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1.
Semin Cell Dev Biol ; 128: 103-111, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-34481710

RESUMO

The colon mucosa is lined with crypts of circa 300 cells, forming a continuous barrier whose roles include absorption of water, recovery of metabolic energy sources (notably short chain fatty acids), secretion of a protective mucus barrier, and physiological signalling. There is high turnover and replenishment of cells in the mucosa, disruption of this may lead to bowel pathologies including cancer and inflammatory bowel disease. Keratins have been implicated in the processes of cell death, epithelial integrity, response to inflammation and as a result are often described as guardians of the colonic epithelium. Keratin proteins carry extensive post-translational modifications, the cofactors for kinases, acetyl transferases and other modification-regulating enzymes are themselves products of metabolism. A cluster of studies has begun to reveal a bidirectional relationship between keratin form and function and metabolism. In this paper we hypothesise a mechanistic interaction between keratins and metabolism is governed through regulation of post-translational modifications and may contribute significantly to the normal functioning of the colon, placing keratins at the centre of a nutrition-metabolism-health triangle.


Assuntos
Colo , Queratinas , Reto , Colo/fisiologia , Neoplasias Colorretais , Humanos , Doenças Inflamatórias Intestinais , Mucosa Intestinal/fisiologia , Queratinas/fisiologia , Reto/fisiologia
2.
Mol Biol Cell ; 31(11): 1103-1111, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32213122

RESUMO

Mitochondria fulfill essential roles in ATP production, metabolic regulation, calcium signaling, generation of reactive oxygen species (ROS), and additional determinants of cellular health. Recent studies have highlighted a role for mitochondria during cell differentiation, including in skin epidermis. The observation of oxidative stress in keratinocytes from Krt16 null mouse skin, a model for pachyonychia congenita (PC)-associated palmoplantar keratoderma, prompted us to examine the role of Keratin (K) 16 protein and its partner K6 in regulating the structure and function of mitochondria. Electron microscopy revealed major anomalies in mitochondrial ultrastructure in late stage, E18.5, Krt6a/Krt6b null embryonic mouse skin. Follow-up studies utilizing biochemical, metabolic, and live imaging readouts showed that, relative to controls, skin keratinocytes null for Krt6a/Krt6b or Krt16 exhibit elevated ROS, reduced mitochondrial respiration, intracellular distribution differences, and altered movement of mitochondria within the cell. These findings highlight a novel role for K6 and K16 in regulating mitochondrial morphology, dynamics, and function and shed new light on the causes of oxidative stress observed in PC and related keratin-based skin disorders.


Assuntos
Queratinas/metabolismo , Mitocôndrias/metabolismo , Pele/metabolismo , Animais , Proteínas do Citoesqueleto , Epiderme , Feminino , Queratina-16/genética , Queratina-16/metabolismo , Queratina-6/genética , Queratina-6/metabolismo , Queratinócitos/metabolismo , Queratinócitos/fisiologia , Queratinas/fisiologia , Ceratodermia Palmar e Plantar , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/fisiologia , Mutação , Paquioníquia Congênita
3.
Skin Res Technol ; 26(3): 422-430, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31802538

RESUMO

BACKGROUND: Nowadays, nail care products are extremely important both in medical and cosmetic fields. Actually, there are only a very few "in vivo" methods to evaluate the safety and the efficacy of nail products. METHODS: The new apparatus, based on a recently patented technology, is developed for the "in vivo" evaluation of nails in terms of thickness, structural firmness, flattening, and bending properties. The device analyzes nails by an "in vivo" non-invasive methodology in a timely way and with high accuracy. The assessment of the resistance to compression measures the cohesion of the nail matrix (nail firmness), while the evaluation of the resistance to transversal deformation detects the elasticity of the nail plate. Furthermore, the apparatus is able to assess the nail thickness and the flexibility of their distal edge. RESULTS: The instrument provides nail thickness and several parameters reflecting mechanical properties of nail plate: Viscoelasticity expressed as viscoelasticity index (VI), structural strength/ firmness expressed as Firmness Index (FI), and viscoelasticity of the distal edge expressed as Bending Index (BI). CONCLUSIONS: The instruments described in this work represent an innovative apparatus for the safety and efficacy evaluation of nail products in several fields: cosmetics, pharmaceuticals, and medical devices.


