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Circulating miRNA signatures of early pregnancy in cattle

BMC GENOMICS 2016年第1期17卷 184-184页

作者： Ioannidis, Jason Donadeu, F. Xavier Univ Edinburgh Roslin Inst & RD SVS Easter Bush EH25 9RG Midlothian Scotland

Background: Low fertility remains a leading cause of poor productivity in dairy cattle. In this context, there is significant interest in developing novel tools for accurate early diagnosis of pregnancy. MicroRNAs (miRNAs) are short RNA molecules which are critically involved in regulating gene expression during both health and disease. MiRNAs have been shown to regulate ovarian function, uterine receptivity, embryonic development and placental function. Circulating miRNAs can provide useful biomarkers of tissue function and disease;importantly, differential miRNA profiles have been linked to pregnancy and preeclampsia in humans. This study sought to establish the potential of circulating miRNAs as biomarkers of early pregnancy in cattle. Results: We applied Illumina small-RNA sequencing to profile miRNAs in plasma samples collected from eight nonpregnant heifers on Days 0, 8 and 16 of the oestrous cycle and 11 heifers on Days 16 and 24 of pregnancy. We sequenced a total of 46 samples and generated 9.2 million miRNA reads per sample. There were no differences in miRNA read abundance between any of the pregnant and non-pregnant time-points (FDR > 0.1). As a complementary approach, we analysed sample pools (3-4 samples/pool) corresponding to Days 0, 8 and 16 of the oestrous cycle and Day 24 of pregnancy (n = 3 pools/group) using Qiagen pcr arrays. A total of 16 miRNAs were differentially expressed (FDR < 0.1) in plasma between pregnant and non-pregnant animals. RT-qpcr validation using the same plasma samples confirmed that miR-26a was differentially upregulated on Day 16 pregnant relative to non-pregnant heifers (1.7-fold;P = 0.043), whereas miR-1249 tended to be upregulated in Day 16 pregnant heifers (1.6-fold;P = 0.081). Further validation in an independent group of heifers confirmed an increase in plasma miR-26a levels during early pregnancy, which was significant only on Day 24 (2.0-fold;P = 0.027). Conclusions: Through genome-wide analyses we have successfully p

关键词： microRNA Circulating Biomarker Early pregnancy Bovine miRNA sequencing pcr array

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Expression of cardiac insulin signalling genes and proteins in rats fed a high-sucrose diet: effect of bilberry anthocyanin extract

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GENES AND NUTRITION 2016年第1 p8-8期11卷 8-8页

作者： Singh, Shamjeet Netticadan, Thomas Ramdath, D. Dan Univ W Indies Fac Med Sci Dept Preclin Sci St Augustine Campus St Augustine Trinidad Tobago Canadian Ctr Agri Food Res Hlth & Med 351 Tache Ave Winnipeg MB Canada Agr & Agri Food Canada Guelph Food Res Ctr 93 Stone Rd West Guelph ON N1G 5C9 Canada

Scope: Insulin resistance is associated with impaired cardiac function, but the underlying molecular abnormalities are largely unexplained. Bilberry anthocyanin (BAcn) may be protective, as it appears to potentiate insulin action. Methods: Rats were randomly allocated to control, sucrose-fed (SF) or sucrose-fed + BAcn diets (SF-A) for 15 weeks. Cardiac insulin signalling genes and proteins were quantified using reverse transcription quantitative real-time polymerase chain reaction and western blots. Results: Glucose tolerance was not different with treatment. SF showed lower (p < 0.05) ferric reducing antioxidant power, which increased with BAcn. SF resulted in significantly decreased (p < 0.05) expression of 10 genes: acetyl-coenzyme A carboxylase alpha;V-Akt murine thymoma viral oncogene homolog 1;Bcl2-like 1;cytosine-cytosine-adenosine-adenosine-thymidine/enhancer binding protein;FK506 binding protein 12-rapamycin associated;glycerol-3-phosphate dehydrogenase 1 (soluble);solute carrier family 2 (facilitated glucose transporter), member 1, 4;hexokinase 2;and thyroglobulin. SF-A prevented these changes. Compared to SF-A, SF up-regulated (p < 0.05) complement factor D and phosphoinositide-3-kinase, regulatory subunit1 (a);sterol regulatory element binding transcription factor 1 was down-regulated (p < 0.05). SF increased (p < 0.05) cardiac phospholamban and decreased phosphorylated troponin I, which were not attenuated by BAcn. Compared to control or SF, SF-A resulted in significantly lower (p < 0.05) 5'-AMP-activated protein kinase. Conclusions: SF lowered antioxidant capacity and changed the expression of insulin signalling genes, which were modulated by BAcn.

