IL-12, pivotal to the development of Th1 cells and formed by association of p35 and p40 subunits, is made by macrophages and the macrophage cell line RAW264.7. In this study, the promoter for p35 was cloned and analyzed. The murine IL-12 p35 gene has promoters upstream from each of the first two exons. The exon 1 and exon 2 promoters, cloned into a reporter vector, were responsive to LPS or IFN-γ/CD40 ligation in transfected RAW264.7 cells. The exon 2 promoter containing bp −809 to +1 has significant homology to the human p35 promoter. Thus, deletion analysis was performed to determine the regions required for responsiveness to LPS, CD40, and/or IFN-γ. Base pairs −809 to −740 influenced responsiveness to LPS. In contrast, bp −740to −444 and bp −122 to −100 were required for responses to IFN-γ, IFN-γ/LPS, or IFN-γ/CD40 ligation. Removal of bp −444 to −392 increased the response of the exon 2 promoter to each stimulant. IFN regulatory factor (IRF)-1 is involved in the activity of this promoter at bp −108 to −103 because levels of nuclear IRF-1 correlated with exon 2 promoter activity in response to IFN-γ and IRF-1 overexpression stimulated and enhanced exon 2 promoter activity. Also, site or deletion mutation of the IRF-1 element at bp −108 to −103 reduced the responsiveness of the promoter and IRF-1 bound to an oligonucleotide containing bp −108 to −103. The data suggest that the response of the p35 promoter to IFN-γ requires a distinct IRF-1 positive regulatory element at bp −108 to −103.
Small cell osteosarcoma is a rare bone tumor of high-grade malignancy that most often arises in the metaphysis of long bones in the second decade of life. Cytogenetic and molecular genetic findings in small cell osteosarcoma are poorly defined. Conventional cytogenetic analysis of a small cell osteosarcoma arising in the proximal tibia of a 9-year-old male revealed a diploid chromosomal complement with complex structural rearrangements involving chromosomes 6, 16, and 17. Immunohistochemical assessment of p53 protein expression revealed nuclear p53 immunoreactivity in approximately 15% of the neoplastic cells. Subsequent fluorescence in situ hybridization (FISH) analyses confirmed loss of the p53 gene locus on the derivative chromosome 17 homolog and were negative for amplification of the MDM2, CDK4, c-MYC, HER-2/neu, CCND1, and COPS3 gene loci. To the best of our knowledge, this represents the first demonstration of monoallelic deletion of p53 in small cell osteosarcoma, suggesting that p53 alterations may play an important role in the development of small cell osteosarcoma as well as conventional osteosarcoma.
We report the fourth case of oral tongue ITAC, and present the first histologic evidence of metaplasia of oral cavity salivary epithelium. We also discuss adjuvant therapy recommendations given the lack of clarity for treatment of this rare disease.
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