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Proper lung function requires the maintenance of a tight endothelial barrier while simultaneously permitting the exchange of macromolecules and fluids to underlying tissue. Disruption of this barrier results in an increased vascular permeability in the lungs, leading to acute lung injury. In this study, we set out to determine whether transcriptional targets of Notch signaling function to preserve vascular integrity. We tested the in vivo requirement for Notch transcriptional signaling in maintaining the pulmonary endothelial barrier by using two complementary endothelial-specific Notch loss-of-function murine transgenic models. Notch signaling was blocked using endothelial-specific activation of an inhibitor of Notch transcriptional activation, Dominant Negative Mastermindlike (DNMAML; CDH5CreERT2), or endothelial-specific loss of Notch1 (Notch1f/f; CDH5CreERT2). Both Notch mutants increased vascular permeability with pan-Notch inhibition by DNMAML showing a more severe phenotype in the lungs and in purified endothelial cells. RNA sequencing of primary lung endothelial cells (ECs) identified novel Notch targets, one of which was transmembrane O-mannosyltransferase targeting cadherins 1 (tmtc1). We show that tmtc1 interacts with vascular endothelial cadherin (VE-cadherin) and regulates VE-cadherin egress from the endoplasmic reticulum through direct interaction. Our findings demonstrate that Notch signaling maintains endothelial adherens junctions and vascular homeostasis by a transcriptional mechanism that drives expression of critical factors important for processing and transport of VE-cadherin.
Assuntos
Antígenos CD , Caderinas , Células Endoteliais , Homeostase , Pulmão , Transdução de Sinais , Animais , Caderinas/metabolismo , Caderinas/genética , Camundongos , Células Endoteliais/metabolismo , Pulmão/metabolismo , Pulmão/irrigação sanguínea , Antígenos CD/metabolismo , Antígenos CD/genética , Humanos , Receptores Notch/metabolismo , Receptores Notch/genética , Camundongos Transgênicos , Permeabilidade Capilar , Receptor Notch1/metabolismo , Receptor Notch1/genética , Junções Aderentes/metabolismo , Camundongos Endogâmicos C57BLRESUMO
The mechanical environment generated through the adhesive interaction of endothelial cells (ECs) with the matrix controls nuclear tension, preventing aberrant gene synthesis and the transition from restrictive to leaky endothelium, a hallmark of acute lung injury (ALI). However, the mechanisms controlling tension transmission to the nucleus and EC-restrictive fate remain elusive. Here, we demonstrate that, in a kinase-independent manner, focal adhesion kinase (FAK) safeguards tension transmission to the nucleus to maintain EC-restrictive fate. In FAK-depleted ECs, robust activation of the RhoA-Rho-kinase pathway increased EC tension and phosphorylation of the nuclear envelope protein, emerin, activating DNMT3a. Activated DNMT3a methylates the KLF2 promoter, impairing the synthesis of KLF2 and its target S1PR1 to induce the leaky EC transcriptome. Repleting FAK (wild type or kinase dead) or inhibiting RhoA-emerin-DNMT3a activities in damaged lung ECs restored KLF2 transcription of the restrictive EC transcriptome. Thus, FAK sensing and control of tension transmission to the nucleus govern restrictive endothelium to maintain lung homeostasis.
