scholarly journals Differential Expression and Localization of Branchial AQP1 and AQP3 in Japanese Medaka (Oryzias latipes)

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 422 ◽  
Author(s):  
Laura V. Ellis ◽  
Rebecca J. Bollinger ◽  
Hannah M. Weber ◽  
Steffen S. Madsen ◽  
Christian K. Tipsmark
Keyword(s):  
Epithelial Cells ◽  
Ex Vivo ◽  
Gill Filament ◽  
Oryzias Latipes ◽  
Japanese Medaka ◽  
Dependent Manner ◽  
Protein Levels ◽  
Low Intensity ◽  
Aqp1 Expression ◽  
Gill Lamellae

Aquaporins (AQPs) facilitate transmembrane water and solute transport, and in addition to contributing to transepithelial water transport, they safeguard cell volume homeostasis. This study examined the expression and localization of AQP1 and AQP3 in the gills of Japanese medaka (Oryzias latipes) in response to osmotic challenges and osmoregulatory hormones, cortisol, and prolactin (PRL). AQP3 mRNA was inversely regulated in response to salinity with high levels in ion-poor water (IPW), intermediate levels in freshwater (FW), and low levels in seawater (SW). AQP3 protein levels decreased upon SW acclimation. By comparison, AQP1 expression was unaffected by salinity. In ex vivo gill incubation experiments, AQP3 mRNA was stimulated by PRL in a time- and dose-dependent manner but was unaffected by cortisol. In contrast, AQP1 was unaffected by both PRL and cortisol. Confocal microscopy revealed that AQP3 was abundant in the periphery of gill filament epithelial cells and co-localized at low intensity with Na+,K+-ATPase in ionocytes. AQP1 was present at a very low intensity in most filament epithelial cells and red blood cells. No epithelial cells in the gill lamellae showed immunoreactivity to AQP3 or AQP1. We suggest that both AQPs contribute to cellular volume regulation in the gill epithelium and that AQP3 is particularly important under hypo-osmotic conditions, while expression of AQP1 is constitutive.

scholarly journals Cross-talk between LAM cells and fibroblasts may influence alveolar epithelial cell behavior in Lymphangioleiomyomatosis

2021 ◽  
Author(s):  
Debbie Clements ◽  
Suzanne Miller ◽  
Roya Babaei-Jadidi ◽  
Mike Adam ◽  
S. Steven Potter ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cross Talk ◽  
Ex Vivo ◽  
Alveolar Epithelial Cell ◽  
Alveolar Epithelial Cells ◽  
Dependent Manner ◽  
Epithelial Migration ◽  
Alveolar Epithelial

Lymphangioleiomyomatosis (LAM) is a female specific cystic lung disease in which TSC2 deficient LAM cells, LAM-Associated Fibroblasts (LAFs) and other cell types infiltrate the lungs. LAM lesions can be associated with type II alveolar epithelial cells (AT2 cells). We hypothesised that the behaviour of AT2 cells in LAM is influenced locally by LAFs. We tested this hypothesis in patient samples and in vitro. In human LAM lung, nodular AT2 cells show enhanced proliferation when compared to parenchymal AT2 cells, demonstrated by increased Ki67 expression. Further, nodular AT2 cells express proteins associated with epithelial activation in other disease states including Matrix Metalloproteinase 7, and Fibroblast Growth Factor 7 (FGF7). In vitro, LAF conditioned medium is mitogenic and positively chemotactic for epithelial cells, increases the rate of epithelial repair and protects against apoptosis. In vitro, LAM patient-derived TSC2 null cells cocultured with LAFs upregulate LAF expression of the epithelial chemokine and mitogen FGF7, which is a potential mediator of fibroblast-epithelial crosstalk, in an mTOR dependent manner. In a novel in vitro model of LAM, ex vivo cultured LAM lung-derived microtissues promote both epithelial migration and adhesion. Our findings suggest that AT2 cells in LAM display a proliferative, activated phenotype and that fibroblast accumulation following LAM cell infiltration into the parenchyma contributes to this change in AT2 cell behaviour. Fibroblast-derived FGF7 may contribute to the cross-talk between LAFs and hyperplastic epithelium in vivo, but does not appear to be the main driver of the effects of LAFs on epithelial cells in vitro.