Assuntos
Fenômenos Biomecânicos/fisiologia , Doenças da Unha/psicologia , Unhas/anatomia & histologia , Adulto , Cosméticos , Elasticidade/fisiologia , Matriz Extracelular , Feminino , Humanos , Queratinas/metabolismo , Queratinas/fisiologia , Pessoa de Meia-Idade , Doenças da Unha/epidemiologia , Patentes como Assunto
4.
Skin Res Technol ; 25(3): 258-269, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30345567

RESUMO

BACKGROUND: Natures and amounts of water retained at the surface of stratum corneum (SC) of human skins, affecting skin health and penetration of chemical components, remain unclear. METHODS: A keratin film, a main component of human SC surface, was measured by IR micro-spectroscopy combined with a quartz crystal microbalance (QCM) and a relative humidity (RH) control system. RESULTS: Water contents increased with RH up to about 19 wt% and were correlated linearly with the OH + NH band areas in IR spectra of the keratin film. The OH + NH band areas for the triple helix collagen film are about twice as large as those for the keratin film (double helix). The free water component increases with RH by keeping the bound water component minor for the keratin film. About twice of water retention capacity of the collagen film can be due to increasing adsorption of free water, interacting possibly with hydrophobic aliphatic CH surfaces. CONCLUSION: The present results suggest relatively low water contents less than about 19 wt% of outermost SC layers of human skin composed mostly of keratin exposed to ambient RH conditions. The triple helix collagen can be used as an effective moisturizing agent.


Assuntos
Colágeno/fisiologia , Epiderme/fisiologia , Queratinas/fisiologia , Fenômenos Fisiológicos da Pele , Água/química , Adsorção , Água Corporal/fisiologia , Colágeno/química , Epiderme/química , Humanos , Umidade , Queratinas/química , Espectrofotometria Infravermelho
5.
J Craniomaxillofac Surg ; 47(1): 80-86, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30503606

RESUMO

Bone morphogenetic protein 6 (BMP6) has unique properties regarding structure and function in supporting bone formation during development and adult life. Despite its known role in various malignant tumors, the prognostic significance of BMP6 expression in oral squamous cell carcinoma (OSCC) remains unknown. The aim of the study was to investigate immunohistochemical expression of BMP6 in OSCC in correlation with clinical and pathological parameters, disease recurrence and survival. In addition, we investigated other parameters in order to identify prognosticators of neck metastases and final outcome. The study included 120 patients with clinically T1-3N0 OSCC who were primarily surgically treated between 2003 and 2008. There were 99 (82.5%) male and 21 (17.5%) female patients. The five-year disease-specific survival for the whole cohort was 79.7%. Tumors smaller than 2 cm in diameter showed higher incidence of strong BMP6 expression. No statistical correlation was observed between other clinico-pathological factors and BMP6 expression. Expression of BMP6 was not associated with disease recurrence and survival. BMP6 may not serve as prognosticator of final outcome or recurrence in clinically node-negative OSCC subjects. In multivariate analysis predictors of poorer survival were positive surgical margin, moderate tumor cell differentiation and pathological involvement of levels IV and/or V.


Assuntos
Proteína Morfogenética Óssea 6/metabolismo , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Neoplasias Bucais/metabolismo , Neoplasias Bucais/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Neoplasias de Cabeça e Pescoço , Humanos , Imuno-Histoquímica , Queratinas/fisiologia , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Metástase Neoplásica , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/patologia , Prognóstico
6.
J Periodontal Res ; 53(5): 793-800, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29858875

