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      Inflammation-related microRNA expression level in the bovine milk is affected by mastitis

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          Abstract

          MicroRNA (miRNA) in tissue and liquid samples have been shown to be associated with many diseases including inflammation. We aimed to identify inflammation-related miRNA expression level in the bovine mastitis milk. Expression level of inflammation-related miRNA in milk from mastitis-affected and normal cows was analyzed using qPCR. We found that expression level of miR-21, miR-146a, miR-155, miR-222, and miR-383 was significantly upregulated in California mastitis test positive (CMT+) milk. We further analyzed these miRNA using a chip-based QuantStudio Digital PCR System. The digital PCR results correlated with those of qPCR, demonstrating upregulation of miR-21, miR-146a, miR-155, miR-222, and miR-383 in CMT+ milk. In conclusion, we identified miRNA that are upregulated in CMT+ milk. These miRNA exhibited sensitivity and specificity greater than 80% for differentiating between CMT+ milk and normal milk. Our findings suggest that inflammation-related miRNA expression level in the bovine milk was affected by mastitis, and miRNA in milk have potential for use as biomarkers of bovine mastitis.

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          Most cited references42

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          Negative regulation of TLR4 via targeting of the proinflammatory tumor suppressor PDCD4 by the microRNA miR-21.

          The tumor suppressor PDCD4 is a proinflammatory protein that promotes activation of the transcription factor NF-kappaB and suppresses interleukin 10 (IL-10). Here we found that mice deficient in PDCD4 were protected from lipopolysaccharide (LPS)-induced death. The induction of NF-kappaB and IL-6 by LPS required PDCD4, whereas LPS enhanced IL-10 induction in cells lacking PDCD4. Treatment of human peripheral blood mononuclear cells with LPS resulted in lower PDCD4 expression, which was due to induction of the microRNA miR-21 via the adaptor MyD88 and NF-kappaB. Transfection of cells with a miR-21 precursor blocked NF-kappaB activity and promoted IL-10 production in response to LPS, whereas transfection with antisense oligonucleotides to miR-21 or targeted protection of the miR-21 site in Pdcd4 mRNA had the opposite effect. Thus, miR-21 regulates PDCD4 expression after LPS stimulation.
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            MicroRNA as Biomarkers and Diagnostics.

            MicroRNAs (miRNAs) are a group of small non-coding RNAs that are involved in regulating a range of developmental and physiological processes; their dysregulation has been associated with development of diseases including cancer. Circulating miRNAs and exosomal miRNAs have also been proposed as being useful in diagnostics as biomarkers for diseases and different types of cancer. In this review, miRNAs are discussed as biomarkers for cancer and other diseases, including viral infections, nervous system disorders, cardiovascular disorders, and diabetes. We summarize some of the clinical evidence for the use of miRNAs as biomarkers in diagnostics and provide some general perspectives on their use in clinical situations. The analytical challenges in using miRNAs in cancer and disease diagnostics are evaluated and discussed. Validation of specific miRNA signatures as biomarkers is a critical milestone in diagnostics.
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              Optimal cut-point and its corresponding Youden Index to discriminate individuals using pooled blood samples.