scholarly journals Involvement of the Extracellular Matrix Proteins Periostin and Tenascin C in Nasal Polyp Remodeling by Regulating the Expression of MMPs

2020 ◽  
Author(s):  
Kun Du ◽  
Min Wang ◽  
Nan Zhang ◽  
Ping Wang ◽  
Pei Yu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Chronic Rhinosinusitis ◽  
Nasal Polyp ◽  
Nasal Polyps ◽  
Ex Vivo ◽  
Tissue Remodeling ◽  
Tenascin C ◽  
Protein Levels ◽  
Control Subjects

Abstract Background: Tissue remodeling caused by increased MMPs is involved in the pathogenesis of chronic rhinosinusitis with nasal polyposis (CRSwNP). We previously found higher levels of periostin and tenascin C in CRSwNPs, but whether they are associated with the dysregulation of MMPs is unknown. Therefore, the present study aimed to investigate the regulatory roles of two ECM proteins in the expression of MMPs in nasal polyps.Methods：The concentrations of MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, MMP-12, MMP-13, TIMP-1, TIMP-2, TIMP-3, TIMP-4, periostin, and tenascin C in tissue homogenates of 51 patients with chronic rhinosinusitis with and without nasal polyps and 15 control subjects were measured and their correlations were analyzed. Primary human nasal polyp fibroblasts and epithelial cells were stimulated ex vivo with periostin and tenascin C and the gene expression of MMPs and TIMPs was determined by means of real-time PCR.Results: The protein levels of MMP-3, MMP-7, MMP-8, MMP-9, TIMP-1, TIMP-2, periostin, and tenascin C were significantly higher in patients with CRSwNPs than in healthy control subjects. Periostin was positively correlated with MMP-3 and TIMP-2, and tenascin C was positively correlated with MMP-3, MMP-7, MMP-8, MMP-9 and TIMP-2. Periostin stimulated the gene expression of MMP-3, MMP-7, and MMP-9 in fibroblasts and MMP-7 in epithelial cells ex vivo. Tenascin C stimulated the expression of MMP-3, MMP-8, and MMP-9 in epithelial cells, but not in fibroblasts. The expression of TIMPs in fibroblasts and epithelial cells was affected by neither periostin nor tenascin C. Conclusions：Periostin and tenascin C might be involved in the remodeling of nasal polyps by regulating the expression of different MMPs in epithelial cells and fibroblasts. Our findings have the potential to identify key factors of tissue remodeling in CRSwNPs.

Ex vivo maintenance of hematopoietic stem cells by quiescence induction through Fbxw7α overexpression

Blood ◽  
2011 ◽  
Vol 117 (8) ◽  
pp. 2373-2377 ◽  
Author(s):  
Hirono Iriuchishima ◽  
Keiyo Takubo ◽  
Sahoko Matsuoka ◽  
Ichiro Onoyama ◽  
Keiichi I. Nakayama ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
Dependent Manner ◽  
Essential Factor ◽  
Hematopoietic Stem ◽  
Protein Levels ◽  
Lineage Marker

Abstract Cell-cycle quiescence in hematopoietic stem cells (HSCs) is essential for maintaining stemness by protecting cells from differentiation or senescence. F-box and WD-40 domain protein 7 (Fbxw7) maintains HSCs and suppresses leukemogenesis by mediating ubiquitin-dependent degradation of cell-cycle activators and oncoproteins. Fbxw7α was shown to be the preferentially expressed Fbxw7 isoform in primitive HSCs. Forced Fbxw7α expression in lineage marker Sca-1+c-Kit+ cells led to cell-cycle dormancy by reducing the protein levels of the Fbxw7 substrates c-Myc, Notch1, and phosphorylated S6 (a key downstream element of mTOR). Hypoxia, an essential factor for HSC quiescence, suppressed c-Myc in an Fbxw7α-dependent manner. Fbxw7α-overexpressing lineage marker Sca-1+c-Kit+ cells sustained high reconstitution capacities during in vitro culture. These data suggest that Fbxw7α sustains HSC dormancy through c-Myc, Notch1, and the mTOR pathways. The modulation of Fbxw7α expression or activity represents a promising new tool for ex vivo HSC maintenance.

