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Masashi Akiyama - One of the best experts on this subject based on the ideXlab platform.
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apparent homozygosity due to compound heterozygosity of one point mutation and an overlapping exon deletion mutation in ABCA12 a genetic diagnostic pitfall
Journal of Dermatological Science, 2015Co-Authors: Akitaka Shibata, Kazumitsu Sugiura, Atsushi Suzuki, Takashi Ichiki, Masashi AkiyamaAbstract:Abstract Background Harlequin ichthyosis (HI), one of the most severe genetic skin disorders, is autosomal recessively inherited. Mutations in ABCA12 , which encodes ATP-binding cassette transporter A12 (ABCA12), are known to be the cause of HI. It is very difficult to make precise genetic diagnosis when an exon deletion mutation overlaps the site of another causative point mutation. This combination of mutations may lead us to conclude incorrectly that the patient has the point mutation homozygously, a phenomenon called "apparent homozygosity". Objective To demonstrate that the present HI patient has apparent homozygosity in ABCA12 mutations. Methods We performed direct sequencing of gDNA in the entire coding region, including exon–intron boundaries, of ABCA12 in the HI patient and her parents. To further elucidate the mutations in the patient, parental mutation segregation study was done and SNP analysis was conducted on the region flanking ABCA12 in the patients and her parents. Quantitative PCR of gDNA in exon 11 of ABCA12 was also performed. Direct sequencing of cDNA from exon 9 to exon 13 and of gDNA between intron 9 and intron 11 of ABCA12 was done in the HI patient and her parents. Results Direct sequencing of gDNA in the entire coding region, including exon–intron boundaries, of ABCA12 seemed to indicate that the patient had the novel homozygous nonsense mutation c.1216A>T (p.Lys406X) in exon 11. However, mutation segregation analysis, SNP analysis, qRTPCR of gDNA in exon 11 of ABCA12 and direct sequencing of cDNA from exon 9 to exon 12 of ABCA12 and of gDNA between intron 9 and intron 11 of ABCA12 in the HI patient and her parents demonstrated that the present patient was compound heterozygous for two ABCA12 mutations: c.1216A>T (p.Lys406X) in exon 11 and g.111346_113217del1872 (p.Leu355_Lys428del, Gln354fs7*) which was overlapping exon deletion mutations involving exons 10 and 11. Conclusion When direct sequencing indicates that a patient from a non-consanguineous family has an apparently homozygous non-founder point mutation, the homozygosity may be "apparent homozygosity", and we should keep in mind the possibility of overlapping exon deletion mutation.
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a palindromic motif in the 2084 to 2078 upstream region is essential for ABCA12 promoter function in cultured human keratinocytes
Scientific Reports, 2015Co-Authors: Yoshitaka Shimizu, Hiroshi Shimizu, Teruki Yanagi, Daisuke Sawamura, Kazumitsu Sugiura, Yasushi Ogawa, Junichi Takeda, Kaori Sakaisawada, Atsushi Kon, Masashi AkiyamaAbstract:ATP-binding cassette transporter family A member 12 (ABCA12) is a keratinocyte transmembrane lipid transporter that plays a critical role in preserving the skin permeability barrier. Biallelic loss of function of the ABCA12 gene is causative of some forms of recessive congenital ichthyosis, an intractable disease marked by dry, thickened and scaly skin on the whole body. Genetic diagnosis is essential, although the results may occasionally be inconclusive, because some patients with low ABCA12 expression have one mutant allele and one apparently intact allele. Aside from aberrant splicing or deletion mutations, one possible explanation for such discrepancy is loss of promoter function. This study aims to elucidate the promoter region of ABCA12 and to locate the essential elements therein, thus providing the necessary information for genetic diagnostic screening of congenital ichthyosis. Close examination of the 2980-bp upstream regions of the ABCA12 gene revealed that a palindromic motif (tgagtca) at -2084 to -2078 is essential for the promoter function, and a short fragment of -2200/-1934 alone has potent promoter activity. Identification of the key promoter element of ABCA12 in this study may provide relevant information for genetic diagnosis of recessive congenital ichthyosis.
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The roles of ABCA12 in epidermal lipid barrier formation and keratinocyte differentiation.