关键词： Anthocyanins Heart Insulin resistance pcr array Type 2 diabetes mellitus

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Activation of RAW264.7 macrophages by the polysaccharide from the roots of Actinidia eriantha and its molecular mechanisms

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CARBOHYDRATE POLYMERS 2015年 121卷 388-402页

作者： Sun, Hongxiang Zhang, Juan Chen, Fengyang Chen, Xiangfeng Zhou, Zhihua Wang, Hui Zhejiang Univ Coll Anim Sci Minist Agr Key Lab Anim Virol Hangzhou 310058 Zhejiang Peoples R China Zhejiang Acad Med Sci Inst Mat Med Hangzhou 310013 Zhejiang Peoples R China

The polysaccharide from the roots of Actinidia eriantha (AEPS), a potent antitumor agent and immunological adjuvant, was investigated for the immunomodulatory effects on RAW264.7 macrophages and its molecular mechanisms. AEPS could significantly enhance the pinocytic and phagocytic activity, induce the production of NO, TNF-alpha, IL-10, IL-1 beta and IL-6, and promote the expression of accessory and costimulatory molecules in RAW264.7 cells. pcr array assay revealed that AEPS up-regulated 28 genes including TLRs (TLR2, TLR8, TLR9), proinflammatory factors (IL-1 beta, G-CSF, IL-1 alpha, GM-CSF, IL-6, COX-2, TNF-alpha, IFN-beta, CXCL10, CCL2, TNF-beta, IL-10), and the genes involved in NF-kappa B signaling pathway, and down-regulated 6 genes such as TLR3, TLR4, PGLYRP1, EIF2 alpha K2, MAP3K1 and IRF1. AEPS was further showed to promote cytoplasmic I kappa B-alpha degradation and increase nuclear NF-kappa B p65 levels in RAW264.7 cells. These results suggested that AEPS activated RAW264.7 macrophages and elicited a M1 and M2 response through TLRs/NF-kappa B signaling pathway. (C) 2014 Elsevier Ltd. All rights reserved.

关键词： Actinidia eriantha Polysaccharides RAW264.7 cell Toll-like receptor (TLR) NF-kappa B signaling pathway pcr array

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COMPARING THE EFFECTS OF TETRABROMOBISPHENOL-A, BISPHENOL A, AND THEIR POTENTIAL REPLACEMENT ALTERNATIVES, TBBPA-BIS(2,3-DIBROMOPROPYL ETHER) AND BISPHENOL S, ON CELL VIABILITY AND MESSENGER RIBONUCLEIC ACID EXPRESSION IN CHICKEN EMBRYONIC HEPATOCYTES

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ENVIRONMENTAL TOXICOLOGY AND CHEMISTRY 2015年第2期34卷 391-401页

作者： Ma, Melissa Crump, Doug Farmahin, Reza Kennedy, Sean W. Environm Canada Natl Wildlife Res Ctr Ottawa ON K1A 0H3 Canada Univ Ottawa Dept Biol Ottawa ON Canada

