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1.
DNA Cell Biol ; 39(4): 690-699, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32027181

RESUMO

The aim of this study was to identify genes with clinical significance in colorectal cancer (CRC). Gene expression profiles of 585 CRC tissues and 61 normal colorectal tissues from Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases were used to identify differentially expressed genes (DEGs) between CRC and normal colorectal tissues. DAVID and KOBAS tools were used to explore Gene Ontology (GO) and KEGG pathways enriched by DEGs, respectively. In addition, TCGA data sets were also used to identify prognostic factors and develop a prognostic prediction model for CRC. A total of 353 DEGs including 117 upregulated and 236 downregulated genes in CRC were identified based on GSE32323 data set. These DEGs were significantly enriched in the biological process related to the regulation of cell proliferation and 50 signaling pathways, such as "TGF-beta signaling pathway," "Wnt signaling pathway," and "Jak-STAT signaling pathway." GCG, ADH1B, SLC4A4, ZG16, and CLCA4 were the top five downregulated in CRC. FOXQ1, LGR5, CLDN1, KRT23, and DPEP1 were the top five upregulated in CRC. KRT23 expression could affect tumor stage and regional lymph node metastasis in CRC patients. FOXQ1 expression could affect tumor distant metastasis in CRC patients. Survival analysis indicated that SLC4A4 expression was associated with the prognosis of CRC patients. Prognostic prediction model developed based on age, tumor stage, and SLC4A4 expression exhibited an efficient performance in predicting 1-, 3-, and 5-year overall survival of CRC patients. In conclusion, the current study identified several genes and pathways related to CRC, which provided new insight in understanding molecular mechanism of tumorigenesis and development of CRC.


Assuntos
Biomarcadores Tumorais/genética , Transformação Celular Neoplásica/genética , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica/genética , Transcriptoma/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Biologia Computacional , Bases de Dados Genéticas , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Perfilação da Expressão Gênica , Humanos , Queratinas Tipo I/genética , Queratinas Tipo I/metabolismo , Pessoa de Meia-Idade , Prognóstico , Simportadores de Sódio-Bicarbonato/genética , Simportadores de Sódio-Bicarbonato/metabolismo
2.
BMC Cancer ; 19(1): 824, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31429720

RESUMO

BACKGROUND: Genomic initiatives such as The Cancer Genome Atlas (TCGA) contain data from -omics profiling of thousands of tumor samples, which may be used to decipher cancer signaling, and related alterations. Managing and analyzing data from large-scale projects, such as TCGA, is a demanding task. It is difficult to dissect the high complexity hidden in genomic data and to account for inter-tumor heterogeneity adequately. METHODS: In this study, we used a robust statistical framework along with the integration of diverse bioinformatic tools to analyze next-generation sequencing data from more than 1000 patients from two different lung cancer subtypes, i.e., the lung adenocarcinoma (LUAD) and the squamous cell carcinoma (LUSC). RESULTS: We used the gene expression data to identify co-expression modules and differentially expressed genes to discriminate between LUAD and LUSC. We identified a group of genes which could act as specific oncogenes or tumor suppressor genes in one of the two lung cancer types, along with two dual role genes. Our results have been validated against other transcriptomics data of lung cancer patients. CONCLUSIONS: Our integrative approach allowed us to identify two key features: a substantial up-regulation of genes involved in O-glycosylation of mucins in LUAD, and a compromised immune response in LUSC. The immune-profile associated with LUSC might be linked to the activation of three oncogenic pathways, which promote the evasion of the antitumor immune response. Collectively, our results provide new future directions for the design of target therapies in lung cancer.


Assuntos
Adenocarcinoma de Pulmão/genética , Carcinoma de Células Escamosas/genética , Neoplasias Pulmonares/genética , Glicoproteínas de Membrana/genética , Mucina-5B/genética , Mucinas/genética , Adenocarcinoma de Pulmão/imunologia , Carcinoma de Células Escamosas/imunologia , Estudos de Coortes , Bases de Dados de Ácidos Nucleicos , Regulação Neoplásica da Expressão Gênica , Glicosilação , Humanos , Interleucina-6/genética , Queratinas Tipo I/genética , Neoplasias Pulmonares/imunologia , Glicoproteínas de Membrana/metabolismo , Mucina-5B/metabolismo , Mucinas/metabolismo , Família Multigênica/genética , Modelos de Riscos Proporcionais , Transcriptoma , Microambiente Tumoral/imunologia
3.
Cells ; 8(6)2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31216713

