Transcriptome Profiling of Atlantic Salmon Adherent Head Kidney Leukocytes Reveals That Macrophages Are Selectively Enriched During Culture

The Atlantic salmon (Salmo salar) is an economically important fish, both in aquaculture and in the wild. In vertebrates, macrophages are some of the first cell types to respond to pathogen infection and disease. While macrophage biology has been characterized in mammals, less is known in fish. Our previous work identified changes in the morphology, phagocytic ability, and miRNA profile of Atlantic salmon adherent head kidney leukocytes (HKLs) from predominantly “monocyte-like” at Day 1 of in vitro culture to predominantly “macrophage-like” at Day 5 of culture. Therefore, to further characterize these two cell populations, we examined the mRNA transcriptome profile in Day 1 and Day 5 HKLs using a 44K oligonucleotide microarray. Large changes in the transcriptome were revealed, including changes in the expression of macrophage and immune-related transcripts (e.g. csf1r, arg1, tnfa, mx2), lipid-related transcripts (e.g. fasn, dhcr7, fabp6), and transcription factors involved in macrophage differentiation and function (e.g. klf2, klf9, irf7, irf8, stat1). The in silico target prediction analysis of differentially expressed genes (DEGs) using miRNAs known to change expression in Day 5 HKLs, followed by gene pathway enrichment analysis, supported that these miRNAs may be involved in macrophage maturation by targeting specific DEGs. Elucidating how immune cells, such as macrophages, develop and function is a key step in understanding the Atlantic salmon immune system. Overall, the results indicate that, without the addition of exogenous factors, the adherent HKL cell population differentiates in vitro to become macrophage-like.

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Impact of IL-27 on hepatocyte antiviral gene expression and function

Wellcome Open Research ◽

10.12688/wellcomeopenres.9917.1 ◽

2016 ◽

Vol 1 ◽

pp. 17 ◽

Cited By ~ 3

Author(s):

Narayan Ramamurthy ◽

Sara Boninsegna ◽

Rebecca Adams ◽

Natasha Sahgal ◽

Helen Lockstone ◽

...

Keyword(s):

Cell Culture ◽

Antigen Presentation ◽

Microarray Analysis ◽

Enrichment Analysis ◽

Cell Types ◽

Antiviral Effect ◽

Gene Set Enrichment Analysis ◽

And Function ◽

The Impact

Background: Interleukin (IL)-27 is a member of the IL-6/IL-12 family of cytokines. It is a potent cytokine, with potential antiviral impact, and has been shown to play a role in modulating functions of diverse cell types, including Th1, Th2, and NK and B cells, demonstrating both pro- and anti-inflammatory roles. In hepatocytes, it is capable of inducing signal transducer and activator of transcription (STAT)1, STAT3 and interferon-stimulated genes. Methods: To address its role in viral hepatitis, the antiviral activity of IL-27 against hepatitis C virus (HCV) and hepatitis B virus (HBV) was tested in vitro using cell-culture-derived infectious HCV (HCVcc) cell culture system and the HepaRG HBV cell culture model. To further investigate the impact of IL-27 on hepatocytes, Huh7.5 cells were treated with IL-27 to analyse the differentially expressed genes by microarray analysis. Furthermore, by quantitative PCR, we analyzed the up-regulation of chemokine (CXCL)-10 in response to IL-27. Results: In both HCV and HBV infection models, we observed only a modest direct antiviral effect. Microarray analysis showed that the up-regulated genes mostly belonged to antigen presentation and DNA replication pathways, and involved strong up-regulation of CXCL-10, a gene associated with liver inflammation. Overall, gene set enrichment analysis showed a striking correlation of these genes with those up-regulated in response to related cytokines in diverse cell populations. Conclusion: Our data indicate that IL-27 can have a significant pro-inflammatory impact in vitro, although the direct antiviral effect is modest. It may have a potential impact on hepatocyte function, especially chemokine expression and antigen presentation.

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Network Pharmacology and Molecular Docking Reveal the Mechanism of Isodon Ternifolius (D. Don) Kudo Against Liver Fibrosis

10.21203/rs.3.rs-1088473/v1 ◽

2021 ◽

Author(s):

Le Qin ◽

Zhipin Zhou ◽

Hui Huang ◽

Jingjing Wang ◽

Yong Chen ◽

...

