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Wang Y.M.,Peking University | Wang Y.M.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | Huang Y.S.,University of British Columbia | Ma Z.H.,Heilongjiang University | And 6 more authors.
Clinical and Experimental Dermatology | Year: 2014

Background. Basal cell carcinoma (BCC) is a prevalent form of nonmelanoma skin cancer. Although numerous studies in white populations suggest that mutations in the TP53 gene play an important role in the development of BCC, it is not clear whether this is also the case in East Asian populations such as in China. Aim. To investigate the frequency and the features of TP53 mutation in sporadic BCC in a Chinese population. Methods. In total, 30 patients with sporadic BCC, who had previously taken part in a study on PTCH1 mutations, were enrolled. BCC and control cells were obtained by laser-capture microdissection, and DNA was amplified and sequenced for analysis of TP53 mutations. Results. In the 30 BCC samples, 6 TP53 point mutations were found (frequency of 20%), and 4 of these 6 mutations had ultraviolet (UV)-specific alterations. Combining these results with those of the previous study on PTCH1 mutations, we found that two patients with had three types of genetic alterations (each had two PTCH1 mutations and one TP53 point mutation). A further two patients each had one PTCH1 mutation and one UV signature TP53 mutation. In addition, the total number of UV-specific mutations of PTCH1 and TP53 accounted for 20% of the total patient group. Conclusions. The incidence of TP53 mutation in BCC in our Chinese subjects was lower than that reported for white populations. Many of the patients carried mutations of other genes in addition to of TP53. The majority of TP53 mutations were UV-induced specific alterations. However, the results of the two studies on TP53 and PTCH1 indicated that the incidence of UV-specific mutations is much lower in Chinese than in white populations. © 2014 British Association of Dermatologists.

Lee M.,Peking University | Lee M.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | Xu G.,Peking University | Xu G.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | And 13 more authors.
Clinical and Experimental Dermatology | Year: 2016

Background Recessive dystrophic epidermolysis bullosa (RDEB) is a rare heritable blistering skin condition caused by loss-of-function mutations in the COL7A1 gene. Incongruent gene transmission is occasionally reported in recessive diseases, and its underlying mechanism is often uniparental disomy (UPD). Aim To understand the genetic basis of incongruent gene transmission in a Chinese family with RDEB, in which a discrepancy of COL7A1 genotyping was encountered during our mutation analysis. Methods We used a pCAS2 minigene-based in vitro splicing assay to confirm the pathogenicity of the splicing variant we identified in the proband. Next, a combination of genetic tools, including whole-genome SNP array analysis and multiplex ligation-dependent probe amplification copy number analysis, was used to unravel the cause of the discrepancy in the COL7A1 genotyping. Results Sanger sequencing identified a novel, single-peak mutation, c.4980+5G>C, in COL7A1 in the proband, which was heterozygous in his father and wild type in his mother. In vitro splicing assay showed that c.4980+5G>C was pathogenic and led to skipping of COL7A1 exon 53. SNP array analysis and multiplex ligation-dependent probe amplification of the proband's DNA revealed a maternally derived, de novo, interstitial deletion on chromosome 3p21.31, which removed COL7A1 and 15 flanking genes, excluding the possibility of UPD. Conclusion Our findings favour an exceptionally rare event, namely a de novo COL7A1 microdeletion in concurrence with an inherited mutation in trans. This study should aid molecular diagnosis and genetic counselling of RDEB and possibly other recessive diseases in which genotyping discrepancy is encountered. © 2016 British Association of Dermatologists.

Lin Z.,Peking University | Lin Z.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | Zhao J.,Peking University | Zhao J.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | And 28 more authors.
American Journal of Human Genetics | Year: 2015

Calpastatin is an endogenous specific inhibitor of calpain, a calcium-dependent cysteine protease. Here we show that loss-of-function mutations in calpastatin (CAST) are the genetic causes of an autosomal-recessive condition characterized by generalized peeling skin, leukonychia, acral punctate keratoses, cheilitis, and knuckle pads, which we propose to be given the acronym PLACK syndrome. In affected individuals with PLACK syndrome from three families of different ethnicities, we identified homozygous mutations (c.607dup, c.424A>T, and c.1750delG) in CAST, all of which were predicted to encode truncated proteins (p.Ile203Asnfs∗8, p.Lys142∗, and p.Val584Trpfs∗37). Immunohistochemistry shows that staining of calpastatin is reduced in skin from affected individuals. Transmission electron microscopy revealed widening of intercellular spaces with chromatin condensation and margination in the upper stratum spinosum in lesional skin, suggesting impaired intercellular adhesion as well as keratinocyte apoptosis. A significant increase of apoptotic keratinocytes was also observed in TUNEL assays. In vitro studies utilizing siRNA-mediated CAST knockdown revealed a role for calpastatin in keratinocyte adhesion. In summary, we describe PLACK syndrome, as a clinical entity of defective epidermal adhesion, caused by loss-of-function mutations in CAST. © 2015 The American Society of Human Genetics.

Wang H.J.,Peking University | Wang H.J.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | Lin Z.M.,Peking University | Lin Z.M.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | And 6 more authors.
Clinical and Experimental Dermatology | Year: 2014

Diffuse cutaneous mastocytosis (DCM) is an extremely rare disease characterized by massive proliferation of mast cells infiltrating the entire skin. We report a Chinese family with indolent DCM, and detection of a new germline KIT mutation located in the fifth immunoglobulin-like loop of the KIT protein, which probably results in a gain-of-function effect and consequent overactivation of mast cells. Our report expands the knowledge of correlations between the genotype of KIT mutations and the phenotype of DCM. © 2013 British Association of Dermatologists.

Gong J.,Peking University | Gong J.,Beijing Key Laboratory of Molecular Diagnosis on Dermatoses | Ran M.,Peking University | Wang X.,Peking University | And 5 more authors.
Mycopathologia | Year: 2016

An accurate diagnosis of tinea unguium is necessary for the selection of antimycotics and successful treatment. To rapidly and accurately identify the aetiological agents causing tinea unguium, we improved upon the conventional boiling method for DNA extraction and developed a novel real-time PCR detection system that includes two assays. The two assays, based on the amplification of ribosomal internal transcribed spacer regions and 28S rDNA, were designed to detect pan-dermatophyte and Trichophyton rubrum, respectively. The analytical sensitivities of both assays permitted the detection of ten copies of plasmid DNA template. The analytical specificity of the detection system was confirmed using 11 dermatophyte strains and 25 non-dermatophyte strains. In total, 165 nail specimens were examined by microscopy, culture, conventional PCR, and the novel real-time PCR method. Real-time PCR gave positive results in 47.3 % of the specimens (78), a rate exceeding those obtained using microscopy (72, 43.6 %), conventional PCR (69, 41.8 %), and culture (49, 29.7 %). All conventional PCR-positive specimens were detected by real-time PCR, and real-time PCR detected nine specimens that were missed by conventional PCR. The results from latent class analysis, and further calculations, showed that real-time PCR was the most sensitive method, but the diagnostic specificity of the four approaches was equivalent. In particular, molecular approaches may be more effective than microscopy and culture when the clinical symptoms of tinea unguium are not evident. © 2015, Springer Science+Business Media Dordrecht.

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