RESUMO

Keratin proteins form intermediate filaments, which provide structural support for many tissues. Multiple keratin family members are reported to be associated with the progression of liver disease of multiple etiologies. For example, keratin 23 (KRT23) was reported as a stress-inducible protein, whose expression levels correlate with the severity of liver disease. Hepatitis C virus (HCV) is a human pathogen that causes chronic liver diseases including fibrosis, cirrhosis, and hepatocellular carcinoma. However, a link between KRT23 and hepatitis C virus (HCV) infection has not been reported previously. In this study, we investigated KRT23 mRNA levels in datasets from liver biopsies of chronic hepatitis C (CHC) patients and in primary human hepatocytes experimentally infected with HCV, in addition to hepatoma cells. Interestingly, in each of these specimens, we observed an HCV-dependent increase of mRNA levels. Importantly, the KRT23 protein levels in patient plasma decreased upon viral clearance. Ectopic expression of KRT23 enhanced HCV infection; however, CRIPSPR/Cas9-mediated knockout did not show altered replication efficiency. Taken together, our study identifies KRT23 as a novel, virus-induced host-factor for hepatitis C virus.


Assuntos
Hepatite C/metabolismo , Fatores Celulares Derivados do Hospedeiro/metabolismo , Queratinas Tipo I/metabolismo , Carcinoma Hepatocelular/genética , Linhagem Celular , Células HEK293 , Hepacivirus/genética , Hepacivirus/metabolismo , Hepatite C/genética , Hepatite C/fisiopatologia , Hepatite C Crônica/metabolismo , Hepatócitos/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Queratinas/metabolismo , Queratinas Tipo I/genética , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/metabolismo , Neoplasias Hepáticas/genética , RNA Mensageiro/metabolismo , Transcriptoma/genética , Replicação Viral
4.
Hum Genomics ; 12(1): 27, 2018 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-29784039

RESUMO

BACKGROUND: Mutations in keratin proteins have been vastly associated with a wide array of genodermatoses; however, mutations of keratins in psoriasis have not been fully investigated. The main aim of the current research was to identify the mutation in K14, K10, K16, and K17 genes in two stages of psoriasis patients. METHODS: Ninety-six psoriatic skin biopsies were collected. mRNA transcript of K14, K10, K16, and K17 was prepared, amplified, and sequenced. Sanger sequences of all keratins were further validated for mutational analysis using Mutation Surveyor and Alamut Visual. Then, in silico analysis of protein stability and protein and gene expression of all keratins was performed and validated. RESULTS: Out of 44 mutations, about 75% of keratins are highly pathogenic and deleterious. Remaining 25% mutations are less pathogenic and tolerated in nature. In these 33 deleterious mutations were immensely found to decrease keratin protein stability. We also found a correlation between keratin and Psoriasis Area and Severity Index score which added that alteration in keratin gene in skin causes severity of psoriasis. CONCLUSIONS: We strongly concluded that acanthosis and abnormal terminal differentiation was mainly due to the mutation in epidermal keratins. In turn, disease severity and relapsing of psoriasis are mainly due to the mutation of hyperproliferative keratins. These novel keratin mutations in psoriatic epidermis might be one of the causative factors for psoriasis.


Assuntos
Queratinas Tipo I/genética , Queratinas/genética , Mutação/genética , Psoríase/genética , Acantose Nigricans/genética , Acantose Nigricans/fisiopatologia , Adolescente , Adulto , Idoso , Biópsia , Diferenciação Celular , Proliferação de Células/genética , Análise Mutacional de DNA , Epiderme/metabolismo , Epiderme/fisiopatologia , Feminino , Humanos , Queratinas/classificação , Masculino , Pessoa de Meia-Idade , Estabilidade Proteica , Psoríase/patologia , Índice de Gravidade de Doença , Pele/metabolismo , Pele/patologia , Adulto Jovem
5.
Artigo em Inglês | MEDLINE | ID: mdl-29610398

RESUMO

Keratins-types I and II-are the intermediate-filament-forming proteins expressed in epithelial cells. They are encoded by 54 evolutionarily conserved genes (28 type I, 26 type II) and regulated in a pairwise and tissue type-, differentiation-, and context-dependent manner. Here, we review how keratins serve multiple homeostatic and stress-triggered mechanical and nonmechanical functions, including maintenance of cellular integrity, regulation of cell growth and migration, and protection from apoptosis. These functions are tightly regulated by posttranslational modifications and keratin-associated proteins. Genetically determined alterations in keratin-coding sequences underlie highly penetrant and rare disorders whose pathophysiology reflects cell fragility or altered tissue homeostasis. Furthermore, keratin mutation or misregulation represents risk factors or genetic modifiers for several additional acute and chronic diseases.


