Chitin, a homopolymer consisting of 2-1,4-linked repeating units of N-acetylglucosamine, is the second most abundant polysaccharide in nature. It is an integral component of fungal cell walls, insect and crustacean exoskeletons and cephalopod beaks. Chitosan is chitin that has been deacetylated >50%. Human contact with chitin and chitosan is very common due to the ubiquitous presence of fungi in the environment and the ingestion of chitin/chitosan in food. Chitin and/or chitosan are used in cosmetics and biomaterials, have been postulated to play a role in asthma, and have been proposed as vaccine adjuvants. Despite its potential importance, the immunological consequences of exposure to chitin and chitosan are poorly understand and controversial. This is due, in large part, to the heterogeneous mix of glucosamine and N-acetylglucosamine in the polymers, their relative insolubility and frequent contamination of available preparations with other immune modulator substances. This application addresses the central hypothesis that the immunological effects of chitin and chitosan varies as a function of its physicochemical properties (e.g., size, acetylation). There are two interconnected specific aims.
Aim 1 is """"""""Isolation and characterization of ultrapure chitin and chitosan for use in immunological assays"""""""". In this aim, ultrapure preparations of chitin and chitosan will be made of varying sizes, shapes, and degree of acetylation. These preparations will be utilized in the studies proposed in Aim 2, which is to determine the """"""""Mechanistic basis and biological implications for differential activation of the inflammatory response by chitin and chitosan"""""""". A focus in this aim will be on the requirements for activation of the inflammasome (multi-protein caspase-1-activating platforms that control maturation and secretion of the potent proinflammatory cytokines IL-2 and IL-18) and other proinflammatory cytokines. In preliminary studies, it was found that chitosan, but not chitin, potently stimulates the inflammasome. Yet, both glycans stimulate other proinflammatory cytokines, such as TNF1 which are not dependent upon inflammasome assembly. Using the reagents generated as part of Aim 1, in Aim 2, we will determine the physicochemical requirements for stimulation of the inflammasome and other cytokine responses. The capacity of chitin and chitosan to synergize with other components of the fungal cell wall will be studied. Finally, in vivo correlates of the in vitro observations will be sought. Completion of these studies will begin to define, in a systematic manner, the properties of chitin and chitosan that lead to activation of immune responses. The results have potential impact on many research fields, including: i) the role of cell wall chitin and chitosan in innate and adaptive immune responses to fungi, ii) the use of chitosan as an adjuvant, iii) elucidating the role of chitin in asthma, iv) smarter engineering of chitin- containing biomaterials, and v) mechanisms of inflammasome activation.
Human exposure to chitin and the closely related polysaccharide chitosan is ubiquitous but the immunological consequences of such contact are poorly understood and controversial. In this proposal, highly purified chitin and chitosan particles will be manufactured and then tested for their capacity to stimulate selected inflammatory responses in vitro and in vivo. These studies will begin to define, in a systematic manner, the properties of chitin and chitosan that lead to activation of immune responses.
| Wiesner, Darin L; Specht, Charles A; Lee, Chrono K et al. (2015) Chitin recognition via chitotriosidase promotes pathologic type-2 helper T cell responses to cryptococcal infection. PLoS Pathog 11:e1004701 |
| Bueter, Chelsea L; Lee, Chrono K; Wang, Jennifer P et al. (2014) Spectrum and mechanisms of inflammasome activation by chitosan. J Immunol 192:5943-51 |
| Huang, Haibin; Ostroff, Gary R; Lee, Chrono K et al. (2013) Characterization and optimization of the glucan particle-based vaccine platform. Clin Vaccine Immunol 20:1585-91 |
| Bueter, Chelsea L; Specht, Charles A; Levitz, Stuart M (2013) Innate sensing of chitin and chitosan. PLoS Pathog 9:e1003080 |
| Huang, Haibin; Ostroff, Gary R; Lee, Chrono K et al. (2012) Relative contributions of dectin-1 and complement to immune responses to particulate ýý-glucans. J Immunol 189:312-7 |
| Brown, Gordon D; Denning, David W; Gow, Neil A R et al. (2012) Hidden killers: human fungal infections. Sci Transl Med 4:165rv13 |
| Levitz, Stuart M; Golenbock, Douglas T (2012) Beyond empiricism: informing vaccine development through innate immunity research. Cell 148:1284-92 |
| Bueter, Chelsea L; Lee, Chrono K; Rathinam, Vijay A K et al. (2011) Chitosan but not chitin activates the inflammasome by a mechanism dependent upon phagocytosis. J Biol Chem 286:35447-55 |
| Mansour, Michael K; Vyas, Jatin M; Levitz, Stuart M (2011) Dynamic virulence: real-time assessment of intracellular pathogenesis links Cryptococcus neoformans phenotype with clinical outcome. MBio 2: |
