title: iTRAQ-proteomics and bioinformatics analyses of mammary tissue from cows with clinical mastitis due to natural infection with Staphylococci aureus.
abstract: BACKGROUND
  Proteomics and bioinformatics may help us better understand the biological adaptations occurring during bovine mastitis. This systems approach also could help identify biomarkers for monitoring clinical and subclinical mastitis. The aim of the present study was to use isobaric tags for relative and absolute quantification (iTRAQ) to screen potential proteins associated with mastitis at late infectious stage.
  
  
  RESULTS
  Healthy and mastitic cows' mammary gland tissues were analyzed using iTRAQ combined with two-dimensional liquid chromatography-tandem mass spectrometry (2D-LC-MS/MS). Bioinformatics analyses of differentially expressed proteins were performed by means of Gene Ontology, metabolic pathways, transcriptional regulation networks using Blast2GO software, the Dynamic Impact Approach and Ingenuity Pathway Analysis. At a false discovery rate of 5%, a total of 768 proteins were identified from 6,499 peptides, which were matched with 15,879 spectra. Compared with healthy mammary gland tissue, 36 proteins were significantly up-regulated (>1.5-fold) while 19 were significantly down-regulated (<0.67-fold) in response to mastitis due to natural infections with Staphylococci aureus. Up-regulation of collagen, type I, alpha 1 (COL1A1) and inter-alpha (Globulin) inhibitor H4 (ITIH4) in the mastitis-infected tissue was confirmed by Western blotting and Immunohistochemistry.
  
  
  CONCLUSIONS
  This paper is the first to show the protein expression in the late response to a mastitic pathogen, thus, revealing mechanisms associated with host tissue damage. The bioinformatics analyses highlighted the effects of mastitis on proteins such as collagen, fibrinogen, fibronectin, casein alpha and heparan sulfate proteoglycan 2. Our findings provide additional clues for further studies of candidate genes for mastitis susceptibility. The up-regulated expression of COL1A1 and ITIH4 in the mastitic mammary gland may be associated with tissue damage and repair during late stages of infection.
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, No,159 North of Industry Road, Jinan, Shandong 250131, China. huangjinm@sina.com.
authors.forename: Jinming
authors.initials: J
authors.lastname: Huang
authors.forename: Guojing
authors.initials: G
authors.lastname: Luo
authors.forename: Zijing
authors.initials: Z
authors.lastname: Zhang
authors.forename: Xiuge
authors.initials: X
authors.lastname: Wang
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Ju
authors.forename: Chao
authors.initials: C
authors.lastname: Qi
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.forename: Changfa
authors.initials: C
authors.lastname: Wang
authors.forename: Rongling
authors.initials: R
authors.lastname: Li
authors.forename: Jianbin
authors.initials: J
authors.lastname: Li
authors.forename: Weijun
authors.initials: W
authors.lastname: Yin
authors.forename: Yinxue
authors.initials: Y
authors.lastname: Xu
authors.forename: Sonia J
authors.initials: SJ
authors.lastname: Moisá
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Loor
authors.forename: Jifeng
authors.initials: J
authors.lastname: Zhong
mesh_terms: D000818:Animals; D002417:Cattle; D024042:Collagen Type I / Q000378:metabolism; D019295:Computational Biology; D015180:Electrophoresis, Gel, Two-Dimensional; D005260:Female; D053263:Gene Regulatory Networks / Q000235:genetics; D008321:Mammary Glands, Animal / Q000378:metabolism* / Q000473:pathology; D008414:Mastitis, Bovine / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D053858:Metabolic Networks and Pathways / Q000235:genetics; D058977:Molecular Sequence Annotation; D053491:Proteinase Inhibitory Proteins, Secretory / Q000378:metabolism; D040901:Proteomics; D012984:Software; D013203:Staphylococcal Infections / Q000378:metabolism* / Q000473:pathology / Q000662:veterinary; D013211:Staphylococcus aureus / Q000472:pathogenicity; D053719:Tandem Mass Spectrometry; D015854:Up-Regulation
chemical_list: D024042:Collagen Type I; D053491:Proteinase Inhibitory Proteins, Secretory