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1.
Artigo em Inglês | MEDLINE | ID: mdl-32145468

RESUMO

BACKGROUND & AIMS: Recent evidence has suggested that the intact intestinal epithelial barrier protects our body from a range of immune-mediated diseases. The epithelial layer has an impressive ability to reconstitute and repair upon damage and this process of repair increasingly is seen as a therapeutic target. In vitro models to study this process in primary intestinal cells are lacking. METHODS: We established and characterized an in vitro model of intestinal damage and repair by applying γ-radiation on small-intestinal organoids. We then used this model to identify novel regulators of intestinal regeneration. RESULTS: We identified hepatocyte nuclear factor 4α (HNF4α) as a pivotal upstream regulator of the intestinal regenerative response. Organoids lacking Hnf4a were not able to propagate in vitro. Importantly, intestinal Hnf4a knock-out mice showed impaired regeneration after whole-body irradiation, confirming intestinal organoids as a valuable alternative to in vivo studies. CONCLUSIONS: In conclusion, we established and validated an in vitro damage-repair model and identified HNF4α as a crucial regulator of intestinal regeneration. Transcript profiling: GSE141515 and GSE141518.

2.
Mol Cell Proteomics ; 2020 Mar 02.
Artigo em Inglês | MEDLINE | ID: mdl-32123031

RESUMO

HNF4α is a nuclear receptor produced as 12 isoforms from two promoters by alternative splicing. In order to characterize the transcriptional capacities of all 12 HNF4α isoforms, stable lines expressing each isoform were generated. The entire transcriptome associated with each isoform was analyzed as well as their respective interacting proteome. Major differences were noted in the transcriptional function of these isoforms. The α1 and α2 isoforms were the strongest regulators of gene expression while the α3 isoform exhibited significantly reduced activity. The α4, α5 and α6 isoforms, which use an alternative first exon, were characterized for the first time, and showed a greatly reduced transcriptional potential with an inability to recognize the consensus response element of HNF4α. Several transcription factors and coregulators were identified as potential specific partners for certain HNF4α isoforms. An analysis integrating the vast amount of omics data enabled the identification of transcriptional regulatory mechanisms specific to certain HNF4α isoforms, hence demonstrating the importance of considering all isoforms given their seemingly diverse functions.

4.
Cancers (Basel) ; 11(5)2019 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-31060309

RESUMO

Hepatocyte nuclear factor 4α (HNF4α) is a transcription factor that acts as a master regulator of genes for several endoderm-derived tissues, including the intestine, in which it plays a central role during development and tumorigenesis. To better define the mechanisms by which HNF4α can influence these processes, we identified proteins interacting with HNF4α using stable isotope labelling with amino acids in cell culture (SILAC)-based quantitative proteomics with either immunoprecipitation of green fluorescent protein (GFP) or with proximity-dependent purification by the biotin ligase BirA (BioID), both fused to HNF4α. Surprisingly, these analyses identified a significant enrichment of proteins characterized with a role in DNA repair, a so far unidentified biological feature of this transcription factor. Several of these proteins including PARP1, RAD50, and DNA-PKcs were confirmed to interact with HNF4α in colorectal cancer cell lines. Following DNA damage, HNF4α was able to increase cell viability in colorectal cancer cells. Overall, these observations identify a potential role for this transcription factor during the DNA damage response.