Assuntos
Queratinas Tipo II/fisiologia , Queratinas Tipo I/fisiologia , Apoptose , Movimento Celular , Proliferação de Células , Homeostase , Queratinas Tipo I/genética , Queratinas Tipo II/genética , Processamento de Proteína Pós-Traducional , Estresse Fisiológico
7.
Pediatr Surg Int ; 34(2): 237-244, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29039047

RESUMO

AIMS AND OBJECTIVES: The use of autologous tissue-engineered skin substitutes is a promising approach to cover large skin defects in patients. Preclinical investigation is pivotal to test and improve the quality of these bio-engineered substitutes. In the skin, the epidermis, formed mainly by keratinocytes, provides the first physical barrier protecting from the environment. Proper keratinocyte differentiation and, thus, formation of a stratified epidermis is essential for this function. Keratins, the main structural support of keratinocytes, play a vital role regarding differentiation of keratinocytes. Here, we examined the expression pattern of a recently described keratinocyte differentiation marker, namely Keratin 24, in our skin substitutes. MATERIALS AND METHODS: Human epidermal keratinocytes, melanocytes, dermal fibroblasts, palmar fibroblasts or sweat gland cells were used to prepare skin substitutes. Fibroblast-containing collagen hydrogels were prepared, and keratinocytes or sweat gland cells and melanocytes were seeded onto the hydrogels. The generated tissue-engineered dermo-epidermal skin analogs were transplanted onto full-thickness skin wounds created on the back of immuno-incompetent rats. The skin substitutes were excised at different time points and histologically examined with regard to Keratin 24 expression. RESULTS: We observed the expression of Keratin 24 in keratinocytes of the upper stratum spinosum of the epidermis. In particular, we observed an intensified expression of Keratin 24 13 weeks after transplantation compared to 4 weeks after transplantation. Importantly, we noticed a markedly higher presence of Keratin 24 in more spinous layers if we used palmar fibroblasts or sweat gland cells in our skin substitutes compared non-palmar fibroblasts or epidermal keratinocytes. CONCLUSION: Our observations prove that the keratinocyte differentiation marker Keratin 24 is expressed in our dermo-epidermal skin substitutes in a normal pattern. This highlights that our bio-engineered skin analogs mature and reach homeostasis in an in vivo assay. These findings harbor favorable implications regarding future clinical application.


Assuntos
Derme/transplante , Epiderme/transplante , Regulação da Expressão Gênica , Queratinas Tipo I/genética , Transplante de Pele/métodos , Engenharia Tecidual/métodos , Ferimentos e Lesões/genética , Adolescente , Animais , Diferenciação Celular , Células Cultivadas , Criança , Pré-Escolar , Derme/citologia , Células Epidérmicas , Feminino , Humanos , Lactente , Queratinas Tipo I/biossíntese , Masculino , RNA/genética , Ratos , Pele/lesões , Pele Artificial , Ferimentos e Lesões/patologia , Ferimentos e Lesões/cirurgia
8.
Biotechniques ; 63(3): 131-134, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28911317

RESUMO

Biological evaluation of hair growth/differentiation activity in vitro has been a formidable challenge, primarily due to the lack of relevant model cell systems. To solve this problem, we generated a stable model cell line in which successive differentiation via epidermal progenitors to hair components is easily inducible and traceable. Mouse induced pluripotent stem (iPS) cell-derived cells were selected to stably express a tetracycline (Tet)-inducible bone morphogenic protein-4 (BMP4) expression cassette and a luciferase reporter driven by a hair-specific keratin 31 gene (krt31) promoter (Tet-BMP4-KRT31-Luc iPS). While Tet- BMP4-KRT31-Luc iPS cells could be maintained as stable iPS cells, the cells differentiated to produce luciferase luminescence in the presence of all-trans retinoic acid (RA) and doxycycline (Dox), and addition of a hair differentiation factor significantly increased luciferase fluorescence. Thus, this cell line may provide a reliable cell-based screening system to evaluate drug candidates for hair differentiation activity.


Assuntos
Alopecia/terapia , Diferenciação Celular , Engenharia Celular/métodos , Cabelo/citologia , Cabelo/crescimento & desenvolvimento , Células-Tronco Pluripotentes Induzidas/citologia , Animais , Proteína Morfogenética Óssea 4/genética , Proteína Morfogenética Óssea 4/metabolismo , Linhagem Celular , Doxiciclina/farmacologia , Avaliação Pré-Clínica de Medicamentos , Células-Tronco Pluripotentes Induzidas/metabolismo , Queratinas Específicas do Cabelo/genética , Queratinas Específicas do Cabelo/metabolismo , Queratinas Tipo I/genética , Queratinas Tipo I/metabolismo , Luciferases/metabolismo , Substâncias Luminescentes/metabolismo , Camundongos , Regiões Promotoras Genéticas , Tetraciclina/farmacologia , Tretinoína/farmacologia
9.
Cell Death Dis ; 8(7): e2961, 2017 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-28749462

