Form Of Osteogenesis Imperfecta Research Articles

A new lethal brittle bone syndrome with increased amount of type V collagen in a patient

A new lethal brittle bone disease is described in three patients with slender long bones, thin ribs, hypomineralized calvaria, and normal facial appearance. In spite of several limb fractures this syndrome can be differentiated from the lethal forms of osteogenesis imperfecta and is better related to the thin-bone group of lethal dysplasias. Biochemical investigation of collagen from one of the patients by the use of gel electrophoresis and high-pressure liquid chromatography analyses failed to demonstrate any evident defect in the structure of type I collagen chains. Nevertheless collagen extractability from the dermis was altered owing to an increase in the proportion of acid-soluble material. Tritium-proline labeling of cultured fibroblasts confirmed the reduction in total collagen synthesis. This was attributed to a lower type I and type III amount whereas type V collagen level was markedly increased in the cell layer. RNA analysis of the three collagen types with the appropriate cDNA probes confirmed the protein data. Electron microscopic examination of bone and skin showed morphologically abnormal fibroblasts and osteoblasts with an abundant distended rough endoplasmic reticulum and an altered plasma membrane. Unexpected thin fibrils with a banding pattern and surrounding the type I fibrils were observed. They might represent type V collagen. We suggest that, in this patient, the moderate decrease in type I collagen amount is insufficient to account for the radiological findings and that type V collagen overproduction could play a role in the bone brittleness by interfering with the process of mineralization.

Osteogenesis imperfecta: insufficient collagen synthesis in early childhood as evidenced by analysis of compact bone and fibroblast cultures

We analysed the composition of compact bone from 30 patients suffering from various forms of osteogenesis imperfecta (OI). Collagen and total protein content per cell of controls increased with the age of the donors, but were generally low in OI. In fibroblast cultures controls had a maximum of collagen synthesis between 2 and 9 years of age, an observation which was not seen in OI cells. In bone collagen both OI type II patients showed overhydroxylation of lysyl residues as did some patients with OI type III (25%) and OI type IV (33%). The collagen of OI type I patients was never found to be overmodified. In controls, collagen III was found exclusively during fetal time while it was present in significant amounts in bone tissue of all types of OI. The proportion of collagen V was somewhat higher in OI bones (about twice) than in controls. Our data suggest that the normal increase of collagen synthesis is defective in patients with OI. Perhaps some of these changes are due to specific molecular defects in collagen while others may be due to defective regulation of the maturation process.

Prenatal diagnosis and prevention of inherited abnormalities of collagen

There is now strong evidence for the implication of collagen alpha 1(I), alpha 2(I) and alpha 1(III) mutations in many forms of osteogenesis imperfecta and inherited arterial aneurysms (Ehlers Danlos syndrome type IV). A sizeable proportion of these disorders have detectable abnormalities by conventional protein chemistry, immunofluorescence, or more sophisticated DNA analysis. Everyone of them with specific defects or with linkage to appropriate gene markers is therefore amenable to prevention using conventional prenatal diagnosis by chorionic villus biopsy (with fibroblast culture), fetoscopic biopsy (with fibroblast culture), ultrasound diagnosis of the severely deformed fetus, or gene linkage studies by chorionic villus biopsy or amniocentesis. Already many collagen alpha 1(I), alpha 2(I) and alpha 1(III) mutations have been characterized including point mutations, small and large deletions and regulatory mutations. Many others are likely to be rapidly studied by exploiting recent advances in DNA technology, and other strong candidate genes include collagen II (some chondrodystrophies), collagen VI (certain arterial and cardiovascular diseases) and collagen VII (dystrophic epidermolysis bullosa). Other important common diseases are likely to include osteoporosis, osteoarthritis and cerebral aneurysms. A detailed review is provided of collagen interstitial genes and proteins, together with a description of the various forms of osteogenesis imperfecta and Ehlers Danlos syndrome in which either collagen alpha 1(I), alpha 2(I) or alpha 1(III) mutations have been identified. Appropriate restriction length polymorphisms (RFLPs) useful in identifying carriers of these mutant genes are also described.

The molecular defect in an autosomal dominant form of osteogenesis imperfecta. Synthesis of type I procollagen containing cysteine in the triple-helical domain of pro-alpha 1(I) chains.