5.
Sci Rep ; 9(1): 4200, 2019 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-30862908

RESUMO

Mutations in the HNF4A gene cause MODY1 and are associated with an increased risk of Type 2 diabetes mellitus. On the other hand, incretins are hormones that potentiate reductions in blood glucose levels. Given the established role of incretin-based therapy to treat diabetes and metabolic disorders, we investigated a possible regulatory link between intestinal epithelial HNF4α and glucose-dependent insulinotropic polypeptide (GIP), an incretin that is specifically produced by gut enteroendocrine cells. Conditional deletion of HNF4α in the whole intestinal epithelium was achieved by crossing Villin-Cre and Hnf4αloxP/loxP C57BL/6 mouse models. GIP expression was measured by qPCR, immunofluorescence and ELISA. Gene transcription was assessed by luciferase and electrophoretic mobility shift assays. Metabolic parameters were analyzed by indirect calorimetry and dual-energy X-ray absorptiometry. HNF4α specific deletion in the intestine led to a reduction in GIP. HNF4α was able to positively control Gip transcriptional activity in collaboration with GATA-4 transcription factor. Glucose homeostasis and glucose-stimulated insulin secretion remained unchanged in HNF4α deficient mice. Changes in GIP production in these mice did not impact nutrition or energy metabolism under normal physiology but led to a reduction of bone area and mineral content, a well described physiological consequence of GIP deficiency. Our findings point to a novel regulatory role between intestinal HNF4α and GIP with possible functional impact on bone density.

6.
Sci Rep ; 9(1): 5363, 2019 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-30926862

RESUMO

Both HDAC1 and HDAC2 are class I deacetylases acting as erasers of lysine-acetyl marks on histones and non-histone proteins. Several histone deacetylase inhibitors, either endogenous to the cell, such as the ketogenic ß-hydroxybutyrate metabolite, or exogenous, such as butyrate, a microbial-derived metabolite, regulate HDAC activity. Different combinations of intestinal epithelial cell (IEC)-specific Hdac1 and/or Hdac2 deletion differentially alter mucosal homeostasis in mice. Thus, HDAC1 and HDAC2 could act as sensors and transmitters of environmental signals to the mucosa. In this study, enteroid culture models deleted for Hdac1 or Hdac2 were established to determine IEC-specific function as assessed by global transcriptomic and proteomic approaches. Results show that Hdac1 or Hdac2 deficiency altered differentiation of Paneth and goblet secretory cells, which sustain physical and chemical protection barriers, and increased intermediate secretory cell precursor numbers. Furthermore, IEC Hdac1- and Hdac2-dependent common and specific biological processes were identified, including oxidation-reduction, inflammatory responses, and lipid-related metabolic processes, as well as canonical pathways and upstream regulators related to environment-dependent signaling through steroid receptor pathways, among others. These findings uncover unrecognized regulatory similarities and differences between Hdac1 and Hdac2 in IEC, and demonstrate how HDAC1 and HDAC2 may complement each other to regulate the intrinsic IEC phenotype.

7.
J Cell Sci ; 131(13)2018 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-29898915

RESUMO

HNF4α is a key nuclear receptor for regulating gene expression in the gut. Although both P1 and P2 isoform classes of HNF4α are expressed in colonic epithelium, specific inhibition of P1 isoforms is commonly found in colorectal cancer. Previous studies have suggested that P1 and P2 isoforms might regulate different cellular functions. Despite these advances, it remains unclear whether these isoform classes are functionally divergent in the context of human biology. Here, the consequences of specific inhibition of P1 or P2 isoform expression was measured in a human colorectal cancer cell transcriptome. Results indicate that P1 isoforms were specifically associated with the control of cell metabolism, whereas P2 isoforms globally supported aberrant oncogenic signalization, promoting cancer cell survival and progression. P1 promoter-driven isoform expression was found to be repressed by ß-catenin, one of the earliest oncogenic pathways to be activated during colon tumorigenesis. These findings identify a novel cascade by which the expression of P1 isoforms is rapidly shut down in the early stages of colon tumorigenesis, allowing a change in HNF4α-dependent transcriptome, thereby promoting colorectal cancer progression.This article has an associated First Person interview with the first author of the paper.