Keyword(s):

Liver Fibrosis ◽

Umbilical Vein ◽

Target Prediction ◽

Interaction Network ◽

Enrichment Analysis ◽

Network Pharmacology ◽

Pathway Enrichment Analysis ◽

Active Ingredients ◽

Chemical Structures

Abstract BackgroundIsodon ternifolius (SanYe Xiang ChaCai, SYXCC) is a traditional Chinses medicine commonly used in the treatment of chronic hepatitis B in China. Many studies have demonstrated the hepatoprotective or anti-fibrotic effects of SYXCC, but its pharmacological basis and mechanism remain unclear. In this study, we used in vitro models to validate the predicted results and reveal the potential mechanism of action and active ingredients with the help of network pharmacology methods and molecular docking.MethodsWe obtained the chemical structures of SYXCC by literatures. Potential targets of SYXCC were predicted by Swiss Target Prediction. The disease targets were collected through the databases of Gene Card. PPI protein interaction network was obtained using STRING database. Signaling pathway enrichment analysis was performed on drug-disease targets with of DAVID database. In vitro, human umbilical vein endothelial cell was used to validate the results predicted by network pharmacology.Results90 pathways related to liver fibrosis were obtained by KEGG enrichment. We were interested in the top 25 core target proteins involved in the TLR4, MAPK and PI3K-Akt signaling pathways associated with liver fibrosis. RT-PCR result showed that SYXCC could reduce EGFR, ERK1, Akt1, VEFGR-2, ERK2 mRNA expression. ConclusionIn the study, our results suggest that a multitude of active ingredients of Isodon ternifouis play an important role in the liver fibrosis disease network. Inhibition of angiogenesis was a new potential pharmacological mechanism for the anti-hepatic fibrosis activity of Isodon ternifoius.

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Role of the 807 C/T Polymorphism of the α2 Gene in Platelet GP Ia Collagen Receptor Expression and Function

Thrombosis and Haemostasis ◽

10.1055/s-0037-1614605 ◽

1999 ◽

Vol 81 (06) ◽

pp. 951-956 ◽

Cited By ~ 59

Author(s):

J. Corral ◽

R. González-Conejero ◽

J. Rivera ◽

F. Ortuño ◽

P. Aparicio ◽

...

Keyword(s):

Venous Thrombosis ◽

Cell Types ◽

Receptor Expression ◽

Case Control Studies ◽

Platelet Surface ◽

Vitro Analysis ◽

And Function ◽

In Vitro Analysis

SummaryThe variability of the platelet GP Ia/IIa density has been associated with the 807 C/T polymorphism (Phe 224) of the GP Ia gene in American Caucasian population. We have investigated the genotype and allelic frequencies of this polymorphism in Spanish Caucasians. The T allele was found in 35% of the 284 blood donors analyzed. We confirmed in 159 healthy subjects a significant association between the 807 C/T polymorphism and the platelet GP Ia density. The T allele correlated with high number of GP Ia molecules on platelet surface. In addition, we observed a similar association of this polymorphism with the expression of this protein in other blood cell types. The platelet responsiveness to collagen was determined by “in vitro” analysis of the platelet activation and aggregation response. We found no significant differences in these functional platelet parameters according to the 807 C/T genotype. Finally, results from 3 case/control studies involving 302 consecutive patients (101 with coronary heart disease, 104 with cerebrovascular disease and 97 with deep venous thrombosis) determined that the 807 C/T polymorphism of the GP Ia gene does not represent a risk factor for arterial or venous thrombosis.

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Modelling the Human Placental Interface In Vitro—A Review

Micromachines ◽

10.3390/mi12080884 ◽

2021 ◽

Vol 12 (8) ◽

pp. 884

Author(s):

Marta Cherubini ◽

Scott Erickson ◽

Kristina Haase

Keyword(s):

Drug Testing ◽

Cell Types ◽

In Vitro Models ◽

Placental Development ◽

Scientific Challenge ◽

Induced Pluripotent ◽

And Function ◽

And Control

Acting as the primary link between mother and fetus, the placenta is involved in regulating nutrient, oxygen, and waste exchange; thus, healthy placental development is crucial for a successful pregnancy. In line with the increasing demands of the fetus, the placenta evolves throughout pregnancy, making it a particularly difficult organ to study. Research into placental development and dysfunction poses a unique scientific challenge due to ethical constraints and the differences in morphology and function that exist between species. Recently, there have been increased efforts towards generating in vitro models of the human placenta. Advancements in the differentiation of human induced pluripotent stem cells (hiPSCs), microfluidics, and bioprinting have each contributed to the development of new models, which can be designed to closely match physiological in vivo conditions. By including relevant placental cell types and control over the microenvironment, these new in vitro models promise to reveal clues to the pathogenesis of placental dysfunction and facilitate drug testing across the maternal–fetal interface. In this minireview, we aim to highlight current in vitro placental models and their applications in the study of disease and discuss future avenues for these in vitro models.

