Biomedical subjects
Publications and source records attributed to B R Olsen.
Cherubism (MIM 118400) is an autosomal dominant inherited syndrome characterized by excessive bone degradation of the upper and lower jaws followed by development of fibrous tissue masses, which causes a characteristic facial swelling. Here we describe seven mutations in the SH3-binding protein SH3BP2 (MIM 602104) on chromosome 4p16.3 that cause cherubism.
Venous malformations with glomus cells are localised cutaneous lesions of vascular dysmorphogenesis. They are usually sporadic, but sometimes familial. Using five families, we mapped the locus, VMGLOM, to chromosome 1p21-p22. In order to refine this locus, spanning 4-6 Mbp, we then studied seven additional families. They exhibited linkage to VMGLOM and the combined lod score for all 12 families was 18.41 at theta = 0.0 for marker D1S188. We found a distinct haplotype shared by seven families, comprising seven alleles which are rare in the general population (P < 0.01). This indicates that the haplotype is identical by descent in all seven families, and hence the locus can be refined by inferring ancestral crossovers. Using this approach, we position the causative gene between two markers on the same non-chimeric YAC of 1.48 Mbp, a feasible size for positional cloning. As there is no known gene involved in vasculogenesis and/or angiogenesis in this YAC, the identification of the causative gene is likely to reveal a novel regulator or vascular development.
Mutations in the serine/threonine kinase STK11 lead to Peutz-Jeghers syndrome (PJS) in a subset of affected individuals. Significant evidence for linkage to a second potential PJS disease locus on 19q13.4 has previously been described in one PJS family (PJS07). In the current study, we investigated this second locus for PJS gene candidates. We mapped the main candidate gene in this region, the gene for the transmembrane-type protein tyrosine phosphatase H (PTPRH), within 15 kb telomeric to the marker D19S880. We determined its genomic structure, and performed mutation analysis of all exons and the exon-intron junctions of the PTPRH gene in the PJS07 family. No disease causing mutation was identified in PTPRH in affected individuals, suggesting the existence of an as yet not identified gene on 19q13.4 as a second PJS gene.
Hemangioma, the most common tumor of infancy, is a benign vascular neoplasm of unknown etiology. We show, for the first time to our knowledge, that endothelial cells from proliferating hemangioma are clonal, and we demonstrate that these hemangioma-derived cells differ from normal endothelial cells in their rates of proliferation and migration in vitro. Furthermore, migration of hemangioma endothelial cells is stimulated by the angiogenesis inhibitor endostatin, unlike the inhibition seen with normal endothelial cells. We conclude that hemangiomas constitute clonal expansions of endothelial cells. This is consistent with the possibility that these tumors are caused by somatic mutations in one or more genes regulating endothelial cell proliferation.
OBJECTIVE: The extracellular matrix (ECM) of hyaline cartilage contains an elaborated collagen fibrillar network, which is essential for the mechanical stability and the proper function of the tissue. Cartilage collagen fibrils consist of collagen II, the quantitatively minor collagens IX and XI, and several non-collagenous fibril-associated proteins. To understand the role some of these molecules in skeletal development, we have generated transgenic mouse strains harboring ablated genes for collagens II and IX, and matrilin-1. DESIGN: Mice lacking collagen II, collagen IX and matrilin-1 have been established earlier in our laboratory using standard techniques. To determine the consequences of the null mutations we used skeletal staining, histochemical and immunohistochemical assays, in situ hybridization and ultrastructural analysis. RESULTS: Transgenic mice deficient in collagen II (Col2a1-/-) die at birth and display a severely malformed skeleton characterized by abnormal endochondral ossification and impaired intervertebral disc development. Mice lacking collagen IX (Col9a1-/-) are viable and develop an osteoarthritis-like phenotype in knee joints between 9-12 months of age. To test the possibility that the reduction in collagen II content has an influence on the onset of degenerative changes of articular cartilage, we have generated mice, which are heterozygous for the collagen II null mutation and homozygous for the collagen IX null mutation. Col2a1+/- Col9a1-/- mice show no accelerated development of osteoarthritis compared with the collagen IX knockout animals. Finally, mice lacking matrilin-1, a non-collagenous glycoprotein that binds to both collagen fibrils and aggrecan, develop normally without detectable abnormalities in their skeleton. CONCLUSIONS: Our transgenic mouse strains carrying null mutations in genes encoding cartilage ECM proteins demonstrate that these proteins have different roles during skeletal development. Collagen II is important for cartilage formation, collagen IX for cartilage maintenance and matrilin-1 is redundant.
