The caspase recruitment domain gene CARD15/NOD2, encoding a cellular receptor involved in an NF-jBmediated pathway of innate immunity, was first identified as a major susceptibility gene for Crohn's disease (CD), and more recently, as responsible for Blau syndrome (BS), a rare autosomal-dominant trait characterized by arthritis, uveitis, skin rash and granulomatous inflammation. While CARD15 variants associated with CD are located within or near the C-terminal leucine-rich repeat domain and cause decreased NF-jB activation, BS mutations affect the central nucleotide-binding NACHT domain and result in increased NF-jB activation. In an Italian family with BS, we detected a novel mutation E383K, whose pathogenicity is strongly supported by cosegregation with the disease in the family and absence in controls, and by the evolutionary conservation and structural role of the affected glutamate close to the Walker B motif of the nucleotide-binding site in the NACHT domain. Interestingly, substitutions at corresponding positions in another NACHT family member cause similar autoinflammatory phenotypes.

CARD15 on chromosome 16 is the only IBD susceptibility gene identified among several mapped loci. Its recurrent variants R702W, G908R and L1007fs have shown significant association with Crohn's disease (CD), but not with ulcerative colitis (UC), in different Caucasian populations. We analysed these three variants in 184 CD and 92 UC Italian patients and in 177 healthy controls. L1007fs and G908R were independently associated with CD, while R702W showed a nonsignificant increase. After combining the three variants together, 32.6% of CD patients were positive vs 18.6% of the controls. The association was stronger for homozygotes and compound heterozygotes, OR 13.9 (1.8-108), and weaker but still significant for simple heterozygotes, OR 1.7 (1.0-2.9). An excess of homozygotes/compound heterozygotes also resulted from the comparison with Hardy -Weinberg expectations. Phenotypegenotype correlations were analysed first by univariate logistic regression and then by multivariate analysis, the effect of CARD15 positivity being adjusted according to the status of smoking, familiarity and sex, so as to focus on the predictivity of genetic and environmental risk factors on the clinical phenotype. Significant risk estimates of the CARD15 genotype were obtained for stricturing vs inflammatory behaviour, OR 2.76 (1.2 -6.3), and for penetrating behaviour, 2.59 (1.0 -6.6), and marginally significant for ileal vs colic location, OR 3.0 (0.9 -9.8). Our findings indicate that the association of the CARD15 genotype with behaviour and location of disease holds also for the Italian population.

NOD2 mutations affect the basal characteristics of MoDCs and their response to G- bacteria differently. MoDCs could be involved in CD onset because they have defects in releasing inflammatory cytokines and in polarizing T-cell responses.

The MET oncogene is aberrantly overexpressed in human osteosarcomas. We have previously converted primary cultures of human bone-derived cells into osteosarcoma cells by overexpressing MET. To determine whether MET transforms mesenchymal stem cells or committed progenitor cells, here we characterize distinct MET overexpressing osteosarcoma (MET-OS) clones using genome-wide expression profiling, cytometric analysis, and functional assays. All the MET-OS clones consistently display mesenchymal and stemness markers, but not most of the mesenchymal-stem cell-specific markers. Conversely, the MET-OS clones express genes characteristic of early osteoblastic differentiation phases, but not those of late phases. Profiling of mesenchymal stem cells induced to differentiate along osteoblast, adipocyte, and chondrocyte lineages confirms that MET-OS cells are similar to cells at an initial phase of osteoblastic differentiation. Accordingly, MET-OS cells cannot differentiate into adipocytes or chondrocytes, but can partially differentiate into osteogenic-matrix-producing cells. Moreover, in vitro MET-OS cells form self-renewing spheres enriched in cells that can initiate tumors in vivo. MET kinase inhibition abrogates the self-renewal capacity of MET-OS cells and allows them to progress toward osteoblastic differentiation. These data show that MET initiates the transformation of a cell population that has features of osteo-progenitors and suggest that MET regulates self-renewal and lineage differentiation of osteosarcoma cells. ß