Chitosan alters inactivated respiratory syncytial virus vaccine elicited immune responses without affecting lung histopathology in mice
Introduction
Chitosan is a generic term for a family of linear polysaccharides that is commercially available as a partially deacetylated α-chitin produced from the exoskeletons of crustacean or the cell walls of fungi [1], [2], [3]. It is a biocompatible, non-toxic and non-allergenic material that has been tested for safety and toxicity in a variety of animal species for various applications through various routes of administration [4], [5], [6]. In solution, the positive charge of chitosan confers it with mucoadhesive properties, which are crucial for its use in intranasal applications [7], [8]. When nasally administered with an antigen, it has been shown to augment antigen-specific immune responses in many animal models [9], [10], [11], [12] and in human subjects [9], [13], [14], [15], [16]. Interestingly, chitosan alone is also able to enhance the immune responses against some viral infections [10], [11], [12]. While its activities of immune enhancement have been well documented, the molecular mechanisms remain to be fully understood. It would be of interest to investigate whether chitosan could alter immune responses induced by a vaccine known to be generally less effective or even induce severe adverse reactions.
Formaldehyde-inactivated respiratory syncytial virus (FIRSV) vaccine was initially developed to protect humans against RSV infection known to cause severe disease in young children, elderly and immunocompromised patients [17], [18], [19], [20]. However, instead of protection, the vaccine was found to be associated with severe vaccine-induced enhanced respiratory disease (ERD), with 80% of the participants hospitalized and 2 deaths following subsequent RSV infection [21], [22], [23], [24]. It has been shown that Th2-skewed immune responses and poorly neutralizing antibodies lead to pulmonary inflammation, airway obstruction, and mucus hypersecretion [25], [26], [27], [28], [29]. Moreover, cell-mediated immune responses could also be involved in the development of ERD [30], [31], [32]. Here, we employed chitosan to gain better insight into FIRSV-induced immune responses, specifically its role in inducing regulatory and tissue resident T cells which are known to be critically important in inducing well-balanced and robust immune responses against microbial infections.
Section snippets
Cells, virus and vaccines
Hep-2 (ATCC: CCL-23) were grown in Dulbecco's Modified Eagle Medium (DMEM) supplemented with 1.5 g/l sodium bicarbonate, 2 mM Glutamax, 1 mM HEPES, 20 U/ml Penicillin, 0.02 mg/ml Streptomycin, and 10% FBS.
RSV-A2 (ATCC: VR-1540) was grown in Hep-2 cells according to supplier’s instructions and sucrose-purified for animal studies. FIRSV was prepared with the RSV-A2 strain in Hep-2 cells as described elsewhere [25], [27]. Briefly, Hep-2 cells were infected at a MOI of 0.02 in Hep-2 growth media.
Chitosan enhances induction of resident effector T cells and regulatory T cells following RSV infection
We first investigated whether chitosan administered therapeutically could afford protection against RSV and the underlying mechanism. To that end, Balb/c mice were infected with RSV-A2 and intranasally inoculated with chitosan either one day or three days post-infection [10]. A ‘no chitosan’ control was included where mice were infected with RSV but did not receive any chitosan. Four days post-infection, all mice were injected with a fluorophore-conjugated anti-mouse CD3 antibody [33], [34] and
Discussion
Because of its proven safety record and a variety of immunomodulatory properties, chitosan has been extensively studied for its potential as a vaccine adjuvant. Not only does it have a profound effect on humoral immunity [9], [44], it also affects cell-mediated immunity [49]. Several lines of evidence prompted us to conduct the current study in which FIRSV vaccine was used as a model vaccine to study the mechanisms of chitosan as a therapeutic. First, although the role of chitosan in
Funding statement
This work was funded by Genomics Research and Development Initiative Grant, Government of Canada.
Contributors
AM, XL, SS, ZC, CL, WC, TC, MRM and LW conceived the overall study. AM, MSR, LL and CG performed the experiments and analyzed the data. AM and XL wrote the manuscript. All authors edited and approved the manuscript.
Declaration of Competing Interest
The authors declare that they have no conflict of interests.
Acknowledgements
We thank Bozena Jaentschke, Sathya Raman and Emily Dupuis for technical assistance. We are so grateful to Dr. Martha Navarro and all technicians in the animal facility for help. Thank you to Dr. Don Caldwell, veterinary pathologist at Health Canada, for his expert analysis of the lung tissues. Drs Roger Tam and Michael Johnston are acknowledged for their critical review of the manuscript.
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