RESUMO

BACKGROUND: This study aims to evaluate the use of the leukocyte- and platelet-rich fibrin (L-PRF) membranes in increasing the width of the keratinized mucosa (KM) around implants. MATERIAL AND METHODS: Eight patients in need for bilateral widening of the KM around implants in the lower jaw were recruited for a spit-mouth randomized controlled trial. At the control site, a free gingival graft (FGG) was used, whereas at the other side (test), L-PRF membranes were applied. The primary outcome was the increase in width of KM around the implants. As secondary outcomes, the postoperative pain and surgical time were assessed. The follow-up period was 6 weeks. RESULTS: A significant increase in the total bucco-lingual width of KM in both groups was observed, with 1.3 mm ± 0.9 extra gain (P < .05) for the FGG sites. Shrinkage of the widened areas in this period was 32.1% at the test site and 23.6% at the control site. All values of the postoperative pain scores at the control site were higher than at the test site. The mean surgery time in the test and control group was 29.1 ± 4.8 and 48.1 ± 7.7 minutes, respectively. CONCLUSION: Within the limitations of this randomized controlled trial with split mouth design, it can be concluded that L-PRF can increase the width of KM around implants. Furthermore, the use of L-PRF results in a lower surgical time with less postoperative discomfort and pain for the patients in comparison to the FGG.


Assuntos
Implantes Dentários , Retração Gengival/cirurgia , Gengivoplastia/métodos , Fibrina Rica em Plaquetas , Feminino , Humanos , Queratinas/fisiologia , Leucócitos , Masculino , Mandíbula , Pessoa de Meia-Idade , Duração da Cirurgia , Dor Pós-Operatória , Projetos Piloto , Resultado do Tratamento
7.
J Dermatol Sci ; 89(3): 241-247, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29198699

RESUMO

BACKGROUND: Mechanical stress is an ubiquitous challenge of human cells with fundamental impact on cell physiology. Previous studies have shown that stretching promotes signalling cascades involved in proliferation and tissue enlargement. OBJECTIVE: The present study is dedicated to learn more about cellular structures contributing to perception and signal transmission of cell stretch. In particular, we hypothesized that desmosmal contacts and the adjacent keratin filament build an intercellular matrix providing information about the mechanical load. METHODS: Epidermal cells with different keratin equipment were seeded on flexible silicon dishes and stretched. As read out parameter the activation of PKB/Akt and p44/42 was monitored by Western blotting. Likewise desomosomal contacts were manipulated by depletion or addition of calcium. Moreover, desmoglein 3 and desmocollin 3 were blocked by either specific antibodies or siRNA. RESULTS: It was found that the omission of calcium from the medium, a necessary cofactor for desmosomal cadherins, inhibited stretch mediated activation of PKB/Akt and p44/42. The relevance of desmosomes in this context was further substantiated by experiments using a desmoglein 3 blocking antibody (AK23) and siRNA against desmocollin 3. Moreover, disruption of the keratin filament by sodium orthovanadate also abrogates PKB/Akt and p44/42 activation in response to stretch. Likewise, KEB-7 keratinocytes harbouring a mutation in the keratin 14 gene and genetically modified keratinocytes devoid of any keratin show an altered signalling after stretch indicating the relevance of the keratin filament in this context. CONCLUSION: Besides their important role in cell architecture our results identify desmosomes and keratins as mechanosensing structures.


Assuntos
Desmossomos/fisiologia , Queratinas/fisiologia , Proteína Quinase 1 Ativada por Mitógeno/fisiologia , Proteína Quinase 3 Ativada por Mitógeno/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Cálcio/fisiologia , Células Cultivadas , Desmogleína 3/fisiologia , Ativação Enzimática , Humanos , Estresse Mecânico
8.
Proc Natl Acad Sci U S A ; 115(2): E218-E225, 2018 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-29279398