Biochimica et biophysica acta, 2013Co-Authors: Masashi AkiyamaAbstract:ATP-binding cassette (ABC) transporters form a large superfamily of transporters that bind and hydrolyze ATP to transport various molecules across limiting membranes or into vesicles. The ABCA subfamily members are thought to transport lipid materials. ABCA12 is a keratinocyte transmembrane lipid transporter protein associated with the transport of lipids via lamellar granules. ABCA12 is considered to transport lipids including ceramides to form extracellular lipid layers in the stratum corneum of the epidermis, which is essential for skin barrier function. ABCA12 mutations are known to underlie the three major types of autosomal recessive congenital ichthyoses: harlequin ichthyosis, lamellar ichthyosis and congenital ichthyosiform erythroderma. ABCA12 mutations result in defective lipid transport via lamellar granules in the keratinocytes, leading to ichthyosis phenotypes from malformation of the stratum corneum lipid barrier. Studies on ABCA12-deficient bioengineered models have revealed that lipid transport by ABCA12 is required for keratinocyte differentiation and epidermal morphogenesis. Defective lipid transport due to loss of ABCA12 function leads to the accumulation of intracellular lipids, including glucosylceramides and gangliosides, in the epidermal keratinocytes. The accumulation of gangliosides seems to result in the apoptosis of ABCA12(-/-) keratinocytes. It was reported that AKT activation occurs in ABCA12(-/-) granular-layer keratinocytes, which suggests that AKT activation serves to prevent the cell death of ABCA12(-/-) keratinocytes. This article is part of a Special Issue entitled The Important Role of Lipids in the Epidermis and their Role in the Formation and Maintenance of the Cutaneous Barrier. Guest Editors: Kenneth R. Feingold and Peter Elias.
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The roles of ABCA12 in keratinocyte differentiation and lipid barrier formation in the epidermis
Dermato-endocrinology, 2011Co-Authors: Masashi AkiyamaAbstract:ABCA12 is a member of the large superfamily of ATP-binding cassette (ABC) transporters, which bind and hydrolyze ATP to transport various molecules across limiting membranes or into vesicles. The ABCA subfamily members are thought to be lipid transporters. ABCA12 is a keratinocyte transmembrane lipid transporter protein associated with the transport of lipids in lamellar granules to the apical surface of granular layer keratinocytes. Extracellular lipids, including ceramide, are thought to be essential for skin barrier function. ABCA12 mutations are known to underlie the three main types of autosomal recessive congenital ichthyoses: harlequin ichthyosis, lamellar ichthyosis and congenital ichthyosiform erythroderma. ABCA12 mutations lead to defective lipid transport via lamellar granules in the keratinocytes, resulting in malformation of the epidermal lipid barrier and ichthyosis phenotypes. Studies of ABCA12-deficient model mice indicate that lipid transport by ABCA12 is also indispensable for intact differentiation of keratinocytes.
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ABCA12 mutations and autosomal recessive congenital ichthyosis a review of genotype phenotype correlations and of pathogenetic conceptsa
Human Mutation, 2010Co-Authors: Masashi AkiyamaAbstract:Mutations in ABCA12 have been described in autosomal recessive congenital ichthyoses (ARCI) including harlequin ichthyosis (HI), congenital ichthyosiform erythroderma (CIE), and lamellar ichthyosis (LI). HI shows the most severe phenotype. CIE and LI are clinically characterized by fine, whitish scales on a background of erythematous skin, and large, thick, dark scales over the entire body without serious background erythroderma, respectively. To date, a total of 56 ABCA12 mutations have been reported in 66 ARCI families including 48 HI, 10 LI, and 8 CIE families of African, European, Pakistani/Indian, and Japanese origin (online database: http://www.derm-hokudai.jp/ABCA12/). A total of 62.5% of reported ABCA12 mutations are expected to lead to truncated proteins. Most mutations in HI are truncation mutations and homozygous or compound heterozygous truncation mutations always results in HI phenotype. In CIE families, at least one mutation on each allele is typically a missense mutation. Combinations of missense mutations in the first ATP-binding cassette of ABCA12 underlie the LI phenotype. ABCA12 is a keratinocyte lipid transporter associated with lipid transport in lamellar granules, and loss of ABCA12 function leads to a defective lipid barrier in the stratum corneum, resulting in an ichthyotic phenotype. Recent work using mouse models confirmed ABCA12 roles in skin barrier formation. Hum Mutat 31:1–7 2010. © 2010 Wiley-Liss, Inc.
Hiroshi Shimizu - One of the best experts on this subject based on the ideXlab platform.
