Biotechnology Bulletin ›› 2018, Vol. 34 ›› Issue (1): 215-222.doi: 10.13560/j.cnki.biotech.bull.1985.2017-0731

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Effects of Lectin Subunit Alpha on the Expression of Staphylococcus aureus-stimulated Inflammatory Cytokines Mediated

WEI Rui-yang1,2,3, BAI Jie1,3, XU Bin1,3, XI Yan-yan1,3, WANG Lin-yi1,3, WANG Gai-li1,3, FU Chen1,3, WEI Feng-xian1,3, LI Shao-yu1,3   

  1. 1. Institute of Animal Husbandry and Veterinary Science,Henan Academy of Agricultural Sciences,Zhengzhou 450002;
    2. College of Animal Science and Veterinary Medicine,Henan Agricultural University,Zhengzhou 450002;
    3. Henan Center for Feed and Aquaculture Environment Control Engineering Techniques Research,Henan Key Laboratory of Farm Animal Breeding and Nutritional Regulation,Zhengzhou 450002
  • Received:2017-09-05 Online:2018-01-26 Published:2018-01-22

Abstract: The attempt of this work is to investigate the anti-inflammatory activity of Musca domestica lectin subunit alpha(Lsa)toward Staphylococcus aureus. The well-constructed pLEX-Lsa and pLEX were transfected into RAW264.7 cells with recombinant lentiviruses,then the stably-expressed RAW-pLEX-Lsa and RAW-pLEX were obtained after screening,and RAW-pLEX was as a negative control. S. aureus was used to stimulate RAW-pLEX-Lsa cells and RAW-pLEX cells,then both un-stimulated cells and stimulated cells for 3 h,6 h,12 h,and 24 h were collected,and the cell culture supernatants of 6 h un-stimulated and stimulated cells were also collected. Real-time quantitative reverse transcription PCR(RT-PCR)was performed to analyze the mRNA expression of TNF-a,IL-1β,NFκB-1 and NFκB-2,and the protein level of TNF-a and IL-1β in the supernatant were detected by ELISA. The results of RT-PCR showed that the stimulation of RAW-pLEX-Lsa by S. aureussignificantly down-regulated the mRNA expression of TNF-a transcript variant 1(TNF-a-tv-1)at 6 h(P<0.05)and TNF-a transcript variant 2(TNF-a-tv-2)at 6 h and 12 h(P < 0.05),compared with the RAW-pLEX cells. The stable transfection of Lsa in RAW264.7 cells stimulated by S. aureus significantly down-regulated the mRNA expression of IL-1β-T at 3 h,6 h and 12 h(P < 0.05)and the mRNA expression of IL-1β at 3 h and 6 h(P < 0.05),compared with the RAW-pLEX cells,while the mRNA expression of NFκB-1 and NFκB-2 in RAW-pLEX-Lsa had insignificant decreasing. The results of ELISA demonstrated that the protein level of TNF-a in the supernatants of RAW-pLEX-Lsa cells at 6 h after stimulation by S. aureus significantly down-regulated(P < 0.05). In conclusion,Lsa possesses potent anti-inflammatory activity,and stable transfection of Lsa in RAW264.7 cells inhibits the expression and production of TNF-α and IL-1β. However,the inhibition of TNF-α and IL-1β is not resulted from blocking the activation of NF-κB signal pathway.

Key words: lectin, RAW264.7, inflammatory cytokines, NFκB, transcript variant