RESUMO

Hybrid speciation is rare in vertebrates, and reproductive isolation arising from hybridization is infrequently demonstrated. Here, we present evidence supporting a hybrid-speciation event involving the genetic admixture of the snow-capped (Lepidothrix nattereri) and opal-crowned (Lepidothrix iris) manakins of the Amazon basin, leading to the formation of the hybrid species, the golden-crowned manakin (Lepidothrix vilasboasi). We used a genome-wide SNP dataset together with analysis of admixture, population structure, and coalescent modeling to demonstrate that the golden-crowned manakin is genetically an admixture of these species and does not represent a hybrid zone but instead formed through ancient genetic admixture. We used spectrophotometry to quantify the coloration of the species-specific male crown patches. Crown patches are highly reflective white (snow-capped manakin) or iridescent whitish-blue to pink (opal-crowned manakin) in parental species but are a much less reflective yellow in the hybrid species. The brilliant coloration of the parental species results from nanostructural organization of the keratin matrix feather barbs of the crown. However, using electron microscopy, we demonstrate that the structural organization of this matrix is different in the two parental species and that the hybrid species is intermediate. The intermediate nature of the crown barbs, resulting from past admixture appears to have rendered a duller structural coloration. To compensate for reduced brightness, selection apparently resulted in extensive thickening of the carotenoid-laden barb cortex, producing the yellow crown coloration. The evolution of this unique crown-color signal likely culminated in premating isolation of the hybrid species from both parental species.


Assuntos
Especiação Genética , Hibridização Genética , Passeriformes/genética , Distribuição Animal , Animais , Carotenoides/metabolismo , Plumas/fisiologia , Feminino , Estudo de Associação Genômica Ampla , Queratinas/fisiologia , Masculino , Passeriformes/fisiologia , Polimorfismo de Nucleotídeo Único , Caracteres Sexuais , África do Sul , América do Sul
9.
J Mol Histol ; 49(1): 1-15, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29181608

RESUMO

Embryonic development of the mouse salivary glands begins with epithelial thickening and continues with sequential changes from the pre-bud to terminal bud stages. After birth, morphogenesis proceeds, and the glands develop into a highly branched epithelial structure that terminates with saliva-producing acinar cells at the adult stage. Acinar cells derived from the epithelium are differentiated into serous, mucous, and seromucous types. During differentiation, cytokeratins, intermediate filaments found in most epithelial cells, play vital roles. Although the localization patterns and developmental roles of cytokeratins in different epithelial organs, including the mammary glands, circumvallate papilla, and sweat glands, have been well studied, their stage-specific localization and morphogenetic roles during salivary gland development have yet to be elucidated. Therefore, the aim of this study was to determine the stage and acinar cell type-specific localization pattern of cytokeratins 4, 5, 7, 8, 13, 14, 18, and 19 in the major salivary glands (submandibular, sublingual, and parotid glands) of the mouse at the E15.5, PN0, PN10, and adult stages. In addition, cell physiology, including cell proliferation, was examined during development via immunostaining for Ki67 to understand the cellular mechanisms that govern acinar cell differentiation during salivary gland morphogenesis. The distinct localization patterns of cytokeratins in conjunction with cell physiology will reveal the roles of epithelial cells in salivary gland formation during the differentiation of serous, mucous or seromucous salivary glands.


Assuntos
Células Acinares/citologia , Queratinas/metabolismo , Glândulas Salivares/citologia , Animais , Diferenciação Celular , Células Epiteliais , Queratinas/fisiologia , Camundongos , Organogênese , Glândulas Salivares/embriologia , Glândulas Salivares/crescimento & desenvolvimento
10.
Development ; 144(23): 4363-4376, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-28982683

RESUMO

The coordination of individual cell behaviors is a crucial step in the assembly and morphogenesis of tissues. Xenopus mesendoderm cells migrate collectively along a fibronectin (FN) substrate at gastrulation, but how the adhesive and mechanical forces required for these movements are generated and transmitted is unclear. Traction force microscopy (TFM) was used to establish that traction stresses are limited primarily to leading edge cells in mesendoderm explants, and that these forces are balanced by intercellular stresses in follower rows. This is further reflected in the morphology of these cells, with broad lamellipodial protrusions, mature focal adhesions and a gradient of activated Rac1 evident at the leading edge, while small protrusions, rapid turnover of immature focal adhesions and lack of a Rac1 activity gradient characterize cells in following rows. Depletion of keratin (krt8) with antisense morpholinos results in high traction stresses in follower row cells, misdirected protrusions and the formation of actin stress fibers anchored in streak-like focal adhesions. We propose that maintenance of mechanical integrity in the mesendoderm by keratin intermediate filaments is required to balance stresses within the tissue to regulate collective cell movements.