Assuntos
Núcleo Celular , Células Endoteliais , Fatores de Transcrição Kruppel-Like , Transcriptoma , Proteína rhoA de Ligação ao GTP , Animais , Humanos , Camundongos , Núcleo Celular/metabolismo , DNA Metiltransferase 3A , Células Endoteliais/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Quinase 1 de Adesão Focal/genética , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Proteínas Nucleares/metabolismo , Proteínas Nucleares/genética , Fosforilação , Regiões Promotoras Genéticas/genética , Quinases Associadas a rho/metabolismo , Quinases Associadas a rho/genética , Proteína rhoA de Ligação ao GTP/metabolismo , Proteína rhoA de Ligação ao GTP/genética , Transcriptoma/genética , Masculino , FemininoRESUMO
Neutrophils (PMNs) reside as a marginated pool within the vasculature, ready for deployment during infection. However, how endothelial cells (ECs) control PMN extravasation and activation to strengthen tissue homeostasis remains ill-defined. Here, we found that the vascular ETS-related gene (ERG) is a generalized mechanism regulating PMN activity in preclinical tissue injury models and human patients. We show that ERG loss in ECs rewired PMN-transcriptome, enriched for genes associated with the CXCR2-CXCR4 signaling. Rewired PMNs compromise mice survival after pneumonia and induced lung vascular inflammatory injury following adoptive transfer into naïve mice, indicating their longevity and inflammatory activity memory. Mechanistically, EC-ERG restricted PMN extravasation and activation by upregulating the deubiquitinase A20 and downregulating the NFκB-IL8 cascade. Rescuing A20 in EC-Erg -/- endothelium or suppressing PMN-CXCR2 signaling rescued EC control of PMN activation. Findings deepen our understanding of EC control of PMN-mediated inflammation, offering potential avenues for targeting various inflammatory diseases. Highlights: ERG regulates trans-endothelial neutrophil (PMN) extravasation, retention, and activationLoss of endothelial (EC) ERG rewires PMN-transcriptomeAdopted transfer of rewired PMNs causes inflammation in a naïve mouse ERG transcribes A20 and suppresses CXCR2 function to inactivate PMNs. In brief/blurb: The authors investigated how vascular endothelial cells (EC) control polymorphonuclear neutrophil (PMN) extravasation, retention, and activation to strengthen tissue homeostasis. They showed that EC-ERG controls PMN transcriptome into an anti-adhesive and anti-inflammatory lineage by synthesizing A20 and suppressing PMNs-CXCR2 signaling, defining EC-ERG as a target for preventing neutrophilic inflammatory injury.
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Background: Access to safe drinking water, sanitation, and hygiene (WASH) facilities is crucial for health and human rights, impacting nutrition and weight. Methods: Multiple Indicators Cluster Survey (MICS) 2017-18 has been used in this study to examine the association between WASH and underweight, alongside other factors. Analysis included descriptive statistics, association tests, logistic regression, and population-attributable fractions (PAF). Results: According to results child were 1.8, 1.1 and 1.04 times less likely to be underweight if they had access to improved source of drinking water, improved sanitation and hygiene facilities respectively. The likelihood of child being underweight reduces by 1.4, 1.89, 2.01 and 2.55 times if the household wealth status increases from poorest to second, middle, fourth and richest wealth quintiles, respectively. As the mothers' education level increases from no schooling to primary, middle, secondary, and higher level, the possibility of child being underweight reduces by 1.22, 1.24, 1.60 and 2.01 times, respectively. Moreover, the likelihood of a child being underweight decreases as the education level of the household head improves. If maternal age is less than 20 or more than 35 years the likelihood of the child being underweight is increased by 1.074 and 1.121 times, respectively. A child is 1.1 times more likely to be underweight if birth spacing is less than 2 years. A child's risk of being underweight decreases by 1.1 times if they have not experienced diarrhea. A child who has never been breastfed has 1.3 times higher risk of being underweight. The results of Population Attributable Fraction (PAF) indicate that holding the other factors constant, approximately 36.46% burden of underweight was preventable by access to improved drinking water, sanitation, and hygiene practices. Conclusion: Comprehensive strategy is needed that focuses on improving access to safe drinking water, sanitation infrastructure, and hygiene behaviors.
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Cell surface G protein-coupled receptors (GPCRs), upon agonist binding, undergo serine-threonine phosphorylation, leading to either receptor recycling or degradation. Here, we show a new fate of GPCRs, exemplified by ER retention of sphingosine-1-phosphate receptor 1 (S1PR1). We show that S1P phosphorylates S1PR1 on tyrosine residue Y143, which is associated with recruitment of activated BiP from the ER into the cytosol. BiP then interacts with endocytosed Y143-S1PR1 and delivers it into the ER. In contrast to WT-S1PR1, which is recycled and stabilizes the endothelial barrier, phosphomimicking S1PR1 (Y143D-S1PR1) is retained by BiP in the ER and increases cytosolic Ca2+ and disrupts barrier function. Intriguingly, a proinflammatory, but non-GPCR agonist, TNF-α, also triggered barrier-disruptive signaling by promoting S1PR1 phosphorylation on Y143 and its import into ER via BiP. BiP depletion restored Y143D-S1PR1 expression on the endothelial cell surface and rescued canonical receptor functions. Findings identify Y143-phosphorylated S1PR1 as a potential target for prevention of endothelial barrier breakdown under inflammatory conditions.
