The Gene Analysis Of A Family With Hereditary Hemorrhagic Telangiectasia

Objective: The aim of this study is to investigate the clinical features and gene mutations of a Chinese hereditary hemorrhagic telangiectasia family,intends to explore the possibility gene mutations of this family and in order to propose more references to the molecular mechanisms of the disease.Method: Investigate the clinical data and create the genetic mapping of the family.Whole genomic DNA was extracted from peripheral blood of the family members,and polymerase chain reaction(PCR) amplification and sequencing were performed to detect mutation in gene ENG,ACVRL1 and SMAD4,which had been reported to cause HHT in the documents.The results showed that the pathogenic genes of this family is not the three known pathogenic genes, which suggested that there may be another novel pathogenic gene related to HHT.By using the method of whole genome sequencing, we found 13 suspected candidate genes : NOL10, GRK4, STX18, ABLIM2, NCAPG, SLC12A7, FAM105 A, CARD6, FAT2, FABP6, AMBP, CNTRL and CRLS1 gene.According to the function of each gene, we select the 6 genes most possibly associated with HHT pathogenic to verify.These 6 genes include: AMBP, FABP6, FAT2, NOL10, SLC12A7, and STX18 gene.The exon 9 of AMBP gene,exon 4 of FABP6 gene,exon 9 of FAT2 gene,exon 1 of NOL10 gene,exon 7 of SLC12A7 gene and exon 1 of STX18 gene were amplified by the method of polymerase chain reaction(PCR), the PCR products were electrophoresised through agarose gel,and then were sequenced to verify the results of whole genome sequencing.Results: In the investigated 26 members of the family including a total of 5 generations,we found 4 members with the Significant clinical features of HHT,including two men and two women,in each generation individual incidence.This pedigree is in line with the autosomal dominant genetic disease characteristics.In the sequencing analysis of the ENG, ACVRL1 and SMAD4 gene,we have not found the DNA mutation and phenotype co-segregation in this family,suggesting that the three known genes are not the pathogenic gene for this family.The whole genome sequencing analysis of the 4 patients and 3 normal phenotype members of the family indicate that:(1)A missense mutation c.854C>T was found in exon 9 of AMBP gene in the proband and other patients of the family,which resulted in Alanine 285 to Valine replacement(P.Ala 285 Val),but the similar mutation were not detected in 3 normal phenotype members of the family.(2)A missense mutation c.173T>A was found in exon 4 of FABP6 gene in the proband and other patients of the family,which resulted in Methionine 58 to Lysine replacement(P.Met 58 Lys), but the similar mutation were not detected in 3 normal phenotype members of the family.(3)A missense mutation c.5962G>A was found in exon 9 of FAT2 gene in the proband and other patients of the family,which resulted in Valine 1988 to Methionine replacement(P.Val 1988 Met), but the similar mutation were not detected in 3 normal phenotype members of the family.(4)A missense mutation c.6G>C was found in exon 1 of NOL10 gene in the proband and other patients of the family,which resulted in Glutamine 2 to Histidine replacement(P.Gln 2 His), but the similar mutation were not detected in 3 normal phenotype members of the family.(5)A missense mutation c.908C>T was found in exon 7 of SLC12A7 gene in the proband and other patients of the family,which resulted in Proline 303 to Leucine replacement(P.Pro 303 Leu), but the similar mutation were not detected in 3 normal phenotype members of the family.(6)A missense mutation c.161G>A was found in exon 1 of STX18 gene in the proband and other patients of the family,which resulted in Arginine 54 to Histidine replacement(P.Arg 54 His), while the same mutation were detected in the son of the proband,even though his son shows the normal phenotype,but the similar mutation were not detected in the remaining 2 normal phenotype members of the family.Conclusions:This study excluded the known pathogenic genes of HHT,and verified that the pathogenicity gene of this family is likely to be one of AMBP, FABP6, FAT2, NOL10, SLC12A7 and STX18 gene,but can not excluded that one of the other 7 genes,which have not been verified in this study,is the pathogenic gene of this family.So it could be inferred that there may exist a new pathogenic gene which related to the HHT,suggesting that HHT has a genetic heterogeneity.However, because of the sample size of this family is too small,it still can not identify the pathogenic gene of the family.If there are other pedigrees of HHT,after the exclusion of the known pathogenic genes,maybe can detect the 13 or 6 suspected genes directly.The results of this study enriches the gene mutation database of HHT,also provides relevant information and evidence for the genetic screening and genetic counseling for the other members of the family with HHT.