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a palindromic motif in the 2084 to 2078 upstream region is essential for ABCA12 promoter function in cultured human keratinocytes
Scientific Reports, 2015Co-Authors: Yoshitaka Shimizu, Hiroshi Shimizu, Teruki Yanagi, Daisuke Sawamura, Kazumitsu Sugiura, Yasushi Ogawa, Junichi Takeda, Kaori Sakaisawada, Atsushi Kon, Masashi AkiyamaAbstract:ATP-binding cassette transporter family A member 12 (ABCA12) is a keratinocyte transmembrane lipid transporter that plays a critical role in preserving the skin permeability barrier. Biallelic loss of function of the ABCA12 gene is causative of some forms of recessive congenital ichthyosis, an intractable disease marked by dry, thickened and scaly skin on the whole body. Genetic diagnosis is essential, although the results may occasionally be inconclusive, because some patients with low ABCA12 expression have one mutant allele and one apparently intact allele. Aside from aberrant splicing or deletion mutations, one possible explanation for such discrepancy is loss of promoter function. This study aims to elucidate the promoter region of ABCA12 and to locate the essential elements therein, thus providing the necessary information for genetic diagnostic screening of congenital ichthyosis. Close examination of the 2980-bp upstream regions of the ABCA12 gene revealed that a palindromic motif (tgagtca) at -2084 to -2078 is essential for the promoter function, and a short fragment of -2200/-1934 alone has potent promoter activity. Identification of the key promoter element of ABCA12 in this study may provide relevant information for genetic diagnosis of recessive congenital ichthyosis.
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self improvement of keratinocyte differentiation defects during skin maturation in ABCA12 deficient harlequin ichthyosis model mice
American Journal of Pathology, 2010Co-Authors: Teruki Yanagi, Masashi Akiyama, Kaori Sakai, Hiroshi Nishihara, Shinya Tanaka, Junko Ishikawa, Yuki Miyamura, Ayano Naoe, Takashi Kitahara, Hiroshi ShimizuAbstract:Harlequin ichthyosis (HI) is caused by loss-of-function mutations in the keratinocyte lipid transporter ABCA12. The patients often die in the first 1 or 2 weeks of life, although HI survivors’ phenotypes improve within several weeks after birth. In order to clarify the mechanisms of phenotypic recovery, we studied grafted skin and keratinocytes from ABCA12-disrupted (ABCA12−/−) mice showing abnormal lipid transport. ABCA12−/− neonatal epidermis showed significantly reduced total ceramide amounts and aberrant ceramide composition. Immunofluorescence and immunoblotting of ABCA12−/− neonatal epidermis revealed defective profilaggrin/filaggrin conversion and reduced protein expression of the differentiation-specific molecules, loricrin, kallikrein 5, and transglutaminase 1, although their mRNA expression was up-regulated. In contrast, ABCA12−/− skin grafts kept in a dry environment exhibited dramatic improvements in all these abnormalities. Increased transepidermal water loss, a parameter representing barrier defect, was remarkably decreased in grafted ABCA12−/− skin. Ten-passage sub-cultured ABCA12−/− keratinocytes showed restoration of intact ceramide distribution, differentiation-specific protein expression and profilaggrin/filaggrin conversion, which were defective in primary-cultures. Using cDNA microarray analysis, lipid transporters including four ATP-binding cassette transporters were up-regulated after sub-culture of ABCA12−/− keratinocytes compared with primary-culture. These results indicate that disrupted keratinocyte differentiation during the fetal development is involved in the pathomechanism of HI and, during maturation, ABCA12−/− epidermal keratinocytes regain normal differentiation processes. This restoration may account for the skin phenotype improvement observed in HI survivors.
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harlequin ichthyosis model mouse reveals alveolar collapse and severe fetal skin barrier defects
Human Molecular Genetics, 2008Co-Authors: Teruki Yanagi, Masashi Akiyama, Kaori Sakai, Hiroshi Nishihara, Wataru Nishie, Shinya Tanaka, Hiroshi ShimizuAbstract:: Harlequin ichthyosis (HI), which is the most severe genodermatosis, is caused by loss-of-function mutations in ABCA12, a member of the ATP-binding cassette transporter family. To investigate the pathomechanism of HI and the function of the ABCA12 protein, we generated ABCA12-deficient mice (ABCA12(-/-)) by targeting ABCA12. ABCA12(-/-) mice closely reproduce the human HI phenotype, showing marked hyperkeratosis with eclabium and skin fissure. Lamellar granule abnormalities and defective ceramide distribution were remarkable in the epidermis. Skin permeability assay of ABCA12(-/-) fetuses revealed severe skin barrier dysfunction after the initiation of keratinization. Surprisingly, the ABCA12(-/-) mice also demonstrated lung alveolar collapse immediately after birth. Lamellar bodies in alveolar type II cells of the ABCA12(-/-) mice lacked normal lamellar structures. The level of surfactant protein B, an essential component of alveolar surfactant, was reduced in the ABCA12(-/-) mice. Fetal therapeutic trials with systemic administration of retinoid or dexamethasone, which are effective for HI and respiratory distress, respectively, to the pregnant mother mice neither improved the skin phenotype nor extended the survival period. Our HI model mice reproduce the human HI skin phenotype soon after the initiation of fetal skin keratinization and provide evidence that ABCA12 plays pivotal roles in lung and skin barrier functions.
