Carlo Della Rocca

Transgenic expression in the liver of truncated Met blocks apoptosis and permits immortalization of hepatocytes.

Hepatocyte growth factor induces proliferation, motility and differentiation of epithelial cells through the tyrosine kinase receptor encoded by the MET proto‐oncogene. The cytoplasmic portion of Met (referred to as cyto‐Met) is activated but only weakly transforming. In order to determine the effect of activated Met on hepatocytes, we have targeted truncated Met expression to the liver by incorporating the cDNA into a vector carrying the entire human a‐1‐antitrypsin transcriptional unit. Transgenic expression in the liver of truncated human Met, containing the regulatory and the catalytic cytoplasmic domains, renders hepatocytes constitutively resistant to apoptosis and reproducibly permits immortalization. The emerging stable cell lines are not transformed and maintain a highly differentiated phenotype judged by the retention of epithelial cell polarity and the expression of hepatocyte‐enriched transcription factors as well as hepatic products.

Coexpression of IL-6 and soluble IL-6R causes nodular regenerative hyperplasia and adenomas of the liver

Studies with tumor necrosis factor p55 receptor‐ and interleukin‐6 (IL‐6)‐deficient mice have shown that IL‐6 is required for hepatocyte proliferation and reconstitution of the liver mass after partial hepatectomy. The biological activities of IL‐6 are potentiated when this cytokine binds soluble forms of its specific receptor subunit (sIL‐6R) and the resulting complex interacts with the transmembrane signaling chain gp130. We show here that double transgenic mice expressing high levels of both human IL‐6 and sIL‐6R under the control of liver‐specific promoters spontaneously develop nodules of hepatocellular hyperplasia around periportal spaces and present signs of sustained hepatocyte proliferation. The resulting picture is identical to that of human nodular regenerative hyperplasia, a condition frequently associated with immunological and myeloproliferative disorders. In high expressors, hyperplastic lesions progress with time into discrete liver adenomas. These data strongly suggest that the IL‐6/sIL‐6R complex is both a primary stimulus to hepatocyte proliferation and a pathogenic factor of hepatocellular transformation.

Histopathologic Analysis of Peritumoral Pseudocapsule and Surgical Margin Status after Tumor Enucleation for Renal Cell Carcinoma

BACKGROUND The oncologic safety of blunt tumor enucleation (TE) of renal cell carcinoma (RCC) depends on the presence of a continuous pseudocapsule (PS) around the tumor and on the possibility of obtaining negative surgical margins (SMs). OBJECTIVE To investigate the PS and SMs after TE to define the real need to take a rim of healthy parenchyma around the tumor to avoid the risk of positive SMs. The risk of PS invasion related to other clinical and pathologic variables was also evaluated. DESIGN, SETTING, AND PARTICIPANTS Between September 2006 and December 2007, data were gathered prospectively from 187 consecutive patients who had kidney surgery. Overall, 90 consecutive patients who had TE for RCC were eligible for the study. All specimens were evaluated using an image analyzer by a dedicated uropathologist. INTERVENTION TE was done by blunt dissection using the natural cleavage plane between the tumor and the normal parenchyma. MEASUREMENTS PS, SM, and routinely available clinical and pathologic variables were recorded. RESULTS AND LIMITATIONS In 60 RCC tumors (67%) the PS was intact and free from invasion (PS-) while in 30 (33%) there were signs of penetration within its layers, with or without invasion beyond it. Indeed, 26.6% had PS that had been penetrated on the parenchymal side and 6.6% had penetration on the perirenal fat tissue side. The odds of having PS penetration increased significantly with an increase in clinical tumor size. PS penetration was also significantly associated with pathologic tumor dimensions and grade. In all cases the SMs were negative after TE. The present patients, followed for >2 yr, will enable us to correlate the risk of local recurrence with PS status. CONCLUSIONS The risk of PS penetration is associated with clinical and pathologic tumor dimensions and grade. If there is PS invasion into normal parenchyma, the presence of a thin layer of tissue allows for negative SM even if a TE is performed.

