positivity. Here breast tumor proliferation activity seems mainly to be induced by the stimulus of EGF-R, the %S values of the EGF-R negative tumors in the ER- and ER+ groups being 6.1 and 6.9%, respectively. Instead, the median %S of EGF-R positive tumors was 10% in the ER- class and 14% in the ER+ group. The analysis of the percentages of 5-year patient disease free survival were 84% for patients with EGF-R negative tumors and 61% for patients with EGF-R positive lesions, respectively. The data reported here further show the crucial role of EGF-R in breast cancer cell growth and that the EGF-R overexpression is indicative of a poor prognosis.

[Quantitative evaluation of endocervical cells in women with human papillomavirus infection]. / Valutazione quantitativa di elementi endocervicali in donne affette da Human Papilloma Virus.

UNLABELLED: Human papilloma virus (HPV) DNA in cells of endocervical origin has been demonstrated by in situ hybridization and Polimerase Chain Reaction (PCR). Similarly, the presence of HPV DNA has been shown in many cases of endocervical Adenocarcinoma. Morphological changes in squamous cells, specific for HPV infection, have already been described in previous reports, but similar specific alterations in the endocervical elements have not yet been reported. In this retrospective study we analyzed 30 endocervical smears from women with HPV infection, ascertained colposcopically, cytologically and/or histologically. Thirty endocervical samples have been taken from healthy women of the same age and hormonal status, as a control group. We evaluated maximum and minimum nuclear diameters, anisocariosis, chromatin pattern and cytoplasmic characteristics of at least 60 endocervical cells selected from each specimen. RESULTS: In the cases with HPV infection, the mean nuclear diameter of endocervical cells was increased and anisonucleosis was more evident. However, the most significant parameter was the chromatin distribution with a granular pattern in almost 90% of the cilindrical cells in more than 80% of the cases. The cytoplasm does not show any peculiar alteration of endocervical cells except for an unconspicous "alo" in some elements, but this pattern is not frequent. In conclusion, some of the parameters studied seem to have a diagnostic relevance in confirming HPV infection. This may suggest that this infection could be present even at endocervical site, and should be confirmed by molecular techniques or by microcolpohysteroscopy.

Adjuvant treatment for breast cancer in the elderly.

Eighty one patients with breast cancer stages I, II and III (T3a) were given adjuvant treatment with tamoxifen (Nolvadex), chemotherapy, or both. Most patients had estrogen and progesterone receptors measurement. Twenty one (63.6%) patients of the tamoxifen group were node positive, all 25 (100%) of the chemotherapy plus tamoxifen group and 13 (56.5%) of the chemotherapy alone group. Tamoxifen was administered for at least 3 years. Disease free survival and overall survival were substantially similar after treatment with tamoxifen or chemotherapy plus tamoxifen. Side effects were more numerous and severe in patients given chemotherapy. In receptor positive elderly breast cancer patients tamoxifen represents an excellent adjuvant therapy independently of disease stage. Chemotherapy should be reserved for subjects at high risk of recurrence in controlled clinical trials.

Precancerous lesions of the uterine cervix.

The presence of mucous and non-pathologic elements liable to hamper analysis of the smear is one of the main problems in ploidy assessment of precancerous cervical lesions. For our study, we employed a sampling system guided by microcolpohysteroscopy, which we used to identify the pathologic zone and select the morphologically altered elements only. Fifty-five cervical lesions and 6 negative controls were analyzed. All controls and 24 lesions were diploid, 15 polyploid and 16 aneuploid. All but one aneuploid lesions had a histological diagnosis of GIN. We failed to observe any significant difference in the aneuploid percentage of high and low grade lesions while diploid cases decreased in proportion to the severity of the lesion. The samples obtained had the advantage of being representative, technically valid and at the same time extremely suitable for an automated image analysis of ploidy. Thus this new sampling method could be extensively used.

Biologic characteristics of T1 papillary bladder cancer. Flow cytometric study of paraffin-embedded material.

DNA ploidy, proliferative activity (evaluated in terms of the proliferative index [PI]) and epidermal growth factor receptors (EGF-R) expression in primary bladder cancer in 52 patients were studied by means of flow cytometry and immunohistochemistry. Forty of the 52 tumors yielded evaluable DNA histograms: 22 were diploid (55%) and 18 aneuploid (45%) (median DNA index = 1.5). Ninety-five percent of the tumors were positive for EGF-R expression. The median PI value of the entire case series, estimated with a mathematical model, was 5.4%, or 6.7% in diploid tumors and 12.1% in aneuploid ones. EGF-R expression was higher in aneuploid than diploid tumors. Most of the well-differentiated tumors were diploid, while aneuploid populations and positivity for EGF-R expression were more frequent in poorly and moderately differentiated tumors. No differences in proliferative activity were observed in relation to grade. Overexpression of EGF-R in aneuploid tumors and a relationship between it, proliferative activity and grading were observed. The disease-free survival rates were 72% and 91% for patients with aneuploid and diploid tumors, respectively.

