RESUMEN
K31 was previously considered as one of the hair keratins. During a study on differential markers between hair follicles and eccrine sweat glands, we observed that K31 was expressed in eccrine sweat gland cells in a scattered pattern, similar to the distribution of dark or clear secretory cells. To investigate the precise cell localization of K31 in human eccrine sweat glands and find new marker for eccrine sweat gland cells, human skin samples were fixed, paraffined and sectioned. The serial sections were stained for K31, dark secretory cell marker gross cystic disease fluid protein 15 (GCDFP15) and clear secretory cell marker carbonic anhydrase II (CAII). The exact cell localization of K31 was detected by double immunofluorescence staining of K31 and a serial of cell-specific markers, and further by dual stain using a combination of periodic acid-Schiff (PAS) and immunofluorescence for K31 and GCDFP15. The expression pattern of K31-positive cells was similar to that of CAII-positive cells but was different from that of GCDFP15-positive staining in serial sections. Double immunofluorescent staining showed that K31-positive cells co-expressed K7 and CAII, but not S100P, α-SMA or GCDFP15. Dual stain by combined PAS and immunofluorescence showed that K31-positive cells are negative for PAS staining. We conclude that K31 is a previously unreported eccrine clear cell marker that allows for distinction between clear and dark secretory cells, as well as between secretory coils and ducts of eccrine sweat glands in human eccrine sweat glands.
Asunto(s)
Antígenos de Diferenciación/biosíntesis , Glándulas Ecrinas/metabolismo , Regulación de la Expresión Génica , Queratinas Específicas del Pelo/biosíntesis , Queratinas Tipo I/biosíntesis , Adolescente , Adulto , Niño , Glándulas Ecrinas/citología , Femenino , Humanos , Masculino , Proteínas de Transporte de Membrana/biosíntesis , Persona de Mediana EdadRESUMEN
BACKGROUND: Pilomatricomas are tumors that emulate the differentiation of matrix cells of the hair follicle, showing cortical differentiation, with sequential expression of K35 and K31 keratins. Beta-catenin gene is frequently mutated in pilomatricoma, leading to beta-catenin nuclear accumulation, and to downstream expression of LEF1. Skin matrical tumors other than pilomatricoma are very rare, and comprise purely matrical tumors and focally matrical tumors. We aimed at studying cortical differentiation, beta-catenin pathway and expression of the follicular stem-cell marker PHLDA1 in a series of matrical tumors other than pilomatricoma. METHODS: In 36 prospectively collected tumors, K31, K35, CK17, LEF1, HOXC13, beta-catenin and PHLDA1 expressions were evaluated. Five pilomatricomas were used as controls. RESULTS: In 18 purely matrical tumors (11 matrical carcinomas, 4 melanocytic matricomas, 3 matricomas) and 18 focally matrical tumors (11 basal cell carcinomas, 3 trichoepithelioma/trichoblastomas, 4 others), sequential K35, HOXC13 and K31 expressions were found, indicating cortical differentiation. Germinative matrix cells were always CK17-, and showed nuclear beta-catenin accumulation, with LEF1 and PHLDA1 expressions. CONCLUSIONS: Nuclear beta-catenin and LEF1 expression was highly conserved in matrical tumors, and suggested a common tumorigenesis driven by Wnt pathway activation. PHLDA1 was consistently expressed in matrical tumors and in areas of matrical differentiation.
Asunto(s)
Enfermedades del Cabello/patología , Queratinas Específicas del Pelo/biosíntesis , Neoplasias Cutáneas/patología , Factores de Transcripción/biosíntesis , beta Catenina/biosíntesis , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Femenino , Enfermedades del Cabello/metabolismo , Humanos , Inmunohistoquímica , Queratinas Específicas del Pelo/análisis , Masculino , Persona de Mediana Edad , Neoplasias Cutáneas/metabolismo , Factores de Transcripción/análisis , Adulto Joven , beta Catenina/análisisRESUMEN
Aurora kinase-A (Aurora-A) promotes timely entry into mitosis, centrosome maturation, and formation of bipolar spindles. To address the role of Aurora-A in skin development and homeostasis, we interbred a floxed Aurora-A (Aurora-A(fl)) mouse with the Cre-deleter strain, K14.Cre. Aurora-A(fl/fl);Krt14.Cre (Aurora-A(-/-)) mice died shortly after birth. These mice had translucent skin, and histological evaluation showed that the dorsal skin was very thin and fragile with frank erosions. Although the expression of the basal layer marker keratin 14 and the differentiation marker keratin 1 was evident in Aurora-A(-/-) epidermis, there was a marked reduction in the number of suprabasal layers and basal keratinocytes. Dye exclusion assays also showed defects in barrier function. Unlike wild-type cells, Aurora-A(-/-) basal progenitors were delayed in forming two layers at embryonic day (E)13.5 when embryonic skin begins to stratify. Increased numbers of mitotic cells, apoptotic bodies, and polyploid keratinocytes were evident in Aurora-A(-/-) epidermis, indicating that a deficiency in Aurora-A promotes aberrant mitosis, mitotic slippage, and cell death. Finally, Aurora-A(-/-) keratinocytes displayed centrosomal abnormalities that included centrosomes located at nonapical sites in basal cells. Thus, the deletion of Aurora-A in the developing epidermis alters centrosome function of basal keratinocytes and markedly impairs their ability to divide and stratify.