Assuntos
Gastrulação/fisiologia , Queratinas/fisiologia , Proteínas de Xenopus/fisiologia , Xenopus/embriologia , Xenopus/fisiologia , Actinas/fisiologia , Animais , Fenômenos Biomecânicos , Miosinas Cardíacas/antagonistas & inibidores , Miosinas Cardíacas/metabolismo , Movimento Celular/fisiologia , Endoderma/citologia , Endoderma/embriologia , Endoderma/fisiologia , Adesões Focais/fisiologia , Técnicas de Silenciamento de Genes , Filamentos Intermediários/fisiologia , Queratina-8/antagonistas & inibidores , Queratina-8/genética , Queratina-8/fisiologia , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/fisiologia , Modelos Biológicos , Morfogênese/fisiologia , Cadeias Leves de Miosina/antagonistas & inibidores , Cadeias Leves de Miosina/metabolismo , Transdução de Sinais , Estresse Mecânico , Xenopus/genética , Proteínas de Xenopus/antagonistas & inibidores , Proteínas de Xenopus/genética , Proteínas rac1 de Ligação ao GTP/antagonistas & inibidores , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/fisiologia
11.
Artigo em Inglês | MEDLINE | ID: mdl-28572456

RESUMO

SUMMARYThe establishment and continuous cell type-specific adaptation of cytoplasmic intermediate filament (IF) networks are linked to various types of IF motility. Motor protein-driven active transport, linkage to other cellular structures, diffusion of small soluble subunits, and intrinsic network elasticity all contribute to the motile behavior of IFs. These processes are subject to regulation by multiple signaling pathways. IF motility is thereby connected to and involved in many basic cellular processes guarding the maintenance of cell and tissue integrity. Disturbances of IF motility are linked to diseases that are characterized by cytoplasmic aggregates containing IF proteins together with other cellular components.


Assuntos
Movimento Celular/fisiologia , Proteínas de Filamentos Intermediários/fisiologia , Filamentos Intermediários/fisiologia , Citoesqueleto de Actina/fisiologia , Actinas/fisiologia , Animais , Axônios/fisiologia , Linhagem Celular Tumoral , Citoplasma/fisiologia , Difusão , Proteínas de Fluorescência Verde/fisiologia , Homeostase , Humanos , Queratinas/fisiologia , Microscopia Confocal , Microtúbulos/fisiologia , Processamento de Proteína Pós-Traducional , Transdução de Sinais , Solubilidade
12.
Int J Oral Maxillofac Surg ; 46(6): 774-781, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28292550

RESUMO

The aim of this overview was to assess the methods, quality, and outcomes of systematic reviews conducted to evaluate the importance of keratinized mucosa (KM) for the maintenance of peri-implant tissue health in humans. An electronic search was conducted without date or language restriction using the MEDLINE/PubMed, Cochrane Library, Web of Science, and Embase databases up to December 2015. The eligibility criteria included systematic reviews with/without meta-analysis and with a focus on the influence of KM on peri-implant health around implants. Two independent authors performed the quality analysis of the reviews with the AMSTAR guidelines and another checklist proposed in 2003. After screening, four systematic reviews were selected. The present study demonstrated the existence of structural and methodological variability among the systematic reviews with/without meta-analysis. None of the systematic reviews that were included in the study obtained the maximum score in the two quality analyses performed. All systematic reviews included reported a positive association between an adequate KM width (≥2mm) and peri-implant health. There is still insufficient data on the long-term survival and success rates of dental implants. Prospective studies evaluating the importance of KM for the long-term maintenance of dental implants are needed.