The angiogenesis inhibitor endostatin is a fragment of the NC1 domain of collagen XVIII. The generation of endostatin has been investigated only in murine hemangioendothelioma cell cultures and was ascribed to cathepsin L. Distinct endostatin-like fragments were detected in human tissues and serum. To identify proteinases able to generate such fragments, we incubated human NC1 with proteinases of all classes, including cathepsin L. Eleven out of 12 generate fragments with an N-terminus within the same 15 residue stretch as those occurring physiologically, indicating that this region is sensitive to many proteinases. None correspond to mouse endostatin. However, the efficiencies of these proteinases differed markedly. Some proteinases also proved to degrade endostatin, pointing to another regulatory loop of angiogenesis.
BACKGROUND: Malignant tumours may produce substances with both stimulatory and inhibitory effect on angiogenesis. MATERIAL AND METHODS: A protein fragment with angiogenesis-inhibiting potential was recently identified in conditioned media from a murine endothelial tumour cell line. RESULTS: The angiogenesis inhibitor, endostatin, is a 20 kDa C-terminal fragment of collagen XVIII, a proteoglycan/collagen found in vessel walls and basement membranes. The generation of endostatin or endostatin-like collagen XVIII fragments is catalyzed by proteolytic enzymes, including cathepsin L and matrix metalloproteases, that cleave peptide bonds within the protease-sensitive hinge region of the C-terminal domain. INTERPRETATION: The physiological processing of collagen XVIII to endostatin may represent a local control mechanism for the regulation of angiogenesis. The outcome of ongoing clinical trials will determine the role of endostatin as a possible angiogenesis-inhibiting drug in the future.
Vascular anomalies are congenital lesions that usually occur sporadically, but can be inherited. Previously, we have described that venous malformations, localized bluish-purple skin lesions, are caused by an activating mutation in the TIE2/TEK receptor. Moreover, we mapped another locus to chromosome 1p21-p22, for venous malformations with glomus cells (VM-GLOM). Here we report a physical map, based on 18 overlapping YAC clones, spanning this 5-Mb VMGLOM locus, from marker GATA63C06 to D1S2664. In addition, we report a sequence-ready PAC map of 46 clones covering 1.48 Mb within the YAC contig, a region to which we have restricted VMGLOM. We describe 21 new STSs and nine novel CA repeats, seven of which are polymorphic. These data will enable positional cloning of genes for diseases mapped to this locus, including the VMGLOM gene, likely a currently unknown regulator of vasculogenesis and/or angiogenesis.
Endostatin, an inhibitor of angiogenesis and tumor growth, was identified originally in conditioned media of murine hemangioendothelioma (EOMA) cells. N-terminal amino acid sequencing demonstrated that it corresponds to a fragment of basement membrane collagen XVIII. Here we report that cathepsin L is secreted by EOMA cells and is responsible for the generation of endostatin with the predicted N-terminus, while metalloproteases produce larger fragments in a parallel processing pathway. Efficient endostatin generation requires a moderately acidic pH similar to the pericellular milieu of tumors. The secretion of cathepsin L by a tumor cell line of endothelial origin suggests that this cathepsin may play a role in angiogenesis. We propose that cleavage within collagen XVIII's protease-sensitive region evolved to regulate excessive proteolysis in conditions of induced angiogenesis.