Assuntos
Neoplasias Colorretais/metabolismo , Fator 4 Nuclear de Hepatócito/genética , Regiões Promotoras Genéticas , beta Catenina/genética , Animais , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Fator 4 Nuclear de Hepatócito/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transcriptoma , beta Catenina/metabolismo
8.
Biochim Biophys Acta Mol Basis Dis ; 1864(5 Pt A): 1539-1551, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29454075

RESUMO

Colorectal tumors are immersed in an array of tumor-promoting factors including extracellular nucleotides such as uridine 5'­diphosphate (UDP). UDP is the endogenous agonist of the G protein-coupled P2Y6 receptor (P2Y6R), which may contribute to the formation of a tumor-promoting microenvironment by coordinating resistance to apoptosis. Colorectal cancer (CRC) was chemically induced in P2ry6 knockout (P2ry6-/-) mice using azoxymethane and dextran sulfate sodium challenges. Mice were euthanatized and their tumor load determined. Fixed tissues were stained for histological and immunohistochemistry analysis. Tumoroids were also prepared from CRC tumors resected from P2ry6+/+ mice to determine the role of P2Y6R in resistance to apoptosis, whereas HT29 carcinoma cells were used to elucidate the signaling mechanism involved in P2Y6R anti-apoptotic effect. P2ry6-/- mice developed a reduced number of colorectal tumors with apparent tumors having smaller volumes. Overall dysplastic score was significantly lower in P2ry6-/- animals. Stimulation of P2Y6R with the selective agonist MRS2693 protected HT-29 cells from TNFα-induced apoptosis. This protective effect was mediated by the stabilizing phosphorylation of the X-linked inhibitor of apoptosis protein (XIAP) by AKT. Using CRC-derived tumoroids, P2Y6R activation was found to contribute to chemoresistance since addition of the P2Y6R agonist MRS2693 significantly prevented the cytotoxic effect of 5-fluorouracil. The present study shows that sustained activation of P2Y6R may contribute to intestinal tumorigenesis by blocking the apoptotic process and by contributing to chemoresistance, a substantial concern in the treatment of patients with CRC. These results suggest that P2Y6R may represent a prime target for reducing colorectal carcinogenesis.


Assuntos
Apoptose , Transformação Celular Neoplásica/metabolismo , Neoplasias Colorretais/metabolismo , Proteínas de Neoplasias/metabolismo , Receptores Purinérgicos P2/metabolismo , Animais , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Receptores Purinérgicos P2/genética , Fator de Necrose Tumoral alfa/metabolismo
9.
Health Psychol Open ; 5(1): 2055102917750331, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29372066

RESUMO

Physical activity plays a crucial role in the prevention and treatment of type 2 diabetes. Therefore, it is important to understand why so few adults with type 2 diabetes regularly engage in physical activity. The role of self-regulation in the context of health-related behavior adherence, especially in terms of physical activity engagement and adherence, has largely been reviewed based on the strength energy model. Building on this line of research, the aim of this theoretical work was to highlight how self-regulation and ego depletion can influence the lower rate of physical activity participation among adults with type 2 diabetes, compared to adults from the general population.

10.
Biochim Biophys Acta Mol Cell Res ; 1864(12): 2347-2355, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28893557

RESUMO

Intestinal epithelial cells form a protective barrier in limiting gut luminal content potentially harmful to the host. Upon gut epithelium injury, several signals instruct epithelial cells to undergo a rapid healing process. Defects in this process induce inflammatory responses and can further evolve into chronic gut inflammatory diseases. We previously identified the transcription factor CUX1 as crucial for protecting against experimental colitis in mice. However, the precise molecular mechanisms by which CUX1 intervenes during this biological process are unknown. Our aim was to evaluate CUX1 biological and functional roles during intestinal epithelial cell wound healing. RNAi knockdown of CUX1 in intestinal epithelial cells revealed a crucial role for this regulator in migratory response following wounding assays. Gene expression profiling identified several gene transcripts modulated in absence of CUX1 during wound healing for which a significant number was associated with cell motility and cytoskeleton function. Chromatin immunoprecipitation assays identified the guanine nucleotide exchange factor Vav2 gene as a direct target for CUX1. Coincidently, reduction of VAV2 in absence of CUX1 was associated with a significant decrease of RAC1 activity in response to epithelial wounding. Our results identify a novel pathway by which CUX1 regulates normal intestinal epithelial cell restitution.


