180 research outputs found

    Mesenteric lymph node transcriptome profiles in BALB/c mice sensitized to three common food allergens

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    <p>Abstract</p> <p>Background</p> <p>Food allergy is a serious health concern among infants and young children. Although immunological mechanism of food allergy is well documented, the molecular mechanism(s) involved in food allergen sensitization have not been well characterized. Therefore, the present study analyzed the mesenteric lymph node (MLN) transcriptome profiles of BALB/c mice in response to three common food allergens.</p> <p>Results</p> <p>Microarray analysis identified a total of 1361, 533 and 488 differentially expressed genes in response to β-lactoglobulin (BLG) from cow's milk, ovalbumin (OVA) from hen's egg white and peanut agglutinin (PNA) sensitizations, respectively (p < 0.05). A total of 150 genes were commonly expressed in all antigen sensitized groups. The expression of seven representative genes from microarray experiment was validated by real-time RT-PCR. All allergens induced significant ear swelling and serum IgG1 concentrations, whereas IgE concentrations were increased in BLG- and PNA-treated mice (p < 0.05). Treatment with OVA and PNA significantly induced plasma histamine concentrations (p < 0.05). The PCA demonstrated the presence of allergen-specific IgE in the serum of previously sensitized and challenged mice.</p> <p>Conclusions</p> <p>Immunological profiles indicate that the allergen dosages used are sufficient to sensitize the BALB/c mice and to conduct transcriptome profiling. Microarray studies identified several differentially expressed genes in the sensitization phase of the food allergy. These findings will help to better understand the underlying molecular mechanism(s) of food allergen sensitizations and may be useful in identifying the potential biomarkers of food allergy.</p

    Involvement of Cox-2 in the metastatic potential of chemotherapy-resistant breast cancer cells

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    <p>Abstract</p> <p>Background</p> <p>A major problem with the use of current chemotherapy regimens for several cancers, including breast cancer, is development of intrinsic or acquired drug resistance, which results in disease recurrence and metastasis. However, the mechanisms underlying this drug resistance are unknown. To study the molecular mechanisms underlying the invasive and metastatic activities of drug-resistant cancer cells, we generated a doxorubicin-resistant MCF-7 breast cancer cell line (MCF-7/DOX).</p> <p>Methods</p> <p>We used MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays, flow cytometry assays, DNA fragmentation assays, Western blot analysis, cell invasion assays, small interfering RNA (siRNA) transfection, reverse transcription-polymerase chain reaction, experimental lung metastasis models, and gelatin and fibrinogen/plasminogen zymography to study the molecular mechanism of metastatic activities in MCF-7/DOX cells.</p> <p>Results</p> <p>We found that MCF-7/DOX acquired invasive activities. In addition, Western blot analysis showed increased expression of epidermal growth factor receptor (EGFR) and Cox-2 in MCF-7/DOX cells. Inhibition of Cox-2, phosphoinositide 3-kinase (PI3K)/Akt, or mitogen-activated protein kinase (MAPK) pathways effectively inhibited the invasive activities of MCF-7/DOX cells. Gelatin and fibrinogen/plasminogen zymography analysis showed that the enzymatic activities of matrix metalloproteinase-2 (MMP-2), MMP-9, and urokinase-type plasminogen activator were markedly higher in MCF-7/DOX cells than in the MCF-7 cells. <it>In vitro </it>invasion assays and mouse models of lung metastasis demonstrated that MCF-7/DOX cells acquired invasive abilities. Using siRNAs and agonists specific for prostaglandin E (EP) receptors, we found that EP1 and EP3 played important roles in the invasiveness of MCF-7/DOX cells.</p> <p>Conclusions</p> <p>We found that the invasive activity of MCF-7/DOX cells is mediated by Cox-2, which is induced by the EGFR-activated PI3K/Akt and MAPK pathways. In addition, EP1 and EP3 are important in the Cox-2-induced invasion of MCF-7/DOX cells. Therefore, not only Cox-2 but also EP1 and EP3 could be important targets for chemosensitization and inhibition of metastasis in breast cancers that are resistant to chemotherapy.</p