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Abstract P748: Transcriptome Profiling of the Neural Stem Cell Niche and the Effect of Exercise in Restoring Neurogenesis in Type II Diabetic Mice

Stroke ◽

10.1161/str.52.suppl_1.p748 ◽

2021 ◽

Vol 52 (Suppl_1) ◽

Author(s):

Gratianne Rabiller ◽

Atsushi Kanoke ◽

Jialing Liu

Keyword(s):

Oxidative Stress ◽

Deleterious Effect ◽

Wheel Running ◽

Enrichment Analysis ◽

Transcriptome Profiling ◽

Cell Types ◽

Neural Regeneration ◽

Diabetic Mice ◽

Middle Aged ◽

Neurogenic Niche

Introduction: Previously we found that mice with type 2 diabetes (T2DM) exhibited an accelerated age-associated decline in neurogenesis during baseline and after ischemic stroke compared to age-matched control mice. The current study sought to delineate the transcriptome landscape involved in the impaired neurogenesis and determine if exercise can prevent the deleterious effect of T2DM on neural regeneration. Hypothesis: We hypothesize that T2DM alters signaling pathways regulating neurogenesis and daily exercise mitigates the deleterious effect on neurogenesis in the T2DM mice. Methods: Transcriptome profiling was performed by single cell RNA sequencing (scRNAseq) of SVZ and DG cells in stroke and non-stroke mice using the 10X Genomics platform. T2DM-induced differential gene expression was analyzed by ClusterProfiler and Wikipathways enrichment analysis. Middle-aged (~260 days old) and old (~700 days old) db/+ or db/db mice were subjected to daily wheel-running exercise for one month. BrdU at 50 mg/kg twice daily for 2 consecutive days was injected i.p. at the end of the experiment to track proliferating neuroprogenitor cells. DCX+ cells and BrDU+ cells were quantified in the dentate gyrus of the hippocampus. Results: The scRNAseq analysis revealed multiple cell types co-existing in the neurogenic niche. GO and Wikipathways enrichment analysis showed that under diabetic condition, genes such as Qdpr, Hsp90ab1, Hsp90aa1, and Sox9 were downregulated in pathways involving eNOS activation; whereas Junb, C1qc, C1qb and C1qa were upregulated in the pathways related to oxidative stress. Exercise, known to increase eNOS expression and reduce oxidative stress-induced cell death, significantly restored the number of DCX+ immature neurons in 8-months-old diabetic mice almost to the level of the control mice without exercise Conclusions: Exercise restores neurogenesis by increasing the number of neuroblasts in the middle-aged diabetic mice. Ongoing experiment will investigate whether exercise promotes neurogenesis by enhancing eNOS and improved blood flow, and inducing genes involved in the survival of the NSC niche of the diabetic mice.

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Secretome Profiling of Atlantic Salmon Head Kidney Leukocytes Highlights the Role of Phagocytes in the Immune Response to Soluble β-Glucan

Frontiers in Immunology ◽

10.3389/fimmu.2021.736964 ◽

2021 ◽

Vol 12 ◽

Author(s):

Dimitar B. Iliev ◽

Guro Strandskog ◽

Mehrdad Sobhkhez ◽

Jack A. Bruun ◽

Jorunn B. Jørgensen

Keyword(s):