Assuntos
Proteínas de Homeodomínio/genética , Inflamação/genética , Neuropeptídeos/genética , Proteínas Nucleares/genética , Proteínas Proto-Oncogênicas c-vav/genética , Proteínas Repressoras/genética , Cicatrização/genética , Proteínas rac1 de Ligação ao GTP/genética , Animais , Células Epiteliais/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Humanos , Inflamação/patologia , Mucosa Intestinal/metabolismo , Intestinos/patologia , Camundongos
11.
Proteomics ; 17(20)2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28643936

RESUMO

The development of 3D cell cultures into self-organizing organ-like structures named organoids provides a model that better reflects in vivo organ physiology and their functional properties. Organoids have been established from several organs, such as the intestine, prostate, brain, liver, kidney and pancreas. With recent advances in high-throughput and -omics profiling technologies, it is now possible to study the mechanisms of cellular organisation at the systems level. It is therefore not surprising that these methods are now used to characterize organoids at the transcriptomic, proteomic, chromatin state and transcription factor DNA-binding levels. These approaches can therefore provide a wealth of information regarding both the mechanisms involved in different diseases, and those involved in cell responses to different conditions, in a more in vivo setting. The authors provide an overview of the potential applications of quantitative mass spectrometry with organoid culture, and how the use of large-scale proteome measurements is emerging in different organoid systems.


Assuntos
Técnicas de Cultura de Órgãos/métodos , Organoides/crescimento & desenvolvimento , Proteômica/métodos , Animais , Encéfalo/citologia , Encéfalo/crescimento & desenvolvimento , Sistema Digestório/citologia , Sistema Digestório/crescimento & desenvolvimento , Humanos , Rim/citologia , Rim/crescimento & desenvolvimento , Espectrometria de Massas , Organogênese , Organoides/citologia , Peptídeos/análise , Fenótipo , Proteoma/análise
12.
PLoS One ; 12(5): e0177110, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28493909

RESUMO

PURPOSE: Hepatocyte nuclear factor 1 alpha (HNF1α) defects cause Mature Onset Diabetes of the Young type 3 (MODY3), characterized by defects in beta-cell insulin secretion. However, HNF1α is involved in many other metabolic pathways with relevance for monogenic or polygenic type 2 diabetes. We aimed to investigate gut hormones, lipids, and insulin regulation in response to a meal test in HNF1α defect carriers (MODY3) compared to non-diabetic subjects (controls) and type 2 diabetes (T2D). METHODS: We administered a standardized liquid meal to each participant. Over 6 hours, we measured post-meal responses of insulin regulation (blood glucose, c-peptide, insulin), gut hormones (ghrelin, glucose-dependent insulinotropic polypeptide, glucagon-like peptide-1) and lipids (non-esterified fatty acids [NEFA] and triglycerides). RESULTS: We found that MODY3 participants had lower insulin secretion indices than controls and T2D participants, showing the expected ß-cell defect. MODY3 had similar glycated hemoglobin levels (HbA1c median [IQR]: 6.5 [5.6-7.6]%) compared to T2D (median: 6.6 [6.2-6.9]%; P<0.05). MODY3 had greater insulin sensitivity (Matsuda index: 71.9 [29.6; 125.5]) than T2D (3.2 [4.0; 6.0]; P<0.05). MODY3 experienced a larger decrease in the ratio of NEFA to insulin (NEFA 30-0 / insulin 30-0: -39 [-78; -30] x104) in the early post-prandial period (0-30 minutes) compared to controls and to T2D (-2.0 [-0.6; -6.4] x104; P<0.05). MODY3 had lower fasting (0.66 [0.46; 1.2] mM) and post-meal triglycerides levels compared to T2D (fasting: 2.3 [1.7; 2.7] mM; P<0.05). We did not detect significant post-meal differences in ghrelin and incretins between MODY3 and other groups. CONCLUSION: In response to a standard meal test, MODY3 showed greater early post-prandial NEFA diminution in response to relatively low early insulin secretion, and they maintained very low post-prandial triglycerides levels.


