1. Academic Validation
  2. Surface-exposed loops L7 and L8 of Haemophilus (Glaesserella) parasuis OmpP2 contribute to the expression of proinflammatory cytokines in porcine alveolar macrophages

Surface-exposed loops L7 and L8 of Haemophilus (Glaesserella) parasuis OmpP2 contribute to the expression of proinflammatory cytokines in porcine alveolar macrophages

  • Vet Res. 2019 Nov 29;50(1):105. doi: 10.1186/s13567-019-0721-4.
Ye Zhou 1 Saixiang Feng 2 Xinyi He 1 Qun Zhou 1 Yuanwei Wang 1 Hua Yue 1 3 Cheng Tang 1 3 Bin Zhang 4 5
Affiliations

Affiliations

  • 1 College of Life Science and Technology, Southwest Minzu University, Chengdu, 610041, China.
  • 2 College of Veterinary Medicine, South China Agricultural University, Guangzhou, 510642, China.
  • 3 Key Laboratory of Ministry of Education and Sichuan Province for Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization, Chengdu, 610041, China.
  • 4 College of Life Science and Technology, Southwest Minzu University, Chengdu, 610041, China. binovy@sina.com.
  • 5 Key Laboratory of Ministry of Education and Sichuan Province for Qinghai-Tibetan Plateau Animal Genetic Resource Reservation and Utilization, Chengdu, 610041, China. binovy@sina.com.
Abstract

Outer membrane protein P2 (OmpP2) of the virulent Haemophilus (Glaesserella) parasuis has been shown to induce the release of proinflammatory cytokines. The OmpP2 protein is composed of eight or nine surface-exposed loops, but it is unclear which of them participates in the OmpP2-induced inflammatory response. In this study, we synthesized linear Peptides corresponding to surface-exposed loops L1-L8 of OmpP2 from the virulent H. parasuis SC096 strain to stimulate porcine alveolar macrophages (PAMs) in vitro. We found that both L7 and L8 significantly upregulated the mRNA expression of interleukin (IL)-1α, IL-1β, IL-6, IL-8, IL-17, and IL-23 and the chemokines CCL-4 and CCL-5 in a time- and dose-dependent manner. Additionally, we constructed ompP2ΔLoop7 and ompP2ΔLoop8 mutant SC096 strains and extracted their native OmpP2 proteins to stimulate PAMs. These mutant proteins induced significantly less mRNA expression of inflammatory cytokines than SC096 OmpP2. Next, the amino acid sequences of L7 and L8 from 15 serovars of H. parasuis OmpP2 were aligned. These sequences were relatively conserved among the most virulent reference strains, suggesting that L7 and L8 are the most active Peptides of the OmpP2 protein. Furthermore, L7 and L8 significantly upregulated the NF-κB and AP-1 activity levels based on luciferase reporter assays in a dose-dependent manner. Therefore, our results demonstrated that both surface-exposed loops L7 and L8 of H. parasuis OmpP2 induced the expression of proinflammatory cytokines possibly by activating the NF-κB and MAPK signalling pathways in cells infected by H. parasuis.

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