A market for alternative brominated flame retardants (BFRs) has emerged recently due to the phase out of persistent and inherently toxic BFRs. Several of these replacement compounds have been detected in environmental matrices, including wild birds. A chicken embryonic hepatocyte (CEH) assay was utilized to assess the effects of the BFR, tetrabromobisphenol-A (TBBPA), and its replacement alternative, tetrabromobisphenol A bis(2,3-dibromopropyl ether [TBBPA-DBPE]) on cell viability and messenger ribonucleic acid (mRNA) expression. Bisphenol A (BPA) and 1 of its replacement alternatives, bisphenol S (BPS), were also screened for effects. Both TBBPA and BPA decreased CEH viability with calculated median lethal concentration (LC50) values of 40.6M and 61.7M, respectively. However, the replacement alternatives, TBBPA-DBPE and BPS, did not affect cell viability (up to 300M). Effects on mRNA expression were determined using an Avian ToxChip polymerse chain reaction (pcr) array and a real-time (RT)-pcr assay for the estrogen-responsive genes, apolipoproteinII (ApoII) and vitellogenin (Vtg). A luciferase reporter gene assay was used to assess dioxin-like effects. Tetrabromobisphenol-A altered mRNA levels of 4 genes from multiple toxicity pathways and increased luciferase activity in the luciferase reporter gene assay, whereas its alternative, TBBPA-DBPE, only altered 1 gene on the array, Cyp1a4, and increased luciferase activity. At 300M, a concentration that decreased cell viability for TBBPA and BPA, the BPA replacement, BPS, altered the greatest number of transcripts, including both ApoII and Vtg. Bisphenol A exposure did not alter any genes on the array but did up-regulate Vtg at 10M. Characterization of the potential toxicological and molecular-level effects of these compounds will ideally be useful to chemical regulators tasked with assessing the risk of new and existing chemicals. Environ Toxicol Chem 2014;9999:1-11. (c) 2014 SETAC

关键词： Bisphenol S Bisphenol A Tetrabromobisphenol A Tetrabromobisphenol A bis(2 3-dibromopropyl ether) pcr array Avian

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An In Vitro alveolar macrophage assay for the assessment of inflammatory cytokine expression induced by atmospheric particulate matter

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ENVIRONMENTAL TOXICOLOGY 2015年第7期30卷 836-851页

作者： Sijan, Zana Antkiewicz, Dagmara S. Heo, Jongbae Kado, Norman Y. Schauer, James J. Sioutas, Constantinos Shafer, Martin M. Univ Wisconsin Dept Environm Chem & Technol Madison WI 53706 USA Univ Wisconsin Wisconsin State Lab Hyg Dept Environm Toxicol Madison WI 53718 USA Univ Calif Davis Dept Environm Toxicol Davis CA 95616 USA Calif Environm Protect Agcy Air Resources Board Sacramento CA USA Univ So Calif Dept Civil & Environm Engn Los Angeles CA 90089 USA Univ Wisconsin Wisconsin State Lab Hyg Madison WI 53718 USA

Exposures to air pollution in the form of particulate matter (PM) can result in excess production of reactive oxygen species (ROS) in the respiratory system, potentially causing both localized cellular injury and triggering a systemic inflammatory response. PM-induced inflammation in the lung is modulated in large part by alveolar macrophages and their biochemical signaling, including production of inflammatory cytokines, the primary mechanism via which inflammation is initiated and sustained. We developed a robust, relevant, and flexible method employing a rat alveolar macrophage cell line (NR8383) which can be applied to routine samples of PM from air quality monitoring sites to gain insight into the drivers of PM toxicity that lead to oxidative stress and inflammation. Method performance was characterized using extracts of ambient and vehicular engine exhaust PM samples. Our results indicate that the reproducibility and the sensitivity of the method are satisfactory and comparisons between PM samples can be made with good precision. The average relative percent difference for all genes detected during 10 different exposures was 17.1%. Our analysis demonstrated that 71% of genes had an average signal to noise ratio (SNR) 3. Our time course study suggests that 4 h may be an optimal in vitro exposure time for observing short-term effects of PM and capturing the initial steps of inflammatory signaling. The 4 h exposure resulted in the detection of 57 genes (out of 84 total), of which 86% had altered expression. Similarities and conserved gene signaling regulation among the PM samples were demonstrated through hierarchical clustering and other analyses. Overlying the core congruent patterns were differentially regulated genes that resulted in distinct sample-specific gene expression fingerprints. Consistent upregulation of Il1f5 and downregulation of Ccr7 was observed across all samples, while TNF was upregulated in half of the samples and downregulated in the other h

