A Holistic Assessment of Polyethylene Fiber Ingestion in Larval and Juvenile Japanese Medaka Fish

Microplastic pollution is of public concern for global environmental health, aquaculture, and fisheries. Toxicity studies have shown that microplastic ingestion may cause intestinal damage, microbiota dysbiosis, and disturb the lipid and energy metabolism in fish. To determine the impact of environmentally relevant, chronic, low dose microplastic fibers on fish health, medaka larvae, and juveniles were exposed to five concentrations of polyethylene (PE) fibers for 21 days through the feed. Fish growth and condition were assessed to determine the overall impact on fish health. To identify impaired energy intake, the gastrointestinal tract (GIT) integrity was evaluated at the molecular and cellular levels. Microbiota analysis was performed by comparing the top seven most abundant phyla present in both larval and juvenile fish exposed to 0, 1.5, and 3 PE fibers/fish/day. A shift in the phyla Proteobacteria and Bacteroidetes were observed. Larval samples demonstrated decreased proteobacteria abundance, while juvenile samples displayed an increase in abundance. Relative gene expression of key digestive genes from GIT tissue was quantified using real time-quantitative polymerase chain reaction. An effect on digestive gene expression potentially affecting nutrient absorption and antioxidant production was indicated via a significant decrease of solute carrier family 6 member 6 expression in larvae exposed to 6 fibers/fish/day. No significant molecular changes were observed in juvenile GIT tissue, although a non-monotonous dose-response was observed. GIT morphology was analyzed using histomorphological observations of the GIT mucus and cell types. No significant impairment of the GIT epithelial layers was observed in larvae or juveniles. To assess growth and condition, Fulton’s condition factor was measured. No differences were observed in larval or juvenile growth. Comparisons of different developmental stages allowed for identifying vulnerable developmental stages for microplastic exposure; larvae were more susceptible to molecular changes, while shifts in juvenile microbial communities were similar to changes reported post-polystyrene microplastic sphere exposure. This study is one of the first to provide toxicological data on the risk of PE fiber ingestion during fish development stages. Results indicate no imminent threat to fish condition at current measured environmental levels of microplastics; however, close monitoring of vital spawning grounds for commercially important fishes is recommended.

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Abstract 353: Mechanical (or “Gravitational”) Unloading Reduces Inflammatory and Cell Adhesion Molecule Gene Expression in Human Endothelial Cells

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.32.suppl_1.a353 ◽

2012 ◽

Vol 32 (suppl_1) ◽

Author(s):

S Marlene Grenon ◽

Jesus Aguado-Zuniga ◽

Michael Conte ◽

Millie Hughes-Fulford

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Adhesion Molecule ◽

Cell Function ◽

Quantitative Polymerase Chain Reaction ◽

Cellular Interactions ◽

Gravitational Unloading ◽

Experimental Conditions ◽

Adhesion Molecule Gene ◽

The Impact

Objectives: Mechanical forces including gravity affect mechanotransduction and subsequent cell function. The goal of this study was to investigate the impact of mechanical unloading (MU) and loading (ML) of endothelial cells (ECs) with microgravity and hypergravity respectively, with the hypothesis that MU alters expression of inflammatory and adhesion molecule gene expression and these changes are reversed by ML. Methods: Human umbilical vascular endothelial cells (HUVECs) grown to confluency were studied. A desktop random positioning machine and a gravitational cell-loading apparatus provided MU and ML conditions, respectively. The experimental conditions included: 1) controls exposed to 1-gravity environment for 24 h (CL), 2) MU for 24 hours, 3) MU for 24 hours with three 30-minutes periods of ML of 12-gravity (MU/ML). Gene expression was studied with reverse transcription followed by real-time quantitative polymerase chain reaction (qRTPCR). Results: MU led to a significant decrease in gene expression of the adhesion molecules ICAM-1, VCAM-1, E-Selectin, as well as TNF-α, IL-6 and VEGF. In contrast, NOS-3, Caveolin-1 and -2 were significantly increased with MU. The changes observed in gene expression with MU were reversed by gravitational mechanical loading (MU/ML). Conclusions: Gravitational MU decreases inflammatory and adhesion molecule gene expression and these changes are reversed by short periods of ML. This points towards the importance of gravitational loading in ECs function and cellular interactions.

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Evaluation of Appropriate Reference Genes For Investigating Gene Expression in Chlorops oryzae (Diptera: Chloropidae)

Journal of Economic Entomology ◽

10.1093/jee/toz142 ◽

2019 ◽

Vol 112 (5) ◽

pp. 2207-2214 ◽

Cited By ~ 1

Author(s):

Ping Tian ◽

Lin Qiu ◽

Ailin Zhou ◽

Guo Chen ◽

Hualiang He ◽

...

