Porcine Beta-Defensin 2 Provides Protection Against Bacterial Infection by a Direct Bactericidal Activity and Alleviates Inflammation via Interference With the TLR4/NF-κB Pathway

Abstract

Porcine beta-defensin 2 (PBD-2) which is a member of the family of antimicrobial peptides, is widely expressed in pig organs with a broad spectrum of bactericidal activities confirmed in vitro . We previously demonstrated that transgenic (TG) pigs overexpressing PBD-2 could resist the infection by the porcine pathogen Actinobacillus pleuropneumoniae . In this study, the roles of PBD-2 in protecting against bacterial infection were further investigated. The biochemical indexes of the blood sample, body weights, histological morphologies, and weights of the organs of TG mice expressing PBD-2 were measured. Results confirmed that these mice showed normal physiological features. An assay of Salmonella Typhimurium infection was conducted on wild-type (WT) and TG mice. The TG mice possessed higher survival rate, less body weight loss, and pathological changes and smaller recovery rates of bacteria after infection with S . Typhimurium. The in vitro synthetic PBD-2 and the serum and tissue homogenates from the TG mice displayed a direct bactericidal activity. Moreover, PBD-2 could inhibit the release of the proinflammatory cytokines, including IL-6, TNF-α, IL-1β, and IL-12, in the TG mice infected with S . Typhimurium or treated with lipopolysaccharide (LPS). The WT mice treated with PBD-2 and S . Typhimurium or LPS showed reduced levels of proinflammatory cytokines. The mouse macrophage cell line RAW 264.7 which expressed PBD-2 was constructed to detect the signal pathways affected by PBD-2. The suppressing effect of PBD-2 on the release of the proinflammatory cytokines was confirmed using RAW 264.7 either expressing PBD-2 or supplemented with PBD-2. The promoter activity and mRNA level of NF-κB were detected, and PBD-2 was shown to significantly inhibit the activation of the NF-κB pathway induced by LPS. The direct interaction of PBD-2 with TLR4 was revealed by isothermal titration calorimetry and far-Western blot in vitro and the coimmunoprecipitation of PBD-2 with TLR4 on RAW 264.7 cells. This interaction indicates one reason for the interference of NF-κB activation. Overall, this study showed that PBD-2 protected against bacterial infection through a direct bactericidal activity and alleviated inflammation by interfering with the TLR4/NF-κB pathway.

Porcine β-defensin 2 inhibits proliferation of pseudorabies virus in vitro and in transgenic mice

“…Chen et al argued that higher expression levels of PBD-2 and other β-defensins in Meishan pigs than those in crossbred pigs conferred stronger immunity of Meishan pigs against diseases [36]. Our previous study revealed that PBD-2 overexpressing pigs and PBD-2 TG mice showed enhanced resistance to bacterial infection [10,13]. In this study, we further verified that PBD-2 TG mice displayed increased resistance to viral infection.…”

“…Although swine serves as the natural host and reservoir of PRV, mice are often utilized as a preferred model organism to study PRV infection because of their susceptibility to PRV. In this study, the effects of PBD-2 on the proliferation of PRV were investigated both in PK-15 cells and transgenic (TG) mice generated in our previous study [13]. The results confirmed the effectiveness of PBD-2 to suppress virus proliferation and provided novel ideas to control PRV infection.…”

“…Mice grouping and viability test of PRV infection C57/BL TG mice expressing PBD-2 were generated by our lab previously [13]. Genomic DNA from mouse tail samples was extracted and used for identification of TG mice expressing PBD-2 by using primers NP03 (5′-GCTGGTTGTTGTGCTGTCTC-3′) and NP04 (5′-AGGTCCCTTCAATCCTGTTG-3′).…”

“…PBD-2 also exhibited immunomodulatory properties [11,12]. Our recent research demonstrated that PBD-2 could alleviate inflammation through interacting with toll-like receptor 4 and suppressing the downstream NF-κB signaling pathway [13]. Additionally, it has been found that PBD-2 can attenuate proliferation of porcine reproductive and respiratory syndrome virus (PRRSV) in MA-104 cells [8].…”

Knockdown of TRIM52 alleviates LPS-induced inflammatory injury in human periodontal ligament cells through the TLR4/NF-κB pathway

“…TLR4, a transmembrane receptor, is responsible for the activation of a wide range of signal pathways involved in the bactericidal response [26]. Accumulating evidence has shown that the NF-κB signaling pathway serves as a crucial role in the progression of periodontitis [27].…”