              Costs can hamper the evaluation of the effectiveness of new biomarkers. Analysis of smaller numbers of pooled specimens has been shown to be a useful cost-cutting technique. The Youden index (J), a function of sensitivity (q) and specificity (p), is a commonly used measure of overall diagnostic effectiveness. More importantly, J is the maximum vertical distance or difference between the ROC curve and the diagonal or chance line; it occurs at the cut-point that optimizes the biomarker's differentiating ability when equal weight is given to sensitivity and specificity. Using the additive property of the gamma and normal distributions, we present a method to estimate the Youden index and the optimal cut-point, and extend its applications to pooled samples. We study the effect of pooling when only a fixed number of individuals are available for testing, and pooling is carried out to save on the number of assays. We measure loss of information by the change in root mean squared error of the estimates of the optimal cut-point and the Youden index, and we study the extent of this loss via a simulation study. In conclusion, pooling can result in a substantial cost reduction while preserving the effectiveness of estimators, especially when the pool size is not very large.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                17 May 2017
                2017
                : 12
                : 5
                : e0177182
                Affiliations
                [1 ]Veterinary Teaching Hospital, Joint Faculty of Veterinary Medicine, Kagoshima University, Kagoshima, Japan
                [2 ]The United Graduate School of Veterinary Science, Yamaguchi University, Yamaguchi, Japan
                [3 ]Laboratory of Veterinary Theriogenology, Joint Faculty of Veterinary Medicine, Kagoshima University, Kagoshima, Japan
                [4 ]Department of Veterinary Sciences, Faculty of Agriculture, University of Miyazaki, Miyazaki, Japan
                [5 ]Laboratory of Small Animal Internal Medicine, Joint Faculty of Veterinary Medicine, Kagoshima University, Kagoshima, Japan
                [6 ]Laboratory of Veterinary Clinical Pathology, Joint Faculty of Veterinary Medicine, Kagoshima University, Kagoshima, Japan
                [7 ]School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido, Japan
                Gustave Roussy, FRANCE
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                • Conceptualization: YL NM.

                • Formal analysis: YE OY NM.

                • Funding acquisition: CK NM.

                • Investigation: YL TF TM.

                • Project administration: NM.

                • Resources: TA GK MK.

                • Supervision: CK NM.

                • Validation: YL NM.

                • Writing – original draft: YL TM.

                • Writing – review & editing: YL NM.

                Author information
                http://orcid.org/0000-0002-4625-5231
                Article
                PONE-D-17-00948
                10.1371/journal.pone.0177182
                5435311
                28520748
                1d5bc3f8-72a2-461a-873a-d170d8e38ecf
                © 2017 Lai et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 8 January 2017
                : 23 April 2017
                Page count
                Figures: 5, Tables: 0, Pages: 13
                Funding
                Funded by: the Project of the Bio-oriented Technology Research Advancement Institution, NARO (the special scheme project on advanced research and development for next-generation technology)
                Award Recipient :
                Funded by: the Project of the Bio-oriented Technology Research Advancement Institution, NARO (the special scheme project on advanced research and development for next-generation technology)
                Award Recipient :
                Funded by: the JSPS KAKENHI
                Award ID: 25660243
                Award Recipient :
                Funded by: the Morinaga Houshikai Research Grant
                This work was supported by the Project of the Bio-oriented Technology Research Advancement Institution, NARO (the special scheme project on advanced research and development for next-generation technology) to CK and NM, and by the the JSPS KAKENHI-25660243 to NM, and by the the Morinaga Houshikai Research Grant to NM.
                Categories
                Research Article
                Biology and life sciences
                Genetics
                Gene expression
                Gene regulation
                MicroRNAs
                Biology and life sciences
                Biochemistry
                Nucleic acids
                RNA
                Non-coding RNA
                MicroRNAs
                Biology and Life Sciences
                Zoology
                Animal Diseases
                Bovine Mastitis
                Medicine and Health Sciences
                Women's Health
                Maternal Health
                Mastitis
                Biology and Life Sciences
                Immunology
                Immune Response
                Inflammation
                Medicine and Health Sciences
                Immunology
                Immune Response
                Inflammation
                Medicine and Health Sciences
                Diagnostic Medicine
                Signs and Symptoms
                Inflammation
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Signs and Symptoms
                Inflammation
                Medicine and Health Sciences
                Inflammatory Diseases
                Biology and Life Sciences
                Biochemistry
                Biomarkers
                Research and analysis methods
                Extraction techniques
                RNA extraction
                Biology and Life Sciences
                Veterinary Science
                Veterinary Diseases
                Custom metadata
                All relevant data are within the paper and its Supporting Information files.

                Uncategorized
                Uncategorized

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