RESUMO

The overexpression of human telomerase reverse transcriptase (hTERT) has been associated with the proliferation and migration of colorectal cancer (CRC) cells. We investigated the roles of KRT23 and hTERT in promoting CRC cell proliferation and migration. We verified the relationship between KRT23 and hTERT in CRC using streptavidin-agarose pulldown and chromatin immunoprecipitation (ChIP) assays. One hundred and fifty-four human CRC specimens were analyzed using immunohistochemistry. The roles of KRT23 and hTERT in cell growth and migration were studied using siRNA and lentiviruses in vivo and in vitro. Western blot and wound scratch analyses were used to determine the signaling pathway for KRT23-mediated activation of CRC growth and migration. Telomerase activity was measured by using the TeloTAGGG Telomerase PCR ELISA PLUS Kit. We identified KRT23 as a new hTERT promoter-binding protein. Patients with high KRT23 and hTERT expression had markedly shorter overall survival. Overexpression of KRT23 upregulated the expression of hTERT protein, hTERT promoter-driven luciferase and telomerase activity in CRC. Conversely, inhibition of KRT23 by a KRT23-specific siRNA repressed the endogenous hTERT protein, the expression of hTERT promoter-driven luciferase and telomerase activity. Overexpression of KRT23 also promoted CRC proliferation and migration. By contrast, KRT23 inhibition significantly inhibited tumor cell growth in vitro and in vivo. KRT23 promoted cancer stem cell properties and increased the expression of CD133 and CD44. These results demonstrate that KRT23 is an important cellular factor that promotes CRC growth by activating hTERT expression and that KRT23 is a potential novel therapeutic target for CRC.


Assuntos
Neoplasias Colorretais/enzimologia , Neoplasias Colorretais/metabolismo , Queratinas Tipo I/metabolismo , Telomerase/metabolismo , Animais , Western Blotting , Linhagem Celular Tumoral , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Imunoprecipitação da Cromatina , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Imuno-Histoquímica , Queratinas Tipo I/genética , Masculino , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sincalida/genética , Sincalida/metabolismo , Telomerase/genética
10.
PLoS One ; 12(3): e0174626, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28362807

RESUMO

Keratin 24 (K24) is a new kind of keratin genes, which encodes a novel keratin protein, K24 that bears high similarity to the type I keratins and displays a unique expression profile. However, the role of K24 is incompletely understood. In our study, we investigated the localization of K24 within the epidermis and possible functions. Keratin 24 was found to be modestly overexpressed in senescent keratinocytes and was mainly restricted to the upper stratum spinosum of epidermis. The protein was required for terminal differentiation upon CaCl2-induced differentiation. In vitro results showed that increased K24 in keratinocytes dramatically changed the differentiation of primary keratinocytes. It also inhibited cell survival by G1/S phase cell cycle arrest and induced senescence, autophagy and apoptosis of keratinocytes. In addition, K24 activated PKCδ signal pathway involving in cellular survival. In summary, K24 may be suggested as a potential differentiation marker and anti-proliferative factor in the epidermis.


Assuntos
Células Epidérmicas , Queratinócitos/metabolismo , Queratinas Tipo I/metabolismo , Adolescente , Adulto , Apoptose/genética , Apoptose/fisiologia , Western Blotting , Ciclo Celular/genética , Ciclo Celular/fisiologia , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Células Cultivadas , Feminino , Imunofluorescência , Humanos , Queratinas Tipo I/genética , Masculino , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Pessoa de Meia-Idade , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Adulto Jovem
11.
Sci Rep ; 6: 35610, 2016 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-27752144

RESUMO

Alcoholic hepatitis (AH) is the most severe form of alcoholic liver disease for which there are no effective therapies. Patients with AH show impaired hepatocyte proliferation, expansion of inefficient ductular cells and high lipopolysaccharide (LPS) levels. It is unknown whether LPS mediates ductular cell expansion. We performed transcriptome studies and identified keratin 23 (KRT23) as a new ductular cell marker. KRT23 expression correlated with mortality and LPS serum levels. LPS-TLR4 pathway role in ductular cell expansion was assessed in human and mouse progenitor cells, liver slices and liver injured TLR4 KO mice. In AH patients, ductular cell expansion correlated with portal hypertension and collagen expression. Functional studies in ductular cells showed that KRT23 regulates collagen expression. These results support a role for LPS-TLR4 pathway in promoting ductular reaction in AH. Maneuvers aimed at decreasing LPS serum levels in AH patients could have beneficial effects by preventing ductular reaction development.


