| BackgroundEctodermal dysplasia-skin fragility syndrome?EDSF syndrome,MIM 604536?is a rare single-gene autosomal recessive inheritable disease.This syndrome is characterized by skin fragility,hyperkeratosis of palms and soles,hypotrichosis,hyperhidrosis and nail dystrophy,also including tooth decay and growth retardation et al.EDSF syndrome can involve both male and female,and its symptom usually show at birth,and it can cause sever septicemia and even death.Ectodermal dysplasia-skin fragility syndrome is due to mutations in the PKP1 gene.There are only 21 patients of ectodermal dysplasia-skin fragility syndrome were reported since McGrath et al.first reported this disease in 1997.The PKP1 gene is mapped at 1q32 which consists of 15 exons and its cDNA is 2.7kb.The plakophilin 1,which is coding by PKP1 gene,belongs to the armadillo protein family and is also an important part of desmosome proteins.This protein exists in at least two different isoforms:726 amino acids?aa?,plakophilin 1a;and 747 aa,plakophilin 1b.Mainly related to ectodermal dysplasia-skin fragility syndrome is plakophilin 1a,which is distributed in the epidermal,outer root sheaths,and sweat glandular epithelium.Plakophilin 1b is mainly distributed in the nucleus,which may be related to signal transduction.Ectodermal dysplasia-skin fragility syndrome is one of inheritable desmosome disease.Desmosome proteins are abundant in the heart and epidermis,and autosomal dominant or recessive mutations in genes encoding desmosome proteins may cause skin and/or heart abnormalities.For example,Naxos disease and Carvajal syndrome,which affect both heart and skin.Their skin abnormalities are mainly palmoplantar keratoderma and wool-like hair.This is consistent with some of the symptoms in EDSF syndrome.Mutations in the DSP gene that have been reported in the literature can cause similar clinical manifestations.Therefore,we infer that mutations in the PKP1 gene may cause abnormal expression of proteins which interact with PKP1,leading to the onset of ectodermal dysplasia-skin fragility syndrome.Objectives1.To Summarize of one case of ectodermal dysplasia-skin fragility syndrome patient with medical history,symptoms,signs and examinations;2.The PKP1 gene sequence analysis of the patients and his parents.Review and analyze the mutation sites of PKP1 gene reported in previous literature.3.To investigate the effect of PKP1 gene on the expression of PKP1-related proteins PKP2 and DSP.Methods1.Collecting the medical history,physical examination,laboratory tests results of a 2-month-old Chinese male patient with ectodermal dysplasia-skin fragility syndrome in the dermatology clinic of the First Affiliated Hospital of Zhengzhou University.Taking patient's skin tissue for Pathological biopsy using HE staining,and using anti-Plakophilin 1 antibody?ab183512?for immunofluorescence test,with normal skin tissue as control.Searching relevant literatures of EDSF syndrome cases by Pubmed,China Knowledge Network,and Wanfang Medical Network.2.Genomic DNA samples were collected from patient's and his parents' blood.Detecting the mutations of PKP1 gene by using polymerase chain reaction?PCR?and Sanger sequencing,and reference to Human Gene Mutation Database?http://www.hgmd.cf.ac.uk/ac/index.php?to obtain suspicious mutation sites.3.The target fragment was obtained by PCR amplification using homo PKP1 as a template.The plasmid pIRES2-ZsGreenl and the target fragment homo PKP1?EcoRI/XhoI?were digested with EcoRI and XhoI.SiRNAs were synthesized with the target sequence?NM001005337?by in vitro transcription as interference fragments.The regulation of PKP1 expression was achieved by transfection of overexpression vectors and interference fragments in HaCAT cells.Real-time quantitative PCR was used to detect PKP1 mRNA in each group.Then using Western blot to detect PKP1,PKP2,and DSP protein in each group.Finally,the expression of PKP2 and DSP was detected by immunohistochemistry in the skin tissue of patient and compared with healthy adults.Results1.It was clinically found that the patient had shown skin fragility,sparse and curly hair,nail dystrophy and shedding and mild palmoplantar keratoderma.The parents of the child had no similar clinical manifestations.Pathological biopsy of this patient showed that thicken spinous layer,widening of spaces between adjacent keratinocytes and formation of small crack.Immunofluorescence results showed the absence of immunoreactive PKP1 protein in the skin of patient as opposed to normal membranal staining in the epidermis of a healthy control individual.A total of 21 cases of EDSF syndrome were reported in the literature.They have the clinical features including skin fragility,abnormal hair,palmoplantar keratoderma,nail dystrophy,and hyperhidrosis.2.Gene sequencing revealed two compound heterozygous mutations in PKP1 gene of our patient:c.95-96insAC?p.T33Qfs*14?and c.1492G>T?p.E498X?.c.95-96insAC?p.T33Qfs*14?is a spontaneous mutation site and c.1492G>T?p.E498X?is inherited from his mother.The above two mutation sites have not been reported in the HGMD professional database.3.There were no significant differences in the expression levels of PKP1,PKP2,and DSP in the normal control group and the negative control group in HaCAT cells.The expression of PKP2 and DSP protein which are PKP1-related proteins,was consistent with expression level of PKP1 gene.Immunohistochemistry showed that there was no significant difference in the expression of DSP in the epidermis of patient and normal control,and there was weakly expression of PKP2 in the epidermis of out case while there was no expression of PKP2 in normal control group.Conclusion1.The second Chinese ectodermal dysplasia-skin fragility syndrome patient was diagnosed;2.Two PKP1 gene mutation sites c.95-96insAC,c.1492G>T were found,which expanded the mutation spectrum of PKP1 gene;3.The expression level of PKP1 gene has a co-regulatory effect on the expression of PKP1-related proteins PKP2 and DSP.Mutations in PKP1 gene may cause compensatory expression of PKP2 protein in the epidermis. |
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