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compound heterozygous ABCA12 mutations including a novel nonsense mutation underlie harlequin ichthyosis
Dermatology, 2007Co-Authors: Masashi Akiyama, Maki Goto, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Toshihiro Sato, Hiroshi ShimizuAbstract:Recently, it has been reported that several harlequin ichthyosis (HI) patients survive the neonatal period and their condition subsequently improves. Here we describe a 2-year-old Japanese boy who exhibited typical clinical features of HI at birth. He survived beyond the neonatal period after oral retinoid treatment and, at the age of 2 years, showed moderately thick, lamellar scales and erythroderma over his whole body. The patient is a compound heterozygote for 2 ABCA12 mutations, a paternal deletion mutation c.2021_2022del (p.Lys674ArgfsX63) and a novel maternal nonsense mutation c.7444C → T (p.Arg2482X). Electron microscopic observation of a skin biopsy specimen from the perinatal period revealed epidermal ultrastructural features consistent with HI. Immunofluorescence labeling using antiserum against a C-terminal ABCA12 epitope showed loss of expression in the patient’s epidermis. The present patient demonstrates that rapid diagnosis of HI by ABCA12 expression analysis and mutation detection, and early commencement of systemic retinoid therapy are crucial to significantly improving an HI patient’s prognosis.
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expression of the keratinocyte lipid transporter ABCA12 in developing and reconstituted human epidermis
American Journal of Pathology, 2007Co-Authors: Yasuko Yamanaka, Yoriko Sugiyamanakagiri, Maki Goto, Masashi Akiyama, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Mitsuhito Ota, Hiroshi ShimizuAbstract:Serious defects in the epidermal keratinocyte lipid transporter ABCA12 are known to result in a deficient skin lipid barrier, leading to harlequin ichthyosis (HI). HI is the most severe inherited keratinizing disorder and is frequently fatal in the perinatal period. To clarify the role of ABCA12, ABCA12 expression was studied in developing human skin and HI lesions artificially reconstituted in immunodeficient mice. By immunofluorescent study, ABCA12 was expressed in the periderm of the early stage two-layered human fetal epidermis. After formation of a three-layered epidermis, ABCA12 staining was seen throughout the entire epidermis. ABCA12 mRNA expression significantly increased during human skin development and reached 62% of the expression in normal adult skin, whereas the expression rate of transglutaminase 1, loricrin, and kallikrein 7 remained low. We transplanted keratinocytes from patients with HI and succeeded in reconstituting HI skin lesions in immunodeficient mice. The reconstituted lesions showed similar changes to those of patients with HI. Our findings demonstrate that ABCA12 is highly expressed in fetal skin and suggest that ABCA12 may play an essential role under both the wet and dry conditions, including the dramatic turning point from a wet environment of the amniotic fluid to a dry environment after birth.
Kaori Sakai - One of the best experts on this subject based on the ideXlab platform.
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self improvement of keratinocyte differentiation defects during skin maturation in ABCA12 deficient harlequin ichthyosis model mice
American Journal of Pathology, 2010Co-Authors: Teruki Yanagi, Masashi Akiyama, Kaori Sakai, Hiroshi Nishihara, Shinya Tanaka, Junko Ishikawa, Yuki Miyamura, Ayano Naoe, Takashi Kitahara, Hiroshi ShimizuAbstract:Harlequin ichthyosis (HI) is caused by loss-of-function mutations in the keratinocyte lipid transporter ABCA12. The patients often die in the first 1 or 2 weeks of life, although HI survivors’ phenotypes improve within several weeks after birth. In order to clarify the mechanisms of phenotypic recovery, we studied grafted skin and keratinocytes from ABCA12-disrupted (ABCA12−/−) mice showing abnormal lipid transport. ABCA12−/− neonatal epidermis showed significantly reduced total ceramide amounts and aberrant ceramide composition. Immunofluorescence and immunoblotting of ABCA12−/− neonatal epidermis revealed defective profilaggrin/filaggrin conversion and reduced protein expression of the differentiation-specific molecules, loricrin, kallikrein 5, and transglutaminase 1, although their mRNA expression was up-regulated. In contrast, ABCA12−/− skin grafts kept in a dry environment exhibited dramatic improvements in all these abnormalities. Increased transepidermal water loss, a parameter representing barrier defect, was remarkably decreased in grafted ABCA12−/− skin. Ten-passage sub-cultured ABCA12−/− keratinocytes showed restoration of intact ceramide distribution, differentiation-specific protein expression and profilaggrin/filaggrin conversion, which were defective in primary-cultures. Using cDNA microarray analysis, lipid transporters including four ATP-binding cassette transporters were up-regulated after sub-culture of ABCA12−/− keratinocytes compared with primary-culture. These results indicate that disrupted keratinocyte differentiation during the fetal development is involved in the pathomechanism of HI and, during maturation, ABCA12−/− epidermal keratinocytes regain normal differentiation processes. This restoration may account for the skin phenotype improvement observed in HI survivors.
