Clinical And Genetic Diagnosis Of Three Hereditary Bone Diseases

Hereditary bone disease, is a kind of skeletal system diseases,which characterized by clinical and genetic heterogeneity. Most of them are rare and low incidence. The common clinical manifestations include short statures,deformities,disproportionate growth,and a single or a group of skeletal dysplasia. According to clinical features, radiographs and genetic diagnosis and other characteristics, "The Classification Standard of International Genetic Bone Disease (2015)"[1] divided hereditary bone disease into 42 groups, 436 hereditary osteodystrophy were collected in and 361 genes were confirmed associated with these diseases. There are some genes related to cartilage dysplasia gene, such as cartilage oligomeric matrix protein gene (COMP) associated with calcium ion binding, type Ⅰ collagen gene (COL1A1,COL1A2), type II collagen gene (COL2A1) and so on.The study was carried on molecular genetics research in three pedigrees with Legg-Calve-Perthes disease (LCPD), osteogenesis imperfect (OI),pseudoachondroplastic dysplasia (PSACH) and multiple epiphyseal dysplasia (MED).Pedigree 1: A LCPD family was collected. Up to date, several literatures have been identified and only one pathogenic gene (COL2A1) has been found.Methods: We extract blood samples from the proband and his parents.Next-generation sequencing was performed and seleceted the target gene and corresponding mutation site. Primers were designed for polymerase chain reaction (PCR)and bioinformatics was used to analyse the mutation.Results: There was a heterozygous mutation (c.3392G>T, P.Gly1131Val) in COL2A1 gene. We found that the proband’s parents and the control group did not carry this mutation, which suggest that the mutation was new and de novo. The Gly at position 1131 in the triple helical domain of type II collagen, was highly conserved.Conclusion: We have discovered a novel substitution mutation (c.3392G>T,p.Gly1131Val), which also as a de novo mutation in his family. This mutation not only extends the mutation spectrum associated with collagenopathies, but also may be helpful in early molecular diagnosis and prenatal diagnosis.Pedigree 2: An OI family was collected. There were 15 subtypes and this family was type I . Pathogenic gene of OI type Ⅰ is COL1A1 .Methods: We extract blood. samples from the proband. Next-generation sequencing was performed to seleceted the corresponding mutation site. Primers were designed for polymerase chain reaction (PCR) and sequencing in all patients and non-patients in the family.Results: There was a splicing mutation (c.471+1G>A, IVS5+1G>A) in COL1A1 gene. The mutation was completely cosegregated in the large family and it had been reported in 2005. However, it was rare that the patients in the family had different severity.Conclusion: Our study revealed that a recurrent mutation c.471+1G>A in COL1A1 played a pathogenic role in a large Chinese family and we identified different phenotypes of this variant. The identification of the mutation not only provides genetic counseling and prenatal diagnosis for other members in the family,but also extends the evidence for genetic and phenotype heterogeneity in OI.Pedigree 3: A PSACH or MED family was collected.These two conditions are part of the same bone dysplasia family. The common pathogenic gene was COMP.Methods: We extract blood samples from the proband and his parents.Whole-exone sequencing (WES) was performed in all patients to screen the mutation.According to the genotype-phenotype analysis, some pathogenic sites were screened and primers were designed for polymerase chain reaction (PCR) and bioinformatics was used to analyse the mutation.Results: We observed a c.1201G>A (p.Asp401Asn) heterozygous mutation in exon 11 in all patient of the family. The mutation was not found in controls and other unaffected members in the family and it had been reported in 2009 in a MED patient.The asp at position 401 was highly conserved.Conclusion: We have discovered a missense mutation (c.1201G>A,p.Asp401Asn) in COMP gene, which produces two similar diseases. This mutation extends the genotype-phenotype spectrum associated with chondrodysplasia and may be helpful in early molecular diagnosis and prenatal diagnosis.Our study described the clinical and molecular genetics of each family. Provide evidence for genetic counseling,clear diagnosis,and prenatal diagnosis in these pedigrees and extend the genotype-phenotype spectrum.