Assuntos
Ducto Hepático Comum/patologia , Hepatite Alcoólica/imunologia , Hepatócitos/metabolismo , Queratinas Tipo I/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Proliferação de Células , Células Cultivadas , Colágeno/genética , Colágeno/metabolismo , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Hepatócitos/patologia , Humanos , Queratinas Tipo I/genética , Lipopolissacarídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Pessoa de Meia-Idade , Transdução de Sinais , Receptor 4 Toll-Like/genética
12.
Acta Dermatovenerol Croat ; 24(2): 116-23, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27477171

RESUMO

Palmoplantar keratoderma (PPK) is a heterogeneous group of hereditary and acquired disorders characterized by abnormal thickening of the palms and soles. There are three clinical patterns: diffuse, focal, and punctuate. Palmoplantar keratodermas can be divided into the following functional subgroups: disturbed gene functions in structural proteins (keratins), cornified envelope (loricrin, transglutaminase), cohesion (plakophilin, desmoplakin, desmoglein 1), cell-to-cell communication (connexins) and transmembrane signal transduction (cathepsin C). Unna-Thost disease is the most common variety of hereditary PPK. Mutations in keratin 1 have been reported in Unna-Thost disease. We report 12 cases in which Unna-Thost disease was diagnosed. Genealogical study demonstrated that the genodermatosis was a familial disease inherited as an autosomal dominant disorder. Dermatological examination revealed yellowish hyperkeratosis on the palms and soles. Oral mucosa, teeth, and nails remained unchanged. Histopathological examination of the biopsy sample taken from the soles of the patients showed orthokeratotic keratosis, hypergranulosis, and acanthosis without epidermolysis.


Assuntos
Ceratodermia Palmar e Plantar/genética , Ceratodermia Palmar e Plantar/patologia , Adolescente , Feminino , Humanos , Queratinas Tipo I/genética , Masculino , Pessoa de Meia-Idade , Mutação , Linhagem , Adulto Jovem
13.
Int J Obes (Lond) ; 40(8): 1250-9, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27089994

RESUMO

BACKGROUND/OBJECTIVES: Consumption of fat-rich foods is associated with obesity and related alterations. However, there is a group of individuals, the metabolically obese normal-weight (MONW) subjects, who present normal body weight but have metabolic features characteristic of the obese status, including fat deposition in critical tissues such as liver, recognized as a major cause for the promotion of metabolic diseases. Our aim was to better understand metabolic alterations present in liver of MONW rats applying whole genome transcriptome analysis. METHODS: Wistar rats were chronically fed a high-fat diet isocaloric relative to Control animals to avoid the hyperphagia and overweight and to mimic MONW features. Liver transcriptome analysis of both groups was performed. RESULTS: Sustained intake of an isocaloric high-fat diet had a deep impact on the liver transcriptome, mainly affecting lipid metabolism. Although serum cholesterol levels were not affected, circulating triacylglycerols were lower, and metabolic adaptations at gene expression level indicated adaptation toward handling the increased fat content of the diet, an increased triacylglycerol and cholesterol deposition in liver of MONW rats was observed. Moreover, gene expression pointed to increased risk of liver injury. One of the top upregulated genes in this tissue was Krt23, a marker of hepatic disease in humans that was also increased at the protein level. CONCLUSION: Long-term intake of a high-fat diet, even in the absence of overweight/obesity or increase in classical blood risk biomarkers, promotes a molecular environment leading to hepatic lipid accumulation and increasing the risk of suffering from hepatic diseases.


Assuntos
Dieta Hiperlipídica/efeitos adversos , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Metabolismo dos Lipídeos , Fígado/metabolismo , Adiposidade , Fenômenos Fisiológicos da Nutrição Animal , Animais , Western Blotting , Peso Corporal , Colesterol/sangue , Colesterol/metabolismo , Modelos Animais de Doenças , Regulação para Baixo , Ingestão de Energia , Fígado Gorduroso/genética , Fígado Gorduroso/patologia , Imuno-Histoquímica , Queratinas Tipo I/análise , Queratinas Tipo I/genética , Metabolismo dos Lipídeos/genética , Fígado/química , Masculino , Obesidade/metabolismo , Ratos , Ratos Wistar , Transcriptoma , Regulação para Cima
14.
Mol Med Rep ; 12(6): 8071-6, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26497548