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harlequin ichthyosis model mouse reveals alveolar collapse and severe fetal skin barrier defects
Human Molecular Genetics, 2008Co-Authors: Teruki Yanagi, Masashi Akiyama, Kaori Sakai, Hiroshi Nishihara, Wataru Nishie, Shinya Tanaka, Hiroshi ShimizuAbstract:: Harlequin ichthyosis (HI), which is the most severe genodermatosis, is caused by loss-of-function mutations in ABCA12, a member of the ATP-binding cassette transporter family. To investigate the pathomechanism of HI and the function of the ABCA12 protein, we generated ABCA12-deficient mice (ABCA12(-/-)) by targeting ABCA12. ABCA12(-/-) mice closely reproduce the human HI phenotype, showing marked hyperkeratosis with eclabium and skin fissure. Lamellar granule abnormalities and defective ceramide distribution were remarkable in the epidermis. Skin permeability assay of ABCA12(-/-) fetuses revealed severe skin barrier dysfunction after the initiation of keratinization. Surprisingly, the ABCA12(-/-) mice also demonstrated lung alveolar collapse immediately after birth. Lamellar bodies in alveolar type II cells of the ABCA12(-/-) mice lacked normal lamellar structures. The level of surfactant protein B, an essential component of alveolar surfactant, was reduced in the ABCA12(-/-) mice. Fetal therapeutic trials with systemic administration of retinoid or dexamethasone, which are effective for HI and respiratory distress, respectively, to the pregnant mother mice neither improved the skin phenotype nor extended the survival period. Our HI model mice reproduce the human HI skin phenotype soon after the initiation of fetal skin keratinization and provide evidence that ABCA12 plays pivotal roles in lung and skin barrier functions.
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compound heterozygous ABCA12 mutations including a novel nonsense mutation underlie harlequin ichthyosis
Dermatology, 2007Co-Authors: Masashi Akiyama, Maki Goto, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Toshihiro Sato, Hiroshi ShimizuAbstract:Recently, it has been reported that several harlequin ichthyosis (HI) patients survive the neonatal period and their condition subsequently improves. Here we describe a 2-year-old Japanese boy who exhibited typical clinical features of HI at birth. He survived beyond the neonatal period after oral retinoid treatment and, at the age of 2 years, showed moderately thick, lamellar scales and erythroderma over his whole body. The patient is a compound heterozygote for 2 ABCA12 mutations, a paternal deletion mutation c.2021_2022del (p.Lys674ArgfsX63) and a novel maternal nonsense mutation c.7444C → T (p.Arg2482X). Electron microscopic observation of a skin biopsy specimen from the perinatal period revealed epidermal ultrastructural features consistent with HI. Immunofluorescence labeling using antiserum against a C-terminal ABCA12 epitope showed loss of expression in the patient’s epidermis. The present patient demonstrates that rapid diagnosis of HI by ABCA12 expression analysis and mutation detection, and early commencement of systemic retinoid therapy are crucial to significantly improving an HI patient’s prognosis.
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expression of the keratinocyte lipid transporter ABCA12 in developing and reconstituted human epidermis
American Journal of Pathology, 2007Co-Authors: Yasuko Yamanaka, Yoriko Sugiyamanakagiri, Maki Goto, Masashi Akiyama, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Mitsuhito Ota, Hiroshi ShimizuAbstract:Serious defects in the epidermal keratinocyte lipid transporter ABCA12 are known to result in a deficient skin lipid barrier, leading to harlequin ichthyosis (HI). HI is the most severe inherited keratinizing disorder and is frequently fatal in the perinatal period. To clarify the role of ABCA12, ABCA12 expression was studied in developing human skin and HI lesions artificially reconstituted in immunodeficient mice. By immunofluorescent study, ABCA12 was expressed in the periderm of the early stage two-layered human fetal epidermis. After formation of a three-layered epidermis, ABCA12 staining was seen throughout the entire epidermis. ABCA12 mRNA expression significantly increased during human skin development and reached 62% of the expression in normal adult skin, whereas the expression rate of transglutaminase 1, loricrin, and kallikrein 7 remained low. We transplanted keratinocytes from patients with HI and succeeded in reconstituting HI skin lesions in immunodeficient mice. The reconstituted lesions showed similar changes to those of patients with HI. Our findings demonstrate that ABCA12 is highly expressed in fetal skin and suggest that ABCA12 may play an essential role under both the wet and dry conditions, including the dramatic turning point from a wet environment of the amniotic fluid to a dry environment after birth.