Immune System ◽

Atlantic Salmon ◽

Primary Cultures ◽

Head Kidney ◽

Cell Types ◽

Mononuclear Phagocytes ◽

Immunostimulatory Activity ◽

Complement Receptor 3 ◽

Bacteria And Fungi

β‐Glucans (BG) are glucose polymers which are produced in bacteria and fungi but not in vertebrate organisms. Being recognized by phagocytic leukocytes including macrophages and neutrophils through receptors such as dectin-1 and Complement receptor 3 (CR3), the BG are perceived by the innate immune system of vertebrates as foreign substances known as Pathogen Associated Molecular Patterns (PAMPs). The yeast-derived BG has been recognized for its potent biological activity and it is used as an immunomodulator in human and veterinary medicine. The goal of the current study was to characterize the immunostimulatory activity of soluble yeast BG in primary cultures of Atlantic salmon (Salmo salar) head kidney leukocytes (HKLs) in which phagocytic cell types including neutrophils and mononuclear phagocytes predominate. The effect of BG on the secretome of HKL cultures, including secretion of extracellular vesicles (EVs) and soluble protein55s was characterized through western blotting and mass spectrometry. The results demonstrate that, along with upregulation of proinflammatory genes, BG induces secretion of ubiquitinated proteins (UbP), MHCII-containing EVs from professional antigen presenting cells as well as proteins derived from granules of polymorphonuclear granulocytes (PMN). Among the most abundant proteins identified in BG-induced EVs were beta-2 integrin subunits, including CD18 and CD11 homologs, which highlights the role of salmon granulocytes and mononuclear phagocytes in the response to soluble BG. Overall, the current work advances the knowledge about the immunostimulatory activity of yeast BG on the salmon immune system by shedding light on the effect of this PAMP on the secretome of salmon leukocytes.

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A single nuclei transcriptomic analysis of the Atlantic salmon gill through smoltification and seawater transfer

10.1101/2020.09.03.281337 ◽

2020 ◽

Author(s):

Alexander C. West ◽

Yasutaka Mizoro ◽

Shona H. Wood ◽

Louise M. Ince ◽

Marianne Iversen ◽

...

Keyword(s):

Gene Expression ◽

Atlantic Salmon ◽

Direct Evidence ◽

Cell Types ◽

Prior Work ◽

In The Wild ◽

Induced Changes ◽

Freshwater Environments ◽

Transcriptomic Studies

AbstractAnadromous salmonids begin life adapted to the freshwater environments of their natal streams before a developmental transition, known as smoltification, transforms them into marine-adapted fish. In the wild, the extending photoperiods of spring stimulates smoltification, typified by radical reprogramming of the gill from an ion-absorbing organ to ion-excreting organ. Prior work has highlighted the role of specialized “mitochondrion-rich” cells in delivering this phenotype. However, transcriptomic studies identify thousands of smoltification-driven differentially regulated genes, indicating that smoltification causes a multifaceted, multicellular change; but direct evidence of this is lacking.Here, we use single-nuclei RNAseq to characterize the Atlantic salmon gill during smoltification and seawater transfer. We identify 20 distinct clusters of nuclei, including known, but also novel gill cell types. These data allow us to isolate cluster-specific, smoltification-induced changes in gene expression. We also show how cellular make-up of the gill changes through smoltification. As expected, we noted an increase in the proportion of seawater mitochondrion-rich cells, however, we also identify a reduction of several immune-related cells. Overall, our results provide unrivaled detail of the cellular complexity in the gill and suggest that smoltification triggers unexpected immune reprogramming directly preceding seawater entry.

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High expression of MCM10 is predictive of poor outcomes in lung adenocarcinoma

PeerJ ◽

10.7717/peerj.10560 ◽

2021 ◽

Vol 9 ◽

pp. e10560

Author(s):

Mingrui Shao ◽

Shize Yang ◽

Siyuan Dong

Keyword(s):

Lung Adenocarcinoma ◽

Complex Disease ◽

Enrichment Analysis ◽

Integrated Approach ◽

Replication Initiation ◽

Prognostic Biomarkers ◽

Normal Lung ◽

Pathway Enrichment Analysis ◽

Therapeutic Value

Backgrounds Lung adenocarcinoma is a complex disease that results in over 1.8 million deaths a year. Recent advancements in treating and managing lung adenocarcinoma have led to modest decreases in associated mortality rates, owing in part to the multifactorial etiology of the disease. Novel prognostic biomarkers are needed to accurately stage the disease and act as the basis of adjuvant treatments. Material and Methods The microarray datasets GSE75037, GSE31210 and GSE32863 were downloaded from the Gene Expression Omnibus (GEO) database to identify prognostic biomarkers for lung adenocarcinoma and therapy. The differentially expressed genes (DEGs) were identified by GEO2R. Functional and pathway enrichment analysis were performed by Kyoto Encyclopedia of Genes and Genomes and Gene Ontology (GO). Validation was performed based on 72 pairs of lung adenocarcinoma and adjacent normal lung tissues. Results Results showed that the DEGs were mainly focused on cell cycle and DNA replication initiation. Forty-one hub genes were identified and further analyzed by CytoScape. Here, we provide evidence which suggests MCM10 is a potential target with prognostic, diagnostic and therapeutic value. We base this on an integrated approach of comprehensive bioinformatics analysis and in vitro validation using the A549 lung adenocarcinoma cell line. We show that MCM10 overexpression correlates with a poor prognosis, while silencing of this gene decreases aberrant growth by 2-fold. Finally, evaluation of 72 clinical biopsy samples suggests that overexpression of MCM10 in the lung adenocarcinoma highly correlates with larger tumor size. Together, this work suggests that MCM10 may be a clinically relevant gene with both predictive and therapeutic value in lung adenocarcinoma.