Assuntos
Retículo Endoplasmático/genética , Inflamação/genética , Receptores de Esfingosina-1-Fosfato/genética , Fator de Necrose Tumoral alfa/genética , Citosol/metabolismo , Endocitose/genética , Chaperona BiP do Retículo Endoplasmático/química , Chaperona BiP do Retículo Endoplasmático/genética , Células Endoteliais/metabolismo , Humanos , Inflamação/patologia , Fosforilação/genética , Proteólise , Receptores Acoplados a Proteínas G/genética , Receptores de Esfingosina-1-Fosfato/metabolismo , Tirosina/genéticaRESUMO
Increased lung vascular permeability and neutrophilic inflammation are hallmarks of acute lung injury. Alveolar macrophages (AMÏ), the predominant sentinel cell type in the airspace, die in massive numbers while fending off pathogens. Recent studies indicate that the AMÏ pool is replenished by airspace-recruited monocytes, but the mechanisms instructing the conversion of recruited monocytes into reparative AMÏ remain elusive. Cyclic AMP (cAMP) is a vascular barrier protective and immunosuppressive second messenger in the lung. Here, we subjected mice expressing GFP under the control of the Lysozyme-M promoter (LysM-GFP mice) to the LPS model of rapidly resolving lung injury to address the impact of mechanisms determining cAMP levels in AMÏ and regulation of mobilization of the reparative AMÏ-pool. RNA-seq analysis of flow-sorted MÏ identified phosphodiesterase 4b (PDE4b) as the top LPS-responsive cAMP-regulating gene. We observed that PDE4b expression markedly increased at the time of peak injury (4 h) and then decreased to below the basal level during the resolution phase (24 h). Activation of transcription factor NFATc2 was required for the transcription of PDE4b in MÏ. Inhibition of PDE4 activity at the time of peak injury, using intratracheal rolipram, increased cAMP levels, augmented the reparative AMÏ pool, and resolved lung injury. This response was not seen following conditional depletion of monocytes, thus establishing airspace-recruited PDE4b-sensitive monocytes as the source of reparative AMÏ. Interestingly, adoptive transfer of rolipram-educated AMÏ into injured mice resolved lung edema. We propose suppression of PDE4b as an effective approach to promote reparative AMÏ generation from monocytes for lung repair.
Assuntos
Lesão Pulmonar Aguda/patologia , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Macrófagos Alveolares/citologia , Monócitos/citologia , Fatores de Transcrição NFATC/metabolismo , Transferência Adotiva/métodos , Animais , Permeabilidade Capilar/fisiologia , Diferenciação Celular/fisiologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Feminino , Inflamação , Lipopolissacarídeos/farmacologia , Macrófagos Alveolares/transplante , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Inibidores da Fosfodiesterase 4/farmacologia , Rolipram/farmacologia , Ativação Transcricional/genéticaRESUMO
We have recently uncovered that endothelial cell (EC) S1PR1 controls the effectiveness of VEGFR2 driven tumor angiogenesis. By using tumor ECs, EC-S1PR1-/- mice and S1PR1 antagonist, we showed that VEGF-VEGFR2 pathway requires EC-S1PR1-induced signaling to efficiently drive tumor vascularization and growth, indicating combining S1PR1 antagonist with anti-VEGF/VEGFR2 therapy may eradicate resistant tumors.
RESUMO
Sphingosine-1-phosphate receptor-1 (S1PR1), a G-protein coupled receptor that is expressed in endothelium and activated upon ligation by the bioactive lipid sphingosine-1-phosphate (S1P), is an important vascular-barrier protective mechanism at the level of adherens junctions (AJ). Loss of endothelial barrier function is a central factor in the pathogenesis of various inflammatory conditions characterized by protein-rich lung edema formation, such as acute respiratory distress syndrome (ARDS). While several S1PR1 agonists are available, the challenge of arresting the progression of protein-rich edema formation remains to be met. In this review, we discuss the role of S1PRs, especially S1PR1, in regulating endothelial barrier function. We review recent findings showing that replenishment of the pool of cell-surface S1PR1 may be crucial to the effectiveness of S1P in repairing the endothelial barrier. In this context, we discuss the S1P generating machinery and mechanisms that regulate S1PR1 at the cell surface and their impact on endothelial barrier function.