Assuntos
Implantes Dentários , Queratinas/fisiologia , Mucosa Bucal/fisiologia , Projetos de Pesquisa , Falha de Restauração Dentária , Gengiva/fisiologia , Humanos
13.
Methods Enzymol ; 569: 261-85, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26778563

RESUMO

Epiplakin is a large member (>700 kDa) of the plakin protein family and exclusively expressed in epithelial cell types. Compared to other plakin proteins epiplakin exhibits an unusual structure as it consists entirely of a variable number of consecutive plakin repeat domains (13 in humans, 16 in mice). The only binding partners of epiplakin identified so far are keratins of simple as well as of stratified epithelia. Epiplakin-deficient mice show no obvious spontaneous phenotype. However, ex vivo studies using epiplakin-deficient primary cells indicated protective functions of epiplakin in response to stress. Recent studies using stress models for organs of the gastrointestinal tract revealed that epiplakin-deficient mice develop more pronounced pancreas and liver injuries than their wild-type littermates. In addition, impaired stress-induced keratin network reorganization was observed in the affected organs, and primary epiplakin-deficient hepatocytes showed reduced tolerance for forced keratin overexpression which could be rescued by a chemical chaperone. These findings indicate protective functions of epiplakin in chaperoning disease-induced keratin reorganization. In this review, we describe some of the methods we used to analyze epiplakin's function with the focus on biochemical and ex vivo techniques.


Assuntos
Autoantígenos/fisiologia , Células Epiteliais/metabolismo , Animais , Modelos Animais de Doenças , Expressão Gênica , Hepatócitos/metabolismo , Humanos , Queratinas/fisiologia , Fígado/metabolismo , Fígado/patologia , Camundongos Knockout , Cultura Primária de Células
14.
Clin Exp Dermatol ; 41(4): 410-2, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-26685052

RESUMO

We present a case of a patient with long-standing hyperpigmented macules and erythematous papules over his chest, abdomen, back and arms, suggestive of Dowling-Degos disease (DDD). In addition, there were hyperkeratotic papules, alternating red and white nail-bed discolouration, and V-shaped nail notching consistent with Darier disease (DD). Histology showed findings consistent with DDD and DD on separate specimens. The lack of acantholysis in areas of filiform hyperpigmented rete ridges ruled out Galli-Galli disease (GGD). DDD results from mutations in the genes encoding keratin 5 (KRT5), protein O-glucosyltransferase 1 (POGLUT1) or protein O-fucosyltransferase 1 (POFUT1), while DD results from mutations in the ATP2A2 gene. Both genes are present on chromosome 12. In this case, the patient presented with features of both DDD and DD, which suggests that either a cooperating mutation or a mutation in an unrelated gene locus may underlie the findings in this patient.


Assuntos
Doença de Darier/complicações , Doença de Darier/diagnóstico , Doença de Darier/patologia , Hiperpigmentação/complicações , Hiperpigmentação/diagnóstico , Hiperpigmentação/patologia , Dermatopatias Genéticas/complicações , Dermatopatias Genéticas/diagnóstico , Dermatopatias Genéticas/patologia , Dermatopatias Papuloescamosas/complicações , Dermatopatias Papuloescamosas/diagnóstico , Dermatopatias Papuloescamosas/patologia , Acantólise/etiologia , Acantólise/patologia , Erupções Acneiformes/patologia , Cromossomos Humanos Par 12/genética , Doença de Darier/genética , Humanos , Hiperpigmentação/genética , Queratinas/genética , Queratinas/fisiologia , Masculino , Pessoa de Meia-Idade , Mutação , Doenças da Unha , Linhagem , Dermatopatias Genéticas/genética , Dermatopatias Papuloescamosas/genética
15.
Int Endod J ; 49(5): 471-82, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26016886

