Journal of Medical Virology 75:130–136 (2005) Protection Against Influenza Virus Infection by Intranasal Administration of Hemagglutinin Vaccine With Chitin Microparticles as an Adjuvant Hideki Hasegawa, 1 * Takeshi Ichinohe, 1,2 Peter Strong, 3 Izumi Watanabe, 1,2 Satoshi Ito, 1,2 Shin-ichi Tamura, 4 Hidehiro Takahashi, 1 Hirofumi Sawa, 5 Joe Chiba, 2 Takeshi Kurata, 1 and Tetsutaro Sata 1 1 Department of Pathology, National Institute of Infectious Diseases, Musashimurayama-shi, Tokyo, Japan 2 Department of Biological Science and Technology, Tokyo University of Science, Yamazaki, Noda, Chiba, Japan 3 Medical Research Council Immunochemistry Unit, University of Oxford, Oxford, United Kingdom 4 Laboratory of Prevention of Viral Diseases, Research Institute for Microbial Diseases, Osaka University, Yamadaoka, Osaka, Japan 5 Laboratory of Molecular and Cellular Pathology, 21st Century COE Program for Zoonosis Control, Hokkaido University School of Medicine, and CREST, JST, Sapporo, Japan Chitin in the form of microparticles (chitin micro- particles, CMP) has been demonstrated to be a potent stimulator of macrophages, promoting T-helper-1 (Th1) activation and cytokine response. In order to examine the mucosal adjuvant effect of CMP co-administered with influenza hemag- glutinin (HA) vaccine against influenza infection, CMP were intranasally co-administered with in- fluenza HA vaccine prepared from PR8 (H1N1) virus. Inoculation of the vaccine with CMP induc- ed primary and secondary anti-HA IgA responses in the nasal wash and anti-HA IgG responses in the serum, which were significantly higher than those of nasal vaccination without CMP, and provided a complete protection against a homo- logous influenza virus challenge in the nasal in- fection influenza model. In addition, CMP-based immunization using A/Yamagata (H1N1) and A/ Guizhou (H3N2) induced PR8 HA-reactive IgA in the nasal washes and specific-IgG in the serum. The immunization with A/Yamagata and CMP resulted in complete protection against a PR8 (H1N1) challenge in A/Yamagata (H1N1)-vac- cinated mice, while that with A/Guizhou (H3N2) and CMP exhibited a 100-fold reduction of nasal virus titer, demonstrating the cross-protective effect of CMP and influenza vaccine. It is sug- gested that CMP provide a safe and effective adjuvant for nasal vaccination with inactivated influenza vaccine. J. Med. Virol. 75:130–136, 2005. ß 2005 Wiley-Liss, Inc. KEY WORDS: influenza; chitin microparticles; nasal vaccine; adjuvant; IgA INTRODUCTION Effectiveness and safety are important issues to be considered in the development of a vaccine. The mucosal immune system is usually the first immunological barrier against influenza virus infections [Mestecky and McGhee, 1987]. The respiratory tract mucosa is the primary site of infection and the immunological com- partment where the host immune system attacks the influenza virus. Secretory IgA antibodies are major effectors providing a front-line defense against influenza viruses in the respiratory tract mucosa [Shvartsman and Zykov, 1976; Underdown and Schiff, 1986; Murphy, 1994]. The influenza virus causes annual epidemics of influenza, largely due to the selection of new variants with mutations in the surface hemagglutinin (HA). The surface HA determines the antigenic properties of the virus and combines with sialic acid residues on epithe- lial cells during cell attachment [Renegar and Small, 1994]. Inactivated vaccines against the influenza virus are administered parenterally to induce serum anti-HA IgG antibodies that are highly protective against homo- logous virus infection, but are less effective against heterologous virus infection [Renegar and Small, 1994; Murphy and Webster, 1996]. In contrast, a large number of studies have shown that the mucosal immunity *Correspondence to: Hideki Hasegawa, Department of Pathol- ogy, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama-shi, Tokyo 208-0011, Japan. E-mail: hasegawa@nih.go.jp Accepted 9 August 2004 DOI 10.1002/jmv.20247 Published online in Wiley InterScience (www.interscience.wiley.com) ß 2005 WILEY-LISS, INC.