Assuntos
Fator 1-alfa Nuclear de Hepatócito/metabolismo , Insulina/metabolismo , Adolescente , Adulto , Idoso , Glicemia/metabolismo , Peptídeo C/sangue , Peptídeo C/metabolismo , Criança , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Ácidos Graxos não Esterificados/sangue , Ácidos Graxos não Esterificados/metabolismo , Feminino , Polipeptídeo Inibidor Gástrico/sangue , Polipeptídeo Inibidor Gástrico/metabolismo , Grelina/sangue , Grelina/metabolismo , Peptídeo 1 Semelhante ao Glucagon/sangue , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Fator 1-alfa Nuclear de Hepatócito/genética , Humanos , Insulina/genética , Masculino , Pessoa de Meia-Idade , Período Pós-Prandial , Triglicerídeos/sangue , Triglicerídeos/metabolismo , Adulto Jovem
13.
FASEB J ; 31(8): 3512-3526, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28465325

RESUMO

Shp-1 (Src homology region 2 domain-containing protein tyrosine phosphatase-1) is a phosphatase that is highly expressed in hematopoietic and epithelial cells. Whereas its function is largely characterized in hematopoietic cells, its role in epithelial cells, such as intestinal epithelial cells (IECs), is not well known. Here, we generated mice with an IEC-specific knockout of Shp-1 (Src homology region 2 domain-containing phosphatase-1; Shp-1IEC-KO). We showed that the loss of epithelial Shp-1 leads to an intestinalomegaly that is associated with an increase in epithelial cell proliferation and size. Histologic analysis demonstrates significant perturbation of the crypt-villus architecture with an apparent increase in the number of goblet and Paneth cells and increased expression of their respective markers {Muc2 (mucin 2), αDef, and Sox9 [SRY (sex determining region Y)-box 9]}. Expansion of intermediate cells-common progenitors of goblet and Paneth cell lineages-is also observed in Shp-1IEC-KO mice. Although sustained activation of Wnt/ß-catenin and PI3K/Akt/mammalian target of rapamycin signaling is observed, Shp-1IEC-KO mice fail to develop any intestinal tumors after 15 mo; however, the loss of Shp-1 in IECs markedly enhances tumor load ApcMin/+ mice. These findings show a novel role for Shp-1 in the regulation of IEC growth and secretory lineage allocation, possibly via modulation of PI3K/Akt-dependent signaling pathways. Finally, Shp-1 does not function as a classic tumor suppressor gene in the intestinal epithelium.-Leblanc, C., Langlois, M.-J., Coulombe, G., Vaillancourt-Lavigueur, V., Jones, C., Carrier, J. C., Boudreau, F., Rivard, N. Epithelial Src homology region 2 domain-containing phosphatase-1 restrains intestinal growth, secretory cell differentiation, and tumorigenesis.


Assuntos
Neoplasias do Colo/metabolismo , Regulação da Expressão Gênica/fisiologia , Intestinos/crescimento & desenvolvimento , Proteína Tirosina Fosfatase não Receptora Tipo 6/metabolismo , Animais , Cateninas/genética , Cateninas/metabolismo , Células Epiteliais/fisiologia , Humanos , Mucosa Intestinal/metabolismo , Intestinos/citologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 6/genética , Proteínas Proto-Oncogênicas c-akt , Proteínas Wnt/genética , Proteínas Wnt/metabolismo
14.
Biomed Res Int ; 2017: 4092304, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28393077

