Activation of toll-like receptor signaling pathway for protection against influenza virus infection

Abstract

This study aims to evaluate the antiviral role of nucleic acid-based agonists for the activation of toll-like receptor (TLR) signaling pathways, and its protective role in respiratory influenza A virus infections. TLR-3 is expressed on myeloid dendritic cells, respiratory epithelium, and macrophages, and appears to play a central role in mediating both the antiviral and inflammatory responses of the innate immunity in combating viral infections. Influenza viruses can effectively inhibit the host's ability to produce interferons, and thereby suppress the immune system's antiviral defence mechanisms. Poly ICLC is a synthetic double stranded RNA comprising of polyriboinosinic-poly ribocytidylic acid (Poly IC) stabilized with l-lysine (L) and carboxymethylcellulose (C). Poly ICLC and liposome-encapsulated Poly ICLC (LE Poly ICLC) are TLR-3 agonists and are potent inducer of interferons and natural killer cells. Intranasal pre-treatment of mice with Poly ICLC and LE Poly ICLC provided high level of protection against lethal challenge with a highly lethal avian H5N1 influenza (HPAI) strain (A/H5N1/chicken/Henan clade 2), and against lethal seasonal influenza A/PR/8/34 [H1N1] and A/Aichi/2 [H3N2] virus strains. The duration of protective antiviral immunity to multiple lethal doses of influenza virus A/PR/8/34 virus had been previously found to persist for up to 3 weeks in mice for LE Poly ICLC and 2 weeks for Poly ICLC. Similarly, pre-treatment of mice with CpG oligonucleotides (TLR-9 agonist) was also found to provide complete protection against influenza A/PR/8/34 infection in mice. RT-PCR analysis of lung tissues of mice treated with Poly ICLC and LE Poly ICLC revealed upregulation of TLR-3 mRNAs gene expression. Taken together, these results do support the potential role of TLR-3 and TLR-9 agonists such as Poly ICLC and LE Poly ICLC in protection against lethal seasonal and HPAI virus infection.

Fig. 1

TLR-3 mRNA expression in the lungs of mice which received two intranasal administered doses of Poly ICLC, LE Poly ICLC (1 mg/kg body weight), sham liposomes and PBS were analyzed using qRT-PCR as described in Section 2. Results were normalized to β-actin as endogenous control, and are shown as fold increases relative to the PBS control group. Error bars represent standard error of the mean.

Fig. 2

Efficacy of CpG ODN prophylaxis against influenza viral challenge. Mice were intranasally given 5 μg (0.25 mg/kg body weight) of free or liposome-encapsulated CpG ODN (Lip-CPG ODN) 4 days prior to infection with influenza A/PR/8/34. Liposomes were composed of dimyristoltrimethylammonium propane/cholesterol/dioleylphosphatidyl choline (molar ratio of 25:50:25). Mice (n = 5) were monitored daily for appearance, weight and survival. Graph shows the percentage of mice surviving 14 days after challenge.