RESUMO

AIM: To fabricate a keratin hydrogel, characterize its functionality as a biomaterial and investigate the effects of keratin on growth and differentiation of odontoblast-like cells. METHODOLOGY: Keratins were extracted from sheep wool using a well-established technique. The extracted proteins were purified by dialysis, quantified by gel electrophoresis, mass spectrometry, amino acid analysis and inductively coupled mass spectrometry. The microstructure of the fabricated keratin hydrogels was studied by scanning electron microscopy, flow characteristics by rheometer, hydrolytic stability and cytocompatibility by Live/Dead(®) cell assay. Furthermore, the influence of keratin on odontoblast-like cells (MDPC-23) was assessed to confirm their bioactivity at different dilutions. Cell proliferation was studied using alamarBlue(®) assay and differentiation by alkaline phosphatase enzyme activity, alizarin red staining and calcium quantification, reverse transcription polymerase chain reaction (rt-PCR) and immunocytochemical staining for dentine matrix protein- 1 (DMP-1) expression. anova with Tukey's tests was performed for statistical comparison. RESULTS: The characterized hydrogel was injectable with a highly porous architecture that underwent slow degradation, and its cytocompatibility was statistically equivalent to collagen hydrogel (P > 0.05). Cell proliferation and differentiation were enhanced at the optimal keratin concentration of 0.1 mg mL(-1) . At this concentration, the influence of keratin on cell differentiation was demonstrated by marked elevation in alkaline phosphatase activity (P < 0.05), calcium deposition (P < 0.01), gene expression (P < 0.01) and positive immunostaining for DMP-1. CONCLUSION: The presence of keratin enhanced odontoblast cell behaviour. Keratin hydrogels may be a potential scaffold for pulp-dentine regen-eration.


Assuntos
Diferenciação Celular , Queratinas/fisiologia , Odontoblastos/fisiologia , Animais , Proliferação de Células , Polpa Dentária/citologia , Ovinos ,
16.
Carcinogenesis ; 36(11): 1372-80, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26271098

RESUMO

Solar ultraviolet irradiation is an environmental carcinogen that causes skin cancer. Caspase-7 is reportedly expressed at reduced levels in many cancers. The present study was designed to examine the role of caspase-7 in solar-simulated light (SSL)-induced skin cancer and to elucidate its underlying molecular mechanisms. Our study revealed that mice with genetic deficiency of caspase-7 are highly susceptible to SSL-induced skin carcinogenesis. Epidermal hyperplasia, tumor volume and the average number of tumors were significantly increased in caspase-7 knockout (KO) mice compared with SKH1 wild-type mice irradiated with SSL. The expression of cell proliferation markers, such as survivin and Ki-67, was elevated in SSL-irradiated skin of caspase-7 KO mice compared with those observed in SSL-exposed wild-type SKH1 mouse skin. Moreover, SSL-induced apoptosis was abolished in skin from caspase-7 KO mice. Two-dimensional gel electrophoresis, followed by matrix-assisted laser desorption/ionization-time-of-flight analysis of skin tissue lysates from SSL-irradiated SKH1 wild-type and caspase-7 KO mice revealed an aberrant induction of keratin-17 in caspase-7 KO mice. Immunohistochemical analysis of skin tumors also showed an increase of keratin-17 expression in caspase-7 KO mice compared with SKH1 wild-type mice. The expression of keratin-17 was also elevated in SSL-irradiated caspase-7 KO keratinocytes as well as in human basal cell carcinomas. The in vitro caspase activity assay showed keratin-17 as a substrate of caspase-7, but not caspase-3. Overall, our study demonstrates that genetic loss of caspase-7 promotes SSL-induced skin carcinogenesis by blocking caspase-7-mediated cleavage of keratin-17.


Assuntos
Carcinoma de Células Escamosas/enzimologia , Caspase 7/genética , Queratinas/fisiologia , Lesões Experimentais por Radiação/enzimologia , Neoplasias Cutâneas/enzimologia , Luz Solar/efeitos adversos , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Caspase 7/metabolismo , Células Cultivadas , Epiderme/enzimologia , Epiderme/patologia , Epiderme/efeitos da radiação , Feminino , Técnicas de Inativação de Genes , Queratinócitos/enzimologia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteólise , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Carga Tumoral
17.
PLoS One ; 10(3): e0121090, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25822661

RESUMO

Keratin are among the most abundant proteins in epithelial cells. Functions of the keratin network in cells are shaped by their dynamical organization. Using a collection of experimentally-driven mathematical models, different hypotheses for the turnover and transport of the keratin material in epithelial cells are tested. The interplay between turnover and transport and their effects on the keratin organization in cells are hence investigated by combining mathematical modeling and experimental data. Amongst the collection of mathematical models considered, a best model strongly supported by experimental data is identified. Fundamental to this approach is the fact that optimal parameter values associated with the best fit for each model are established. The best candidate among the best fits is characterized by the disassembly of the assembled keratin material in the perinuclear region and an active transport of the assembled keratin. Our study shows that an active transport of the assembled keratin is required to explain the experimentally observed keratin organization.