scholarly journals Galectin-9 Produced by Intestinal Epithelial Cells Enhances Aldehyde Dehydrogenase Activity in Dendritic Cells in a PI3K- and p38-Dependent Manner

2017 ◽  
Vol 9 (6) ◽  
pp. 609-620 ◽  
Author(s):  
Sander de Kivit ◽  
Atanaska I. Kostadinova ◽  
JoAnn Kerperien ◽  
Veronica Ayechu Muruzabal ◽  
Mary E. Morgan ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Epithelial Cells ◽  
Aldehyde Dehydrogenase ◽  
Intestinal Epithelial Cells ◽  
Ex Vivo ◽  
Dependent Manner ◽  
Intestinal Epithelial ◽  
Aldh Activity ◽  
Cpg Dna ◽  
Gm Csf

Intestinal epithelial cells (IEC) drive regulatory T cell (Treg) responses by promoting the differentiation of aldehyde dehydrogenase (ALDH)-expressing CD103+ dendritic cells (DC). Apical stimulation of TLR9 by CpG DNA on IEC supports galectin-9 expression by IEC, which is promoted by short-chain galacto-oligosaccharides and long-chain fructo-oligosaccharides (GF). While galectin-9 can induce the maturation of monocyte-derived DC (moDC), the contribution of galectin-9 on the induction of ALDH activity in DC is not known. To this end, DC were stimulated with galectin-9, and ALDH activity and the expression of CD103 were assessed. ALDH activity was increased by moDC exposed to galectin-9, while the expression of CD103 remained unaltered. Galectin-9 secreted by IEC apically exposed to CpG DNA and GF enhanced ALDH activity, but not CD103 expression by moDC, which was abrogated upon galectin-9 neutralization. Similar observations were found in murine GM-CSF-cultured bone marrow-derived DC (BMDC). Using Flt3L-cultured BMDC and ex vivo murine splenic DC, it was observed that galectin-9 only enhanced ALDH activity in the presence of GM-CSF in CD103- cells. The induction of ALDH activity in BMDC was dependent on p38 and PI3K signaling. These data indicate a novel role for galectin-9 in modulating innate immunity by inducing ALDH activity in DC.

scholarly journals Elevated microRNA-21 Is a Brake of Inflammation Involved in the Development of Nasal Polyps

2021 ◽  
Vol 12 ◽  
Author(s):  
Ruowu Liu ◽  
Jintao Du ◽  
Jiao Zhou ◽  
Bing Zhong ◽  
Luo Ba ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Ex Vivo ◽  
Luciferase Reporter ◽  
Reporter System ◽  
Nasal Epithelial Cells ◽  
Protein Levels ◽  
Control Subjects ◽  
Luciferase Reporter System ◽  
Suppressive Cytokine

