Date Published: January 28, 2019
Publisher: Public Library of Science
Author(s): Shanjana Awasthi, Bhupinder Singh, Vijay Ramani, Jun Xie, Stanley Kosanke, Samithamby Jeyaseelan.
Interaction between surfactant protein-A (SP-A) and toll-like receptor (TLR)4 plays a critical role in host defense. In this work, we studied the host defense function of SPA4 peptide (amino acids GDFRYSDGTPVNYTNWYRGE), derived from the TLR4-interacting region of SP-A, against Pseudomonas aeruginosa. We determined the binding of SPA4 peptide to live bacteria, and its direct antibacterial activity against P. aeruginosa. Pro-phagocytic and anti-inflammatory effects were investigated in JAWS II dendritic cells and primary alveolar macrophages. The biological relevance of SPA4 peptide was evaluated in a mouse model of acute lung infection induced by intratracheal challenge with P. aeruginosa. Our results demonstrate that the SPA4 peptide does not interact with or kill P. aeruginosa when cultured outside the host. The SPA4 peptide treatment induces the uptake and localization of bacteria in the phagolysosomes of immune cells. At the same time, the secreted amounts of TNF-α are significantly reduced in cell-free supernatants of SPA4 peptide-treated cells. In cells overexpressing TLR4, the TLR4-induced phagocytic response is maintained, but the levels of TLR4-stimulated TNF-α are reduced. Furthermore, our results demonstrate that the therapeutic administration of SPA4 peptide reduces bacterial burden, inflammatory cytokines and chemokines, intracellular signaling, and lactate levels, and alleviates lung edema and tissue damage in P. aeruginosa-infected mice. Together, our results suggest that the treatment with SPA4 peptide can help control the bacterial burden, inflammation, and tissue injury in a P. aeruginosa lung infection model.
Antimicrobial resistance and the acquisition of new virulence traits have contributed to a worldwide increase in the incidence of infections and associated morbidity and mortality.[1–3] Therefore, new therapeutic approaches are urgently needed to control difficult-to-treat infections. One way of addressing this need is to harness natural immune defenses of the host to develop therapeutic entities.
The amino acid sequence of the SPA4 peptide (GDFRYSDGTPVNYTNWYRGE) is derived from the C-terminal TLR4-interacting region of human SP-A. We solved the structure of the SPA4 peptide by nuclear magnetic resonance spectroscopy, and studied the physicochemical features, which demonstrated its structural adaptability for binding to TLR4.
Using multidisciplinary approaches, we identified the SPA4 peptide as the lead peptide from TLR4-interacting regions of SP-A.[18, 20] Our results demonstrated that the SPA4 peptide binds to TLR4 and reduces LPS-MYD88-NF-κB activity, TLR4-priming of the inflammasome, and the inflammatory cytokine response.[18, 20, 22] In this report, we investigated the effect of SPA4 peptide on phagocytosis and clearance of P. aeruginosa and the associated inflammatory response in immune cells, and bacterial burden and inflammatory and lung injury parameters in a mouse model of P. aeruginosa-induced lung infection. All parameters were analyzed on a short-term basis specifically to delineate the effect of SPA4 peptide on early events of TLR4 signaling, which may be critical in orchestrating the long-term immunity, local homeostasis, and biological effect.