RESUMO
BACKGROUND: Psoriasis is characterized by aberrant activation of several pro-inflammatory circuits as well as abnormal hyperproliferation and dysregulated apoptosis of keratinocytes (KCs). Most currently available therapeutic options primarily target psoriasis-associated immunological defects rather than epidermal abnormalities. OBJECTIVE: To investigate the efficacy of the histone deacetylase (HDAC) inhibitor, Vorinostat, in targeting hyperproliferation and impaired apoptosis in psoriatic skin. METHODS: Vorinostat effect was investigated in primary KCs cell cultures using cell cycle analysis by flow cytometry, apoptosis assays (Annexin V-FICH and caspase-3/7) and antibody arrays, qRT-PCR and immunohistochemistry. Vorinostat impact on clinical manifestations of psoriasis was investigated in a chimeric mouse model. RESULTS: Vorinostat was found to inhibit KCs proliferation and to induce their differentiation and apoptosis. Using a chimeric mouse model, vorinostat was found to result in marked attenuation of a psoriasiform phenotype with a significant decrease in epidermal thickness and inhibition of epidermal proliferation. CONCLUSIONS: Our results support the notion that vorinostat, a prototypic HDAC inhibitor, may be of potential use in the treatment of psoriasis and other hyperproliferative skin disorders.
Assuntos
Inibidores de Histona Desacetilases , Psoríase , Animais , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Inibidores de Histona Desacetilases/farmacologia , Inibidores de Histona Desacetilases/uso terapêutico , Camundongos , Psoríase/tratamento farmacológico , Vorinostat/farmacologia , Vorinostat/uso terapêuticoRESUMO
BACKGROUND: In peripheral artery disease (PAD), blockage of the blood supply to the limbs, most frequently the legs, leads to impaired blood flow and tissue ischemia. Pluristem's PLX-PAD cells are placenta-derived mesenchymal stromal-like cells currently in clinical trials for the treatment of peripheral artery diseases. METHODS: In this work, the hind limb ischemia (HLI) mouse model was utilized to study the efficacy and mechanism of action of PLX-PAD cells. ELISA assays were performed to characterize and quantitate PLX-PAD secretions in vitro. RESULTS: PLX-PAD cells administered intramuscularly rescued blood flow to the lower limb after HLI induction in a dose-dependent manner. While rescue of blood flow was site-dependent, numerous administration regimes enabled rescue of blood flow, indicating a systemic effect mediated by PLX-PAD secretions. Live PLX-PAD cells were more efficacious than cell lysate in rescuing blood flow, indicating the importance of prolonged cytokine secretion for maximal blood flow recovery. In vitro studies showed a multifactorial secretion profile including numerous pro-angiogenic proteins; these are likely involved in the PLX-PAD mechanism of action. DISCUSSION: Live PLX-PAD cells were efficacious in rescuing blood flow after the induction of HLI in the mouse model in a dose- and site-dependent manner. The fact that various administration routes of PLX-PAD rescued blood flow indicates that the mechanism of action likely involves one of systemic secretions which promote angiogenesis. Taken together, the data support the further clinical testing of PLX-PAD cells for PAD indications.
Assuntos
Membro Posterior/irrigação sanguínea , Doença Arterial Periférica/terapia , Placenta/citologia , Células Estromais/transplante , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Isquemia/fisiopatologia , Isquemia/terapia , Masculino , Células-Tronco Mesenquimais/citologia , Camundongos Endogâmicos C57BL , Gravidez , Fluxo Sanguíneo RegionalRESUMO
The coexistence of abnormal keratinization and aberrant pigmentation in a number of cornification disorders has long suggested a mechanistic link between these two processes. Here, we deciphered the genetic basis of Cole disease, a rare autosomal-dominant genodermatosis featuring punctate keratoderma, patchy hypopigmentation, and uncommonly, cutaneous calcifications. Using a combination of exome and direct sequencing, we showed complete cosegregation of the disease phenotype with three heterozygous ENPP1 mutations in three unrelated families. All mutations were found to affect cysteine residues in the somatomedin-B-like 2 (SMB2) domain in the encoded protein, which has been implicated in insulin signaling. ENPP1 encodes ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1), which is responsible for the generation of inorganic pyrophosphate, a natural inhibitor of mineralization. Previously, biallelic mutations in ENPP1 were shown to underlie a number of recessive conditions characterized by ectopic calcification, thus providing evidence of profound phenotypic heterogeneity in ENPP1-associated genetic diseases.