Assuntos
Endotélio/metabolismo , Receptores de Esfingosina-1-Fosfato/metabolismo , Animais , Humanos , Lisofosfolipídeos/metabolismo , Processamento de Proteína Pós-Traducional , Esfingosina/análogos & derivados , Esfingosina/metabolismoRESUMO
BACKGROUND: A huge array of function is played by the Wnt/ß-catenin signaling pathway in development by balancing gene expression through the modulation of cell-specific DNA binding downstream effectors such as T-cell factor/lymphoid enhancer factor (TCF/LEF). The ß-catenin/TCF-4 complex is a central regulatory switch for differentiation and proliferation of intestinal cells (both normal and malignant). Thus, in the present study we evaluated each of 60 cases of sporadic adenocarcinoma, alongside adjoining and normal mucosa specimens of colorectum in humans, for mutation and expression analysis of the gene coding for TCF-4 protein. METHODS: DNA sequencing following PCR amplification and SSCP analysis (single strand conformation polymorphism) was employed to detect TCF-4 gene mutations in the case of exon 1. Quantitative real-time (qRT) PCR, immunohistochemistry (IHC), confocal microscopy and western blot analysis were used to detect TCF-4 gene/protein expression. RESULTS: Sequencing analysis confirmed 5/60 patients with a point mutation in exon 1 of the TCF-4 gene in tumor samples. mRNA expression using qRT-PCR showed approximately 83% decreased TCF-4 mRNA expression in tumor tissue and adjoining mucosa compared to normal mucosa. Similarly, a significant decrease in protein expression using IHC showed decreased TCF-4 protein expression in tumor tissue and adjoining mucosa compared to normal mucosa, which also corresponds to some important clinicopathological factors, including disease metastasis and tumor grade. Mutational alterations and downregulation of TCF-4 mRNA and hence decreased expression of TCF-4 protein in tumors suggest its involvement in the pathogenesis of CRC. CONCLUSIONS: A remarkable decrease in TCF-4 mRNA and protein expression was detected in tumorous and adjoining tissues compared to normal mucosa. Hence the alterations in genomic architecture along with downregulation of TCF-4 mRNA and decreased expression of TCF-4 protein in tumors, which is in accordance with clinical features, suggest its involvement in the pathogenesis of CRC. Thus, deregulation and collaboration of TCF-4 with CRC could be a concrete and distinctive feature in the prognosis of the disease at an early stage of development.
Assuntos
Adenocarcinoma/metabolismo , Neoplasias Colorretais/metabolismo , Fator de Transcrição 4/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/patologia , Adenocarcinoma/secundário , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/genética , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Intervalo Livre de Doença , Regulação para Baixo , Éxons , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Imuno-Histoquímica , Masculino , Microscopia Confocal , Pessoa de Meia-Idade , Gradação de Tumores , Mutação Puntual , Prognóstico , Fator de Transcrição 4/genética , Proteínas Supressoras de Tumor/genética , beta Catenina/metabolismoRESUMO
The vascular endothelial growth factor-A (VEGF-A)-VEGFR2 pathway drives tumor vascularization by activating proangiogenic signaling in endothelial cells (ECs). Here, we show that EC-sphingosine-1-phosphate receptor 1 (S1PR1) amplifies VEGFR2-mediated angiogenic signaling to enhance tumor growth. We show that cancer cells induce S1PR1 activity in ECs, and thereby, conditional deletion of S1PR1 in ECs (EC-S1pr1-/- mice) impairs tumor vascularization and growth. Mechanistically, we show that S1PR1 engages the heterotrimeric G-protein Gi, which amplifies VEGF-VEGFR2 signaling due to an increase in the activity of the tyrosine kinase c-Abl1. c-Abl1, by phosphorylating VEGFR2 at tyrosine-951, prolongs VEGFR2 retention on the plasmalemma to sustain Rac1 activity and EC migration. Thus, S1PR1 or VEGFR2 antagonists, alone or in combination, reverse the tumor growth in control mice to the level seen in EC-S1pr1-/- mice. Our findings suggest that blocking S1PR1 activity in ECs has the potential to suppress tumor growth by preventing amplification of VEGF-VEGFR2 signaling.