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dna based prenatal diagnosis of harlequin ichthyosis and characterization of ABCA12 mutation consequences
Journal of Investigative Dermatology, 2007Co-Authors: Masashi Akiyama, James R. Mcmillan, Kaori Sakai, Alain Hovnanian, M Titeux, Patrick Calvas, Frederique Jossic, Laure Tonasso, Hiroshi ShimizuAbstract:Until the identification of ABCA12 as the causative gene, prenatal diagnosis (PD) for harlequin ichthyosis (HI) had been performed by electron microscopic observation of fetal skin biopsy samples. We report the first case of HI DNA-based PD. Direct sequence analysis of ABCA12 revealed that the deceased proband was a compound heterozygote for two novel mutations. The maternal nonsense mutation p.Ser1249Term likely leads to nonsense-mediated messenger RNA decay. The paternal mutation c.7436G>A affects the last codon of exon 50 and was expected to be a splice site mutation. For their third pregnancy, the parents requested PD. Direct sequence analysis of fetal genomic DNA from amniotic fluid cells at 17 weeks gestation revealed the fetus was a compound heterozygote for both mutations. The parents requested the pregnancy to be terminated. Analysis of ABCA12 transcripts of cultured keratinocytes from the abortus showed the presence of six abnormally spliced products from the allele carrying the splice site mutation. Four of them lead to premature termination codons whereas the two others produced shortened proteins missing 21 and 31 amino acids from the second ATP-binding cassette. This report provides evidence for residual ABCA12 expression in HI, and demonstrates the efficiency of early DNA-based PD of HI.
James R. Mcmillan - One of the best experts on this subject based on the ideXlab platform.
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compound heterozygous ABCA12 mutations including a novel nonsense mutation underlie harlequin ichthyosis
Dermatology, 2007Co-Authors: Masashi Akiyama, Maki Goto, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Toshihiro Sato, Hiroshi ShimizuAbstract:Recently, it has been reported that several harlequin ichthyosis (HI) patients survive the neonatal period and their condition subsequently improves. Here we describe a 2-year-old Japanese boy who exhibited typical clinical features of HI at birth. He survived beyond the neonatal period after oral retinoid treatment and, at the age of 2 years, showed moderately thick, lamellar scales and erythroderma over his whole body. The patient is a compound heterozygote for 2 ABCA12 mutations, a paternal deletion mutation c.2021_2022del (p.Lys674ArgfsX63) and a novel maternal nonsense mutation c.7444C → T (p.Arg2482X). Electron microscopic observation of a skin biopsy specimen from the perinatal period revealed epidermal ultrastructural features consistent with HI. Immunofluorescence labeling using antiserum against a C-terminal ABCA12 epitope showed loss of expression in the patient’s epidermis. The present patient demonstrates that rapid diagnosis of HI by ABCA12 expression analysis and mutation detection, and early commencement of systemic retinoid therapy are crucial to significantly improving an HI patient’s prognosis.
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expression of the keratinocyte lipid transporter ABCA12 in developing and reconstituted human epidermis
American Journal of Pathology, 2007Co-Authors: Yasuko Yamanaka, Yoriko Sugiyamanakagiri, Maki Goto, Masashi Akiyama, James R. Mcmillan, Kaori Sakai, Daisuke Sawamura, Mitsuhito Ota, Hiroshi ShimizuAbstract:Serious defects in the epidermal keratinocyte lipid transporter ABCA12 are known to result in a deficient skin lipid barrier, leading to harlequin ichthyosis (HI). HI is the most severe inherited keratinizing disorder and is frequently fatal in the perinatal period. To clarify the role of ABCA12, ABCA12 expression was studied in developing human skin and HI lesions artificially reconstituted in immunodeficient mice. By immunofluorescent study, ABCA12 was expressed in the periderm of the early stage two-layered human fetal epidermis. After formation of a three-layered epidermis, ABCA12 staining was seen throughout the entire epidermis. ABCA12 mRNA expression significantly increased during human skin development and reached 62% of the expression in normal adult skin, whereas the expression rate of transglutaminase 1, loricrin, and kallikrein 7 remained low. We transplanted keratinocytes from patients with HI and succeeded in reconstituting HI skin lesions in immunodeficient mice. The reconstituted lesions showed similar changes to those of patients with HI. Our findings demonstrate that ABCA12 is highly expressed in fetal skin and suggest that ABCA12 may play an essential role under both the wet and dry conditions, including the dramatic turning point from a wet environment of the amniotic fluid to a dry environment after birth.