Assuntos
Calcificação Fisiológica/genética , Hipopigmentação/genética , Ceratose/genética , Mutação , Diester Fosfórico Hidrolases/genética , Poroceratose/genética , Pirofosfatases/genética , Dermatopatias/genética , Exoma , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Linhagem , Transdução de Sinais/genética , Dermatopatias Genéticas/genética , Somatomedinas/genéticaRESUMO
Pityriasis rubra pilaris (PRP) is a papulosquamous disorder phenotypically related to psoriasis. The disease has been occasionally shown to be inherited in an autosomal-dominant fashion. To identify the genetic cause of familial PRP, we ascertained four unrelated families affected by autosomal-dominant PRP. We initially mapped PRP to 17q25.3, a region overlapping with psoriasis susceptibility locus 2 (PSORS2 [MIM 602723]). Using a combination of linkage analysis followed by targeted whole-exome sequencing and candidate-gene screening, we identified three different heterozygous mutations in CARD14, which encodes caspase recruitment domain family, member 14. CARD14 was found to be specifically expressed in the skin. CARD14 is a known activator of nuclear factor kappa B signaling, which has been implicated in inflammatory disorders. Accordingly, CARD14 levels were increased, and p65 was found to be activated in the skin of PRP-affected individuals. The present data demonstrate that autosomal-dominant PRP is allelic to familial psoriasis, which was recently shown to also be caused by mutations in CARD14.
Assuntos
Proteínas Adaptadoras de Sinalização CARD/genética , Guanilato Ciclase/genética , Proteínas de Membrana/genética , Mutação , Pitiríase Rubra Pilar/genética , Adulto , Sequência de Bases , Criança , Pré-Escolar , Feminino , Humanos , Masculino , Linhagem , Pele/metabolismoRESUMO
Disproportionate short stature refers to a heterogeneous group of hereditary disorders that are classified according to their mode of inheritance, clinical skeletal and nonskeletal manifestations, and radiological characteristics. In the present study, we report on an autosomal-recessive osteocutaneous disorder that we termed SOFT (short stature, onychodysplasia, facial dysmorphism, and hypotrichosis) syndrome. We employed homozygosity mapping to locate the disease-causing mutation to region 3p21.1-3p21.31. Using whole-exome-sequencing analysis complemented with Sanger direct sequencing of poorly covered regions, we identified a homozygous point mutation (c.512T>C [p.Leu171Pro]) in POC1A (centriolar protein homolog A). This mutation was found to cosegregate with the disease phenotype in two families. The p.Leu171Pro substitution affects a highly conserved amino acid residue and is predicted to interfere with protein function. Poc1, a POC1A ortholog, was previously found to have a role in centrosome stability in unicellular organisms. Accordingly, although centrosome structure was preserved, the number of centrosomes and their distribution were abnormal in affected cells. In addition, the Golgi apparatus presented a dispersed morphology, cholera-toxin trafficking from the plasma membrane to the Golgi was aberrant, and large vesicles accumulated in the cytosol. Collectively, our data underscore the importance of POC1A for proper bone, hair, and nail formation and highlight the importance of normal centrosomes in Golgi assembly and trafficking from the plasma membrane to the Golgi apparatus.