Assuntos
Neoplasias Experimentais/metabolismo , Neovascularização Patológica/metabolismo , Transdução de Sinais , Receptores de Esfingosina-1-Fosfato/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Animais , Células Cultivadas , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Endotélio Vascular/metabolismo , Endotélio Vascular/patologia , Células HEK293 , Humanos , Masculino , Camundongos , Neoplasias Experimentais/patologia , Neuropeptídeos/metabolismo , Proteínas Proto-Oncogênicas c-abl/metabolismo , Proteínas rac1 de Ligação ao GTP/metabolismoRESUMO
Pakistani farmers are using groundwater at an increasing rate to supplement their irrigation needs. This practice has led to overexploitation of groundwater in the country, resulting in many negative externalities and increased resource costs. In response to the growing water shortage, the informal groundwater markets in the arid and semi-arid regions of Punjab have gradually emerged. These markets are believed to improve the fair distribution of groundwater and encourage more efficient use of agricultural water. This study aims to investigate these claims through conducting a field survey of 120 farmers that are further divided into three groups i.e. buyers, self-users cum sellers, and self-users (control group). Further, the study employed a Data Envelopment Analysis (DEA) approach to estimate the water use efficiency of all three type of groundwater actors. The study findings show that water buyers are mostly small farmers who do not own tube wells, hence buy water from tube well owners (large farmers). The study also found that groundwater markets improve the equity of water access to some extent, as water is transferred from large farmers to small farmers. The results of DEA analysis show water buyers and water sellers are more efficient in using water than the control group, making buyers the most efficient of all groups. Therefore, participation in water markets appears to be improving the WUE of farmers. The results of single bootstrapped truncated regression show that participation in water markets and access to extension services can improve WUE, while off-farm income and the diesel tube wells can reduce WUE in the study area. However, government could play an important role here through introducing groundwater regulations and improving water use efficiency for sustainable and equitable distribution of water among water users.
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Alveolar macrophages (AMs), upon sensing pathogens, trigger host defense by activating toll-like receptor 4 (TLR4), but the counterbalancing mechanisms that deactivate AM inflammatory signaling and prevent lethal edema, the hallmark of acute lung injury (ALI), remain unknown. Here, we demonstrate the essential role of AM protease-activating receptor 2 (PAR2) in rapidly suppressing inflammation to prevent long-lasting injury. We show that thrombin, released during TLR4-induced lung injury, directly activates PAR2 to generate cAMP, which abolishes Ca2+ entry through the TRPV4 channel. Deletion of PAR2 and thus the accompanying cAMP generation augments Ca2+ entry via TRPV4, causing sustained activation of the transcription factor NFAT to produce long-lasting TLR4-mediated inflammatory lung injury. Rescuing thrombin-sensitive PAR2 expression or blocking TRPV4 activity in PAR2-null AMs restores their capacity to resolve inflammation and reverse lung injury. Thus, activation of the thrombin-induced PAR2-cAMP cascade in AMs suppresses TLR4 inflammatory signaling to reinstate tissue integrity.
Assuntos
Sinalização do Cálcio , AMP Cíclico/metabolismo , Inflamação/prevenção & controle , Macrófagos Alveolares/metabolismo , Receptor PAR-2/metabolismo , Canais de Cátion TRPV/metabolismo , Receptor 4 Toll-Like/metabolismo , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Animais , Cálcio/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Inflamação/metabolismo , Lipopolissacarídeos/toxicidade , Macrófagos Alveolares/citologia , Macrófagos Alveolares/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Transcrição NFATC/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Receptor PAR-2/deficiência , Receptor PAR-2/genética , Canais de Cátion TRPV/antagonistas & inibidores , Canais de Cátion TRPV/genética , Trombina/metabolismoRESUMO
BACKGROUND: Fungal endophytes are the living symbionts which cause no apparent damage to the host tissue. The distribution pattern of these endophytes within a host plant is mediated by environmental factors. This study was carried out to explore the fungal endophyte community and their distribution pattern in Asparagus racemosus and Hemidesmus indicus growing in the study area. RESULTS: Foliar endophytes were isolated for 2 years from A. racemosus and H. indicus at four different seasons (June-August, September-November, December-February, March-May). A total of 5400 (675/season/year) leaf segments harbored 38 fungal species belonging to 17 genera, 12 miscellaneous mycelia sterile from 968 isolates and 13 had yeast like growth. In A. racemosus, Acremonium strictum and Phomopsis sp.1, were dominant with overall relative colonization densities (RCD) of 7.11% and 5.44% respectively, followed by Colletotrichum sp.3 and Colletotrichum sp.1 of 4.89% and 4.83% respectively. In H. indicus the dominant species was A. strictum having higher overall RCD of 5.06%, followed by Fusarium moniliforme and Colletotrichum sp.2 with RCD of 3.83% and 3%, respectively. Further the overall colonization and isolation rates were higher during the wet periods (September-November) in both A. racemosus (92.22% and 95.11%) and H. indicus (82% and 77.11%). CONCLUSION: Study samples treated with 0.2% HgCl2 and 75% EtOH for 30 s and 1 min, respectively, confirmed most favorable method of isolation of the endophytes. Owing to high mean isolation and colonization rates, September-November season proved to be the optimal season for endophyte isolation in both the study plants. Assessing the bioactive potential of these endophytes, may lead to the isolation of novel natural products and metabolites.