关键词： air pollution particulate matter inflammation toxicity cytokines gene expression pcr array oxidative stress ROS macrophages

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Safety assessment of potential food ingredients in canine hepatocytes

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FOOD AND CHEMICAL TOXICOLOGY 2015年 78卷 105-115页

作者： Zhang, Leshuai W. Koci, Juraj Jeffery, Brett Riviere, Jim E. Monteiro-Riviere, Nancy A. Kansas State Univ Coll Vet Med Dept Anat & Physiol Manhattan KS 66506 USA Mars Global Food Safety Ctr Beijing 101407 Peoples R China

This research aimed to develop in vitro methods to assess hazard of canine food ingredients. Canine hepatocytes were harvested and cell viability of clove-leaf oil (CLO), eugenol (EUG), lemongrass oil (LGO), guanosine monophosphate (GMP), inosine monophosphate (IMP), sorbose, ginger-root extract (GRE), cinnamon-bark oil (CBO), cinnamaldehyde (CINA), thymol oil (TO), thymol (THYM), and citric acid were assessed with positive controls: acetaminophen (APAP), aflatoxin B1 and xylitol. Molecular Toxicology PathwayFinder array (MTPF) analyzed toxicity mechanisms for LGO. LC50 for APAP was similar among human (3.45), rat (2.35), dog (4.26 mg/ml). Aflatoxin B1 had an LC50 of 4.43 (human), 5.78 (rat) and 6.05 (dog) mu g/ml;xylitol did not decrease viability. LC50 of CLO (0.185 +/- 0.075(SD)), EUG (0.165 +/- 0.112), LGO (0.220 +/- 0.012), GRE (1.54 +/- 0.31) mg/m1;GMP (166.03 +/- 41.83), GMP + IMP (208.67 +/- 15.27) mM;CBO (0.08 +/- 0.03), CINA (0.11 +/- 0.01), TO (0.21 +/- 0.03), THYM (0.05 +/- 0.01), citric acid (1.58 +/- 0.08) mg/ml, while sorbose was non-toxic. LGO induced upregulation of 16 and down-regulation of 24 genes, which CYP and heat shock most affected. These results suggest that in vitro assays such as this may be useful for hazard assessment of food ingredients for altered hepatic function. (C) 2015 Elsevier Ltd. All rights reserved.

关键词： Hepatocytes In vitro Toxicity Food ingredient pcr array Dog

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Biological tests for major depressive disorder that involve leukocyte gene expression assays

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JOURNAL OF PSYCHIATRIC RESEARCH 2015年 66-67卷 1-6页

作者： Watanabe, Shin-ya Iga, Jun-ichi Ishii, Kazuo Numata, Shusuke Shimodera, Shinji Fujita, Hirokazu Ohmori, Tetsuro Univ Tokushima Sch Med Dept Psychiat Course Integrated Brain Sci Tokushima 7708503 Japan Tokyo Univ Agr & Technol Fac Agr Dept Appl Biol Sci Fuchu Tokyo 1838509 Japan Kochi Univ Kochi Med Sch Dept Neuropsychiat Kochi 780 Japan