Assuntos
Anormalidades Múltiplas/genética , Cromossomos Humanos Par 3/genética , Hipotricose/genética , Proteínas/genética , Anormalidades Múltiplas/patologia , Sequência de Aminoácidos , Sequência de Bases , Proteínas de Ciclo Celular , Mapeamento Cromossômico , Proteínas do Citoesqueleto , Exoma/genética , Feminino , Complexo de Golgi/patologia , Humanos , Indóis , Masculino , Microscopia Confocal , Dados de Sequência Molecular , Linhagem , Mutação Puntual/genética , Polimorfismo de Fragmento de Restrição/genética , Transporte Proteico/genética , Análise de Sequência de DNARESUMO
Alopecia-neurological defects-endocrinopathy (ANE) syndrome is a rare inherited hair disorder, which was shown to result from decreased expression of the RNA-binding motif protein 28 (RBM28). In this study, we attempted to delineate the role of RBM28 in hair biology. First, we sought to obtain evidence for the direct involvement of RBM28 in hair growth. When RBM28 was downregulated in human hair follicle (HF) organ cultures, we observed catagen induction and HF growth arrest, indicating that RBM28 is necessary for normal hair growth. We also aimed at identifying molecular targets of RBM28. Given that an RBM28 homologue was recently found to regulate miRNA biogenesis in C. elegans and given the known pivotal importance of miRNAs for proper hair follicle development, we studied global miRNA expression profile in cells knocked down for RBM28. This analysis revealed that RBM28 controls the expression of miR-203. miR-203 was found to regulate in turn TP63, encoding the transcription factor p63, which is critical for hair morphogenesis. In conclusion, RBM28 contributes to HF growth regulation through modulation of miR-203 and p63 activity.
Assuntos
Alopecia/metabolismo , Doenças do Sistema Endócrino/metabolismo , Regulação da Expressão Gênica , Folículo Piloso/metabolismo , Deficiência Intelectual/metabolismo , MicroRNAs/fisiologia , Proteínas de Ligação a RNA/fisiologia , Fatores de Transcrição/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Alopecia/fisiopatologia , Células Cultivadas , Doenças do Sistema Endócrino/fisiopatologia , Genes Reporter , Cabelo/crescimento & desenvolvimento , Folículo Piloso/crescimento & desenvolvimento , Humanos , Deficiência Intelectual/fisiopatologia , Queratinócitos/metabolismo , Morfogênese , Técnicas de Cultura de Órgãos , Interferência de RNA , RNA Interferente Pequeno/genética , Proteínas de Ligação a RNA/genética , Transfecção , Regulação para CimaRESUMO
Monogenic disorders offer unique opportunities for researchers to shed light upon fundamental physiological processes in humans. We investigated a large family affected with autosomal-dominant adermatoglyphia (absence of fingerprints) also known as the "immigration delay disease." Using linkage and haplotype analyses, we mapped the disease phenotype to 4q22. One of the genes located in this interval is SMARCAD1, a member of the SNF subfamily of the helicase protein superfamily. We demonstrated the existence of a short isoform of SMARCAD1 exclusively expressed in the skin. Sequencing of all SMARCAD1 coding and noncoding exons revealed a heterozygous transversion predicted to disrupt a conserved donor splice site adjacent to the 3' end of a noncoding exon uniquely present in the skin-specific short isoform of the gene. This mutation segregated with the disease phenotype throughout the entire family. Using a minigene system, we found that this mutation causes aberrant splicing, resulting in decreased stability of the short RNA isoform as predicted by computational analysis and shown by RT-PCR. Taken together, the present findings implicate a skin-specific isoform of SMARCAD1 in the regulation of dermatoglyph development.