Previous linkage analysis (Briggs, M.D., Choi, H.-C., Warman, M.L. et al., 1994. Genetic mapping of a locus for multiple epiphyseal dysplasia (EDM 2) to a region of chromosome 1 containing a type IX collagen gene. Am. J. Hum. Genet. 55, 678-684) in a large English family with multiple epiphyseal dysplasia established the EDM2 locus, a region of chromosome 1 containing the COL9A2 collagen gene. We now report that affected members of this family are heterozygous for a single base transversion (T-->G) at the sixth position of the intron 3 splice donor of COL9A2. The mutation leads to skipping of exon 3 during splicing, and results in a 36-nucleotide deletion in COL9A2 transcripts derived from the mutant allele. Skipping of exon 3 predicts an in-frame deletion of 12 amino acid residues within the COL3 domain of the alpha2(IX) chain. This is the fifth instance of an exon 3 deletion within the COL3 region of collagen IX heterotrimers causing the MED phenotype, as yet the only type IX collagen defect identified in this disorder. Electron microscopy (EM) of chondrocytes obtained from articular cartilage of one affected individual in the family demonstrated normal appearing rough endoplasmic reticulum (RER). In addition, the articular cartilage matrix did not show any gross abnormalities in the quantity or caliber of collagen fibrils.
Early development of the vertebrate skeleton depends on genes that pattern the distribution and proliferation of cells from cranial neural crest, sclerotomes, and lateral plate mesoderm into mesenchymal condensations at sites of future skeletal elements. Within these condensations, cells differentiate to chondrocytes or osteoblasts and form cartilages and bones under the control of various transcription factors. In most of the skeleton, organogenesis results in cartilage models of future bones; in these models cartilage is replaced by bone by the process of endochondral ossification. Lastly, through a controlled process of bone growth and remodeling the final skeleton is shaped and molded. Significant and exciting insights into all aspects of vertebrate skeletal development have been obtained through molecular and genetic studies of animal models and humans with inherited disorders of skeletal morphogenesis, organogenesis, and growth.
Collagen XII has been postulated to organize the extracellular matrix (ECM) architecture of dense connective tissues such as the periodontal ligament (PDL) and skin. The objective of this study was to test this hypothesis in transgenic mice carrying a dominant interference mutation of collagen XII. The truncated alpha1(XII) collagen minigene construct MXIINC3(-), driven by the mouse alpha2(I) collagen promoter, was prepared and used to generate transgenic mouse lines. The PDL matrix fibers of molar teeth lost the ordered architecture characteristic of ligament tissue without noticeable inflammation. Cellular cement appeared to be disrupted at the PDL insertion. By confocal laser scanning microscopy, the PDL of transgenic mice demonstrated swollen and irregularly arranged collagen fibers associated with internal porosity. The skin of transgenic mice revealed the lack of matrix fiber structure in the papillary dermis. These results indicated that the dominant interference mutation of collagen XII disorganized the ECM architecture of PDL and skin.
The angiogenesis inhibitor endostatin is a 20 kDA C-terminal fragment of collagen XVIII, a proteoglycan/collagen found in vessel walls and basement membranes. The endostatin fragment was originally identified in conditioned media from a murine endothelial tumor cell line. Endostatin inhibits endothelial cell migration in vitro and appears to be highly effective in murine in vivo studies. The molecular mechanisms behind the inhibition of angiogenesis have not yet been elucidated. Studies of the crystal structure of endostatin have shown a compact globular fold, with one face particularly rich in arginine residues acting as a heparin-binding epitope. It was initially suggested that zinc binding was essential for the antiangiogenic mechanism but later studies indicate that zinc has a structural rather than a functional role in endostatin. The generation of endostatin or endostatin-like collagen XVIII fragments is catalyzed by proteolytic enzymes, including cathepsin L and matrix metalloproteases, that cleave peptide bonds within the protease-sensitive hinge region of the C-terminal domain. The processing of collagen XVIII to endostatin may represent a local control mechanism for the regulation of angiogenesis.
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Endostatin, produced as recombinant protein in human 293-EBNA cells, inhibits the migration of human umbilical vein endothelial cells (HUVECs) in response to vascular endothelial growth factor (VEGF) in a dose-dependent manner and prevents the subcutaneous growth of human renal cell carcinomas in nude mice at concentrations and in doses that are from 1000- to 100 000-fold lower than those previously reported. The inhibition of migration is not affected by mutations which eliminate Zn or heparin binding and inhibition of tumor growth does not depend on Zn binding. The results of the migration assays suggest that endostatin causes a block at one or more steps in VEGF-induced migration, while VEGF in turn can cause a block of the inhibition by endostatin of VEGF-induced migration of HUVECs.