RESUMO

The present study aimed to investigate the gene expression profiles of rats brain tissues treated with halothane compared with untreated controls to improve current understanding of the mechanism of action of the inhaled anesthetic. The GSE357 gene expression profile was dowloaded from the Gene Expression Omnibus database, and included six gene chips of samples repeatedly exposed to halothane and 12 gene chips of untreated controls. The differentially expressed genes (DEGs) between these two groups were identified using the Limma package in R language. Subsequently, the Database for Annotation, Visualization and Integrated Discovery was used to annotate the function of these DEGs. In addition, the most significantly upregulated gene and downregulated gene were annotated, to reveal the functional interactions with other associated genes, in FuncBase database. A total of 44 DEGs were obtained between The control and halothane exposure samples. Following Gene Ontology functional classification, these DEGs were found to be involved predominantly in the circulatory system, regulation of cell proliferation and response to endogenous stimulus and corticosteroid stimulus processes. KRT31 and HMGCS2, which were identified as the most significantly downregulated and upregulated DEGs, respectively, were associated with the lipid metabolic process and T cell activation, respectively. These results provided a basis for the development of improved inhalational anesthetics with minimal side effects and are essential for optimization of inhaled anesthetic techniques for advanced surgical procedures.


Assuntos
Anestésicos Inalatórios/farmacologia , Encéfalo/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Halotano/farmacologia , Regulação para Cima/efeitos dos fármacos , Animais , Encéfalo/metabolismo , Biologia Computacional , Bases de Dados Genéticas , Perfilação da Expressão Gênica , Redes Reguladoras de Genes/efeitos dos fármacos , Hidroximetilglutaril-CoA Sintase/metabolismo , Queratinas Tipo I/genética , Queratinas Tipo I/metabolismo , Ratos
15.
J Med Genet ; 52(10): 676-80, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26160856

RESUMO

BACKGROUND: Woolly hair (WH) is a hair abnormality that is primarily characterised by tightly curled hair with abnormal growth. METHODS: In two unrelated consanguineous Pakistani families with non-syndromic autosomal recessive (AR) WH, homozygosity mapping and linkage analysis identified a locus within 17q21.1-q22, which contains the type I keratin gene cluster. A DNA sample from an affected individual from each family underwent exome sequencing. RESULTS: A homozygous missense variant c.950T>C (p.(Leu317Pro)) within KRT25 segregated with ARWH in both families, and has a combined maximum two-point LOD score of 7.9 at Ï´=0. The KRT25 variant is predicted to result in disruption of the second α-helical rod domain and the entire protein structure, thus possibly interfering with heterodimerisation of K25 with type II keratins within the inner root sheath (IRS) of the hair follicle and the medulla of the hair shaft. CONCLUSIONS: Our findings implicate a novel gene involved in human hair abnormality, and are consistent with the curled, fragile hair found in mice with Krt25 mutations, and further support the role of IRS-specific type I keratins in hair follicle development and maintenance of hair texture.


Assuntos
Doenças do Cabelo/genética , Cabelo/anormalidades , Queratinas Tipo I/genética , Mutação de Sentido Incorreto , Criança , Pré-Escolar , Consanguinidade , Humanos , Masculino , Paquistão , Linhagem , Análise de Sequência de DNA
16.
PLoS One ; 8(9): e73593, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24039993

RESUMO

Keratin 23 (KRT23) is strongly expressed in colon adenocarcinomas but absent in normal colon mucosa. Array based methylation profiling of 40 colon samples showed that the promoter of KRT23 was methylated in normal colon mucosa, while hypomethylated in most adenocarcinomas. Promoter methylation correlated with absent expression, while increased KRT23 expression in tumor samples correlated with promoter hypomethylation, as confirmed by bisulfite sequencing. Demethylation induced KRT23 expression in vitro. Expression profiling of shRNA mediated stable KRT23 knockdown in colon cancer cell lines showed that KRT23 depletion affected molecules of the cell cycle and DNA replication, recombination and repair. In vitro analyses confirmed that KRT23 depletion significantly decreased the cellular proliferation of SW948 and LS1034 cells and markedly decreased the expression of genes involved in DNA damage response, mainly molecules of the double strand break repair homologous recombination pathway. KRT23 knockdown decreased the transcript and protein expression of key molecules as e.g. MRE11A, E2F1, RAD51 and BRCA1. Knockdown of KRT23 rendered colon cancer cells more sensitive to irradiation and reduced proliferation of the KRT23 depleted cells compared to irradiated control cells.