An improved helper-dependent adenoviral vector allows persistent gene expression after intramuscular delivery and overcomes preexisting immunity to adenovirus

Helper-dependent adenoviral vectors deleted of all viral coding sequences have shown an excellent gene expression profile in a variety of animal models, as well as a reduced toxicity after systemic delivery. What is still unclear is whether long-term expression and therapeutic dosages of these vectors can be obtained also in the presence of a preexisting immunity to adenovirus, a condition found in a high proportion of the adult human population. In this study we performed intramuscular delivery of helper-dependent vectors carrying mouse erythropoietin as a marker transgene. We found that low doses of helper-dependent adenoviral vectors can direct long-lasting gene expression in the muscles of fully immunocompetent mice. The best performance—i.e., 100% of treated animals showing sustained expression after 4 months—was achieved with the latest generation helper-dependent backbones, which replicate and package at high efficiency during vector propagation. Moreover, efficient and prolonged transgene expression after intramuscular injection was observed with limited vector load also in animals previously immunized against the same adenovirus serotype. These data suggest that human gene therapy by intramuscular delivery of helper-dependent adenoviral vectors is feasible.

Degenerative changes of porcine intervertebral disc induced by vertebral endplate injuries.

Study Design. Graded endplate injuries were performed in porcine lumbar discs. The effects of such injuries were compared to control animals in which a sham operation was performed. Objectives. To investigate the effects of endplate injuries on disc tissue. Summary of Background Data. Studies have shown that injuries of vertebral endplates are frequently found at autopsy. However, little is known on the effects of acute injuries of vertebral endplates in vivo. Methods. Ten domestic pigs were included in the study group. Under general anesthesia, the lower three discs of the lumbar spine were exposed and randomly submitted to multiple endplate injuries, isolated endplate injury, and no treatment. A sham operation was performed in 5 pigs used as control group. Animals were killed 7 months after surgery and the harvested lumbar spine submitted to MRI investigations, histologic, and biochemical analysis. Results. MRI showed that all but one discs treated with multiple endplate injuries were markedly degenerated while, of the discs treated with an isolated injury, one was markedly degenerated, five slightly degenerated and two were normal (P = 0.01). Histologic analysis showed severe changes in discs treated with multiple injuries. In those who had an isolated injury, changes were less severe and essentially limited to the posterior anulus or the inner anterior anulus. Biochemical analysis showed an inverse correlation between uronate content in the nucleus pulposus and severity of endplate injuries. Conclusions. Injuries of vertebral endplates in porcine discs were found to cause degenerative changes in the disc tissue on MRI, histologic, and biochemical investigations. The severity of such degenerative changes was related to the severity of endplate injuries. Injuries of vertebral endplate may be one of the pathomechanisms leading to early changes in the disc matrix and eventually to abnormal biomechanical behavior of the whole disc. The present animal model seems to be a suitable experimental model for disc degeneration.

Evolution of precancerous laryngeal lesions: A clinicopathologic study with long-term follow-up on 259 patients

A wide spectrum of lesions ranging from dysplasia to in situ carcinoma have to be considered when dealing with laryngeal precancerous conditions. Recently the concept of laryngeal intraepithelial neoplasia (LIN) was introduced.

Bone histomorphometric reference values in 88 normal Italian subjects

The study deals with bone histomorphometric results obtained from 88 normal subjects (38 women and 50 men; range: 20-89 years), in order to establish control values in an Italian population. Bone specimens were obtained at autopsy from a standardized area of the iliac crest. The following indicators were measured: bone volume (BV/TV), osteoid volume, osteoid surface, osteoblast surface, eroded surface, osteoclast surface, osteoid thickness. Dependence of histomorphometric indicators on sex and age was evaluated by multiple regression analysis, including sex, age, and also a quadratic term (age2) and two interaction terms (sex x age, sex x age2). BV/TV was mainly affected by age. In fact, a decrease in the amount of bone was found with increasing age in both males and females. The reduction appeared rather regular, with negligible differences between males and females. The other indicators were found to be age- and sex-independent. As a consequence, they did not give information on the possible changes of bone apposition and resorption processes due to aging. On the whole, histomorphometric indicators of trabecular bone of the normal Italian population do not greatly differ from those reported for most other caucasian people.

