The defined attenuated Listeria monocytogenes Δmpl2 mutant is an effective oral vaccine carrier to trigger a long-lasting immune response against a mouse fibrosarcoma

Listeria monocytogenes has been proposed as a carrier to elicit major histocompatibility complex class-I restricted immune responses able to protect against tumor challenge. In this study the properties of the attenuated L. monocytogenes Δmpl2 mutant has been evaluated in vivo against a highly aggressive mouse fibrosarcoma which expresses β-galactosidase (β-gal) as a tumor-associated antigen (TAA). Immunization with the vaccine prototypes resulted in both elicitation of specific antibodies and generation of cytotoxic lymphocytes (CTL). Oral vaccination protected 55–64% of the immunized animals from tumor take (p < 0.01) and strongly reduced the average size of the tumor in the other 34–45% (p < 0.01). Vaccinated mice developed a long-lasting response, which resulted in 100% protection from a subsequent tumor challenge. Substitution of the whole TAA by its CTL-defined immunodominant epitope resulted in 43% protection, suggesting a contribution of the humoral response to the observed antitumor effect. No statistically significant differences were observed in the antitumor response when mice were immunized with strains expressing the immunodominant TAA epitope in the context of carrier proteins which were either exported or restricted to the bacterial cytoplasm. This suggests that the topology of the recombinant antigen does not play a major role in the outcome of the protective response.     

Expression of insulin-like growth factor II (IGF–II) in human hepatocellular carcinomas: An immunohistochemical study

Recent results obtained using molecular biology techniques have suggested a possible role for insulin-like growth factor II (IGF-II) in the pathogenesis of hepatocellular carcinoma (HCC). To investigate this phenomenon, a monoclonal anti-body was used against IGF-II to study 54 patients with HCC. The presence of HBsAg was also tested both in serum and liver tissue. A positive immunoreaction was found in 9/15 (60%) of the HCC arising in cirrhotic livers of patients who had serum markers for HBV (HBV+ positive patients). These results provide further evidence that HBV might play a role in the expression of IGF-II. In HCC of patients without any markers of HBV infection (HBV- negative patients), IGF–II was detected in 10/39 (25.6%) of the tumors, and in some benign neoplastic lesions. It was found not only in neoplastic cells but also in some dysplastic nodules. The speculation arises that IGF–II expression may play a role in some steps of hepato-carcinogenesis.     

Could mitochondrial haplogroups play a role in sporadic amyotrophic lateral sclerosis?

Mitochondrial impairment has been implicated in the pathogenesis of the amyotrophic lateral sclerosis (ALS). Furthermore, mitochondrial-specific polymorphisms were previously related to other neurodegenerative diseases, such as Parkinson, Friedreich and Alzheimer disease. To investigate if specific genetic polymorphisms within the mitochondrial genome (mtDNA) could act as susceptibility factors and contribute to the clinical expression of sporadic ALS (sALS), we have genotyped predefined European mtDNA haplogroups in 222 Italian patients with sALS and 151 matched controls. Individuals classified as haplogroup I demonstrated a significant decrease in risk of ALS versus individuals carrying the most common haplogroup, H (odds ratio 0.08, 95% confidence interval 0.04-0.4, p < 0.01). Further stratification of the dataset by sex, age and site of onset of disease and survival failed to reach significance for association. Our study provides evidence of the contribution of mitochondrial variation to the risk of ALS development in Caucasians. Further it may help elucidate the mechanism of the mitochondrial dysfunction detectable in ALS, and may be of relevance in development of strategies for the treatment of this disease.     

A novel patient with White–Sutton syndrome refines the mutational and clinical repertoire of the POGZ‐related phenotype and suggests further observations

A rare developmental delay (DD)/intellectual disability (ID) syndrome with craniofacial dysmorphisms and autistic features, termed White–Sutton syndrome (WHSUS, MIM#614787), has been recently described, identifying truncating mutations in the chromatin regulator POGZ (KIAA0461, MIM#614787). We describe a further WHSUS patient harboring a novel nonsense de novo POGZ variant, which afflicts a protein domain with transposase activity less frequently impacted by mutational events (DDE domain). This patient displays additional physical and behavioral features, these latter mimicking Smith–Magenis syndrome (SMS, MIM#182290). Considering sleep–wake cycle anomalies and abnormal behavior manifested by this boy, we reinforced the clinical resemblance between WHSUS and SMS, being both chromatinopathies. In addition, using the DeepGestalt technology, we identified a different facial overlap between WHSUS patients with mutations in the DDE domain (Group 1) and individuals harboring variants in other protein domains/regions (Group 2). This report further delineates the clinical and molecular repertoire of the POGZ‐related phenotype, adding a novel patient with uncommon clinical and behavioral features and provides the first computer‐aided facial study of WHSUS patients.     

