Abstract
Extract: We have examined control subjects and patients in an effort to discover a metabolic basis for dominantly inherited osteogenesis imperfecta (OI). Studies were carried out in vitro with cultured skin fibroblasts obtained from OI patients, and in vivo on peptide-bound hydroxyproline excretion in urine. Urinary hydroxyproline excretion (milligrams/24 hr) adjusted for age is essentially normal in OI patients, although the mean excretion rate is below average. The latter finding is presumably a reflection of the smaller body mass of OI patients.
The OI skin fibroblasts, matched for age of donor, site of biopsy, phase of growth, and generation number in culture, incorporate L-proline into hot trichloroacetic acid (TCA)-soluble protein (collagen) at normal rates. The rate of conversion of proline to hydroxyproline in the nascent polypeptide is also normal in OI. Incorporation of L-lysine was also normal in OI. These findings indicate that peptide synthesis of collagen is not impaired in OI.
Rates of galactose incorporation into collagen and the extractability of collagen into normal saline or 0.2 M citric acid were all normal both in OI cells and in the culture medium recovered from the monolayer. These findings, in combination with the urinary data on hydroxyproline excretion in vivo reveal that cross-linking and export of collagen in OI is essentially normal.
The elution profile after ion exchange chromatography of fibroblast collagen on carboxymethyl (CM)-Sephadex was also examined. The normal 2/1 ratio of peak 1 (largely α1(I) chains) to peak 2 (largely α2 chains) was found in OI fibroblast extracts, which implies that synthesis and initial aggregation of the two types of polypeptide to yield [α1(I)]2α2 collagen composition is not abnormal in OI.
Despite the negative biochemical findings, a consistent defect in the morphology of OI cells was identified in the log phase and the confluent phase of monolayer cultures. The finding is characterized by irregular packing of the aggregated cells and by an irregular tessellated appearance of the individual OI fibroblast. This observation reassures us that the inherited defect is expressed in vitro.
Speculation: An abnormality in the primary sequence of polypeptide chain in collagen would be compatible with all of our findings and with the genetics of OI. The mutant allele would affect only about half the products, under the control of only one of the loci determining the polypeptide sequences in collagen chains. Because the OI allele is not expressed in cartilage, a tissue without α2 collagen chains, the defect in OI would perhaps be found in the α2 polypeptide. However, since the α1(II) chain of cartilage differs in amino acid composition and in hydroxylysine-linked carbohydrate from the α1(I) chains of noncartilagenous structures, a defect in α1(I) chains at the nonhomologous residues will also require investigation.
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Lancaster, G., Goldman, H., Scriver, C. et al. Dominantly Inherited Osteogenesis Imperfecta in Man: An Examination of Collagen Biosynthesis. Pediatr Res 9, 83–88 (1975). https://doi.org/10.1203/00006450-197502000-00005
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DOI: https://doi.org/10.1203/00006450-197502000-00005