Background: Development of easy-to-use biological diagnostic tests for major depressive disorder (MDD) may facilitate MDD diagnosis and delivery of optimal treatment. Here, we examined leukocyte gene expression to develop a biological diagnostic test for MDD. Methods: 25 drug-naive MDD patients (MDDs) and 25 age- and sex-matched healthy subjects (Controls) participated in a pilot study. A subsequent replication study involved 20 MDDs and 18 Controls. We used custom-made pcr array plates to examine mRNA levels of 40 candidate genes in leukocyte samples to assess whether any combination of these genes could be used to differentiate MDDs from Controls based on expression profiles. Results: Among 40 candidate genes, we identified a set of seven genes (PDGFC, SLC6A4, PDLIM5, ARKGA P24, PRNP, HDAC5, and IL1R2), each of which had expression levels that differed significantly between MDD and Control samples in the pilot study. To identify genes whose expression best differentiated between MDDs and Controls, a linear discriminant function was developed to discriminate between MDDs and Controls based on the standardized values of gene expression after Z-score transformation. Ultimately, five genes (PDGFC, SLC6A4, ARHGAP24, PRNP, and HDAC5) were selected for a multi-assay diagnostic test. In the pilot study, this diagnostic test demonstrated sensitivity and specificity of 80% and 92%, respectively. The replication study yielded nearly identical results, sensitivity of 85% and specificity. of 89%. Conclusions: Using leukocyte gene expression profiles, we could differentiate MDDs from Controls with adequate sensitivity and specificity. Additional markers not yet identified might further improve the performance of this test. (C) 2015 Elsevier Ltd. All rights reserved.

关键词： Major depressive disorder Healthy subjects Leukocyte gene expression Biomarker Diagnostic test pcr array

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The apoptotic thanatotranscriptome associated with the liver of cadavers

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FORENSIC SCIENCE MEDICINE AND PATHOLOGY 2015年第4期11卷 509-516页

作者： Javan, Gulnaz T. Can, Ismail Finley, Sheree J. Soni, Shivani Alabama State Univ Forens Sci Program Montgomery AL 36104 USA Alabama State Univ Montgomery AL 36104 USA Alabama State Univ Dept Biol Sci Montgomery AL 36101 USA

Gene expression investigations are well-established components of ante mortem studies with broad applications ranging from elucidating basic mechanisms responsible for normal physiological processes to discovering therapeutic targets in pathophysiological conditions. However, gene expression studies and their application in the medico-legal field are still in their infancy. Therefore, the present study focuses on RNA using pcr array in the analysis of gene expression associated with tissues taken from actual criminal cases. RNA was extracted from the liver tissues of bodies with PMIs between 6 and 48 h. The results demonstrated that mRNA was stable up to 48 h postmortem. Further, as cell death is an indispensable and necessary part of the biological life cycle, apoptotic gene expression profiles were investigated. The gene expression related to the programmed cell death found in body tissues after death is defined as the apoptotic thanatotranscriptome (thanatos-, Greek for death). On comparison of control and decaying tissues, the results show that with time, pro-apoptotic genes such as caspases are up-regulated and the expression of genes responsible for anti-apoptosis such as BCL2 and BAG3 were down-regulated. Thus, this current work gives a unique perspective of the apoptotic thanatotranscriptome that is affected after death. Up to the present time, gene expression in bodies from criminal cases has not been reported in literature using pcr array techniques. Thus, this thanatotranscriptome study provides insight into postmortem gene activity with potential applications in medico-legal investigations.

关键词： Thanatotranscriptome Gene expression Cadaver liver pcr array

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Resveratrol prevents hepatic steatosis and endoplasmic reticulum stress and regulates the expression of genes involved in lipid metabolism, insulin resistance, and inflammation in rats

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NUTRITION RESEARCH 2015年第7期35卷 576-584页

作者： Pan, Qing-Rong Ren, Yan-Long Liu, Wen-Xian Hu, Yan-Jin Zheng, Jin-Su Xu, Yuan Wang, Guang Capital Med Univ Beijing Chaoyang Hosp Dept Endocrinol Beijing 100020 Peoples R China Capital Med Univ Beijing Anzhen Hosp Dept Cardiol Beijing Inst Heart Lung & Blood Vessel Dis Beijing 100029 Peoples R China Capital Med Univ Beijing Chaoyang Hosp Dept Tradit Chinese Med Beijing 100020 Peoples R China

