Differential expression of myc, max and RB1 genes in human gliomas and glioma cell lines.

Deregulated expression of myc proto-oncogenes is implicated in several human neoplasias. We analysed the expression of c-myc, N-myc, L-myc, max and RB1 mRNAs in a panel of human gliomas and glioma cell lines and compared the findings with normal neural cells. The max and RB1 genes were included in the study because their protein products can interact with the Myc proteins, being thus putative modulators of Myc activity. Several gliomas contained c/L-myc mRNAs at levels higher than those in fetal brain, L-myc predominantly in grade II/III and c-myc in grade III gliomas. High-level N-myc expression was detected. In one small-cell glioblastoma and lower levels in five other gliomas. In contrast, glioma cell lines totally lacked N/L-myc expression. The in situ hybridisations revealed mutually exclusive topographic distribution of myc and glial fibrillary acidic protein (GFAP) mRNAs, and a lack of correlation between myc expression and proliferative activity, max and RB1 mRNAs were detected in most tumours and cell lines. The glioma cells displayed interesting alternative splicing patterns of max mRNAs encoding Max proteins which either suppress (Max) or augment (delta Max) the transforming activity of Myc. We conclude that (1) glioma cells in vivo may coexpress several myc genes, thus resembling fetal neural cells; but (2) cultured glioma cells expression only c-myc; (3) myc, max and RB1 are regulated independently in glioma cells; and (4) alternative processing of max mRNA in some glioma cells results in delta Max encoding mRNAs not seen in normal fetal brain.

Co-expression of collagens II and XI and alternative splicing of exon 2 of collagen II in several developing human tissues.

Northern analyses, RNAase protection assays and in situ hybridizations were used to study the expression of the mRNA for the alpha 2 chain of collagen XI and the two different mRNAs generated from the collagen II gene through alternative splicing of exon 2 in several different tissues of 15-19-week-old fetuses. The highest expression levels of procollagen alpha 2(XI) and alpha 1(II) mRNAs were detected in cartilage, but, using long exposure times, Northern hybridization revealed the presence of the approximately 5.3 kb procollagen alpha 1(II) mRNA in most tissues analysed: calvarial and diaphyseal bone, striated and cardiac muscle, skin, brain, lung, kidney, liver, small intestine and colon. Both alternatively spliced forms of the mRNA were present in these tissues. In cartilage, the short form of the procollagen alpha 1(II) mRNA (without exon 2 sequences) was clearly more abundant, whereas in most of the non-cartilaginous tissues the long form was the predominant one. Low levels of procollagen alpha 2(XI) mRNA were also seen in non-cartilaginous tissues: calvarial and diaphyseal bone, kidney, skin, muscle, intestine, liver, brain, and lung. In all the other positive tissues except brain cortex, both collagen II and XI transcripts were observed. The localization of collagen II and XI signals was identical in cartilage, kidney and skin. However, in cartilage the signal with collagen II probe was much higher than that with the collagen alpha 2(XI) probe. In epidermis the situation was reversed. Our results show considerable co-expression and co-localization of procollagen alpha 1(II) and alpha 2(XI) mRNAs in many tissues of developing human fetuses. Since the collagen alpha 1(II) gene also codes for the alpha 3(XI) chain of collagen XI we propose that some, but not all, of the expression of the collagen II gene in non-cartilaginous tissues relates to collagen XI production.

Gene expression during bone repair.

Detailed understanding of the basic events in fracture healing constitutes a foundation for the development of new approaches to stimulate bone healing. Since the fracture healing process repeats, in an adult organism, several stages of skeletal growth in the same temporal order, it offers an interesting model for developmental regulation of cellular phenotypes and tissue-specific genes. Molecular biology has introduced new methods to study the regulatory phenomena during the process of fracture repair. Gene technology has also produced purified growth factors for research, which will help to understand their roles in fracture healing. This review summarizes data on the regulation of genes coding for extracellular matrix components and growth regulatory molecules during fracture healing. The information available focuses on the sequential expression of genes coding for collagens, proteoglycans, and some other matrix proteins during secondary (callus) healing. The temporal and spatial appearance of the different connective tissue components, mesenchyme, cartilage, and bone, are closely linked to the expression of genes coding for their characteristic constituents. Members of the transforming growth factor-beta superfamily, such as the bone morphogenetic proteins (BMP), are currently the most interesting ones among the factors that regulate chondrogenesis and osteogenesis. In the coming years, the availability of new cloned probes combined with sensitive analytical methods, as reviewed here, will add greatly to our understanding of the various aspects of gene expression during bone repair. This information should provide answers to some of the unresolved questions in fracture callus development.

Matrix in cartilage and bone development: current views on the function and regulation of major organic components.

Study of the growth and development of cartilage and bone has been difficult because the structure of the tissues makes biological experiments hard to conduct. Recent advances in molecular biology have offered new possibilities for studying these processes. Many cartilage and bone specific cDNAs have been cloned and characterized and consequently used to localize the corresponding mRNAs in tissue sections. Developing cartilage and bone serve as a model for the study of extracellular matrix gene regulation during the proliferation, growth and differentiation of connective tissue cells. Normal skeletal growth and development are regulated by both systemic and local factors. The effects of many systemic hormones on bone metabolism have been studied extensively, but the pathways triggered by these hormones in the target cells are less well known. Recent evidence suggests that some growth factors, such as TGF-beta, IGFs and PDGF, act as local regulators of cartilage and bone metabolism. The different extracellular matrix components, e.g. collagens, are expressed differently in distinct cell types and developmental stages during cartilage and bone development. This model, therefore, facilitates the study of relations between the production of the various extracellular matrix components and the growth factors and the proto-oncogenes which may regulate them. Existing knowledge of the expression of major cartilage and bone components and their regulation during growth, differentiation and development is reviewed. An understanding of the normal growth and development of cartilage and bone is fundamental for elucidating the mechanisms underlying the various diseases -- both hereditary and acquired -- affecting the human skeleton.