Nitrosative stress in the bronchial mucosa of severe chronic obstructive pulmonary disease

BACKGROUND: Reactive nitrogen species, formed via the reaction of nitric oxide (NO) with superoxide anion and via (myelo)peroxidase-dependent oxidation of NO(2)(-), have potent proinflammatory and oxidizing actions. Reactive nitrogen species formation and nitrosative stress are potentially involved in chronic obstructive pulmonary disease (COPD) pathogenesis. OBJECTIVES: To investigate the expression of markers of nitrosative stress, including nitrotyrosine (NT), inducible NO synthase (iNOS), endothelial NO synthase (eNOS), myeloperoxidase (MPO), and xanthine oxidase (XO) in bronchial biopsies and bronchoalveolar lavage from patients with mild to severe stable COPD compared with control groups (smokers with normal lung function and nonsmokers). METHODS: The expression of NT, iNOS, eNOS, MPO and XO in the bronchial mucosa and bronchoalveolar lavage of patients was measured by using immunohistochemistry, Western blotting, and ELISA and correlated with the inflammatory cell profile. RESULTS: Patients with severe COPD in stable phase had higher numbers of NT(+) and MPO(+) cells in their bronchial submucosa compared with mild/moderate COPD, smokers with normal lung function, and nonsmokers (P < .01). iNOS(+) and eNOS(+) but not XO(+) cells were significantly increased in smokers with COPD or normal lung function compared with nonsmokers (P < .05 and P < .01, respectively). In patients with COPD, the number of MPO(+) cells was significantly correlated with the number of neutrophils (r = +0.61; P < .0025) in the bronchial submucosa. Furthermore, the number of NT(+) and MPO(+) cells was negatively correlated with postbronchodilator FEV(1). CONCLUSION: These data suggest that nitrosative stress, mainly mediated by MPO and neutrophilic inflammation, may contribute to the pathogenesis of severe COPD.     

Multiple pathways of cell invasion are regulated by multiple families of serine proteases

The complex process of tumor invasion requires the coordinated expression and activity of cell-substratum adhesive interactions and of cell-associated protease systems, which destroy the extracellular matrix (ECM), in order to enable the invading cells to simultaneously grip and destroy the anatomical barriers that control cell spreading. A number of data indicate that such a `grip and go' process may be performed by an enlarging series of cell membrane-associated serine proteases and serine protease receptors, which provide the invasive cells with a functional unit (the protease and its receptor), able to mediate cell-substratum adhesion through specific receptor domains, to proteolytically degrade ECM and to deliver into the cell signals that up-regulate the expression either of the protease/receptor complex, or of other adhesion molecules, such as integrins. There is evidence that some proteases and protease receptor expression are under the control of tumor hypoxia, which is the result of an imbalance in oxygen supply and demand. The urokinase-type plasminogen activator (u-PA) receptor (u-PAR) is under hypoxic control and cooperates with other serine proteases of the blood coagulation pathways that may extravasate in the tumor milieu as a result of hypoxia-simulated increase of vessel permeability. Other serine proteases and their receptors cooperate with the cell-associated fibrinolytic system to promote cell invasion. Among these, tissue factor and its ligand coagulation factor VII, thrombin and its protease-activated receptors, and type II trans-membrane serine proteases seem to play a crucial role. This Review takes into consideration the complex scenario of the single serine proteases and related receptors that are involved in cell invasion, as well as the protease receptor/adhesion molecule interplay which is necessary to focus the cell surface-driven proteolysis where adhesion provides a grip to the invading cell. This revised version was published online in July 2006 with corrections to the Cover Date.     

Clinical,immunological, and molecular analysis in a large cohort of patients with X-linked agammaglobulinemia: an Italian multicenter study

A questionnaire-based retrospective clinical and immunological survey was conducted in 73 males with a definite diagnosis of X-linked agammaglobulinemia based on BTK sequence analysis. Forty-four were sporadic and 29 familial cases. At December 2000, the patients' ages ranged from 2 to 33 years; mean age at diagnosis and mean duration of follow-up were 3.5 and 10 years respectively. After the mid-1980s all but 2 were on intravenous immunoglobulin (IVIG) substitution therapy, with residual IgG >500 mg/dl in 94% of the patients at the time of enrollment. Respiratory infections were the most frequent manifestation both prior to diagnosis and over follow-up. Chronic lung disease (CLD) was present in 24 patients, in 15 already at diagnosis and in 9 more by 2000. The cumulative risk to present at diagnosis with CLD increased from 0.17 to 0.40 and 0.78 when the diagnosis was made at the ages of 5, 10, and 15 years respectively. For the 9 patients who developed CLD during follow-up, the duration of follow-up, rather than age at diagnosis; previous administration of intramuscular immunoglobulin; and residual IgG levels had a significant effect on the development of CLD. Chronic sinusitis was present in 35 patients (48%), in 15 already at diagnosis and in 20 by 2000. Sistemic infections such as sepsis and meningitis/meningoencephalitis decreased over follow-up, probably due to optimal protection provided by high circulating IgG levels reached with IVIG.     

Successful xenografting of cryopreserved primary pancreatic cancers

In order to assess the suitability of cryopreserved neoplastic tissues for xenografting into nude (nu/nu) mice, we compared the take rate in 28 samples of pancreatic ductal carcinoma. Eleven fresh samples were implanted in nu/nu mice, and 17 were frozen in cryopreserving solution and implanted at a later time. All samples were examined for the presence of neoplastic tissue in cryostat sections. A total of 15 tumors grew in the animals; five from the freshly implanted samples and ten from those cryopreserved. Ten xenografted tumors were characterized for alterations in p53, K-ras, and p16 genes, which were found in six, eight, and nine cases, respectively. Our results demonstrate that the take rate for xenografting is comparable between cryopreserved and fresh tissue samples. The procedure allows for the exchange of tumor material between institutions and permits the establishment of centralized facilities for the storage of an array of different primary tumor samples suitable for the production of in vivo models of cancers.