Assuntos
Genes Dominantes/genética , Mutação/genética , Dermatopatias/genética , Pele/patologia , Sequência de Bases , Mapeamento Cromossômico , DNA Helicases/genética , DNA Helicases/metabolismo , Análise Mutacional de DNA , Feminino , Perfilação da Expressão Gênica , Haplótipos/genética , Células HeLa , Humanos , Escore Lod , Masculino , Dados de Sequência Molecular , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Conformação de Ácido Nucleico , Especificidade de Órgãos/genética , Linhagem , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Pele/metabolismoRESUMO
Autosomal-recessive congenital ichthyoses represent a large and heterogeneous group of disorders of epidermal cornification. Recent data suggest that most of these disorders might result from defective lipid transport and metabolism. In the present study, we describe a late-onset form of recessive ichthyosis in a large consanguineous pedigree. By using a combination of homozygosity mapping and positional candidate-gene screening, we identified a 2 bp deletion in LIPN that segregated with the disease phenotype throughout the family. LIPN encodes one of six acid lipases known to be involved in triglyceride metabolism in mammals . LIPN was found to be exclusively expressed in the epidermis and to be strongly induced during keratinocyte differentiation.
Assuntos
Ictiose/enzimologia , Ictiose/genética , Lipase/genética , Deleção de Sequência , Adolescente , Sequência de Bases , Consanguinidade , Primers do DNA/genética , Feminino , Genes Recessivos , Haplótipos , Homozigoto , Humanos , Ictiose/patologia , Masculino , Linhagem , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de RestriçãoRESUMO
Autosomal recessive congenital ichthyosis refers to a heterogeneous group of cornification disorders of major impact on patients' life. The disease has been linked so far to mutations in 8 distinct genes. We report a consanguineous family of Arab Muslim origin with several members displaying a severe form of congenital ichthyosiform erythroderma. Using a panel of polymorphic microsatellite markers, we identified a region of homozygosity shared by all patients on 2q34, in a region harbouring the ABCA12 gene. Direct sequencing of genomic DNA derived from a patient failed to reveal any obviously pathogenic change in the coding sequence of this gene. In contrast, cDNA sequence analysis revealed the existence of a 163-bp-long deletion in exon 24, thus pointing to a splicing defect. Careful reanalysis of the genomic DNA sequence revealed apart from several known single-nucleotide polymorphisms, a hitherto unreported homozygous synonymous mutation in exon 24 (c.3456G>A; p.S1152S), which was found to lead to the formation of a novel splicing acceptor site. Synonymous mutations have been shown to uncommonly cause inherited disorders in humans. Here, we present the first example of a congenital form of ichthyosis resulting from such a genetic defect.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Eritrodermia Ictiosiforme Congênita/genética , Mutação , Adolescente , Árabes/genética , Cromossomos Humanos Par 2/genética , Consanguinidade , Análise Mutacional de DNA , Feminino , Genes Recessivos , Homozigoto , Humanos , Israel , Masculino , LinhagemRESUMO
3-Methylglutaconic aciduria (3-MGCA) type IV is defined as a heterogeneous group of inborn errors featuring in common 3-MGCA and associated with primary mitochondrial dysfunction leading to a spectrum of multisystem conditions. We studied four patients who presented at birth with a clinical picture simulating a primary mitochondrial hepatic disorder consistent with the MEGDEL syndrome including 3-MGCA, sensorineural deafness, encephalopathy and a brain magnetic resonance imaging with signs of Leigh disease. All affected children displayed biochemical features consistent with mitochondrial OXPHOS dysfunction including hepatic mitochondrial DNA depletion in one patient. Homozygosity mapping identified a candidate locus on 6q25.2-6q26. Using whole exome sequencing, we identified two novel homozygous mutations in SERAC1 recently reported to harbor mutations in MEGDEL syndrome. Both mutations were found to lead to decreased or absent expression of SERAC1. The present findings indicate that infantile hepatopathy is a cardinal feature of MEGDEL syndrome. We thus propose to rename the disease MEGDHEL syndrome.
