RESUMEN
Sebaceous gland carcinoma (SGC) is a rare, but life-threatening condition with a predilection for the periocular region. Eyelid SGC can be broadly categorised into two subtypes, namely either nodular or pagetoid with the latter being more aggressive and requiring radical excision to save life. We have identified key altered microRNAs (miRNA) involved in SGC shared by both subtypes, hsa-miR-34a-5p and hsa-miR-16-5p. However, their gene targets BCL2 and MYC were differentially expressed with both overexpressed in pagetoid but unchanged in nodular suggesting different modes of action of these two miRNAs on BCL/MYC expression. Hsa-miR-150p is nodular-specifically overexpressed, and its target ZEB1 was significantly downregulated in nodular SGC suggesting a tumour suppressor role. Invasive pagetoid subtype demonstrated specific overexpression of hsa-miR-205 and downregulation of hsa-miR-199a. Correspondingly, miRNA gene targets, EZH2 (by hsa-miR-205) and CD44 (by hsa-miR-199a), were both overexpressed in pagetoid SGC. CD44 has been identified as a potential cancer stem cell marker in head and neck squamous cell carcinoma and its overexpression in pagetoid cells represents a novel treatment target. Aberrant miRNAs and their gene targets have been identified in both SGC subtypes, paving the way for better molecular understanding of these tumours and identifying new treatment targets.
Asunto(s)
Neoplasias de los Párpados/genética , Perfilación de la Expresión Génica/métodos , Redes Reguladoras de Genes , MicroARNs/genética , Neoplasias de las Glándulas Sebáceas/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-myc/genética , Homeobox 1 de Unión a la E-Box con Dedos de Zinc/genéticaRESUMEN
PURPOSE: Graves' orbitopathy (GO) is a sight-threatening autoimmune disorder causing extraocular muscle fibrosis, upper lid retraction and eye bulging due to orbital fat expansion. These clinical features are mediated by aspects of orbital fibroblasts differentiation, including adipogenesis and fibrosis. Our previous work suggested that this dual phenotype might be a manifestation of mixed cell populations, partially linked to the expression of mesenchymal stem cell (MSC) marker CD90. Thus, we set out to determine whether GO orbital fibroblasts displayed MSC properties. METHODS: Control and GO orbital fibroblasts previously characterized for CD90 and CD45 expression were analyzed by flow cytometry for classical MSC positive (CD73, CD105) and negative (CD14, CD19, HLA-DR, and CD34) markers. Graves' orbitopathy fibroblasts were tested further for their ability to undergo lineage specific differentiation following standard protocols. RESULTS: Control and GO fibroblasts strongly expressed CD73 and CD105, with a higher percentage of positive cells and stronger expression levels in GO. Neither cell type expresses CD14, CD19, and HLA-DR. Protein CD34 was expressed at low levels by 45% to 70% of the cells, with its expression significantly lower in GO cells. Graves' orbitopathy fibroblasts displayed features of osteogenesis (calcium deposits, and osteocalcin [BGLAP] and osteonectin [SPARC] expression), chondrogenesis (glycosaminoglycan production; SOX9 and aggrecan [ACAN] expression), myogenesis (α-smooth muscle actin expression), and neurogenesis (ß-III tubulin expression) upon differentiation. CONCLUSIONS: Our findings suggest that orbital fibroblasts contain a population of cells that fulfil the criteria defining MSC. This subpopulation may be increased in GO, possibly underlying the complex differentiation phenotype of the disease.
Asunto(s)
Fibroblastos/metabolismo , Oftalmopatía de Graves/metabolismo , Células Madre Mesenquimatosas/metabolismo , Agrecanos/metabolismo , Antígenos CD/metabolismo , Biomarcadores/metabolismo , Estudios de Casos y Controles , Diferenciación Celular/fisiología , Células Cultivadas , Condrogénesis/fisiología , Fibroblastos/citología , Oftalmopatía de Graves/patología , Antígenos HLA-DR/metabolismo , Humanos , Células Madre Mesenquimatosas/citología , Osteocalcina/metabolismo , Osteogénesis/fisiología , Osteonectina/metabolismo , Factor de Transcripción SOX9/metabolismoRESUMEN
Graves' orbitopathy (GO) is a disfiguring and sometimes blinding disease, characterised by inflammation and swelling of orbital tissues, with fibrosis and adipogenesis being predominant features. Little is known about the disease aetiology and the molecular mechanisms driving the phenotypic changes in orbital fibroblasts are unknown. Using fibroblasts isolated from the orbital fat of undiseased individuals or GO patients, we have established a novel in vitro model to evaluate the dual profile of GO cells in a three-dimensional collagen matrix; this pseudo-physiological 3D environment allows measurement of their contractile and adipogenic properties. GO cells contracted collagen matrices more efficiently than control cells following serum or TGFß1 stimulation, and showed a slightly increased ability to proliferate in the 3D matrix, in accordance with a fibro-proliferative phenotype. GO cells, unlike controls, also spontaneously differentiated into adipocytes in 3D cultures - confirming an intrinsic adipogenic profile. However, both control and GO cells underwent adipogenesis when cultured under pathological pressure levels. We further demonstrate that a Thy-1-low population of GO cells underlies the adipogenic - but not the contractile - phenotype and, using inhibitors, confirm that the contractile and adipogenic phenotypes are regulated by separate pathways. In view of the current lack of suitable treatment for GO, we propose that this new model testing the duality of the GO phenotype could be useful as a preclinical evaluation for the efficacy of potential treatments.
Asunto(s)
Adipogénesis , Fibroblastos/patología , Oftalmopatía de Graves/patología , Modelos Biológicos , Anciano , Células Cultivadas , Matriz Extracelular/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fenotipo , Presión , Antígenos Thy-1/metabolismoRESUMEN
Oral submucous fibrosis (OSF) is characterized by abnormal collagen metabolism in the submucosal connective tissue. Its influence on the overlying epithelium is not known but about 14% of OSF cases undergo malignant transformation to squamous cell carcinoma indicating association with abnormality of the epithelium. Here, we have defined the keratin expression profile, by immunohistochemistry and quantitative image analysis, using a panel of 22 anti-keratin monoclonal antibodies on 28 OSF samples. We observed an increase of K1 and K10 in the suprabasal layers, induction of K6 in the basal layer and complete loss of K19 in the epithelium. Furthermore, there was increased K17 expression in the suprabasal layers, which correlated with disease severity. In a subset of the most severe OSF cases (14%), K17 expression was completely lost in the basal layer which might define them to be at most risk to undergo malignant transformation. There was no detectable expression of K8, K18, K7 and K9 and the expression of K4, K13, K14, K15 and K16 did not change in OSF. We propose that the altered keratin profiles could be useful as histological diagnostic markers and provide important insights into the pathogenesis of the disease and its predisposition to malignancy.