Assuntos
Asparagus/microbiologia , Endófitos/fisiologia , Fungos/fisiologia , Hemidesmus/microbiologia , Microbiota , Endófitos/classificação , Fungos/classificação , Índia , Folhas de Planta/microbiologia , Estações do AnoRESUMO
Radiation enteritis is one of the most feared complications of abdominal and pelvic regions. Thus, radiation to abdominal or pelvic malignancies unavoidably injures the intestine. Because of rapid cell turnover, the intestine is highly sensitive to radiation injury, which is the limiting factor in the permissible dosage of irradiation. Bowel injuries such as fistulas, strictures, and chronic malabsorption are potentially life-threatening complications and have an impact on patient quality of life. The incidence of radiation enteritis is increasing because of the current trend of combined chemotherapy and radiation. The consequences of radiation damage to the intestine may result in considerable morbidity and even mortality. The observed effects of ionizing radiation are mediated mainly by oxygen-free radicals that are generated by its action on water and are involved in several steps of signal transduction cascade, leading to apoptosis. The oxyradicals also induce DNA strand breaks and protein oxidation. An important line of defense against free radical damage is the presence of antioxidants. Therefore, administration of antioxidants may ameliorate the radiation-induced damage to the intestine.
RESUMO
ß-catenin (CTNNB1), an oncogene/onco-protein and an adhesion molecule is a key effector in colorectal cancer (CRC). Its activation, and subsequent up-regulation of Wnt-signaling, is an important event in the development of certain human cancers including CRC. Mutations in the ß-catenin gene in the region of serine-threonine glycogen kinase (GSK)-3ß phosphorylation target sites have been identified in colorectal cancer in humans. In the current study, we investigated 60 sporadic colorectal adenocarcinomas along with adjoining and normal mucosa cases in humans for ß-catenin mutations. Thirteen of sixty colorectal tumors from humans had point mutations with a frequency of 21.66% at codons 24, 26, 27, 32, 34, 35, 41, 42,43, 46, 49, 54, 55, or 67 sites which are mutated in colorectal cancer and some of these sites in other cancers. Thus, there appears to be a key involvement of ß-catenin activation in human colorectal carcinogenesis. mRNA expression analysis using q-Real Time PCR showed 21.5-fold up-regulation of ß-catenin mRNA in tumor tissue compared to normal and adjoining mucosa. Protein expression analysis using immunohistochemistry, confocal microscopy, and Western blot confirmed aberrant accumulation of ß-catenin protein along the nucleus and cytoplasm following mutation. The observed mutations and up-regulation of mRNA in tumors, and the increased expression of ß-catenin protein in CRC suggest that these alterations are early and prognostic events in sporadic colorectal carcinogenesis in humans. © 2015 Wiley Periodicals, Inc.