Assuntos
Anormalidades Múltiplas/genética , Hidrolases de Éster Carboxílico/genética , Perda Auditiva Neurossensorial/genética , Doença de Leigh/genética , Hepatopatias/genética , Erros Inatos do Metabolismo/genética , Doenças Mitocondriais/genética , Mutação , Anormalidades Múltiplas/diagnóstico , Encéfalo/patologia , Estudos de Casos e Controles , Mapeamento Cromossômico , Consanguinidade , Análise Mutacional de DNA , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Perda Auditiva Neurossensorial/diagnóstico , Homozigoto , Humanos , Recém-Nascido , Doença de Leigh/diagnóstico , Fígado/patologia , Fígado/ultraestrutura , Hepatopatias/diagnóstico , Imageamento por Ressonância Magnética , Masculino , Erros Inatos do Metabolismo/diagnóstico , Repetições de Microssatélites/genética , Mitocôndrias/patologia , Mitocôndrias/ultraestrutura , Doenças Mitocondriais/diagnóstico , Linhagem , Polimorfismo de Nucleotídeo Único , SíndromeRESUMO
Inherited disorders of elastic tissue represent a complex and heterogeneous group of diseases, characterized often by sagging skin and occasionally by life-threatening visceral complications. In the present study, we report on an autosomal-recessive disorder that we have termed MACS syndrome (macrocephaly, alopecia, cutis laxa, and scoliosis). The disorder was mapped to chromosome 20p11.21-p11.23, and a homozygous frameshift mutation in RIN2 was found to segregate with the disease phenotype in a large consanguineous kindred. The mutation identified results in decreased expression of RIN2, a ubiquitously expressed protein that interacts with Rab5 and is involved in the regulation of endocytic trafficking. RIN2 deficiency was found to be associated with paucity of dermal microfibrils and deficiency of fibulin-5, which may underlie the abnormal skin phenotype displayed by the patients.
Assuntos
Alopecia/genética , Cútis Laxa/genética , Fatores de Troca do Nucleotídeo Guanina/deficiência , Escoliose/genética , Crânio/crescimento & desenvolvimento , Adolescente , Adulto , Proteínas de Transporte/genética , Estudos de Casos e Controles , Mapeamento Cromossômico , Cromossomos Humanos Par 20 , Consanguinidade , Cútis Laxa/metabolismo , Procedimentos Cirúrgicos Dermatológicos , Derme/metabolismo , Derme/patologia , Tecido Elástico/metabolismo , Tecido Elástico/ultraestrutura , Proteínas da Matriz Extracelular/metabolismo , Mutação da Fase de Leitura , Genes Recessivos , Fatores de Troca do Nucleotídeo Guanina/genética , Homozigoto , Humanos , Imuno-Histoquímica , Fenótipo , Radiografia , Pele/metabolismo , Pele/patologia , Crânio/diagnóstico por imagem , SíndromeRESUMO
Culturing 3D-expanded human placental-derived adherent stromal cells (ASCs) in the presence of tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) transiently upregulated the secretion of numerous anti-proliferative, anti-angiogenic and pro-inflammatory cytokines. In a 3D-spheroid screening assay, conditioned medium from these induced-ASCs inhibited proliferation of cancer cell lines, including triple-negative breast cancer (TNBC) lines. In vitro co-culture studies of induced-ASCs with MDA-MB-231 human breast carcinoma cells, a model representing TNBC, supports a mechanism involving immunomodulation and angiogenesis inhibition. In vivo studies in nude mice showed that intramuscular administration of induced-ASCs halted MDA-MB-231 cell proliferation, and inhibited tumor progression and vascularization. Thirty percent of treated mice experienced complete tumor remission. Murine serum concentrations of the tumor-supporting cytokines Interleukin-6 (IL-6), Vascular endothelial growth factor (VEGF) and Granulocyte-colony stimulating factor (G-CSF) were lowered to naïve levels. A somatic mutation analysis identified numerous genes which could be screened in patients to increase a positive therapeutic outcome. Taken together, these results show that targeted changes in the secretion profile of ASCs may improve their therapeutic potential.