Clinical And Skin Image Characteristics Of Verrucous Epidermal Nevus And Investigate It's Molecular Mechanisms By TMT-labeling Proteomics Methods

Background Verrucous epidermal nevus(VEN)is one of the most common skin diseases in the Department of Dermatology.VEN are benign hamartomatous growths of skin that may be present at birth or a few months after birth and rapid progress in early childhood,school-age,puberty and gestation.The clinical manifestations of the VEN are mainly dense papillomatous and hyperkeratosis papules.The VEN lesions color can be normal,red,sepia or black,with clear boundaries.The rash is hard to touch,and the rash at the folds of body is often soft due to sweat impregnation.According to the lesion location,VEN can be divided into three clinical variants,including localized,systematized and inflammatory linear verrucous epidermal naevus(ILVEN).The Epidermal Nevus Syndrome(ENS)was defined as verrucous epidermal nevus compliacted with systemic symptoms.Until now,the clinical characteristics of EVN with a large sample size have not been reported.Although most studies considered that K10,FGFR3 and CJA1 gene mutations may be related to the occurrence of VEN,the specific mechanism is still not fully understood.Protein is the last product of gene expression,as well as a direct manifestation of life activity.In recent years,with the rapid development of proteomics technology,and was widely used to investigate the pathogenesis of complex diseases.Proteomics refers to a full set of proteins expressed by the genome,which includes a cell protein and an expression of all living creatures.Through systematic study on proteomics can better clear the molecular mechanisms of the disease.Object A large size sample of children(cases: 121)with verrucous epidermal nevus were included in this study and to investigate the clinical and skin image characteristics and molecular mechanism by using TMT-labeling proteomics.Methods(1)A total of 121 children with verrucous epidermal nevus were enrolled in this study and the basic clinical information such as gender,age,onset-age,family history,clinical type,distribution of skin lesions,accompanied by pruritus,etc.were also collected by using the self-made questionnaire of verrucous epidermal nevus.(2)Dermatoscopy and confocal laser scanning microscopy was performed on 16 and 12 patients with verrucous epidermal nevus,respectively.All skin image results were collected and analyzed.(3)Nine different clinical types children with verrucous epidermal nevus received surgical treatment(excision and direct suture,excision combined with blade-thick skin graft),and the postoperative prognosis was observed.(4)The TMT-labeling proteomics technology was used to explore the differentially expressed proteins in children with verrucous epidermal nevus.The differentially expressed proteins levels were further validation in independent children with verrucous epidermal nevus lesions.About 8 cases of children with verrucous epidermal nevus and8 cases of children with pigmented nevus were enrolled.Lesions(VEN group)and adjacent tissues(VENC group)from 5 children with verrucous epidermal nevus and naevus-adjacent normal skin tissues(C group)from 5 children with pigmented naevus were obtained after surgical removal.The protein expression levels of lesions,adjacent tissues in children with verrucous epidermal nevus(n=5 cases)and naevus-adjacent normal skin tissues in children with pigmented naevus(n=5 cases)were explored by TMT-labeling proteomics technology.We also compared the protein expression levels differences between VEN and C group,VENC and C group,VEN and VENC group.The significantly differentially expressed proteins between VEN and C group was further analyzed by GO,KEGG and Protein Protein Interaction(PPI)network bioinformatics analyzed methods.We selected some proteins which related to pathogenesis of the verrucous epidermal nevus for verification.Finally,9 proteins were selected and we explored and compared it's expression levels in lesions,adjacent tissues in children with verrucous epidermal nevus(n=3 cases)and naevus-adjacent normal skin tissues in children with pigmented naevus(n=3 cases)by Western blotting methods.Results(1)The onset-age of patients with localized verrucous epidermal nevus is more earlier,and the skin lesions are more common in the head,neck,trunk and limbs,and the itching is not obvious,while the onset-age of patients with inflammatory linear verrucous epidermal nevus is later,and the skin lesions are more common in the limbs,accompanied by significant itching.There were significant differences in onset-age,lesions site and pruritus severity between localized and inflammatory linear verrucous epidermal nevus.(2)The dermatoscope characteristics of localized verrucous epidermal nevus was mainly brown round rings,while the pink areas and thick attached scales were the common features of inflammatory linear verrucous epidermal nevus.The gyri like structures was the common features of system verrucous epidermal nevus.Hyperkeratosis and prolongation of epidermal processes were the most common features of confocal laser scanning microscopy manifestations of verrucous epidermal nevus.For inflammatory linear verrucous epidermal nevus,they were often accompanied by inflammatory cell infiltration.(3)Compared with normal skin tissue,399 proteins were significantly up-regulated and 187 proteins were significantly down-regulated in the skin lesions of the epidermal nevus verrucous,while 38 proteins were significantly up-regulated and 43 proteins were significantly down-regulated in the non-skin lesions of the epidermal nevus verrucous.Compared with non-skin lesions,204 proteins were significantly up-regulated and 117 proteins were significantly down-regulated in skin lesions of the epidermal nevus verrucous.(4)The expression of Involucrin,NDUFA4,Loricrin,Filaggrin,S100A7,Desmocolin-3,BRAF,Keratin,and Type II Cytoskeletal 6A protein in skin lesions of patients with verrucous epidermal nevus were significantly increased,which involved pathophysiological mechanisms may be related to skin barrier dysfunction,excessive proliferation of epidermal cells,immune inflammation,oxidative phosphorylation,etc.Conclusions This study found there were significant differences in onset-age,lesions site,pruritus severity between patients with localized and inflammatory linear verrucous epidermal nevus.Brown circles,pinked areas,thick attached scales,and gyri structures are the main manifestations of dermoscopic of verrucous epidermal nevi.Hyperkeratosis,prolongation of epidermal processes and infiltration of inflammatory cells are the main manifestations of confocal laser scanning microscopy of verrucous epidermal nevi.Proteomic techniques can explored the pathogenesis of unknown and complex diseases.In this study,some pathogenic proteins associated with verrucous epidermal nevi were found,providing a theoretical basis for further functional studies.