Assuntos
Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Análise Mutacional de DNA/métodos , Regulação para Cima , beta Catenina/genética , beta Catenina/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Núcleo Celular/metabolismo , Neoplasias Colorretais/metabolismo , Citoplasma/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Mutação Puntual , Prognóstico , Análise de Sobrevida , Adulto JovemRESUMO
BACKGROUND: Lactase activity declines during childhood in the majority of human populations leading to adult-type hypolactasia (AtH). C/T-13910 and G/A-22018 single nucleotide polymorphisms (SNPs) have been suggested to be associated with AtH in different human populations. Coeliac disease (CD) is an autoimmune condition characterized by damage to intestinal cells leading to ultimate deterioration. AIM: This study investigated the association between coeliac disease (CD) and SNPs leading to AtH in children from North India. SUBJECTS AND METHODS: Intestinal biopsies and saliva samples were obtained from 52 children with CD diagnosis and 102 control subjects. Biopsies were assayed for disaccharidase activities and samples were genotyped for given SNPs. RESULTS: Prevalence of C/C and G/G genotypes of AtH was almost equal in the CD and control group. The CD group had low lactase activity compared to the control group, irrespective of genotype at C/T -13910 and G/A -22018 SNPs (p < 0.05). For the control group, lactase activity was high in children with C/T + G/A genotypes compared to C/C + G/G (p < 0.05). CONCLUSION: There appears to be no significant correlation between C/T -13910 or G/A -22018 SNPs of AtH and CD. Children with C/C or G/G genotype of AtH may not be at greater risk of CD.
Assuntos
Doença Celíaca/genética , Lactase/deficiência , Lactase/genética , Intolerância à Lactose/genética , Doença Celíaca/epidemiologia , Doença Celíaca/imunologia , Criança , Pré-Escolar , Estudos de Coortes , Genótipo , Humanos , Índia/epidemiologia , Polimorfismo de Nucleotídeo ÚnicoRESUMO
This is the 12th in a series of articles exploring international trends in health science librarianship. This issue describes developments in health science librarianship in the first decade of the 21st century in South Asia. The three contributors report on challenges facing health science librarians in India, Pakistan and Sri Lanka. There is consensus as to the need for education, training and professional development. Starting in the next issue, the focus will turn to Africa, starting with countries in southern Africa. JM.
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Bibliotecas Médicas/tendências , Biblioteconomia/tendências , Redes de Comunicação de Computadores , Humanos , Índia , Serviços Técnicos de Biblioteca/tendências , Paquistão , Sri LankaRESUMO
To determine the etiological factors of human colorectal cancer (CRC) we assessed the frequency and prognostic significance of hMLH1 and hMSH2 genes in conjunction with hMLH1 and hMSH2 protein expression in 30 Indian CRC patients. The protein expression and promoter methylation of hMLH1 and hMSH2; Mismatch Repair genes (MMR) were analyzed by immunohistochemistry and methylation-specific PCR (MSP), respectively. A loss of hMLH1 expression was recognized in 4(13.3%) and loss of hMSH2 expression was recognized in 2(6.6%) of 30 CRC cases whereas 50% tumors showed reduced expression of hMLH1 and 33.3% showed reduced expression of hMSH2 protein. One tumor showed a loss of both hMLH1 and hMSH2 expression. Normal nuclear staining pattern of hMLH1 and hMSH2 was observed in almost all the adjoining and normal mucosa. Promoter hypermethylation of the hMLH1 gene was detected in 15 of 30 CRC cases (50%) and of hMSH2 gene was only in 3 of 30 CRC cases (10%). No promoter methylation of hMLH1 and hMSH2 genes was observed in adjoining and normal mucosa. Combination of methylation of hMLH1 and hMSH2 gene was observed in two tumors (6.6%). A significant correlation between histological grade of the tumor, methylation and expression of hMLH1 gene (p < 0.05) was observed. Normal expression of hMLH1 and hMSH2 was seen in all of the unmethylated tumors (100%). Nuclear staining and promoter methylation of hMLH1 and hMSH2 did not significantly influence survival. hMLH1 methylation was common and was significantly correlated with loss of hMLH1 protein expression. In contrast, hMSH2 methylation was infrequent. These findings suggest that the inactivation of MMR gene expression probably via hypermethylation may lead to inactivation of their functions which finally leads to tumor aggressiveness and the immunostaining of hMLH1 protein can be used as a prognostic factor for determining the grade of the tumor.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Neoplasias Colorretais/genética , Metilação de DNA , Proteína 2 Homóloga a MutS/genética , Proteínas Nucleares/genética , Regiões Promotoras Genéticas , Proteínas Adaptadoras de Transdução de Sinal/análise , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/mortalidade , Feminino , Humanos , Imuno-Histoquímica , Índia , Masculino , Pessoa de Meia-Idade , Proteína 1 Homóloga a MutL , Proteína 2 Homóloga a MutS/análise , Proteínas Nucleares/análise , Estudos ProspectivosRESUMO