Keyword(s):

Gene Expression ◽

Ribosomal Protein ◽

Reference Genes ◽

Molecular Mechanisms ◽

Developmental Stages ◽

Quantitative Polymerase Chain Reaction ◽

Housekeeping Genes ◽

Future Research ◽

Economic Damage ◽

Physiological Processes

Abstract Reverse transcription quantitative polymerase chain reaction (PCR) has become an invaluable technique for analyzing gene expression in many insects. However, this approach requires the use of stable reference genes to normalize the data. Chlorops oryzae causes significant economic damage to rice crops throughout Asia. The lack of suitable reference genes has hindered research on the molecular mechanisms underlying many physiological processes of this species. In this study, we used quantitative real-time PCR to evaluate the expression of eight C. oryzae housekeeping genes glyceraldehyde-3-phosphate dehydrogenase (GAPDH), β-actin (βACT), beta-tubulin (βTUB), Delta Elongation factor-1 (EF1δ), ribosomal protein S11 (RPS11), RPS15, C-terminal-Binding Protein (CtBP), and ribosomal protein 49 (RP49) in different developmental stages and tissues in both larvae and adults. We analyzed the data with four different software packages: geNorm, NormFinder, BestKeeper, and RefFinder and compared the results obtained with each method. The results indicate that PRS15 and RP49 can be used as stable reference genes for quantifying gene expression in different developmental stages and larval tissues. GAPDH and βACT, which have been considered stable reference genes by previous studies, were the least stable of the candidate genes with respect to larval tissues. GAPDH was, however, the most stable reference gene for adult tissues. We verified the candidate reference genes identified and found that the expression levels of Cadherins (Cads) changed when different reference genes were used to normalize gene expression. This study provides a valuable foundation for future research on gene function, and investigating the molecular basis of physiological processes, in C. oryzae.

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Innate Immune Responses of Skin Mucosa in Common Carp (Cyprinus Carpio) Fed a Diet Supplemented with Galactooligosaccharides

Animals ◽

10.3390/ani10030438 ◽

2020 ◽

Vol 10 (3) ◽

pp. 438 ◽

Cited By ~ 3

Author(s):

Elzbieta Pietrzak ◽

Jan Mazurkiewicz ◽

Anna Slawinska

Keyword(s):

Gene Expression ◽

Immune Responses ◽

Common Carp ◽

Geometric Mean ◽

Juvenile Fish ◽

Innate Immune ◽

Innate Immune Responses ◽

Immune Related Genes ◽

The Impact ◽

Skin Mucosa

Galactooligosaccharides (GOS) are well-known immunomodulatory prebiotics. We hypothesize that GOS supplemented in feed modulates innate immune responses in the skin-associated lymphoid tissue (SALT) of common carp. The aim of this study was to determine the impact of GOS on mRNA expression of the immune-related genes in skin mucosa. During the feeding trial, the juvenile fish (bodyweight 180 ± 5 g) were fed two types of diet for 50 days: control and supplemented with 2% GOS. At the end of the trial, a subset of fish was euthanized (n = 8). Skin mucosa was collected, and RNA was extracted. Gene expression analysis was performed with RT-qPCR to determine the mRNA abundance of the genes associated with innate immune responses in SALT, i.e., acute-phase protein (CRP), antimicrobial proteins (His2Av and GGGT5L), cytokines (IL1β, IL4, IL8, IL10, and IFNγ), lectin (CLEC4M), lyzosymes (LyzC and LyzG), mucin (M5ACL), peroxidase (MPO), proteases (CTSB and CTSD), and oxidoreductase (TXNL). The geometric mean of 40s s11 and ACTB was used to normalize the data. Relative quantification of the gene expression was calculated with ∆∆Ct. GOS upregulated INFγ (p ≤ 0.05) and LyzG (p ≤ 0.05), and downregulated CRP (p ≤ 0.01). We conclude that GOS modulates innate immune responses in the skin mucosa of common carp.

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Effects of histamine and various histamine receptor antagonists on gene expression profiles of macrophages during compressive strain

Journal of Orofacial Orthopedics / Fortschritte der Kieferorthopädie ◽

10.1007/s00056-021-00318-x ◽

2021 ◽

Author(s):

Agnes Schröder ◽

Catharina Petring ◽

Anna Damanaki ◽

Jonathan Jantsch ◽

Peter Proff ◽

...