Synthesis of procollagen was examined in skin fibroblasts from a patient with a moderately severe autosomal dominant form of osteogenesis imperfecta. Proteolytic removal of the propeptide regions of newly synthesized procollagen, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions, revealed the presence of type I collagen in which two alpha 1(I) chains were linked through interchain disulfide bonds. Fragmentation of the disulfide-bonded alpha 1(I) dimers with vertebrate collagenase and cyanogen bromide demonstrated the presence of a cysteine residue in alpha 1(I)CB8, a fragment containing amino acid residues 124-402 of the alpha 1(I) collagen chain. Cysteine residues are not normally found in the triple-helical domain of type I collagen chains. The heterozygous nature of the molecular defect resulted in the formation of three kinds of type I trimers: a normal type with normal pro-alpha(I) chains, a type I trimer with one mutant pro-alpha 1(I) chain and two normal chains, and a type I trimer containing two mutant pro-alpha 1(I) chains and one normal pro-alpha 2(I) chain. The presence of one or two mutant pro-alpha 1(I) chains in trimers of type I procollagen was found to reduce the thermal stability of the protein by 2.5 and 1 degree C, respectively. In addition to post-translational overmodification, procollagen containing one mutant pro-alpha 1(I) chain was also cleared more slowly from cultured fibroblasts. The most likely explanation for these disruptive changes in the physical stability and secretion of the mutant procollagen is that a cysteine residue is substituted for a glycine in half of the pro-alpha 1(I) chains synthesized by the patient's fibroblasts.

The differential symptomatology of errors of collagen metabolism: a tentative classification.

We address the confusion in the current classification of inherited disorders of collagen and the excessive extension of the concept of the Ehlers-Danlos "syndrome" that tends to cover many facts and conditions frequently without strong clinical connection. We propose to subdivide the collagen disorders into four main classes depending on whether skin, joints, bone, or blood vessels are mainly involved. The class with mainly skin involvement includes the different forms of cutis laxa, Ehlers-Danlos syndrome types I and II (autosomal dominant), types V and IX (X-linked recessive), type VI (autosomal recessive), and type VIII (autosomal dominant). The group with mainly articular involvement includes Larsen and related syndromes and other types with a more benign course. The conditions with mainly skeletal involvement include the different forms of osteogenesis imperfecta. The class with mainly blood vessel involvement includes disorders of type III collagen and the Marfan syndrome. This tentative classification proposes a logical clinical framework that will allow easier integration of molecular biology data.

Dentine is biochemically abnormal in osteogenesis imperfecta

In osteogenesis imperfecta the bones are brittle but the teeth, whose dentine contains the same genetic collagen as bone (type I), may be clinically normal. To investigate this paradox we have measured the amino acid composition of insoluble dentine collagen from 16 deciduous and 18 permanent teeth in control subjects and in 59 patients with different forms of osteogenesis imperfecta. In 55 of the patient samples significant differences from normal were found, especially in the number of lysine residues, and in the relative amounts of hydroxylysine to lysine. These results demonstrate the high frequency of biochemical abnormalities in osteogenesis imperfecta. They also suggest that classifications of this disorder based on the presence or absence of clinical dentiogenesis imperfecta are likely to be unsound.

Use of R-loop mapping for the assessment of human collagen mutations.

R-loop mapping of DNA:RNA hybrids formed between mutant pro-alpha 2(I) mRNAs and appropriate human pro-alpha 2(I) genomic clones was employed to define the location of mutations which result in the synthesis of shortened pro-alpha 2(I) chains in skin fibroblasts from two variants of osteogenesis imperfecta. Hybridization of the genomic clone NJ-9 with pro-alpha 2(I) mRNA from a patient with a mild atypical form of the disease resulted in the identification of mutant pro-alpha 2(I) mRNA lacking the sequences which correspond to exon 11 of the pro-alpha 2(I) collagen gene. Exon 11, a 54-base pair exon, encodes amino acids 73 to 90 of the alpha 2(I) chain. Also, electron microscopy of R-loop structures formed between the genomic clone NJ-1 and mRNA from a variant with a perinatal lethal form of osteogenesis imperfecta visualized pro-alpha 2(I) mRNAs which did not hybridize to the sequences of exon 28, a 54-base pair exon coding for amino acids 448 to 465 of the alpha 2(I) chain. Moreover, nuclease S1 mapping of the variant's mutant pro-alpha 2(I) mRNA, employing the human pro-alpha 2(I) cDNA clone Hf-15, confirmed the location of the mismatch to the sequences corresponding to exon 28. Although the data do not determine the exact nature of the mutations, they illustrate the use of R-loop mapping as an alternative approach to S1 mapping analysis for the detection and localization of collagen mRNA deletions.