RESUMO

Background. In the domain of health behavior change, the deployment and utilization of information and communications technologies as a way to deliver interventions appear to be promising. This article describes the development of a web-based tailored intervention, TAVIE en santé, to support people living with HIV in the adoption of healthy behaviors. Methods. This intervention was developed through an Intervention Mapping (IM) framework and is based on the theory of planned behavior. Results. Crucial steps of IM are the selection of key determinants of behavior and the selection of useful theory-based intervention methods to change the targeted determinants (active ingredients). The content and the sequence of the intervention are then created based on these parameters. TAVIE en santé is composed of 7 interactive web sessions hosted by a virtual nurse. It aims to develop and strengthen skills required for behavior change. Based on an algorithm using individual cognitive data (attitude, perceived behavioral control, and intention), the number of sessions, theory-based intervention methods, and messages contents are tailored to each user. Conclusion. TAVIE en santé is currently being evaluated. The use of IM allows developing intervention with a systematic approach based on theory, empirical evidence, and clinical and experiential knowledge.


Assuntos
Comorbidade , Infecções por HIV/epidemiologia , Comportamentos Relacionados com a Saúde , Infecções por HIV/psicologia , Infecções por HIV/terapia , Humanos , Internet , Enfermeiras e Enfermeiros , Autocuidado
15.
Sci Rep ; 6: 38195, 2016 11 30.
Artigo em Inglês | MEDLINE | ID: mdl-27901089

RESUMO

Organoids have the potential to bridge 3D cell culture to tissue physiology by providing a model resembling in vivo organs. Long-term growing organoids were first isolated from intestinal crypt cells and recreated the renewing intestinal epithelial niche. Since then, this technical breakthrough was applied to many other organs, including prostate, liver, kidney and pancreas. We describe here how to apply a SILAC-based quantitative proteomic approach to measure protein expression changes in intestinal organoids under different experimental conditions. We generated SILAC organoid media that allow organoids to grow and differentiate normally, and confirmed the incorporation of isotopically labelled amino acids. Furthermore, we used a treatment reported to affect organoid differentiation to demonstrate the reproducibility of the quantification using this approach and to validate the identification of proteins that correlate with the inhibition of cellular growth and development. With the combined use of quantitative mass spectrometry, SILAC and organoid culture, we validated this approach and showed that large-scale proteome variations can be measured in an "organ-like" system.


Assuntos
Mucosa Intestinal/metabolismo , Organoides/metabolismo , Proteoma/metabolismo , Proteômica/métodos , Aminoácidos/metabolismo , Animais , Western Blotting , Células Cultivadas , Cromatografia Líquida , Células Epiteliais/metabolismo , Intestinos/citologia , Marcação por Isótopo/métodos , Camundongos , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem
16.
Sci Rep ; 6: 36776, 2016 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-27827449

RESUMO

The intestinal epithelial barrier is critical to limit potential harmful consequences from exposure to deleterious luminal contents on the organism. Although this barrier is functionally important along the entire gut, specific regional regulatory mechanisms involved in the maintenance of this barrier are poorly defined. Herein, we identified Gata4 as a crucial regulator of barrier integrity in the mouse proximal intestinal epithelium. Conditional deletion of Gata4 in the intestine led to a drastic increase in claudin-2 expression that was associated with an important increase of gut barrier permeability without causing overt spontaneous inflammation. Administration of indomethacin, a non-steroidal anti-inflammatory drug (NSAID) that causes enteritis, led to rapid and restricted proximal small intestinal injuries in Gata4 mutant mice as opposed to control mice. Comparative analysis of gene transcript profiles from indomethacin-challenged control and Gata4 mutant mice identified defects in epithelial cell survival, inflammatory cell recruitment and tissue repair mechanisms. Altogether, these observations identify Gata4 as a novel crucial regulator of the intestinal epithelial barrier and as a critical epithelial transcription factor implicated in the maintenance of proximal intestinal mucosal integrity after injury.