Keyword(s):

Gene Expression ◽

Gene Expression Profile ◽

Expression Profile ◽

Periodontal Ligament ◽

Tooth Movement ◽

Expression Profiles ◽

Quantitative Polymerase Chain Reaction ◽

Compressive Strain ◽

Orthodontic Tooth Movement ◽

The Impact

Abstract Purpose Tissue hormone histamine can accumulate locally within the periodontal ligament via nutrition or may be released during allergic reactions by mast cells, which may have an impact on orthodontic tooth movement. In addition to periodontal ligament fibroblasts, cells of the immune system such as macrophages are exposed to compressive strain. The aim of this study was thus to investigate the impact of histamine on the gene expression profile of macrophages in the context of simulated orthodontic compressive strain. Methods Macrophages were incubated with different histamine concentrations (50, 100, 200 µM) for 24 h and then either left untreated or compressed for another 4 h. To assess the role of different histamine receptors, we performed experiments with antagonists for histamine 1 receptor (cetirizine), histamine 2 receptor (ranitidine) and histamine 4 receptor (JNJ7777120) under control and pressure conditions. We tested for lactate dehydrogenase release and analyzed the expression of genes involved in inflammation and bone remodeling by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Results Histamine elevated gene expression of tumor necrosis factor under control conditions and in combination with pressure application. Increased prostaglandin-endoperoxide synthase‑2 mRNA was observed when histamine was combined with compressive force. Interleukin‑6 gene expression was not affected by histamine treatment. In macrophages, compressive strain increased osteoprotegerin gene expression. Histamine further elevated this effect. Most of the observed histamine effects were blocked by the histamine 1 receptor antagonist cetirizine. Conclusions Histamine has an impact on the gene expression profile of macrophages during compressive strain in vitro, most likely having an impairing effect on orthodontic tooth movement by upregulation of osteoprotegerin expression.

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Screening of reference genes using real-time quantitative PCR for gene expression studies inNeoseiulus barkeriHughes (Acari: Phytoseiidae)

Bulletin of Entomological Research ◽

10.1017/s000748531800072x ◽

2018 ◽

Vol 109 (4) ◽

pp. 443-452 ◽

Cited By ~ 1

Author(s):

C. Wang ◽

J. Yang ◽

Q. Pan ◽

S. Yu ◽

R. Luo ◽

...

Keyword(s):

Gene Expression ◽

Real Time ◽

Reference Genes ◽

Developmental Stages ◽

Quantitative Polymerase Chain Reaction ◽

Real Time Quantitative Pcr ◽

Expression Studies ◽

Gene Expression Studies ◽

The Stability ◽

Suitable Reference

AbstractA stable reference gene is a key prerequisite for accurate assessment of gene expression. At present, the real-time reverse transcriptase quantitative polymerase chain reaction has been widely used in the analysis of gene expression in a variety of organisms.Neoseiulus barkeriHughes (Acari: Phytoseiidae) is a major predator of mites on many important economically crops. Until now, however, there are no reports evaluating the stability of reference genes in this species. In view of this, we used GeNorm, NormFinder, BestKeeper, and RefFinder software tools to evaluate the expression stability of 11 candidate reference genes in developmental stages and under various abiotic stresses. According to our results, β-ACTandHsp40were the top two stable reference genes in developmental stages. TheHsp60andHsp90were the most stable reference genes in various acaricides stress. For alterations in temperature,Hsp40and α-TUBwere the most suitable reference genes. About UV stress,EF1α and α-TUBwere the best choice, and for the different prey stress, β-ACTand α-TUBwere best suited. In normal conditions, the β-ACT and α-TUB were the two of the highest stable reference genes to respond to all kinds of stresses. The current study provided a valuable foundation for the further analysis of gene expression inN. barkeri.

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A distinguishing gene signature shared by tumor-infiltrating Tie2-expressing monocytes, blood “resident” monocytes, and embryonic macrophages suggests common functions and developmental relationships

Blood ◽

10.1182/blood-2009-01-200931 ◽

2009 ◽

Vol 114 (4) ◽

pp. 901-914 ◽

Cited By ~ 238

Author(s):

Ferdinando Pucci ◽

Mary Anna Venneri ◽

Daniela Biziato ◽

Alessandro Nonis ◽

Davide Moi ◽

...