Assuntos
Proliferação de Células , Reparo do DNA/genética , Queratinas Tipo I/genética , Interferência de RNA , Sequência de Bases , Western Blotting , Ciclo Celular/genética , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Sobrevivência Celular/efeitos da radiação , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Dano ao DNA , Metilação de DNA , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Raios gama , Regulação Neoplásica da Expressão Gênica/genética , Células HCT116 , Células HEK293 , Humanos , Queratinas Tipo I/metabolismo , Proteína Homóloga a MRE11 , Microscopia de Fluorescência , Dados de Sequência Molecular , Análise de Sequência com Séries de Oligonucleotídeos , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transcriptoma/genética
17.
Cancer Prev Res (Phila) ; 6(7): 666-74, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23682078

RESUMO

The length of time required for preinvasive adenoma to progress to carcinoma, the immunogenicity of colorectal cancer (CRC), and the identification of high-risk populations make development and testing of a prophylactic vaccine for the prevention of CRC possible. We hypothesized that genes upregulated in adenoma relative to normal tissue, which maintained increased expression in CRC, would encode proteins suitable as putative targets for immunoprevention. We evaluated existing adenoma and CRC microarray datasets and identified 160 genes that were ≥2-fold upregulated in both adenoma and CRC relative to normal colon tissue. We further identified 23 genes that showed protein overexpression in colon adenoma and CRC based on literature review. Silencing the most highly upregulated genes, CDH3, CLDN1, KRT23, and MMP7, in adenoma and CRC cell lines resulted in a significant decrease in viability (P < 0.0001) and proliferation (P < 0.0001) as compared to controls and an increase in cellular apoptosis (P < 0.05 for CDH3, KRT23). Results were duplicated across cell lines representing microsatellite instability, CpG island methylator, and chromosomal instability phenotypes, suggesting immunologic elimination of cells expressing these proteins could impact the progression of all CRC phenotypes. To determine whether these proteins were immunogens, we interrogated sera from early stage CRC patients and controls and found significantly elevated CDH3 (P = 0.006), KRT23 (P = 0.0007), and MMP7 (P < 0.0001) serum immunoglobulin G in cases as compared to controls. These data show a high throughput approach to the identification of biologically relevant putative immunologic targets for CRC and identified three candidates suitable for vaccine development.


Assuntos
Adenoma/diagnóstico , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/diagnóstico , Regulação Neoplásica da Expressão Gênica , Lesões Pré-Cancerosas/diagnóstico , Adenoma/metabolismo , Adenoma/prevenção & controle , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Western Blotting , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Estudos de Casos e Controles , Proliferação de Células , Claudina-1/antagonistas & inibidores , Claudina-1/genética , Claudina-1/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/prevenção & controle , Metilação de DNA , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Queratinas Tipo I/antagonistas & inibidores , Queratinas Tipo I/genética , Queratinas Tipo I/metabolismo , Masculino , Metaloproteinase 7 da Matriz/química , Metaloproteinase 7 da Matriz/genética , Metaloproteinase 7 da Matriz/metabolismo , Instabilidade de Microssatélites , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Lesões Pré-Cancerosas/metabolismo , Lesões Pré-Cancerosas/prevenção & controle , Prognóstico , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto Jovem
18.
PLoS One ; 7(10): e46584, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23071592

RESUMO

BACKGROUND: Pathogenesis and factors for determining progression of alcoholic and non-alcoholic steatosis to steatohepatitis with risk of further progression to liver cirrhosis and cancer are poorly understood. In the present study, we aimed to identify potential molecular signatures for discrimination of steatohepatitis from steatosis. METHODOLOGY AND RESULTS: Global microarray gene expression analysis was applied to unravel differentially expressed genes between steatohepatitis compared to steatosis and control samples. For functional annotation as well as the identification of disease-relevant biological processes of the differentially expressed genes the gene ontology (GO) database was used. Selected candidate genes (n = 46) were validated in 87 human liver samples from two sample cohorts by quantitative real-time PCR (qRT-PCR). The GO analysis revealed that genes down-regulated in steatohepatitis were mainly involved in metabolic processes. Genes up-regulated in steatohepatitis samples were associated with cancer progression and proliferation. In surgical liver resection samples, 39 genes and in percutaneous liver biopsies, 30 genes were significantly up-regulated in steatohepatitis. Furthermore, immunohistochemical investigation of human liver tissue revealed a significant increase of AKR1B10 protein expression in steatohepatitis. CONCLUSIONS: The development of steatohepatitis is characterized by distinct molecular changes. The most striking examples in this respect were KRT23 and AKR1B10, which we found to be highly differentially expressed in steatohepatitis compared to steatosis and normal liver. We propose that KRT23 and AKR1B10 may serve as future potential biomarkers for steatohepatitis as well as markers for progression to HCC.