    Heterochronic Shift in Hox-Mediated Activation of Sonic hedgehog Leads to Morphological Changes during Fin Development

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    We explored the molecular mechanisms of morphological transformations of vertebrate paired fin/limb evolution by comparative gene expression profiling and functional analyses. In this study, we focused on the temporal differences of the onset of Sonic hedgehog (Shh) expression in paired appendages among different vertebrates. In limb buds of chick and mouse, Shh expression is activated as soon as there is a morphological bud, concomitant with Hoxd10 expression. In dogfish (Scyliorhinus canicula), however, we found that Shh was transcribed late in fin development, concomitant with Hoxd13 expression. We utilized zebrafish as a model to determine whether quantitative changes in hox expression alter the timing of shh expression in pectoral fins of zebrafish embryos. We found that the temporal shift of Shh activity altered the size of endoskeletal elements in paired fins of zebrafish and dogfish. Thus, a threshold level of hox expression determines the onset of shh expression, and the subsequent heterochronic shift of Shh activity can affect the size of the fin endoskeleton. This process may have facilitated major morphological changes in paired appendages during vertebrate limb evolution

    The application of cortical layer markers in the evaluation of cortical dysplasias in epilepsy

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    The diagnostic criteria for focal cortical dysplasia type I (FCD I) remain to be well and consistently defined. Cortical layer-specific markers (CLM) provide a potential tool for the objective assessment of any dyslamination. We studied expression patterns of recognised CLM using immunohistochemistry for N200, ER81, Otx1, Map1b (subsets of V/VI projection neurones), Pax6, Tbr1, Tbr2 (differentially expressed in cortical neurones from intermediate progenitor cells), Cux 1 (outer cortical layers) and MASH1 (ventricular zone progenitors). Dysplasia subtypes included FCD I and II, dysplasias adjacent to hippocampal sclerosis (HS) or dysembryoplastic neuroepithelial tumours (DNTs); all were compared to neonatal and adult controls. Laminar expression patterns in normal cortex were observed with Tbr1, Map1b, N200 and Otx1. FCDI cases in younger patients were characterised by abnormal expression in layer II for Tbr1 and Otx1. FCDII showed distinct labelling of balloon cells (Pax6, ER81 and Otx1) and dysmorphic neurones (Tbr 1, N200 and Map1b) supporting origins from radial glia and intermediate progenitor cells, respectively. In temporal lobe sclerosis cases with dysplasia adjacent to HS, Tbr1 and Map1b highlighted abnormal orientation of neurones in layer II. Dyslamination was not confirmed in the perilesional cortex of DNT with CLM. Finally, immature cell types (Otx1, Pax6 and Tbr2) were noted in varied pathologies. One possibility is activation of progenitor cell populations which could contribute to the pathophysiology of these lesions

    An Essential Regulatory Role of Downstream of Kinase-1 in the Ovalbumin-Induced Murine Model of Asthma

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    The downstream of kinase (DOK)-1 is involved in the protein tyrosine kinase (PTK) pathway in mast cells, but the role of DOK-1 in the pathogenesis of asthma has not been defined. In this study, we have demonstrated a novel regulatory role of DOK-1 in airway inflammation and physiologic responses in a murine model of asthma using lentiviral vector containing DOK-1 cDNA or DOK-1-specific ShRNA. The OVA-induced inflammatory cells, airway hyperresponsiveness, Th2 cytokine expression, and mucus response were significantly reduced in DOK-1 overexpressing mice compared to OVA-challenged control mice. The transgenic introduction of DOK-1 significantly stimulated the activation and expression of STAT-4 and T-bet, while impressively inhibiting the activation and expression of STAT-6 and GATA-3 in airway epithelial cells. On the other hand, DOK-1 knockdown mice enhanced STAT-6 expression and its nuclear translocation compared to OVA-challenged control mice. When viewed in combination, our studies demonstrate DOK-1 regulates allergen-induced Th2 immune responses by selective stimulation and inhibition of STAT-4 and STAT-6 signaling pathways, respectively. These studies provide a novel insight on the regulatory role of DOK-1 in allergen-induced Th2 inflammation and airway responses, which has therapeutic potential for asthma and other allergic diseases