BackgroundCRSwNP is an inflammatory disease but the mechanism is not yet fully understood. MiR-21, a member of miRNAs, has been reported to play roles in mediating inflammation. However, the expression of miR-21 and its role in patients with CRSwNP remain elusive.MethodsTurbinates from control subjects, uncinate processes from CRSsNP, polyp tissues from CRSwNP, and nasal epithelial cells brushed from nasal mucosa were collected. The expression of miR-21 and cytokines in nasal tissues and epithelial cells were detected by qPCR. The localization of miR-21 was detected by ISH, and its target was identified by bioinformation analysis, qPCR, IHC, WB, and luciferase reporter system. The protein and mRNA of PDCD4 and NF-κB P65 were determined by WB and qPCR after miR-21 transfection in HNEpC. The role of miR-21 on cytokines was analyzed in HNEpC and nasal polyp explants.ResultsMiR-21 was upregulated in CRSwNP relative to control subjects by qPCR, which was determined mainly in nasal epithelial cells of CRSwNP by ISH. Both pro-inflammation cytokines (IL-1β, IL-6, IL-8, IL-25, and TSLP) and a suppressive cytokine (IL-10) were overexpressed in the epithelial cells of CRSwNP. The expression of miR-21 was positively correlated with IL-10 and negatively correlated with IL-6, IL-8, IL-33, and TSLP in the epithelial cells of CRSwNP. As a potential target of miR-21, the expression of PDCD4 was negatively correlated with miR-21 in CRSwNP. In HNEpC, miR-21 could reduce the expression of PDCD4 at both mRNA and protein levels, and bioinformation analysis and luciferase reporter system confirmed PDCD4 as one target of miR-21. Furthermore, miR-21 could decrease the activation of NF-κB and increase IL-10 mRNA. Both SEB and LPS could elevate miR-21, with IL-25, IL-33, TSLP induced by SEB and IL-1β, IL-6, IL-8 induced by LPS, while the miR-21 could regulate the expression of IL-33, TSLP, IL-1β, IL- 6 and IL-8 in vitro and ex vivo. Clinically, miR-21 expression was inversely correlated with the Lund-Mackay CT scores and the Lund-Kennedy scores in CRSwNP.ConclusionMiR-21 could be a prominent negative feedback factor in the inflammation process to attenuate the expression of pro-inflammatory cytokines, thereby playing an anti-inflammation role in CRSwNP.

scholarly journals Upregulation of MUC5AC by VEGF in human primary bronchial epithelial cells: implications for asthma

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Sung-Ho Kim ◽  
Qing-Mei Pei ◽  
Ping Jiang ◽  
Juan Liu ◽  
Rong-Fei Sun ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Bronchial Epithelial Cells ◽  
Cholesterol Depletion ◽  
Vegf Receptor ◽  
Dependent Manner ◽  
Bronchial Epithelial ◽  
Caveolin 1 ◽  
Protein Levels ◽  
Rhoa Activation ◽  
Primary Bronchial Epithelial Cells

Abstract Background Airway mucus hypersecretion is an important pathophysiological feature in asthma. Mucins are glycoproteins that are mainly responsible for the viscoelastic property of mucus, and MUC5AC is a major mucin glycoprotein that is overproduced in asthma. Vascular endothelial growth factor (VEGF) has been implicated in inflammatory and airway blood vessel remodeling in asthmatics. Therefore, we sought to investigate the effect of VEGF on MUC5AC expression and study the underlying mechanisms. Methods In order to elucidate the precise mechanism underlying the effect of VEGF on MUC5AC expression, we tested the effects of VEGF on RhoA activation and the association of caveolin-1 and VEGFR2 in Primary Bronchial Epithelial Cells. Results VEGF up-regulated MUC5AC mRNA and protein levels in a dose- and time-dependent manner, and activated RhoA. Additionally, VEGF-induced MUC5AC expression and RhoA activation were enhanced by disrupting caveolae with cholesterol depletion and reversed by cholesterol repletion, and inhibited by a selective VEGF receptor 2 (VEGFR2) inhibitor SU1498. Furthermore, phospho-VEGFR2 expression was decreased via overexpression of caveolin-1. VEGF treatment reduced the association of caveolin-1 and VEGFR2. Conclusion Collectively, our findings suggest that VEGF up-regulates MUC5AC expression and RhoA activation by interaction with VEGFR2, and this phenomenon was related with the association of caveolin-1 and VEGFR2. Further studies on these mechanisms are needed to facilitate the development of treatments for asthma.

