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101 SHORT COMMUNICATION Evaluation of Single Nucleotide Variants in Ethiopian Patients with Atopic Dermatitis Samina ASAD 1 , Isabel TAPIA-PÁEZ 1,2 , Andrea Montano MONTES 1 , Carl-Fredrik WAHLGREN 1 , Kassahun D. BILCHA 3 , Magnus NORDENSKJÖLD 2 and Maria BRADLEY 1,2 Dermatology Unit, Department of Medicine Solna, Karolinska Institutet, 2 Center for Molecular Medicine, Department of Molecular Medicine and Surgery, Karolinska Institutet, SE-171 76, Stockholm, Sweden, and 3 Department of Dermatovenereology, Faculty of Medicine, Gondar University, Gondar, Ethiopia. E-mail: [email protected] 1 Accepted Sep 23, 2018; Epub ahead of print Sep 24, 2018 Atopic dermatitis (AD) is an inflammatory skin disorder characterized by pruritus and dryness of the skin in typical locations. The incidence rate of AD varies worldwide. In Europe the prevalence is up to 20% in children, while reports in Ethiopia vary between 1% and 2% in rural areas and 19% in urban areas (1, 2). The major risk gene for AD in Europeans is the filaggrin (FLG) gene (1q21), where a number of null-mutations as well as variations in gene dose are associated with disease susceptibility (2). FLG mutations have also been identified in the Asian population, although these mutations seem to be more population- or family-specific (2). Little is known about AD susceptibility genes in the African population. Our previous studies indicate a low prevalence of FLG null- mutations in Ethiopian cases of AD (1). This is in line with studies performed in the South African AmaXhosa popula- tion, suggesting that, in the African population, genes other than FLG may be involved in the development of AD (3). In searching for risk genes for AD in the Ethiopian po- pulation, we previously performed whole-exome sequen- cing (WES) of 22 Ethiopian patients with AD/ichthy­osis vulgaris in whom several genetic polymorph­isms were detected (1). This study aimed to elucidate whether some of the single-nucleotide variants (SNVs) identified by Taylan et al. are associated with AD susceptibility in our Ethiopian AD cohort, consisting of 184 cases of AD and 186 healthy controls (Table I) including the 22 previously sequenced individuals (1). METHODS All study participants gave written or oral informed consent, which complies with the principles of the Declaration of Helsinki. This study was approved by the Ethics Review Board of the Univer- sity of Gondar, and the regional ethics committee, Karolinska Institutet, Sweden. All patients were diagnosed with AD according to the UK Working Party’s diagnostic criteria (2) and answered questionn- Table I. Ethiopian cases and controls Cases (n  = 184) Female, % Age, median; range Early age of onset (<2 years old), % Moderate AD (SCORAD 15–40), % Severe AD (SCORAD >40), % Controls (n  = 186) 46 51 17; 2 months–45 years 12; 5–60 years 68 n.a. 30 n.a. 70 n.a. AD: atopic dermatitis; SCORAD: SCOring Atopic Dermatitis; n.a.: not applicable. aires regarding other atopic manifestations (1). The samples were obtained from ALERT Dermatology Hospital, Tikur Anbessa Hos- pital and Gondar University Hospital. We chose to genotype gene variants in: (i) epidermal differentiation complex (EDC) genes; cornulin (CRNN) (4) and hornerin (HRNR) (5); (ii) non-EDC genes that had either previously been associated with AD or candidate genes with a known function in the skin; (iii) gene variants that more than one individual was carrying in the WES. Based on this strategy we genotyped 38 SNVs using TaqMan (Applied Bio- systems Inc., Foster City, CA, USA) in the following 10 genes; transglutaminase (TGM) 1, 3 and 5 (6), desmoglein (DSG) 1 and 3–4 (7), a variant in C11orf30 encoding for the EMSY protein (8), SPINK5 (11), hornerin (HRNR) (5) and cornulin (CRNN) (4). Primers/probes were obtained from Thermo-Fisher Scientific Inc., (Foster City, CA, USA). Genotyping indicated that 28 SNVs were non-polymorphic or did not detect minor allele homozygotes in our material (Table SI 1 ). For the remaining 10 SNVs (Table II and Table SII 1 ), association with AD was calculated using χ 2 tests on both genotype and allele frequencies. Fisher’s exact test was performed in instances where the counts were lower than 5. R program v.2.12.2 was used to calculate logistic-regression using generalized linear modelling between patients and controls. SNV frequencies were checked to follow Hardy-Weinberg equilibrium. RESULTS The genotyping success and concordance rates for SNVs showing 3 genotypes was above 90%. Furthermore, all successfully genotyped SNVs (showing all 3 genotypes) correlated with the previous whole exome-sequence results. In our WES study, 12 SNVs were identified in the HRNR gene, which is a component of the epidermal cornified cell envelope thought to share similar features with filaggrin and demonstrated to have reduced expression in the epi- dermis of patients with AD (5). In the HRNR gene, only 3 SNVs demonstrated 3 genotypes and were analysed further. Two SNVs were analysed in CRNN, which is a marker of late epidermal differentiation and down-regulated in AD skin (4). However, these SNVs were non-polymorphic in our study material and discarded from further studies. In addition, SNVs were genotyped in DSG1-4 and TGM1, 3 and 5, all of which have important roles in forming the epidermis and are associated with altered expression in AD skin (6, 7). One SNV located in the c30orf11 region, which has been significantly associated with AD in pre- vious studies, was also analysed (8). In the SPINK5 gene, 7 SNVs were genotyped. SPINK5 (5q32) encodes for the https://www.medicaljournals.se/acta/content/abstract/10.2340/00015555-3051 1 This is an open access article under the CC BY-NC license. www.medicaljournals.se/acta Journal Compilation © 2019 Acta Dermato-Venereologica. doi: 10.2340/00015555-3051 Acta Derm Venereol 2019; 99: 101–102