Keyword(s):

Gene Expression ◽

Developmental Stages ◽

Expression Profiles ◽

Quantitative Polymerase Chain Reaction ◽

Gene Expression Profiles ◽

Tissue Remodeling ◽

Suppressor Cells ◽

Gene Signature ◽

Blood Monocytes ◽

Inflammatory Monocytes

Abstract We previously showed that Tie2-expressing monocytes (TEMs) have nonredundant proangiogenic activity in tumors. Here, we compared the gene expression profile of tumor-infiltrating TEMs with that of tumor-associated macrophages (TAMs), spleen-derived Gr1+Cd11b+ neutrophils/myeloid-derived suppressor cells, circulating “inflammatory” and “resident” monocytes, and tumor-derived endothelial cells (ECs) by quantitative polymerase chain reaction–based gene arrays. TEMs sharply differed from ECs and Gr1+Cd11b+ cells but were highly related to TAMs. Nevertheless, several genes were differentially expressed between TEMs and TAMs, highlighting a TEM signature consistent with enhanced proangiogenic/tissue-remodeling activity and lower proinflammatory activity. We validated these findings in models of oncogenesis and transgenic mice expressing a microRNA-regulated Tie2-GFP reporter. Remarkably, resident monocytes and TEMs on one hand, and inflammatory monocytes and TAMs on the other hand, expressed coordinated gene expression profiles, suggesting that the 2 blood monocyte subsets are committed to distinct extravascular fates in the tumor microenvironment. We further showed that a prominent proportion of embryonic/fetal macrophages, which participate in tissue morphogenesis, expressed distinguishing TEM genes. It is tempting to speculate that Tie2+ embryonic/fetal macrophages, resident blood monocytes, and tumor-infiltrating TEMs represent distinct developmental stages of a TEM lineage committed to execute physiologic proangiogenic and tissue-remodeling programs, which can be coopted by tumors.

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Selection and Validation of Reference Genes for Gene Expression Analysis in Tuta absoluta Meyrick (Lepidoptera: Gelechiidae)

Insects ◽

10.3390/insects12070589 ◽

2021 ◽

Vol 12 (7) ◽

pp. 589

Author(s):

Xin Yan ◽

Yibo Zhang ◽

Kangkang Xu ◽

Yawei Wang ◽

Wenjia Yang

Keyword(s):

Gene Expression ◽

Reference Genes ◽

Developmental Stages ◽

Quantitative Polymerase Chain Reaction ◽

Housekeeping Genes ◽

Tuta Absoluta ◽

Leaf Miner ◽

Specific Reference ◽

Internal Reference ◽

Experimental Conditions

The tomato leaf miner, Tuta absoluta is a destructive pest of tomato. The leaf-mining activities of its larvae can cause significant yield losses. Real-time quantitative polymerase chain reaction (RT-qPCR) is commonly used to measure gene expression, and the selection of stable reference genes for calibration and standardization is critical for accurate use of RT-qPCR. We studied the stable expression of nine common housekeeping genes in T. absoluta. These were examined at different developmental stages, in larval tissues, as well as those induced by exposure to 20E and insecticides. Four dedicated algorithms (geNorm, BestKeeper, NormFinder, and ΔCt method) and online tool (RefFinder) were used to analyze and rank the tested reference genes. Based on the standardized gene expression data of target gene ecdysone receptor (EcR), the applicability of specific reference genes was verified. The results clarify that the optimal internal reference genes vary greatly under different experimental conditions. GAPDH and RPS11 were the best reference genes for developmental stages; RPL28 and RPL10 for different tissues; EF1α and RPL28 for 20E treatment; EF1α and RPL7A for insecticide treatments. The most suitable reference genes in all experimental conditions are EF1α and RPL28.

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Identification of reference genes for gene expression studies among different developmental stages of murine hearts

BMC Developmental Biology ◽

10.1186/s12861-021-00244-6 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Jie Ren ◽

Ningning Zhang ◽

Xiangjie Li ◽

Xiaogang Sun ◽

Jiangping Song

Keyword(s):

Gene Expression ◽

Reference Gene ◽

Heart Development ◽

Reference Genes ◽

Developmental Stages ◽

Expression Profiles ◽

Quantitative Polymerase Chain Reaction ◽

Expression Patterns ◽

Gene Expression Profiles ◽

Qpcr Data

Abstract Background Real-time quantitative polymerase chain reaction (RT-qPCR) is a widely-used standard assay for assessing gene expression. RT-qPCR data requires reference genes for normalization to make the results comparable. Therefore, the selected reference gene should be highly stable in its expression throughout the experimental datasets. So far, reports about the optimal set of reference genes in murine left ventricle (LV) across embryonic and postnatal stages are few. The objective of our research was to identify the appropriate reference genes in murine LV among different developmental stages. Methods We investigated the gene expression profiles of 21 widely used housekeeping genes in murine LV from 7 different developmental stages (almost throughout the whole period of the mouse lifespan). The stabilities of the potential reference genes were evaluated by five methods: GeNorm, NormFinder, BestKeeper, Delta-Ct and RefFinder. Results We proposed a set of reliable reference genes for normalization of RT-qPCR experimental data in different conditions. Furthermore, our results showed that 6 genes (18S, Hmbs, Ubc, Psmb4, Tfrc and Actb) are not recommended to be used as reference genes in murine LV development studies. The data also suggested that the Rplp0 gene might serve as an optimal reference gene in gene expression analysis. Conclusions Our study investigated the expression stability of the commonly used reference genes in process of LV development and maturation. We proposed a set of optimal reference genes that are suitable for accurate normalization of RT-qPCR data in specific conditions. Our findings may be helpful in future studies for investigating the gene expression patterns and mechanism of mammalian heart development.