Asunto(s)
División Celular/fisiología , Proteínas Serina-Treonina Quinasas/fisiología , Piel/enzimología , Piel/crecimiento & desarrollo , Animales , Apoptosis/fisiología , Aurora Quinasa A , Aurora Quinasas , Diferenciación Celular/genética , Diferenciación Celular/fisiología , División Celular/genética , Centrosoma/enzimología , Eliminación de Gen , Queratina-1 , Queratina-14/biosíntesis , Queratinocitos/fisiología , Queratinas Específicas del Pelo/biosíntesis , Ratones , Poliploidía , Proteínas Serina-Treonina Quinasas/genética , Piel/patología , Células Madre/fisiologíaRESUMEN
The hair follicle is an intricate miniature organ dedicated to the production of the structural hair fiber, which is largely composed of hair keratin (HK) proteins. Many developmental pathways contribute to hair follicle development; however, the molecular control of HK genes is still far from being resolved. Because the nuclear factor (NF)-kappaB pathway is known to be involved in the morphogenesis of the hair follicle, we explored the possibility that it may also regulate HK expression. To this end, we analyzed the effect of p65/RelA, an NF-kappaB effector, on HK regulatory regions using transient transfections into tissue culture cells. Reporter assays on cells transfected with HK promoter constructs and real-time polymerase chain reaction analysis of endogenous HK gene activity demonstrated that p65 induces transcriptional activation of several HK genes of human and mouse origin, primarily that of acidic hair keratin 5 (Ha5). Focusing on the highly responsive human Ha5 gene, we defined the major NF-kappaB/RelA binding sites in its regulatory region and showed the direct binding of p65 to these sites using gel shift assays. We further show, using immunohistochemistry on human hair follicle sections, that p65 is co-expressed with HKs in the hair shaft compartment and may thus be the effector that mediates the NF-kappaB pathway's activity, which recently was genetically demonstrated to be active in the same region. Thus, we provide evidence for a previously unknown function of NF-kappaB in hair formation-direct activation of HK target genes-a function that may shed light on some of the symptoms of ectodermal dysplasias.
Asunto(s)
Folículo Piloso/metabolismo , Queratinas Específicas del Pelo/genética , Queratinas Tipo II/genética , Queratinas Tipo I/genética , Factor de Transcripción ReIA/fisiología , Activación Transcripcional , Animales , Sitios de Unión , Células Cultivadas/metabolismo , ADN/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Células Epiteliales/metabolismo , Regulación de la Expresión Génica , Humanos , Queratinocitos/metabolismo , Queratinas Específicas del Pelo/biosíntesis , Queratinas Tipo I/biosíntesis , Queratinas Tipo II/biosíntesis , Ratones , FN-kappa B/fisiología , Regiones Promotoras Genéticas/genética , Mapeo de Interacción de Proteínas , Proteínas Recombinantes de Fusión/fisiología , Factor de Transcripción ReIA/genéticaRESUMEN
Two classical mouse hair coat mutations, Rex (Re) and Rex wavy coat (Re(wc)), are linked to the type I inner root sheath (IRS) keratin genes of chromosome 11. An N-ethyl-N-nitrosourea-induced mutation, M100573, also maps close to the type I IRS keratin genes. In this study, we demonstrate that Re and M100573 mice bear mutations in the type I IRS gene Krt25; Re(wc) mice bear an additional mutation in the type I IRS gene Krt27. These three mutations are located in the helix termination motif of the 2B alpha-helical rod domain of a type I IRS keratin protein. Immunohistological analysis revealed abnormal foam-like immunoreactivity with an antibody raised to type II IRS keratin K71 in the IRS of Re/+ mice. These results suggest that the helix termination motif is essential for the proper assembly of types I and II IRS keratin protein complexes and the formation of keratin intermediate filaments.