Assuntos
Enterite/genética , Fator de Transcrição GATA4/genética , Indometacina/efeitos adversos , Mucosa Intestinal/metabolismo , Animais , Claudinas/metabolismo , Enterite/induzido quimicamente , Enterite/metabolismo , Fator de Transcrição GATA4/metabolismo , Regulação da Expressão Gênica , Camundongos , Mutação , Salmonella typhi
17.
Proteomics ; 16(23): 3009-3018, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27689624

RESUMO

Studying cell differentiation and transformation allows a better understanding of the mechanisms involved in the initiation and the evolution of cancer. The role of proteins which participate in these processes is dependent on their location within the cell. Determining the subcellular localization of proteins or the changes in localization is, therefore, paramount in elucidating their role. Using quantitative mass spectrometry, we characterized the protein expression and subcellular localization of nearly 5000 proteins from seven different colorectal cancer (CRC) cell lines, as well as normal colon fibroblasts and intestinal epithelial cells. This cellular characterization allowed the identification of colon cancer-associated proteins with differential expression patterns as well as deregulated protein networks and pathways. Indeed, our results demonstrate differential expression of proteins involved in cell adhesion, cytoskeleton, and transcription in colon cancer cells compared to normal colon-derived cells. Pathway analyses identified different cellular functions, including endocytosis and eIF2 signaling, whose deregulation correlates with mutations found in the different CRC phenotypes. Our results provide an unbiased, quantitative and high-throughput approach to measure changes in protein expression and subcellular protein locations in different CRC cell lines.


Assuntos
Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Proteínas/metabolismo , Proteômica/métodos , Linhagem Celular Tumoral , Ensaios de Triagem em Larga Escala/métodos , Humanos , Proteínas/análise , Frações Subcelulares
18.
Sci Rep ; 6: 32759, 2016 09 09.
Artigo em Inglês | MEDLINE | ID: mdl-27609464

RESUMO

Bmps are morphogens involved in various gastric cellular functions. Studies in genetically-modified mice have shown that Bmp disruption in gastric epithelial and stromal cell compartments leads to the development of tumorigenesis. Our studies have demonstrated that abrogation of gastric epithelial Bmp signaling alone was not sufficient to recapitulate the neoplastic features associated with total gastric loss of Bmp signaling. Thus, epithelial Bmp signaling does not appear to be a key player in gastric tumorigenesis initiation. These observations suggest a greater role for stromal Bmp signaling in gastric polyposis initiation. In order to identify the specific roles played by mesenchymal Bmp signaling in gastric homeostasis, we generated a mouse model with abrogation of Bmp signaling exclusively in the gastro-intestinal mesenchyme (Bmpr1a(ΔMES)). We were able to expose an unsuspected role for Bmp loss of signaling in leading normal gastric mesenchyme to adapt into reactive mesenchyme. An increase in the population of activated-fibroblasts, suggesting mesenchymal transdifferentiation, was observed in mutant stomach. Bmpr1a(ΔMES) stomachs exhibited spontaneous benign polyps with presence of both intestinal metaplasia and spasmolytic-polypeptide-expressing metaplasia as early as 90 days postnatal. These results support the novel concept that loss of mesenchymal Bmp signaling cascade acts as a trigger in gastric polyposis initiation.


Assuntos
Pólipos Adenomatosos/genética , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Proteínas Morfogenéticas Ósseas/metabolismo , Transformação Celular Neoplásica/genética , Neoplasias Gástricas/genética , Células Estromais/metabolismo , Pólipos Adenomatosos/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Regulação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Camundongos , Transdução de Sinais , Neoplasias Gástricas/metabolismo
19.
J Cell Physiol ; 231(11): 2529-40, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27100271