Use of the Japanese Medaka ( Oryzias latipes ) and Guppy ( Poecilia reticulata ) in Carcinogenesis Testing Under National Toxicology Program Protocols

2003 ◽  
Vol 31 (1) ◽  
pp. 88-91 ◽  
Author(s):  
William E. Hawkins ◽  
William W. Walker ◽  
John W. Fournie ◽  
C. Steve Manning ◽  
Rena M. Krol
Keyword(s):  
Poecilia Reticulata ◽  
Oryzias Latipes ◽  
Japanese Medaka ◽  
National Toxicology Program

The Natural Flavonoid Naringenin Inhibits the Cell Growth of WilmsTumorinChildren by Suppressing TLR4/NF-κB Signaling

2020 ◽  
Vol 20 ◽  
Author(s):  
Hongtao Li ◽  
Peng Chen ◽  
Lei Chen ◽  
Xinning Wang
Keyword(s):  
Cell Growth ◽  
Western Blot ◽  
Wilms Tumor ◽  
Critical Role ◽  
Time Dependent ◽  
Luciferase Assay ◽  
Dependent Manner ◽  
Tlr4 Expression ◽  
Protein Levels

Background: Nuclear factor kappa B (NF-κB) is usually activated in Wilms tumor (WT) cells and plays a critical role in WT development. Objective: The study purpose was to screen a NF-κB inhibitor from natural product library and explore its effects on WT development. Methods: Luciferase assay was employed to assess the effects of natural chemical son NF-κB activity. CCK-8 assay was conducted to assess cell growth in response to naringenin. WT xenograft model was established to analyze the effect of naringenin in vivo. Quantitative real-time PCR and Western blot were performed to examine the mRNA and protein levels of relative genes, respectively. Results: Naringenin displayed significant inhibitory effect on NF-κB activation in SK-NEP-1 cells. In SK-NEP-1 and G-401 cells, naringenin inhibited p65 phosphorylation. Moreover, naringenin suppressed TNF-α-induced p65 phosphorylation in WT cells. Naringenin inhibited TLR4 expression at both mRNA and protein levels in WT cells. CCK-8 staining showed that naringenin inhibited cell growth of the two above WT cells in dose-and time-dependent manner, whereas Toll-like receptor 4 (TLR4) over expression partially reversed the above phenomena. Besides, naringenin suppressed WT tumor growth in dose-and time-dependent manner in vivo. Western blot found that naringenin inhibited TLR4 expression and p65 phosphorylation in WT xenograft tumors. Conclusion: Naringenin inhibits WT development viasuppressing TLR4/NF-κB signaling

Platelet-Rich and Platelet-Poor Plasma Might Play Supportive Roles in Cancer Cell Culture: A Replacement for Fetal Bovine Serum?

2021 ◽  
Vol 21 ◽  
Author(s):  
Mehdi Talebi ◽  
Mousa Vatanmakanian ◽  
Ali Mirzaei ◽  
Yaghoub Barfar ◽  
Maryam Hemmatzadeh ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Growth ◽  
Cancer Cell ◽  
Human Cancer ◽  
High Price ◽  
Dependent Manner ◽  
Platelet Poor Plasma ◽  
Protein Levels ◽  
Healthy Donors ◽  
Regulatory Functions

Background: Platelet-rich (PRP) and Platelet-poor plasma (PPP) are widely used in research and clinical platforms mainly due to their capacities to enhance cell growth. Although short half-life (5 days) and the high price of platelet products pose challenges regarding their usage, they maintain the growth regulatory functions for weeks. Thus, we aimed to assess the supplementary values of these products in human CCRF-CEM cancer cells. Mechanistically, we also checked if the PRP/PPP treatment enhances YKL-40 expression as a known protein regulating cell growth. Methods: The PRP/PPP was prepared from healthy donors using manual stepwise centrifugation and phase separation. The viability of the cells treated with gradient PRP/PPP concentrations (2, 5, 10, and 15%) was measured by the MTT assay. The YKL-40 mRNA and protein levels were assessed using qRT-PCR and western blotting. The data were compared to FBS-treated cells. Result: Our findings revealed that the cells treated by PRP/PPP not only were morphologically comparable to those treated by FBS but also, they showed greater viability at the concentrations of 10 and 15%. Moreover, it was shown that PRP/PPP induce cell culture support, at least in part, via inducing YKL-40 expression at both mRNA and protein levels in a time- and dose-dependent manner. Conclusion: Collectively, by showing cell culture support comparable to FBS, the PRP/PPP might be used as good candidates to supplement the cancer cell culture and overcome concerns regarding the use of FBS as a non-human source in human cancer research.

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