Screening And Functional Study Of Gene Mutations In Nodular Sclerosis And Pyogenic Sweatadenitis

Part 1 Mutation sreening of TSC pateintsTuberous sclerosis complex (TSC), as a multi-organism involved autosomal dominant neurocutaneous syndrome, characterized by the non-invasive hamartomas, mainly occurred in the brain, skin, heart, lungs, eyes, and kidneys. The Prevalence in adults is 1 per 10,000 and in children is 1 per 6,800.The target criminal gene of TSC are TSC1 (Mendelian Inheritance in Man number 605284) and TSC2 (Mendelian Inheritance in Man number 191092), mutational studies of TSC1 and TSC2 in patients have been drawn greater attention, recently. At least 1480 mutations of TSC1 and 3328 of TSC2 have been reported till April,2013. However, there is no significant mutational hotspot in both genes.TSC1 cods hamartin (TSC1), a 140KD protein; TSC2 cods for tuberin (TSC2), a 200KD protein. Nonhomologous TSC1 and TSC2 form a heterodimer complex (TSC1-TSC2), functional as a GTPase activating protein and participant in mTOR signal pathway regulating cell growth.The therapeutic approaches of TSC is inadequate, a promising rapamycin drugs which inhibits mTOR is still in the stage of clinical trial. Therefore, clinic practice of TSC treatment is mainly focus on accurate diagnosis and periodic review. Mutation screening of criminal gene TSC1 and TSC2 provides evidence of not only pathogenesis diagnosis but also prenatal diagnosis.In the present study, Sanger sequencing and MLPA were used to screen pathogenic mutations of TSC1 and TSC2 in 82 TSC patients. Total of 43 mutation and 4 of large deletion were funded. Then, WES was used to screening DNA of hamartomas tissues from 6 mutation negative patients and 1 case of somatic mutations was examined. For patients both negative in mutation screening and WES,3 highly suspicious mutations in mTOR pathway were located.Part 2Mutation sreening and fuctional analysis of a family with HSHidradenitis suppurativa (HS) is a chronic inflammatory dermatitis, affected areas present with black acne that progressively enlarge to cysts, abscesses, rupture with the release of pus and form sinus. Usually affects areas rich in apocrine glands, such as axillary, breast folds, abdominal folds, groin, buttocks and inner thighs. There are significant variation in involving areas, lesion type and complications. Patients are heavily suffering because pain, chronic suppurative and sustained stench of the disease.In 2006, the pathogenic gene of HS was first located in 80cM area of chromosome 1p21.1-1q25.3. Criminal mutation was target at the gene coding y-secretase. The first mutations were reported on three gene (PSENEN、PSEN1 and NCSTN) from 6 Chinese pedigree, and one of them (NCSTN) located in the linkage analysis targeted area. After that, these mutations were reported successively in 2 English pedigree,5 Chinese pedigree,1 Japanese pedigree,3 French pedigree and 4 sporadic cases.17 mutation of NCSTN,3 of PSENEN and 1 of PSEN1 have been approved so far.3 of them are nonsense mutations,7 frame shifts,4 missence and 7 lead to alternative splicing.Our study focus on a Chinese HS pedigree, and a new mutation was targeted in NCSTN gene. This mutation was located at the junction of intron 4 and exon 5, which might affect splicing. Therefore, we constructed minigene containing mutant DNA fragments, and examined the influence on NCSTN gene splicing by using in vitro splicing analysis. The mutations result in non-cut of intron 4.