RESUMO

Polymorphisms in the PTPN11 gene encoding for the tyrosine phosphatase SHP-2 were described in patients with ulcerative colitis. We have recently demonstrated that mice with an intestinal epithelial cell-specific deletion of SHP-2 (SHP-2(IEC-KO) ) develop severe colitis 1 month after birth. However, the mechanisms by which SHP-2 deletion induces colonic inflammation remain to be elucidated. We generated SHP-2(IEC-KO) mice lacking Myd88 exclusively in the intestinal epithelium. The colonic phenotype was histologically analyzed and cell differentiation was determined by electron microscopy and lysozyme or Alcian blue staining. Microbiota composition was analyzed by 16S sequencing. Results show that innate defense genes including those specific to Paneth cells were strongly up-regulated in SHP-2-deficient colons. Expansion of intermediate cells (common progenitors of the Goblet and Paneth cell lineages) was found in the colon of SHP-2(IEC-KO) mice whereas Goblet cell number was clearly diminished. These alterations in Goblet/intermediate cell ratio were noticed 2 weeks after birth, before the onset of inflammation and were associated with significant alterations in microbiota composition. Indeed, an increase in Enterobacteriaceae and a decrease in Firmicutes were observed in the colon of these mice, indicating that dysbiosis also occurred prior to inflammation. Importantly, loss of epithelial Myd88 expression inhibited colitis development in SHP-2(IEC-KO) mice, rescued Goblet/intermediate cell ratio, and prevented NFκB hyperactivation and inflammation. These data indicate that SHP-2 is functionally important for the maintenance of appropriate barrier function and host-microbiota homeostasis in the large intestine. J. Cell. Physiol. 231: 2529-2540, 2016. © 2016 The Authors. Journal of Cellular Physiology published by Wiley Periodicals, Inc.


Assuntos
Diferenciação Celular , Colo/patologia , Homeostase , Inflamação/patologia , Inflamação/prevenção & controle , Microbiota , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Animais , Animais Recém-Nascidos , Peptídeos Catiônicos Antimicrobianos/metabolismo , Biomarcadores/metabolismo , Células Caliciformes/metabolismo , Células Caliciformes/patologia , Inflamação/genética , Camundongos Endogâmicos C57BL , Muramidase/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Celulas de Paneth/metabolismo , Celulas de Paneth/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/deficiência , Regulação para Cima/genética
20.
JMIR Res Protoc ; 5(1): e20, 2016 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-26869015

RESUMO

BACKGROUND: Type 2 diabetes is a major challenge for Canadian public health authorities, and regular physical activity is a key factor in the management of this disease. Given that less than half of people with type 2 diabetes in Canada are sufficiently active to meet the Canadian Diabetes Association's guidelines, effective programs targeting the adoption of regular physical activity are in demand for this population. Many researchers have argued that Web-based interventions targeting physical activity are a promising avenue for insufficiently active populations; however, it remains unclear if this type of intervention is effective among people with type 2 diabetes. OBJECTIVE: This research project aims to evaluate the effectiveness of two Web-based interventions targeting the adoption of regular aerobic physical activity among insufficiently active adult Canadian Francophones with type 2 diabetes. METHODS: A 3-arm, parallel randomized controlled trial with 2 experimental groups and 1 control group was conducted in the province of Quebec, Canada. A total of 234 participants were randomized at a 1:1:1 ratio to receive an 8-week, fully automated, computer-tailored, Web-based intervention (experimental group 1); an 8-week peer support (ie, Facebook group) Web-based intervention (experimental group 2); or no intervention (control group) during the study period. RESULTS: The primary outcome of this study is self-reported physical activity level (total min/week of moderate-intensity aerobic physical activity). Secondary outcomes are attitude, social influence, self-efficacy, type of motivation, and intention. All outcomes are assessed at baseline and 3 and 9 months after baseline with a self-reported questionnaire filled directly on the study websites. CONCLUSIONS: By evaluating and comparing the effectiveness of 2 Web-based interventions characterized by different behavior change perspectives, findings of this study will contribute to advances in the field of physical activity promotion in adult populations with type 2 diabetes. TRIAL REGISTRATION: International Standard Randomized Controlled Trial Number (ISRCTN): ISRCTN15747108; http://www.isrctn.com/ISRCTN15747108 (Archived by WebCite at http://www.webcitation.org/6eJTi0m3r).

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