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Polymorphonuclear Leukocytes Prepared by Continuous-Flow Filtration Leukapheresis: Viability and Function

Blood ◽

10.1182/blood.v44.5.707.707 ◽

1974 ◽

Vol 44 (5) ◽

pp. 707-713 ◽

Cited By ~ 23

Author(s):

Michael B. Harris ◽

Isaac Djerassi ◽

Elias Schwartz ◽

Richard K. Root

Keyword(s):

Continuous Flow ◽

Polymorphonuclear Leukocytes ◽

Cell Types ◽

High Yield ◽

Trypan Blue Exclusion ◽

Flow Filtration ◽

And Function ◽

Combating Infection

Abstract Preparation of granulocytes for transfusion in high yield and relatively free of contamination by other cell types has been made possible by the technique of continuous-flow filtration leukapheresis (CFFL). Since previous work suggested that granulocytes collected in this manner may have impaired viability and function, a detailed study of the bactericidal, metabolic, and chemotactic properties of such cells was performed and compared to control cells obtained from the same donors prior to CFFL. The granulocyte percentage of the cell suspensions obtained by CFFL averaged 94.5% ± 1.5% compared to 82.5% ± 1.8% for the controls (p < 0.001) with viability of the PMNs determined by trypan blue exclusion being 97.5% ± 0.9% and 98.2% ± 0.5%, respectively. The phogocytic, metabolic (14C-I-glucose oxidation and protein iodination) and chemotactic properties of both cell types were equivalent in suspensions equalized for granulocyte content. These findings indicate that CFFL technique employed does not impair granulocyte viability or function in vitro. Studies of the in vivo survival and function of CFFL granulocytes are necessary to evaluate their efficacy in combating infection in severely leukopenic patients.

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Human Heart Explant-Derived Extracellular Vesicles: Characterization and Effects on the In Vitro Recellularization of Decellularized Heart Valves

International Journal of Molecular Sciences ◽

10.3390/ijms20061279 ◽

2019 ◽

Vol 20 (6) ◽

pp. 1279 ◽

Cited By ~ 4

Author(s):

Amanda Leitolis ◽

Paula Suss ◽

João Roderjan ◽

Addeli Angulski ◽

Francisco da Costa ◽

...

Keyword(s):

Wound Healing ◽

Stem Cell ◽

Mesenchymal Stem Cell ◽

Extracellular Vesicles ◽

Tissue Regeneration ◽

Human Heart ◽

Heart Valves ◽

Enrichment Analysis ◽

Cell Types

Extracellular vesicles (EVs) are particles released from different cell types and represent key components of paracrine secretion. Accumulating evidence supports the beneficial effects of EVs for tissue regeneration. In this study, discarded human heart tissues were used to isolate human heart-derived extracellular vesicles (hH-EVs). We used nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM) to physically characterize hH-EVs and mass spectrometry (MS) to profile the protein content in these particles. The MS analysis identified a total of 1248 proteins. Gene ontology (GO) enrichment analysis in hH-EVs revealed the proteins involved in processes, such as the regulation of cell death and response to wounding. The potential of hH-EVs to induce proliferation, adhesion, angiogenesis and wound healing was investigated in vitro. Our findings demonstrate that hH-EVs have the potential to induce proliferation and angiogenesis in endothelial cells, improve wound healing and reduce mesenchymal stem-cell adhesion. Last, we showed that hH-EVs were able to significantly promote mesenchymal stem-cell recellularization of decellularized porcine heart valve leaflets. Altogether our data confirmed that hH-EVs modulate cellular processes, shedding light on the potential of these particles for tissue regeneration and for scaffold recellularization.

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