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dna based prenatal diagnosis of harlequin ichthyosis and characterization of ABCA12 mutation consequences
Journal of Investigative Dermatology, 2007Co-Authors: Masashi Akiyama, James R. Mcmillan, Kaori Sakai, Alain Hovnanian, M Titeux, Patrick Calvas, Frederique Jossic, Laure Tonasso, Hiroshi ShimizuAbstract:Until the identification of ABCA12 as the causative gene, prenatal diagnosis (PD) for harlequin ichthyosis (HI) had been performed by electron microscopic observation of fetal skin biopsy samples. We report the first case of HI DNA-based PD. Direct sequence analysis of ABCA12 revealed that the deceased proband was a compound heterozygote for two novel mutations. The maternal nonsense mutation p.Ser1249Term likely leads to nonsense-mediated messenger RNA decay. The paternal mutation c.7436G>A affects the last codon of exon 50 and was expected to be a splice site mutation. For their third pregnancy, the parents requested PD. Direct sequence analysis of fetal genomic DNA from amniotic fluid cells at 17 weeks gestation revealed the fetus was a compound heterozygote for both mutations. The parents requested the pregnancy to be terminated. Analysis of ABCA12 transcripts of cultured keratinocytes from the abortus showed the presence of six abnormally spliced products from the allele carrying the splice site mutation. Four of them lead to premature termination codons whereas the two others produced shortened proteins missing 21 and 31 amino acids from the second ATP-binding cassette. This report provides evidence for residual ABCA12 expression in HI, and demonstrates the efficiency of early DNA-based PD of HI.
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Localization of ABCA12 from Golgi apparatus to lamellar granules in human upper epidermal keratinocytes
Experimental dermatology, 2007Co-Authors: Kaori Sakai, Masashi Akiyama, James R. Mcmillan, Yoriko Sugiyama-nakagiri, Daisuke Sawamura, Hiroshi ShimizuAbstract:ABCA12 is an ATP-binding cassette transporter and is thought to act as a transmembrane lipid transporter. We reported that deleterious ABCA12 mutations cause a disturbance in lamellar granule (LG) lipid transport in the epidermal granular layer keratinocytes, resulting in harlequin ichthyosis, a severe genodermatosis. Detailed localization of ABCA12 in comparison with glucosylceramide and Golgi apparatus markers were studied in order to obtain clues to clarify the function(s) of ABCA12 in human skin. We performed double-labelling immunofluorescent staining using antibodies against ABCA12, glucosylceramide and two Golgi apparatus markers (TGN46 and GM130) in normal human skin and cultured keratinocytes. Immunogold electron microscopy for ABCA12 and glucosylceramide was studied on postembedding and cryoultrathin sections of normal human skin. Confocal laser scanning microscopy demonstrated that ABCA12 and glucosylceramide co-localized in the granular layer keratinocytes as well as in keratinocytes cultured in high Ca2+ conditions through the Golgi apparatus to the cell periphery. Postembedding immunogold electron microscopy revealed that both ABCA12 and glucosylceramide labellings were associated with the LG of the uppermost granular layer keratinocytes. Using cryoultramicrotomy, lamellar structures in the LG were more clearly observed, and ultrastructural localization of ABCA12 and glucosylceramide was better demonstrated to LG in the uppermost granular layer cells. These results indicate that ABCA12 plays an important role in lipid transport from the Golgi apparatus to LG in human granular layer keratinocytes.
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dna based prenatal diagnosis of harlequin ichthyosis and characterization of ABCA12 mutation
2007Co-Authors: Consequences Akiyama, James R. Mcmillan, Kaori Sakai, Alain Hovnanian, M Titeux, Patrick Calvas, Frederique Jossic, Laure Tonasso, Hiroshi ShimizuAbstract:Until the identification of ABCA12 as the causative gene, prenatal diagnosis (PD) for harlequin ichthyosis (HI) had been performed by electron microscopic observation of fetal skin biopsy samples. We report the first case of HI DNA-based PD. Direct sequence analysis of ABCA12 revealed that the deceased proband was a compound heterozygote for two novel mutations. The maternal nonsense mutation p.Ser1249Term likely leads to nonsense-mediated messenger RNA decay. The paternal mutation c.7436G4A affects the last codon of exon 50 and was expected to be a splice site mutation. For their third pregnancy, the parents requested PD. Direct sequence analysis of fetal genomic DNA from amniotic fluid cells at 17 weeks gestation revealed the fetus was a compound heterozygote for both mutations. The parents requested the pregnancy to be terminated. Analysis of ABCA12 transcripts of cultured keratinocytes from the abortus showed the presence of six abnormally spliced products from the allele carrying the splice site mutation. Four of them lead to premature termination codons whereas the two others produced shortened proteins missing 21 and 31 amino acids from the second ATPbinding cassette. This report provides evidence for residual ABCA12 expression in HI, and demonstrates the efficiency of early DNA-based PD of HI.
Mason W Freeman - One of the best experts on this subject based on the ideXlab platform.
