Donatella Spina

Neoplastic cells of Hodgkin's disease show differences in EBV expression between Kenya and Italy

The Epstein‐Barr Virus (EBV) has been implicated in the pathogenesis of Hodgkins disease (HD). However, the association of EBV with this disease varies greatly from series to series and from country to country. Epidemiological studies have shown differences in HD occurring in different parts of the world. In particular, it has been reported that HD in developing countries differs from HD in Western countries in terms of epidemiological, pathological and clinical characteristics. These discrepancies among populations suggest an interaction with environmental factors and a direct role of different etiological agents. At present, there are no data on the frequency of association of EBV with HD in equatorial Africa. In this study, a large series of HD cases have been collected at the University of Nairobi, Kenya, and at the Universities of Bologna and Siena, Italy. The cases have been reviewed and classified according to the REAL Classification and the presence of EBV has been assessed by in situ hybridization (ISH). A statistical difference in EBV expression was found between HD from Kenya and HD from Italy. EBV‐positive neoplastic cells were detected in 92% of Kenyan cases, whereas only 48% of Italian cases showed EBER1/2 positivity in the neoplastic cells. Our results suggest that, in Kenya, EBV plays a more direct role in the pathogenesis of HD, as it does for endemic Burkitt lymphoma.

Cellular kinetic and phenotypic heterogeneity in and among Burkitt's and Burkitt-like lymphomas.

This study asks whether the known genotypic heterogeneity within and between endemic or sporadic Burkitts lymphomas (eBLs and sBLs, n=10 each), and Burkitt‐like lymphomas (BLLs, n‐12), is reflected in divergent cytokinetics and related immunophenotypes. There was strong evidence that eBL and BLL grow markedly faster than sBL, as shown by differences in mitotic and apoptotic indices. Furthermore, in BLL, the median percentage of neoplastic cells immunoreactive for the bcl‐2 protein was much higher than that observed in eBL and sBL. The reverse was true for the median fraction of cells containing c‐myc protein. In eBL and sBL, the median fraction of bcl‐6 protein‐positive cells reached values above 50 per cent, while cells of 8/12 BLLs did not contain detectable amounts of this protein. This observation indicates that in this respect, eBL and sBL resemble normal germinal centres of lymphatic tissue much more than do BLL. Evidence for infection of neoplastic cells by the Epstein‐Barr virus (EBV) was observed in 9/10 cases of eBL and in 3/10 of sBL, but not in BLL. EBV‐positive lymphomas were associated with distinctly lower apoptotic indices and smaller median percentages of bcl‐6‐positive cells than EBV‐negative tumours.

The effects of HIV-1 Tat protein on cell cycle during cervical carcinogenesis

The role of HPV in the carcinogenesis of intraepithelial and invasive anogenital lesions is currently well established. E6 and E7 oncoproteins of high risk HPV genotypes are known to inactivate p53 and pRb pathways. Several studies have described an increased prevalence and recurrence of both cervical HPV infection and invasive cervical cancer among HIV-1 positive women compared to HIV-1 negative cases. For these reasons, cervical cancer is considered an AIDS-defining neoplasm. Unlike other AIDS-associated neoplasms, the occurrence of cervical cancer is independent of immune suppression. HIV-1 infection in patients with high grade precancerous lesions and cervical cancers results in a therapy refractory and more aggressive disease phenotype, which is not yet well understood at the molecular level. An upregulation of HPV E6 and E7 gene expressions by HIV-1 proteins such as Tat has been documented by some authors. However, the role of HIV-1 in cervical carcinomas is still unclear. It is already known that HIV-1 Tat protein is able to influence cell cycle progression. Altogether, these facts led us to investigate the effects of Tat on the expression of cell cycle regulator genes. After transfection of HeLa cells with Tat, we analyzed the expression of cell cycle regulators from these cells by IHC and Real-time PCR. A significant reduction in the expression of cell cycle inhibitors of transcription and an increase in the levels of proliferation markers were observed. These results suggest that HIV-1 may enhance cervical carcinogenesis by promoting cell cycle progression. We also found that this HIV-1 Tat-induced cell proliferation was not dependent on the E2F family of transcription factors, and therefore postulate that Sp factors may be involved.

