| 1. Background and ObjectiveKeloid is one of challenging problems in our modern medicine. In order to the make clear the keloid pathogenesis, many studies have been carried out from aspects such as histo-pathology, immunology and genetics, collagen syntheses and dissolution imbalance, the cell growth factor, the fibroblast cell defective apoptosis, the immune response and the genetics were proved to be involved in it, but up till now, the accurate keloid pathogenesis was not clear yet.The keloid patient mostly assumes the sending out morbidity, but has certain family heredity tendency. Cosman and so on confirmed its familial form morbidity rate is approximately 3%, and the deep color skin race has higher occurrence. The keloid heredity pattern has not been clear yet. Some scholars thought it may belong to an autosomal dominant inheritance pattern. Also some scholars thought it belongs to an autosomal recessive inheritance pattern. In 2001 year Harvard University Marneros and so on carried out the genetics investigation and study on 14 keloid pedigrees, and its heredity pattern confirms to be an autosomal dominant inheritance mode with incomplete clinical penetrance and variable expression. In 2004, Marneros and so on having conducted a genome-wide STR scan and linkage analysis to locate locus for genes predisposing to keloid formation in a Japanese and an African-American keloid family with microsatellite markers, Marneros identified linkage to chromosome 2q23 for the Japanese family and linkage to chromosome 7pl 1 for the African-American family.The research explored the keloid predisposing gene for the keloid genetics evidence for the first time. It have the vital significance to locate the keloid predisposing gene in future.Because the keloid morbidity existence remarkable race difference, up till now the Chinese keloid pedigree predisposing gene position is not clear yet, whether is consistent to Marneros's research discovery. Therefore, our study group collected two Chinese Han Nationality keloid big pedigrees to carry on the STR scan and linkage analysis the research.This two pedigrees include many members. They can well fit the research hereditary disease to the pedigree request. They are similar to the Marneros's two pedigrees, also has the very good representation. The two pedigrees lay the good foundation for further the predisposing gene the localization research.Our research in the article on Marneros research foundations, through micro satellite scanning and linkage analysis hope to identify the two Chinese keloid pedigrees whether the predisposing genes linkage relation with chromosome 2p23 are similar or consistent to what Marneros had detected in those keloid families.Our previous study selects the micro satellite marks position spot in chromosome 7pl 1 and its periphery . Through the micro satellite scan and linkage analysis , the results excluded linkage to chromosome 7p11 of A Chinese keloid pedigree by 4 microsatellite markers on around chromosome 7p11 with the known maximal two-point LOD scores. Because the keloid has unique growth characteristic and clinical manifestation, therefore it maybe regarded as one kind of the benign tumor after wounding. Now, fibroblast has been proved to be the functional cell which contributes to the keloid formation. The unbalance of proliferation and apoptosis in fibroblast is the cytological substructure that leads keloid to unceasing accrementition and makes it hard to regress.The Fas gene belongs to NGF/ TNF acceptor family gene, its corresponding ligand and the clone antibody (McabFas) may induce the cell apoptosis fast. Apoptosis mediated by Fas gene has been considered to play an important role in fibroblast apoptosis. Domestic and foreign researches indicate that Keloid Fas signal conduction exists barrier or suppression.In order to be clear about whether the Fas acceptor gene exists changes in the keloid pedigree samples, the research to Fas gene exon 1-9 has been carried on the gene sequence analysis.2. Objects and Methods1. To investigate the genetic susceptibility to keloid , the authors performed STR scan and linkage analysis on chromosome 2q23. Two four-generation Chinese keloid pedigrees were investigated and confirmed. Venous blood samples of 64 members in the two keloid pedigrees were collected and extracted genomic DNA. 11 markers on chromosome 2q23 were selected as microsatellite markers according to foreign literature on the similar study recently . Subsequently, these markers were amplified by PCR, and all PCR products were genotyped and the linkage analyses were performed.2. The experimental samples from A and B keloid pedigrees were selected from Department of Plastic Surgery, Nanfang Hospital, Southern Medical University in 2005. The polymerase chain reaction (PCR) and DNA sequencing analysis technique were used in Shanghai Jikang Company between January and May 2005, A pedigree of two cases respectively keloid tissues for the study, their peripheral vein blood and their surrounding normal skin as their own contrast, their spouses' peripheral vein blood as normal contrast, the two cases of patient's peripheral vein blood in B pedigree as a contrast between different keloid pedigree. Gene orders of Fas gene (exon 1 -9) from all of 10 samples were tested.3. Results1. Among the 11 selected markers, the two point LOD scores were less than 1 atθ= 0～0.5,they were not hinted the linkage to the disease locus.2. Gene mutations in Fas gene exon 1-8 were not identified in all samples A and B Chinese keloid pedigrees.3. Point mutations in Fas gene exon 9 were identified in 11bp,53bp in two keloid tissue samples from A Chinese keloid pedigrees.4. Conclusion1. After linkage analyses,this study provides the genetic evidence that keloid susceptibility loci in the Chinese keloid pedigrees were not on chromosome 2q23, demonstrating locus heterogeneity in familial keloid formation.2. The study results of Fas gene exon 9 point mutations in the two Samples from A and B Chinese keloid pedigrees may indicate some relations in Fas gene point mutations and keliod formation.
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