Assuntos
Células Epiteliais/metabolismo , Queratinas/metabolismo , Queratinas/fisiologia , Modelos Biológicos , Fluorescência , Humanos , Transporte Proteico/fisiologia , Imagem com Lapso de Tempo
18.
Anat Rec (Hoboken) ; 298(4): 703-19, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25737029

RESUMO

Mysticetes have evolved a novel filter feeding apparatus-baleen-an epidermal keratinous tissue composed of keratin that grows as a serial arrangement of transverse cornified laminae from the right and left sides of the palate. The structure and function of baleen varies among extant mysticete clades and this variation likely can be viewed as adaptations related to different filter feeding strategies. In one of the first morphometric studies of the full baleen apparatus, we describe the morphology of complete baleen racks in neonate, yearling and adult gray whales (Eschrichtius robustus), and note morphometric variations between age groups as well as within individual racks. Morphometric data and detailed descriptions were collected from the full baleen apparatus of three frozen specimens of E. robustus using previously derived ecologically significant and broad scale measurements of baleen. Additionally, characters of the baleen apparatus were described based on visible patterns of baleen laminae and plates on the dorsal root of the rack. Results indicate that the longest, widest, and thickest plates and laminae are found toward the posterior half of the rack, resulting in the greatest surface area for filtration of prey occurring in this region. Ontogenetic changes were also documented that reveal a progressive increase in the filter surface area of the developing baleen apparatus as baleen laminae and main plates grow in length and width. Also noted was a progressive posterior shift in the position of greatest filtration area. Histological examination of the epithelial base (Zwischensubstanz) and laminae showed basic epidermal layers, as well as gapping between layers and vacuoles.


Assuntos
Queratinas/fisiologia , Palato/anatomia & histologia , Baleias/anatomia & histologia , Animais , Animais Recém-Nascidos , Feminino , Oceano Pacífico
20.
BMC Ophthalmol ; 15 Suppl 1: 156, 2015 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-26817690

RESUMO

Meibomian gland dysfunction (MGD) is the major cause of evaporative dry eye disease (EDED) and dysfunction is widely thought to mechanistically involve ductal hyperkeratinization, plugging and obstruction. This review re-evaluates the role of hyperkeratinization in MGD based on more recent findings from mouse models. In these studies, eyelids from normal young and old mice or mice exposed to desiccating stress were evaluated by immunofluorescent tomography and 3-dimensional reconstruction to evaluate gland volume, expression of hyperkeratinization markers and cell proliferation or stimulated Raman scattering (SRS) microscopy to assess lipid quality. Results indicate that aging mice show dropout of meibomian glands with loss of gland volume and a forward migration of the mucocutaneous junction anterior to the gland orifice; similar age-related changes that are detected in human subjects. Atrophic glands also showed evidence of epithelial plugging of the orifice without the presence of hyperkeratinization. Mice exposed to desiccating stress showed hyperproliferation of the meibomian gland and ductal dilation suggesting a marked increase in lipid synthesis. Lipid quality was also affected in EDED mice with an increase in the protein content of lipid within the duct of the gland. Overall, age-related changes in the mouse show similar structural and functional correlates with that observed in clinical MGD without evidence of hyperkeratinization suggesting that gland atrophy may be a major cause of EDED. The response of the meibomian gland to desiccating stress also suggest that environmental conditions may accelerate or potentiate age-related changes.


Assuntos
Proteínas do Olho/fisiologia , Doenças Palpebrais/fisiopatologia , Queratinas/fisiologia , Glândulas Tarsais/fisiopatologia , Animais , Atrofia/patologia , Síndromes do Olho Seco/metabolismo , Doenças Palpebrais/patologia , Humanos , Glândulas Tarsais/patologia , Camundongos
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