    Mycobacterium marinum antagonistically induces an autophagic response while repressing the autophagic flux in a TORC1- and ESX-1-dependent manner.

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    Autophagy is a eukaryotic catabolic process also participating in cell-autonomous defence. Infected host cells generate double-membrane autophagosomes that mature in autolysosomes to engulf, kill and digest cytoplasmic pathogens. However, several bacteria subvert autophagy and benefit from its machinery and functions. Monitoring infection stages by genetics, pharmacology and microscopy, we demonstrate that the ESX-1 secretion system of Mycobacterium marinum, a close relative to M. tuberculosis, upregulates the transcription of autophagy genes, and stimulates autophagosome formation and recruitment to the mycobacteria-containing vacuole (MCV) in the host model organism Dictyostelium. Antagonistically, ESX-1 is also essential to block the autophagic flux and deplete the MCV of proteolytic activity. Activators of the TORC1 complex localize to the MCV in an ESX-1-dependent manner, suggesting an important role in the manipulation of autophagy by mycobacteria. Our findings suggest that the infection by M. marinum activates an autophagic response that is simultaneously repressed and exploited by the bacterium to support its survival inside the MCV

    Dysregulation of Gene Expression in a Lysosomal Storage Disease Varies between Brain Regions Implicating Unexpected Mechanisms of Neuropathology

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    The characteristic neurological feature of many neurogenetic diseases is intellectual disability. Although specific neuropathological features have been described, the mechanisms by which specific gene defects lead to cognitive impairment remain obscure. To gain insight into abnormal functions occurring secondary to a single gene defect, whole transcriptome analysis was used to identify molecular and cellular pathways that are dysregulated in the brain in a mouse model of a lysosomal storage disorder (LSD) (mucopolysaccharidosis [MPS] VII). We assayed multiple anatomical regions separately, in a large cohort of normal and diseased mice, which greatly increased the number of significant changes that could be detected compared to past studies in LSD models. We found that patterns of aberrant gene expression and involvement of multiple molecular and cellular systems varied significantly between brain regions. A number of changes revealed unexpected system and process alterations, such as up-regulation of the immune system with few inflammatory changes (a significant difference from the closely related MPS IIIb model), down-regulation of major oligodendrocyte genes even though white matter changes are not a feature histopathologically, and a plethora of developmental gene changes. The involvement of multiple neural systems indicates that the mechanisms of neuropathology in this type of disease are much broader than previously appreciated. In addition, the variation in gene dysregulation between brain regions indicates that different neuropathologic mechanisms may predominate within different regions of a diseased brain caused by a single gene mutation

    Chromosomal organization at the level of gene complexes

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    Metazoan genomes primarily consist of non-coding DNA in comparison to coding regions. Non-coding fraction of the genome contains cis-regulatory elements, which ensure that the genetic code is read properly at the right time and space during development. Regulatory elements and their target genes define functional landscapes within the genome, and some developmentally important genes evolve by keeping the genes involved in specification of common organs/tissues in clusters and are termed gene complex. The clustering of genes involved in a common function may help in robust spatio-temporal gene expression. Gene complexes are often found to be evolutionarily conserved, and the classic example is the hox complex. The evolutionary constraints seen among gene complexes provide an ideal model system to understand cis and trans-regulation of gene function. This review will discuss the various characteristics of gene regulatory modules found within gene complexes and how they can be characterized
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