Previous research demonstrated that resveratrol possesses promising properties for preventing obesity. Endoplasmic reticulum (ER) stress was proposed to be involved in the pathophysiology of both obesity and hepatic steatosis. In the current study, we hypothesized that resveratrol could protect against high-fat diet (HFD)-induced hepatic steatosis and ER stress and regulate the expression of genes related to hepatic steatosis. Rats were fed either a control diet or a HFD for 12 weeks. After 4 weeks, HFD-fed rats were treated with either resveratrol or vehicle for 8 weeks. Body weight, serum metabolic parameters, hepatic histopathology, and hepatic ER stress markers were evaluated. Moreover, an RT2 Profiler Fatty Liver pcr array was performed to investigate the mRNA expressions of 84 genes related to hepatic steatosis. Our work showed that resveratrol prevented dyslipidemia and hepatic steatosis induced by HFD. Resveratrol significantly decreased activating transcription factor 4, C/EBP-homologous protein and immunoglobulin binding protein levels, which were elevated by the HFD. Resveratrol also decreased PKR-like ER kinase phosphorylation, although it was not affected by the HFD. Furthermore, resveratrol increased the expression of peroxisome proliferator-activated receptor delta, while decreasing the expression of ATP citrate lyase, suppressor of cytokine signaling-3, and interleukin-1 beta. Our data suggest that resveratrol can prevent hepatic ER stress and regulate the expression of peroxisome proliferator-activated receptor delta, ATP citrate lyase, suppressor of cytokine signaling-3, tumor necrosis factor alpha, and interleukin-1 beta in diet-induced obese rats, and these effects likely contribute to resveratrol's protective function against excessive accumulation of fat in the liver. (C) 2015 Published by Elsevier Inc.

关键词： Resveratrol Hepatic steatosis Endoplasmic reticulum stress pcr array Rat

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Abnormal expression of miR-1 in breast carcinoma as a potent prognostic factor

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CANCER SCIENCE 2015年第11期106卷 1642-1650页

作者： Minemura, Hiroyuki Takagi, Kiyoshi Miki, Yasuhiro Shibahara, Yukiko Nakagawa, Saki Ebata, Akiko Watanabe, Mika Ishida, Takanori Sasano, Hironobu Suzuki, Takashi Tohoku Univ Grad Sch Med Dept Pathol & Histotechnol Aoba Ku Sendai Miyagi 9808575 Japan Tohoku Univ Grad Sch Med Dept Anat Pathol Sendai Miyagi 9808575 Japan Tohoku Univ Grad Sch Med Dept Surg Oncol Sendai Miyagi 9808575 Japan Tohoku Univ Hosp Dept Pathol Sendai Miyagi Japan

Metastatic breast cancer remains a highly lethal disease, and it is very important to evaluate the biomarkers associated with the distant metastasis. MicroRNA (miRNA) are small non-protein coding RNA that regulate various cellular functions. Recent investigations have demonstrated the importance of some miRNA in breast cancer, but the significance of the great majority of miRNA remains largely unclear in breast cancer metastasis. Therefore, in this study, we first examined expression profiles of miRNA in stage IV breast carcinoma tissues, comparing stage I-III cases by miRNA pcr array, and identified miR-1 as the miRNA which was the most associated with the distant metastasis. However, miR-1 has not yet been examined in breast carcinoma tissue, and its significance remains unknown. Therefore, we further examined miR-1 expression in breast carcinoma using in situ hybridization (ISH). miR-1 was localized in carcinoma cells in 20% of breast carcinoma cases, but it was negligible in non-neoplastic mammary glands or stroma. miR-1 ISH status was significantly associated with stage, pathological T factor, lymph node metastasis, distant metastasis, histological grade, estrogen receptor, progesterone receptor and Ki-67 in breast carcinoma. Moreover, the miR-1 status was demonstrated using multivariate analysis as an independent worse prognostic factor for both disease-free and breast cancer-specific survival. These findings suggest that abnormal miR-1 expression is associated with an aggressive phenotype of breast carcinoma and that miR-1 status is a potent prognostic factor in human breast cancer patients.

关键词： Breast cancer in situ hybridization microRNA pcr array prognosis

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