This paper reports the identification of two structural variations in the NC1 domain of rat and mouse type XII collagen. The long NC1 domain encoding 74 amino acids showed homology to chicken type XII and XIV collagens. The short NC1 domain was composed of 19 amino acids. Through genomic DNA analyses, two alternative exons were identified, each of which contained the variable NC1 sequence. With the amino-terminal NC3 splicing alternatives, we propose here a new descriptive nomenclature: types XIIA-1 and XIIB-1 which include a long NC1 sequence encoded by exon 1 (from the 3'-end), and types XIIA-2 and XIIB-2 which include a short NC1 sequence encoded by exon 2. Types XIIA-1 and XIIB-1, the predominant transcripts in 15-day old mouse embryos, showed decreased expression in 17-day old embryos when type XIIB-2 expression was sustained at constant levels. In adult mice, type XIIB-1 associates with ligament and tendon, whereas type XIIB-2 is expressed in various other tissues. The long NC1 domain contains an extended acidic region (pI = 3.4) followed by a terminal basic region (pI = 13.8). Because the short NC1 domain lacks these features, structural variations in the type XII collagen NC1 domain suggests different functional roles in a tissue-specific fashion.
OBJECTIVE: Proteoglycan-induced arthritis (PGIA) is a murine model of rheumatoid arthritis (RA), both in terms of its pathology and its genetics. PGIA can only be induced in susceptible murine strains and their F2 progeny. As with RA, the genetics are complex, containing both major histocompatibility complex (MHC)-related and non-MHC-related components. Our goal was to identify the underlying non-MHC-related loci that confer PGIA susceptibility. METHODS: We used 106 polymorphic markers to perform simple sequence-length polymorphism analysis on F2 hybrids of susceptible (BALB/c) and nonsusceptible (DBA/2) strains of mice. Because both strains of mice share the H2d haplotype, this cross permits identification and analysis of non-MHC-related genes. RESULTS: We identified a total of 12 separate quantitative trait loci (QTL) associated with PGIA, which we have named Pgia1 through Pgia12. QTLs associated with the inflammatory symptoms of PGIA were linked to chromosomes 7, 9, 15 (2 separate loci), 16, and 19. QTLs associated with autoantibody production were identified on chromosomes 1, 2, 7, 8, 10, 11, 16, and 18. QTLs on chromosomes 7 and 16 showed linkage to both inflammation and autoantibody production, suggesting a shared regulatory component in arthritis induction. The first inflammation QTL on chromosome 15 and the autoantibody QTL on chromosome 7 originate from the DBA/2 background, which indicates that as in RA, susceptibility genes can originate from heterogeneous backgrounds. CONCLUSION: These data demonstrate the complexity of PGIA, where QTLs may be involved in multiple traits or even originate from a genetic background previously determined to be resistant.
Bone wound created in intramembranous alveolar bone heals without the formation of cartilage precursor tissue. However, the expression of cartilage collagen mRNAs has been suggested. In this report, we examined the expression and the potential role of type IX collagen in bone restoration and remodeling. The sequence specific polymerase chain reaction demonstrated the exclusive expression of short transcriptional isoform of alpha1(IX) collagen (Col9a1) in alveolar bone wound healing, while the long isoform of Col9a1 transcript was absent. Type IX collagen was immunolocalized in the preliminary matrix organized in granulation tissue before trabecular bone formation in tooth extraction socket. In Col9a1-null mutant mice, there were considerable variations in alveolar bone wound healing with the absence of or abnormally organized trabecular bone. Occasionally, unusual apposition of cortical-bone-like layers in bone marrow space was observed. The Col9a1-null mice indicated no growth retardation, and their facial and long bones maintained the normal size and shape. However, the primary spongiosa region of adult Col9a1 mutant mice showed an abnormal trabecular bone structure associated with abnormal immunostaining with the hypertrophic cartilage specific type X collagen antibody. These data suggest that type IX collagen short transcriptional variant is involved in the restoration and remodeling processes of trabecular bone.