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Porcine Neural Progenitor Cells Derived from Tissue at Different Gestational Ages Can Be Distinguished by Global Transcriptome

Cell Transplantation ◽

10.1177/0963689717723015 ◽

2017 ◽

Vol 26 (9) ◽

pp. 1582-1595

Author(s):

Jing Yang ◽

Steven Menges ◽

Ping Gu ◽

Ronald Tongbai ◽

Melissa Samuel ◽

...

Keyword(s):

Gene Expression ◽

Progenitor Cells ◽

Neural Progenitor Cells ◽

Expression Profiles ◽

Quantitative Polymerase Chain Reaction ◽

Neural Progenitor ◽

Age Related ◽

Donor Cells ◽

Global Transcriptome ◽

The Impact

The impact of gestational age on mammalian neural progenitor cells is potentially important for both an understanding of neural development and the selection of donor cells for novel cell-based treatment strategies. In terms of the latter, it can be problematic to rely entirely on rodent models in which the gestational period is significantly shorter and the brain much smaller than is the case in humans. Here, we analyzed pig brain progenitor cells (pBPCs) harvested at 2 different gestational ages (E45 and E60) using gene expression profiles, obtained by microarray analysis and quantitative polymerase chain reaction (qPCR), across time in culture. Comparison of the global transcriptome of pBPCs from age-matched transgenic green flourescent protein (GFP)-expressing fetuses versus non- GFP-expressing fetuses did not reveal significant differences between the 2 cell types, whereas comparison between E45 and E60 pBPCs did show separation between the data sets by principle component analysis. Further examination by qPCR showed evidence of relative downregulation of proliferation markers and upregulation of glial markers in the gestationally older (E60) cells. Additional comparisons were made. This study provides evidence of age-related changes in the gene expression of cultured fetal porcine neural progenitors that are potentially relevant to the role of these cells during development and as donor cells for transplantation studies.

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Identification of appropriate reference genes for gene expression studies in mice left ventricles from different developmental stages

10.21203/rs.3.rs-208040/v1 ◽

2021 ◽

Author(s):

Jie Ren ◽

Ningning Zhang ◽

Xiangjie Li ◽

Xiaogang Sun ◽

Jiangping Song

Keyword(s):

Gene Expression ◽

Heart Development ◽

Reference Genes ◽

Developmental Stages ◽

Quantitative Polymerase Chain Reaction ◽

Expression Patterns ◽

Housekeeping Genes ◽

Good Choice ◽

Expression Stability ◽

Gene Expressions

Abstract Aims: Real-time quantitative polymerase chain reaction (RT-qPCR) is the standard assay used for revealing the gene expression characteristics. However, the RT-qPCR studies all need reference genes for normalization to make the results comparable, which should hold a high expression stability during all experimental datasets. So far, there was no optimal set of reference genes identified in mice left ventricles (LV) across embryonic and postnatal stages. The objective of our research was to identify the appropriate reference genes in mice LV from different developmental stages.Methods and Results: we investigated the gene expressions of common 21 candidate housekeeping genes in mice LV from 7 different developmental stages, almost throughout the whole period of the mouse lifespan. The expression of some candidate reference genes, such as 18S and Actb, apparently fluctuated. The stability of potential reference genes was evaluated by a number of methods, such as GeNorm, NormFinder, BestKeeper, Delta-Ct and RefFinder method. we identified a set of optimal reference genes that can be reliably used for normalization of RT-qPCR experiments in different developmental stages of mice LV. Our results showed that following genes should not be used as reference genes in mice LV development studies: 18S, Hmbs, Ubc, Psmb4, Tfrc and Actb. And the Rplp0 appeared to represent a good choice. Conclusions: Our study provides the expression stability of the commonly used reference genes in process of LV development and maturation. We also identified a set of optimal reference genes under different conditions. Our findings may be helpful in future studies to investigate the gene expression patterns and mechanism of mammalian heart development.

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