The role of cytology in differentiation of breast lesions.

The introduction of cytologic examination in to the diagnostic procedure has made it possible to define breast lesions better as early as the preoperative stage. However, there are interpretative problems depending on the nature of the lesions that make a histologic examination necessary. The cytology of nipple discharge or FNA of breast lesions would permit the best possible selection of the cases, indicating not only whether surgery is necessary, but also the basis on which it should be performed (outpatient under local anesthesia or inpatient under general anesthesia). The definitive histologic diagnosis of 447 cases submitted to surgery under general anesthesia and of 379 who were operated under local anesthesia has been correlated with the relative cytologic examination. The sensitivity and specificity of the cytologic examination are respectively 97.7% and 98.8%.

CA 125 in ovarian cysts, serous effusions and peritoneal washings: immunocytochemical expression.

The immunocytochemical expression of CA 125 in ovarian cysts, serous effusions and peritoneal washings was examined in patients with primary ovarian malignancies (22) and benign gynecologic conditions (96). The immunostaining of CA 125 was assessed in serous ovarian cysts (CA 125 was detected in 3 out of 14 serous ovarian cysts), follicular ovarian cysts (0/5), luteinized cysts (3/3), endometriotic cysts (6/11), serous cystoadenomas (4/4) and mucinous cystoadenomas (0/2). Reactive mesothelial cells (6/6) showed a low but homogeneous staining which was never found in negative peritoneal washings (0/47). Neoplastic cells in positive peritoneal washings were highly positive (20/22), except for 2 mucinous cystoadenocarcinomas. The reactivity of OC 125 with endometrial and reactive mesothelial cells shows the importance of employing a combination of MoAbs to improve conventional morphologic diagnosis.

p53 expression and genetic evidence for viral infection in intraepithelial neoplasia of the uterine cervix.

Infections with high-risk strains of human papillomaviruses (HPVs) and with herpes simplex virus type 2 (HSV 2), as well as inactivation of the p53 tumor suppressor gene, are important cofactors in cervical carcinogenesis. We analyzed 41 paraffin-embedded cervical intraepithelial lesions, including 25 cases of low-grade cervical intraepithelia neoplasia (CIN), and 16 cases of high-grade CIN for the presence of HPV 16/18 and HSV 2 genomic sequences and for the nuclear accumulation of the p53 protein. HPV 16 DNA was detected in 24.% of low-grade CINs and in 43.7% of high-grade CINs. HPV 18 was found only in 8.% of low-grade CINs. None of the cases tested scored positive for HSV 2 DNA. Nuclear accumulation of p53 was found in 4% of low-grade CINs, and in 31.2% of high-grade CINs, including 57.1% of the lesions that were positive for HPV 16. These results indicate that HPV 16 infection was over sixfold more common than HPV 18 infection and that p53 overexpression was significantly associated with high-grade lesions.

Integrin beta 4 expression in the neoplastic progression of cervical epithelium.

Integrin alpha 6 beta 4, a putative laminin receptor, is expressed in normal squamous epithelium and in squamous cell carcinomas. We evaluated by immunohistochemistry the topography of the beta 4 subunit in 35 biopsy samples representative of the sequence from normal exocervical epithelium to invasive squamous cell carcinoma. In normal and metaplastic squamous cervical epithelium, in condylomas, and in squamous epithelium with low-grade dysplasia (CIN 1), beta 4 was detected in basal and parabasal cells. In high-grade CIN (2-3) there was enhanced beta 4 expression and upward shift of beta 4 topography to the whole epithelial thickness. The beta 4 chain was diffusely expressed in most invasive squamous cell carcinomas. These observations suggest that profound alterations in the expression of the alpha 6 beta 4 adhesion glycoprotein occur in the intraepithelial phase of cervical carcinogenesis.

[Viral and/or intraepithelial lesions of the lower female genital tract: diagnostic and prognostic value of in situ molecular hybridization]. / Lesioni virali e/o intraepiteliali del basso tratto genitale femminile: valore diagnostico e prognostico dell'ibridizzazione molecolare in situ.

Fourteen patients (mean age 37 years) suffering from viral and/or intraepithelial cervico-vagino-vulvar pathologies underwent colposcopic and cyto-histological tests and molecular hybridization. The following types of HPV were assayed: 6/11-16/18-31/35/51 in cytological tissues (exocervical scraping) and biopsy material immersed in paraffin and fixed in formalin buffered with 10% PBS. The aim of the study was to identify different types of HPV using molecular hybridisation in situ. The paper reports the results and correlations with colposcopic, cytological and histological diagnosis, and then discuss the clinico-biological and prognostic aspects of this method.