Assuntos
Fígado Gorduroso/metabolismo , Neoplasias Hepáticas/metabolismo , Transcriptoma , Adulto , Idoso , Aldeído Redutase/genética , Aldeído Redutase/metabolismo , Aldo-Ceto Redutases , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Estudos de Casos e Controles , Análise por Conglomerados , Feminino , Hepatite C Crônica/metabolismo , Humanos , Queratinas Tipo I/genética , Queratinas Tipo I/metabolismo , Fígado/metabolismo , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Componente Principal , Regulação para Cima , Adulto Jovem
19.
Lab Invest ; 92(5): 688-702, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22330335

RESUMO

Notch is a transmembrane receptor functioning in the determination of cell fate. Abnormal Notch signaling promotes tumor development, showing either oncogenic or tumor suppressive activity. The uncertainty about the exact role of Notch signaling, partially, stems from inconsistencies in descriptions of Notch expression in human cancers. Here, we clarified basal-cell dominant expression of NOTCH1 in squamous epithelium. NOTCH1 was downregulated in squamous neoplasms of oral mucosa, esophagus and uterine cervix, compared with the normal basal cells, although the expression tended to be retained in cervical lesions. NOTCH1 downregulation was observed even in precancers, and there was little difference between cancers and high-grade precancerous lesions, suggesting its minor contribution to cancer-specific events such as invasion. In culture experiments, reduction of NOTCH1 expression resulted in downregulation of keratin 13 and keratin 15, and upregulation of keratin 17, and NOTCH1 knockdown cells formed a dysplastic stratified epithelium mimicking a precancerous lesion. The NOTCH1 downregulation and the concomitant alterations of those keratin expressions were confirmed in the squamous neoplasms both by immunohistochemical and cDNA microarray analyses. Our data indicate that reduction of NOTCH1 expression directs the basal cells to cease terminal differentiation and to form an immature epithelium, thereby playing a major role in the histopathogenesis of epithelial dysplasia. Furthermore, downregulation of NOTCH1 expression seems to be an inherent mechanism for switching the epithelium from a normal and mature state to an activated and immature state, suggesting its essential role in maintaining the epithelial integrity.


Assuntos
Carcinoma de Células Escamosas/patologia , Neoplasias Esofágicas/patologia , Queratinócitos/patologia , Queratinas Tipo I/metabolismo , Neoplasias Bucais/patologia , Lesões Pré-Cancerosas/patologia , Receptor Notch1/metabolismo , Neoplasias do Colo do Útero/patologia , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Regulação para Baixo , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Queratina-13/genética , Queratina-13/metabolismo , Queratina-15/genética , Queratina-15/metabolismo , Queratina-17/genética , Queratina-17/metabolismo , Queratinócitos/citologia , Queratinócitos/metabolismo , Queratinas Tipo I/genética , Masculino , Camundongos , Neoplasias Bucais/genética , Neoplasias Bucais/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/metabolismo , Receptor Notch1/genética , Regulação para Cima , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo
20.
J Dermatol Sci ; 65(2): 141-8, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22261007

RESUMO

BACKGROUND: AHF/trichohyalin is a large structural protein abundant in the inner root sheath (IRS) of anagenic hair follicles, which has been thought to mediate the keratin filamentous assembly. However, its functional mechanism is largely unknown. OBJECTIVE: This study aimed at the identification of the key domain in AHF for keratin association and the establishment of a plausible mechanism for the modulation of the keratin meshwork. METHODS: Several keratinocyte cell lines were introduced with the full length or several mutants of AHF, together with IRS-specific keratin krt31, and the profile of the AHF granules and the cellular behaviors were carefully analyzed. RESULTS: Full length of AHF formed small round granules that clearly bound to and aligned on the exogenous keratin filaments in the keratinocytes, severely affected cellular growth, mobility and shape. Intriguingly, the removal of only 6 amino acids around the C-terminal tail of AHF resulted not only in the complete loss of its keratin adherent ability but also in a dramatic enlargement of the granules. CONCLUSION: We propose a model for cytoskeletal modulation in the IRS of anagenic hair follicles: AHF latches onto the keratin bundles by its C-terminus and rearranges the keratin meshwork by intrinsic cohesive activity for the granule formation.


Assuntos
Citoesqueleto/metabolismo , Folículo Piloso/metabolismo , Proteínas de Filamentos Intermediários/metabolismo , Queratinócitos/metabolismo , Queratinas Específicas do Cabelo/metabolismo , Queratinas Tipo I/metabolismo , Animais , Sítios de Ligação , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Forma Celular , Genótipo , Folículo Piloso/crescimento & desenvolvimento , Humanos , Proteínas de Filamentos Intermediários/química , Proteínas de Filamentos Intermediários/genética , Queratinas Específicas do Cabelo/genética , Queratinas Tipo I/genética , Camundongos , Mutação , Fenótipo , Domínios e Motivos de Interação entre Proteínas , Fatores de Tempo , Transfecção
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