PURPOSE: Intestinal mucosa, a rapidly proliferating tissue, is highly sensitive to radiation and undergoes apoptosis as a consequence of over generation of oxidative free radicals and the lack of the antioxidants. Thus the present study was designed to investigate the intestinal damage induced by radiation and to study if supplementation of the diet with antioxidant vitamins could ameliorate the intestinal damage and its digestive activity, as determined by the expression of various border enzymes. MATERIALS AND METHODS: Swiss Albino rats (150-200 g body weight) were divided into six groups. Group I: Control untreated; Group II: Irradiated; Group III: Irradiated + vitamin A; Group IV: Irradiated + vitamin C; Group V: Irradiated + vitamin E; and Group VI: Irradiated + lycopene. Animals were exposed to whole body γ-radiation from (60)Co at the rate of 8 Gy for 15 min/rat. Intestinal morphology and changes in various digestive enzymes together with, DNA damage was studied in six groups and each group consisted of 18 animals. RESULTS: The gastrointestinal toxicity resulted in malabsorption, diarrhoea, weight loss, loss of appetite, abdominal haemorrhage and hair loss. The activities of sucrase and alkaline phosphatase were elevated and those of lactase, leucine aminopeptidase (LAP) and gamma-glutamyl transpeptidase or tranferase (γ-GTP) were markedly reduced. Antioxidant vitamin A, C or E supplementations prevented changes in brush border enzyme activities as compared to lycopene administration in rat intestine by radiation exposure. Intestinal histology showed that the vitamin supplementation to irradiated rats minimized the intestinal damage in rats. CONCLUSION: These findings suggest that the epithelial lining of the intestine is highly sensitive to radiation exposure and supplementation of antioxidant vitamins is helpful in minimizing the intestinal damage and supplementation by vitamin E was most potent in ameliorating the intestinal aberrations.
Assuntos
Antioxidantes/farmacologia , Mucosa Intestinal/fisiopatologia , Mucosa Intestinal/efeitos da radiação , Intestinos/enzimologia , Animais , Ácido Ascórbico/farmacologia , Carotenoides/farmacologia , Radioisótopos de Cobalto/química , Dano ao DNA , Suplementos Nutricionais , Enzimas/biossíntese , Radicais Livres/química , Mucosa Intestinal/efeitos dos fármacos , Intestinos/efeitos dos fármacos , Intestinos/efeitos da radiação , Licopeno , Microvilosidades/efeitos dos fármacos , Microvilosidades/efeitos da radiação , Oxigênio/química , Lesões por Radiação/prevenção & controle , Radioterapia/efeitos adversos , Ratos , Vitamina A/farmacologia , Vitamina E/farmacologiaRESUMO
The incidence of colorectal cancer (CRC) is increasing rapidly in Asian countries during the past few decades, but no comprehensive analysis has been done to find out the exact cause of this disease. In this study, we investigated the frequencies of mutations and expression pattern of K-ras, APC (adenomatosis polyposis coli) and p53 in tumor, adjoining and distant normal mucosa and to correlate these alterations with patients clinicopathological parameters as well as with the survival. Polymerase chain reaction (PCR)-restriction digestion was used to detect mutations in K-ras and PCR-SSCP (Single Strand Conformation Polymorphism) followed by DNA sequencing was used to detect mutations in APC and p53 genes. Immunohistochemistry was used to detect the expression pattern of K-ras, APC and p53 proteins. The frequencies of mutations of K-ras, APC and p53 in 30 tumor tissues samples were 26.7 %, 46.7 % and 20 %, respectively. Only 3.3 % of tumors contained mutations in all the three genes. The most common combination of mutation was APC and p53 whereas mutation in both p53 and K-ras were extremely rare. There was no association between the mutations and expression pattern of K-ras, APC and p53 (p>0.05). In Indians, the frequency of alterations of K-ras and APC is similar as in Westerns, whereas the frequency of p53 mutation is slightly lower. The lack of multiple mutations in tumor specimens suggests that these genetic alterations might have independent influences on CRC development and there could be multiple alternative genetic pathways to CRC in our present study cohort.