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      The Effectiveness of Dietary Byproduct Antioxidants on Induced CYP Genes Expression and Histological Alteration in Piglets Liver and Kidney Fed with Aflatoxin B1 and Ochratoxin A

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          Abstract

          The purpose of this study was to investigate the potential of a byproduct mixture derived from grapeseed and sea buckthorn oil industry to mitigate the harmful damage produced by ochratoxin A and aflatoxin B1 at hepatic and renal level in piglets after weaning. Forty cross-bred TOPIGS-40 hybrid piglets after weaning were assigned to three experimental groups (E1, E2, E3) and one control group (C), and fed with experimental diets for 30 days. The basal diet was served as a control and contained normal compound feed for starter piglets without mycotoxins. The experimental groups were fed as follows: E1—basal diet plus a mixture (1:1) of two byproducts (grapeseed and sea buckthorn meal); E2—the basal diet experimentally contaminated with mycotoxins (479 ppb OTA and 62ppb AFB1); and E3—basal diet containing 5% of the mixture (1:1) of grapeseed and sea buckthorn meal and contaminated with the mix of OTA and AFB1. After 4 weeks, the animals were slaughtered, and tissue samples were taken from liver and kidney in order to perform gene expression and histological analysis. The gene expression analysis showed that when weaned piglets were fed with contaminated diet, the expression of most analyzed genes was downregulated. Among the CYP450 family, CYP1A2 was the gene with the highest downregulation. According to these results, in liver, we found that mycotoxins induced histomorphological alterations in liver and kidney and had an effect on the expression level of CYP1A2, CYP2A19, CYP2E1, and CYP3A29, but we did not detect important changes in the expression level of CY4A24, MRP2 and GSTA1 genes.

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          Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method.

          The two most commonly used methods to analyze data from real-time, quantitative PCR experiments are absolute quantification and relative quantification. Absolute quantification determines the input copy number, usually by relating the PCR signal to a standard curve. Relative quantification relates the PCR signal of the target transcript in a treatment group to that of another sample such as an untreated control. The 2(-Delta Delta C(T)) method is a convenient way to analyze the relative changes in gene expression from real-time quantitative PCR experiments. The purpose of this report is to present the derivation, assumptions, and applications of the 2(-Delta Delta C(T)) method. In addition, we present the derivation and applications of two variations of the 2(-Delta Delta C(T)) method that may be useful in the analysis of real-time, quantitative PCR data. Copyright 2001 Elsevier Science (USA).
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            A new approach to rapid sequence comparison, basic local alignment search tool (BLAST), directly approximates alignments that optimize a measure of local similarity, the maximal segment pair (MSP) score. Recent mathematical results on the stochastic properties of MSP scores allow an analysis of the performance of this method as well as the statistical significance of alignments it generates. The basic algorithm is simple and robust; it can be implemented in a number of ways and applied in a variety of contexts including straightforward DNA and protein sequence database searches, motif searches, gene identification searches, and in the analysis of multiple regions of similarity in long DNA sequences. In addition to its flexibility and tractability to mathematical analysis, BLAST is an order of magnitude faster than existing sequence comparison tools of comparable sensitivity.
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              The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments.

              Currently, a lack of consensus exists on how best to perform and interpret quantitative real-time PCR (qPCR) experiments. The problem is exacerbated by a lack of sufficient experimental detail in many publications, which impedes a reader's ability to evaluate critically the quality of the results presented or to repeat the experiments. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency. MIQE is a set of guidelines that describe the minimum information necessary for evaluating qPCR experiments. Included is a checklist to accompany the initial submission of a manuscript to the publisher. By providing all relevant experimental conditions and assay characteristics, reviewers can assess the validity of the protocols used. Full disclosure of all reagents, sequences, and analysis methods is necessary to enable other investigators to reproduce results. MIQE details should be published either in abbreviated form or as an online supplement. Following these guidelines will encourage better experimental practice, allowing more reliable and unequivocal interpretation of qPCR results.
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                Author and article information

                Journal
                Toxins (Basel)
                Toxins (Basel)
                toxins
                Toxins
                MDPI
                2072-6651
                15 February 2021
                February 2021
                : 13
                : 2
                : 148
                Affiliations
                [1 ]Department of Biochemistry and Molecular Biology, Faculty of Biology, University of Bucharest, Splaiul Independentei No. 91–95, 050095 Bucharest, Romania; roua.popescu@ 123456drd.unibuc.ro (R.G.P.); anca.dinischiotu@ 123456bio.unibuc.ro (A.D.)
                [2 ]Laboratory of Animal Biology, National Institute for Research and Development for Biology and Animal Nutrition, Calea Bucuresti No. 1, Balotesti, 077015 Ilfov, Romania; cristina.bulgaru@ 123456ibna.ro (C.B.); arabela.untea@ 123456ibna.ro (A.U.); daniela.marin@ 123456ibna.ro (D.M.)
                [3 ]Raluca Ripan Institute for Research in Chemistry, Babeş Bolyai University, 30 Fântânele Street, 400294 Cluj-Napoca, Romania; mihaela.vlassa@ 123456ubbcluj.ro (M.V.); miuta.filip@ 123456ubbcluj.ro (M.F.)
                [4 ]“Aurel Ardelean” Institute of Life Sciences, Vasile Godis Western University of Arad, Rebreanu 86, 310414 Arad, Romania; anca.hermenean@ 123456gmail.com
                Author notes
                [* ]Correspondence: ionelia.taranu@ 123456ibna.ro (I.Ț.); sergiu.georgescu@ 123456bio.unibuc.ro (S.E.G.); Tel.: +40-213181575 (ext. 112) (S.E.G.)
                Author information
                https://orcid.org/0000-0003-3000-5024
                https://orcid.org/0000-0001-8510-6653
                https://orcid.org/0000-0003-0727-5827
                https://orcid.org/0000-0003-2631-9098
                Article
                toxins-13-00148
                10.3390/toxins13020148
                7919288
                5599f5b9-8bf0-4249-9c5f-3c7275cbc445
                © 2021 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 11 January 2021
                : 13 February 2021
                Categories
                Article

                Molecular medicine
                piglets,antioxidant effect,feed additives,mycotoxins,cyps gene expression
                Molecular medicine
                piglets, antioxidant effect, feed additives, mycotoxins, cyps gene expression

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