Urinary-free amino acids in osteogenesis imperfecta

Free urinary amino acids were analyzed in patients with osteogenesis imperfecta to determine whether there were any abnormalities that could be used to improve the diagnosis and classification of this syndrome. The results obtained from 15 patients, who had either the type 1, 2, or 3 form of osteogenesis imperfecta, were compared to the values obtained from 115 age-matched controls. Elevated free amino acid levels were not found in any patient with osteogenesis imperfecta.

Vertebral bone mineral content in osteogenesis imperfecta.

Quantitative computed tomography of the lumbar spine was carried out in 28 patients with osteogenesis imperfecta (OI) in order to measure vertebral trabecular bone mineral concentration (BMC). The patients ranged in age from 6-73 years, and included 3 of the 4 major clinical subtypes of the disease. The findings underscore the heterogeneity of osteogenesis imperfecta even among family members with the same disease type. In addition, cross-sectional analysis of Type I OI patients suggests that BMC during young adulthood averages about 70% of normal, and subsequently falls more rapidly than in normal patients. BMC tends to be lower in the more severe forms of OI. Decreased BMC was not found in a few otherwise normal relatives with scoliosis or joint laxity.

Histological investigation of skin biopsies in otosclerosis and osteogenesis imperfecta.

Histological investigation of skin biopsies in four patients with osteogenesis imperfecta, nine patients with otosclerosis and 13 sex- and age-matched normal controls was carried out blindly. The dermal thickness was markedly reduced in osteogenesis imperfecta and slightly reduced in otosclerosis. Minor degenerative changes in the elastic fibres were seen in otosclerosis, while the elastic fibres were found to be more degenerated and more numerous in osteogenesis imperfecta. Our study does not support the hypothesis of otosclerosis being a localized form of osteogenesis imperfecta. The minor changes in the elastic fibres in otosclerosis indicate that further studies of the elastic fibres and of collagen will be necessary to determine whether otosclerosis is a localized disease or part of a general connective tissue disorder.

The Connective Tissue Perspective

The Connective Tissue Perspective

852 BONEMINERAL TURNOVER IN OSTEOGENESIS IMPERFECTA

A young boy with the severe form of osteogenesis imperfecta was treated with NaF for 8 years (1 mg F/kg/d). The frequency of fractures decreased somewhat, but we could detect no gross improvement in bone density. Based on F balance studies and analysis of a bone biopsy, his body F burden was estimated to be about 5 gm at the end of the treatment period (age 14 yr). NaF treatment was abruptly stopped, and during the next 4 1/2 years a number of 24hr urine samples were analyzed for F. After the first few days, during which only 28 mg F were excreted, fluoride excretion declined exponentially: 10% of the body F burden was lost with a half-time of 5 months, 90% with one of 9 years. This latter value is within the range observed by others for the elimination of F and other “bone-seekers” (Sr, Pb, Ra) in normal subjects. Since approximately 99% of body F is located in the mineral phase of bone this element can serve as a tracer for bone mineral. Based on the extended observations on our patient, it would appear that bone mineral turnover is normal in this disease.

Osteogenesis imperfecta: morphological, histochemical and biochemical aspects. Modifications induced by (+)-catechin.

Two patients affected with two different forms of Osteogenesis Imperfecta were examined in order to study collagen and glycosaminoglycans (GAGs) in skin and iliac crest cartilage. A sharp decrease of the galactosamine to glucosamine ratio due to a reduced content of chondroitin sulfate was evidenced in both patients. Moreover the structure of proteoglycans appeared altered, this being more evident in the severe form of the disease. Morphological examination in light and electron microscopy of cartilage of the less severely diseased patient showed that GAGs in the extracellular matrix did not present regular connection with collagen fibers. Chondrocytes, elongated and disorderly scattered, showed large lipidic inclusions and, on histochemical basis, were devoid of UDPG dehydrogenase activity. Treatment with (+)-catechin produced an improvement, in both patients, of the biochemical pattern of collagen and GAGs. Similarly a shift of the cellular activity and of the matrix morphology towards normality was observed in the investigated cartilage of the less severely affected patient.

Disturbance in the regulation of the type of collagen synthesized in a form of osteogenesis imperfecta.

Fibroblasts derived from skin biopsies of a patient with a congenital defect of connective tissue revealed a disturbance in collagen synthesis. The defect was found in the mechanism that controls the type of collagen synthesized. Biochemical data as well as evaluation of immunofluorescent micrographs using collagen type-specific antibodies, suggested that the fibroblasts of this patient synthesized type III collagen, at a time and rate which was not found in fibroblasts from an age-matched healthy individual.