p53 Accumulation Is a Prognostic Factor in Intestinal-Type Gastric Carcinoma but Not in the Diffuse Type

AbstractBackground: The prognostic value of p53 nuclear accumulation in gastric cancer is still unclear, as shown by the discordant results still reported in the literature. In this study, we evaluated the correlation between p53 accumulation and long-term survival of patients resected for intestinal and diffuse-type gastric cancer. Methods: Eighty-three patients with carcinoma of the intestinal type and 53 patients with carcinoma of the diffuse type were included in the study. Immunohistochemical staining of the paraffin sections was performed by using monoclonal antibody DO1; cases were considered positive when nuclear immunostaining was observed in 10% or more of the tumor cells. Prognostic significance of different variables was investigated by univariate and multivariate analysis. Results: p53 positivity was found in 51.8% of intestinal-type and 50.9% of diffuse-type cases. No significant correlation between the rate of p53 overexpression and age, sex, tumor location, tumor size, depth of invasion, lymph node involvement, distant metastases, and surgical radicality was found in the two groups of patients. A statistically significant difference in survival rate was observed between p53-negative and p53-positive cases in the intestinal type (P < .05), confirmed by multivariate analysis (P < .005; relative risk = 3.09). On the contrary, no correlation with survival was found in diffuse-type cases according to p53 overexpression. Conclusions: These results suggest that the immunohistochemical detection of p53 accumulation is a useful indicator of poor prognosis in the intestinal but not in the diffuse type of gastric cancer, and are indicative of distinct molecular pathways and pattern of progression in the two histotypes.

Analysis of macrophage migration inhibitory factor (MIF) in patients with idiopathic pulmonary fibrosis.

By proteomic approach we previously characterised bronchoalveolar lavage (BAL) protein profiles of patients with idiopathic pulmonary fibrosis (IPF), sarcoidosis and systemic sclerosis. Among differently expressed proteins we identified macrophage migration inhibitory factor (MIF), a multi-function pleiotropic cytokine. This study was performed to validate our findings by a further proteomic approach and ELISA in a larger population of patients and controls. MIF expression in lung tissue was also evaluated by immunohistochemistry. MIF was identified in all 2-DE gels of IPF patients and it was significantly increased compared to controls (p<0.05). This result was confirmed by ELISA: MIF concentrations were significantly higher in IPF patients than controls (p<0.001) and were directly correlated with neutrophil percentages (p=0.0095). Immunohistochemical analysis revealed enhanced expression in bronchiolar epithelium, alveolar epithelium, and fibroblastic foci. In conclusion, MIF is a pleiotropic cytokine that could be involved in the pathogenesis of IPF, being particularly abundant in BAL of these patients and mainly expressed in the areas of active fibrosis.

Inflammatory Lung Disease in Rett Syndrome

Rett syndrome (RTT) is a pervasive neurodevelopmental disorder mainly linked to mutations in the gene encoding the methyl-CpG-binding protein 2 (MeCP2). Respiratory dysfunction, historically credited to brainstem immaturity, represents a major challenge in RTT. Our aim was to characterize the relationships between pulmonary gas exchange abnormality (GEA), upper airway obstruction, and redox status in patients with typical RTT (n = 228) and to examine lung histology in a Mecp2-null mouse model of the disease. GEA was detectable in ~80% (184/228) of patients versus ~18% of healthy controls, with “high” (39.8%) and “low” (34.8%) patterns dominating over “mixed” (19.6%) and “simple mismatch” (5.9%) types. Increased plasma levels of non-protein-bound iron (NPBI), F2-isoprostanes (F2-IsoPs), intraerythrocyte NPBI (IE-NPBI), and reduced and oxidized glutathione (i.e., GSH and GSSG) were evidenced in RTT with consequently decreased GSH/GSSG ratios. Apnea frequency/severity was positively correlated with IE-NPBI, F2-IsoPs, and GSSG and negatively with GSH/GSSG ratio. A diffuse inflammatory infiltrate of the terminal bronchioles and alveoli was evidenced in half of the examined Mecp2-mutant mice, well fitting with the radiological findings previously observed in RTT patients. Our findings indicate that GEA is a key feature of RTT and that terminal bronchioles are a likely major target of the disease.

Cell kinetics, morphology, and molecular IgVH gene rearrangements in Hodgkin's disease.