Prognostic Role of Histological Necrosis for Nonmetastatic Clear Cell Renal Cell Carcinoma: Correlation With Pathological Features and Molecular Markers

PURPOSE We defined the prognostic role of tumor necrosis and its extent in nonmetastatic clear cell renal cell carcinoma. Also, we further investigated its pathogenesis by correlating this tumor feature with other pathological characteristics and molecular markers related to the von Hippel Lindau-hypoxia inducible factor pathway and to tumor proliferation. MATERIALS AND METHODS A total of 213 patients with nonmetastatic clear cell renal cell carcinoma were evaluated. Mean followup was 66 months. The presence and extent of histological necrosis were correlated with clinicopathological factors, Ki-67 antigen expression calculated by the MIB-1 (Ki-67 antibody) index, pVHL, HIF-1alpha, the tumor infiltrating lymphocyte subset and cancer specific survival. RESULTS Histological necrosis was present in 63.8% of clear cell renal cell carcinoma cases. Necrosis was significantly associated with grade and the degree of tumor infiltrating lymphocytes, while its extent correlated significantly with grade, the degree of tumor infiltrating lymphocytes and stage. Tumor necrosis was a significant prognostic factor, which was confirmed even when limiting analysis to patients with intracapsular renal cell carcinoma. On multivariate analysis histological necrosis was not an independent predictor of cancer specific survival. The extent of tumor necrosis was not a significant prognostic factor. The presence and extent of histological necrosis was not associated with high Ki-67 expression and it did not correlate with pVHL expression or with nuclear and cytoplasmic HIF-1alpha expression. CONCLUSIONS Based on our results we cannot support histological necrosis and its extent as prognostic factors for clear cell renal cell carcinoma. Efforts should be made to develop nomograms that use routinely available and objective predictor variables. The precise mechanism that causes tumor necrosis remains unknown but the host immune response might significantly contribute to its development.

Phosphorylated ezrin is located in the nucleus of the osteosarcoma cell

The survival of osteosarcoma patients is connected to metastasis. The ezrin expression is associated with the development of metastasis and poor outcome in osteosarcoma. Ezrin is present in the cytoplasm and after phosphorylation assumes an active form and links F-actin to the cell membrane. This study evaluated ezrin and phosphorylated ezrin at site Tyr354 and Thr567 expression and its subcellular localization in osteosarcoma. We studied 50 osteosarcoma patients (mean follow-up 9.8 years). Ezrin expression was assessed using immunohistochemical and immunofluorescence analysis on tissue microarray and cultured cells of human osteosarcoma 143B. The western blot analysis was carried out on cultured cells. The majority of osteosarcomas, showing cytoplasmic positivity for ezrin, phosphorylated and unphosphorylated, were associated with membranous and nuclear positivity for phosphorylated ezrin Thr567 and phosphorylated ezrin Tyr354, respectively. Ezrin expression was associated with high-grade osteosarcoma (P=0.04), with metastasis (P=0.04) and with tumors that developed metastasis (P=0.04); phosphorylated ezrin Thr567 expression was present mostly in tumors with metastasis (P=0.01) and in osteosarcomas that did not develop metastasis (P=0.002). The osteosarcoma patients with ezrin expression have a short survival. The cytoplasmic ezrin expression in osteosarcoma matches its role of membrane-cytoskeleton linker protein. The subcellular trafficking of ezrin is not blocked and it is linked to ezrin phosphorylation, also in cancer. The phosphorylated ezrin Tyr354 nuclear localization suggests its possible role as a nuclear factor in osteosarcoma. The phosphorylated ezrin Thr567 phosphorylation may not be necessary in osteosarcoma metastatic progression but it was modulated. The ezrin expression is associated with more aggressive osteosarcomas and with metastasis.

Histological and structural study of the adhesive tissue in knee fibroarthrosis: a clinical-pathological correlation.

In fibroarthrosis of the knee, it is still unknown if joint range of motion is affected by anatomopathological differences in adhesive tissue, such as tissue maturity, location, and quantity. A retrospective study of 78 patients who underwent arthoscopic arthrolysis was performed to determine a correlation between location of adhesions and preoperative range of motion (ROM). In another 17 patients, a histological and structural evaluation of adhesive tissue was performed. Based on vascularization, number and shape of cells, and collagen fiber orientation, the adhesive tissue was classified into three groups: low, medium, and high maturity. Preoperative joint ROM and the time of onset of joint stiffness was correlated with the degree of adhesion tissue maturity. A strong and statistically significant correlation between the location of adhesions and joint restriction was found. However, histological and structural evaluation showed no correlation between the degree of tissue maturity, the time of onset of joint stiffness, and the amount of joint ROM.