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endogenous β glucocerebrosidase activity in ABCA12 epidermis elevates ceramide levels after topical lipid application but does not restore barrier function
Journal of Lipid Research, 2014Co-Authors: Jorge F Haller, Ruth Welti, Michael L Fitzgerald, Paul Cavallaro, Nicholas J Hernandez, Lee Dolat, Stephanie J Soscia, Gregory A Grabowski, Mason W FreemanAbstract:ABCA12 mutations disrupt the skin barrier and cause harlequin ichthyosis. We previously showed ABCA12(-/-) skin has increased glucosylceramide (GlcCer) and correspondingly lower amounts of ceramide (Cer). To examine why loss of ABCA12 leads to accumulation of GlcCer, de novo sphingolipid synthesis was assayed using [(14)C]serine labeling in ex vivo skin cultures. A defect was found in β-glucocerebrosidase (GCase) processing of newly synthesized GlcCer species. This was not due to a decline in GCase function. ABCA12(-/-) epidermis had 5-fold more GCase protein (n = 4, P < 0.01), and a 5-fold increase in GCase activity (n = 3, P < 0.05). As with ABCA12(+/+) epidermis, immunostaining in null skin showed a typical interstitial distribution of the GCase protein in the ABCA12(-/-) stratum corneum. Hence, we tested whether the block in GlcCer conversion could be circumvented by topically providing GlcCer. This approach restored up to 15% of the lost Cer products of GCase activity in the ABCA12(-/-) epidermis. However, this level of barrier ceramide replacement did not significantly reduce trans-epidermal water loss function. Our results indicate loss of ABCA12 function results in a failure of precursor GlcCer substrate to productively interact with an intact GCase enzyme, and they support a model of ABCA12 function that is critical for transporting GlcCer into lamellar bodies.
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ABCA12 maintains the epidermal lipid permeability barrier by facilitating formation of ceramide linoleic esters
Journal of Biological Chemistry, 2008Co-Authors: Ying Zuo, Debbie Z Zhuang, Rong Han, Giorgis Isaac, Jennifer J Tobin, Mary Mckee, Ruth Welti, Janice L Brissette, Michael L Fitzgerald, Mason W FreemanAbstract:Harlequin ichthyosis is a congenital scaling syndrome of the skin in which affected infants have epidermal hyperkeratosis and a defective permeability barrier. Mutations in the gene encoding a member of the ABCA transporter family, ABCA12, have been linked to harlequin ichthyosis, but the molecular function of the protein is unknown. To investigate the activity of ABCA12, we generated ABCA12 null mice and analyzed the impact on skin function and lipid content. ABCA12-/- mice are born with a thickened epidermis and die shortly after birth, as water rapidly evaporates from their skin. In vivo skin proliferation measurements suggest a lack of desquamation of the skin cells, rather than enhanced proliferation of basal layer keratinocytes, accounts for the 5-fold thickening of the ABCA12-/- stratum corneum. Electron microscopy revealed a loss of the lamellar permeability barrier in ABCA12-/- skin. This was associated with a profound reduction in skin linoleic esters of long-chain omega-hydroxyceramides and a corresponding increase in their glucosyl ceramide precursors. Because omega-hydroxyceramides are required for the barrier function of the skin, these results establish that ABCA12 activity is required for the generation of long-chain ceramide esters that are essential for the development of normal skin structure and function.
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ABCA12 maintains the epidermal lipid permeability barrier by facilitating formation of ceramide linoleic esters
Journal of Biological Chemistry, 2008Co-Authors: Ying Zuo, Debbie Z Zhuang, Rong Han, Giorgis Isaac, Jennifer J Tobin, Mary Mckee, Ruth Welti, Janice L Brissette, Michael L Fitzgerald, Mason W FreemanAbstract:Harlequin ichthyosis is a congenital scaling syndrome of the skin in which affected infants have epidermal hyperkeratosis and a defective permeability barrier. Mutations in the gene encoding a member of the ABCA transporter family, ABCA12, have been linked to harlequin ichthyosis, but the molecular function of the protein is unknown. To investigate the activity of ABCA12, we generated ABCA12 null mice and analyzed the impact on skin function and lipid content. ABCA12-/- mice are born with a thickened epidermis and die shortly after birth, as water rapidly evaporates from their skin. In vivo skin proliferation measurements suggest a lack of desquamation of the skin cells, rather than enhanced proliferation of basal layer keratinocytes, accounts for the 5-fold thickening of the ABCA12-/- stratum corneum. Electron microscopy revealed a loss of the lamellar permeability barrier in ABCA12-/- skin. This was associated with a profound reduction in skin linoleic esters of long-chain ω-hydroxyceramides and a corresponding increase in their glucosyl ceramide precursors. Because ω-hydroxyceramides are required for the barrier function of the skin, these results establish that ABCA12 activity is required for the generation of long-chain ceramide esters that are essential for the development of normal skin structure and function.