The present study dealt with the question of whether any cellular kinetic patterns correlate with clonal rearrangement of the IgVH gene as revealed by polymerase chain reaction on DNA extracted from lymph nodes with classical Hodgkins disease (HD) and/or from single CD30+ cells (Hodgkin [H] and Reed-Sternberg [RS] cells). In 15/4 cases with H-RS cells of B or Null phenotype, signs of such monoclonality could be detected (group I) but not in the others (group II). CD30+/H-RS cells in group I differed slightly but significantly from those in group II in that they a) exhibited a larger fraction of cells attaining the anaphase/telophase stage of mitosis, and b) produced relatively more mononucleated cells (H) at the expense of multinucleated (RS) cells. In addition, reactive lymphoid cell (CD30-) infiltrates were considerably less dense in group I that in group II. These findings suggest that the cytokinesis of H-RS cells in group I was moderately more efficient than in group II. However, signs of monoclonality were not associated with the normalization of the mitotic process, which also proved to be disturbed in group I.

A morphometric semiautomated method for analyzing cell nuclei in lymph node sections from non-Hodgkin's lymphomas. — Significance of data

A semiautomated electronic system has been employed for sizing nuclear area and for evaluating nuclear form factors in non-Hodgkins lymphomas with the purpose to correlate these parameters with survival and histotype. By mathematical models for best fits correlating the dependent variables with the survival has been demonstrated that an inverse correlation corresponding to a negative exponential function exists between mean nuclear area and survival. It has also been shown that the nuclear form is less irregular (i.e. more similar to an ellipse) when the mean nuclear area ranges from 12.5 to 20 mu2 and from 30 to 37.5 mu2 than when it ranges from 20 to 30 mu2. Lymphomas of low-grade malignancy are characterized by a mean nuclear area which is significantly lower than that of lymphomas of high-grade malignancy. The authors believe that morphometrical analysis of the nuclear area and of the form factors may eliminate subjectivity and give reproducible data to be used in both classification and prognosis of lymphomas.

Cell proliferation, cell death and angiogenesis in early and advanced gastric cancer of intestinal type

Mitotic (MI) and apoptotic index (AI), the sum of the 2, i.e., the turnover index (TI), tumor neovascularization (NV) and p53 expression, as well as tumor grading and node status, are evaluated in early and advanced gastric‐cancer cases. T1 cases show significantly less frequent lymph‐node invasion and lower tumor grade, and, taken together, have significantly lower MI, AI and TI and higher values of NV than the T2–3 cases. However, correlation of the variables shows that the above‐mentioned discrimination is due to a minority of T1 cases (11 out of 33), while the majority of them are allocated in the same 95% ellipse of tolerance of the T2–3 cases. Int. J. Cancer 74:637–641, 1997.© 1997 Wiley‐Liss, Inc.

Novel, contrast gradient-oriented, automated chromatin texture analysis. I: Feasibility study on nuclei from benign and malignant breast epithelial cell lines in fine needle aspirates

SummaryCoarse granularity of nuclear chromatin texture is a prominent feature of most malignant cell lines. We have chosen the abrupt transition from eu to heterochromatic foci (high contrast gradient [CG]) as a novel parameter for coarseness. This feature was quantified using automated image analysis of single nuclei in smears stained by the MayGrünwaldGiemsa technique. The principle of this approach consists of eliminating, with the help of substraction between two image lowpass filters, the small grey level differences among pixels, so that only high CG values are retained on the digitized image. The sum of these distinctive microareas is then taken as a fraction of the area of the peripherally eroded nucleus, and this ratio is designated as contrast gradient index (CGI) per nucleus. This method was tested on fine needle aspirates from 11 patients with benign breast disease (BBD) and 14 with mammary carcinoma (CA). For each specimen, 60 nuclei were analyzed, with a measuring time per nucleus of about 1 min. A highly significant distinction between epithelial cell populations in BBD and CA, respectively, was obtained by variance analysis of all CGIs per nucleus (p = 2 × 10−18). The median and the mean values of CGI per specimen were the next best discriminators, followed by the modes and the standard deviation of CGI per specimen. The percentage of nuclei per specimen with CGI values of >12 was also significantly greater in CA than in BBD.