Some studies have proved it is effective to administer peptide coupled to a potent carrier for eliciting an immune response [5] and [6]. A disadvantage of some carrier molecules is their relatively low immunogenicity and the need for potent adjuvants such as CFA to stimulate the immune response non-specifically. Certain carrier proteins such as the mycobacterial heat shock protein (HSP) 65 also have adjuvant-like properties

and may be used efficiently MLN8237 nmr as carriers in an adjuvant-free system [7] and [8]. Epitope analysis has shown that HSP65 proteins have numerous B and T cell epitopes and the HSP65 from Mycobacterium tuberculosis can evoke a strong T-cell dependent immune response without the need for external adjuvant when used as a carrier molecule coupled to a peptide antigen [9]. We have used HSP65 as carrier to develop anti-cancer vaccine [10], [11], [12] and [13] and anti-atherosclerosis vaccine [14] and [15]. However, PF-06463922 order HSP65 never serve as a carrier for P277-based vaccines. Mucosal administration of autoantigen HSP65 decrease organ-specific inflammation has been tested experimentally in several models of autoimmunity, such as atherogenesis and arthritis [16] and [17]. HSP65 and peptide P277 are all identified as an ideal target antigen to develop type 1 diabetes vaccines [18]. We are interested

in whether the HSP65 serves as an immunogenic carrier for peptide P277 will induce anti-inflammatory immune response in NOD mice by mucosal administration.

Carnitine palmitoyltransferase II It is conceivable that the dual functions of anti-type 1 diabetes of HSP65 and P277 will be obtained. Therefore, an immunotherapy based on the mucosal administration of an adjuvant-free fusion protein comprising Mycobacterium bovis BCG heat shock protein 65 linked to P277 has been developed [19]. The results reported here indicate that prevention of diabetes was associated with a decrease in the degree of insulitis and with down-regulation of spontaneous proliferative T cell responses to the fusion protein HSP65-6 × P277, and the pattern of cytokine secretion to HSP65-6 × P277, showed an increase in IL-10 and a decrease in IFN-γ secretion, compatible with a shift from a Th1-like toward a Th2-like autoimmune response. We conclude that HSP65 may serve as a particularly advantageous carrier for P277-based vaccines and mucosal administration may be a therapeutic approach for treatment of type 1 diabetes. The fusion protein HSP65-6 × P277 and HSP65 were prepared as described [20]. The peptide P277 (VLGGGCALLRCIPALDSLTPANED) was synthesized at the GL Biochem (Shanghai) Ltd. Purified recombinant human VEGF-P277 was gift from Dr. Zhu ai-hua, Key Laboratory of Biotechnology for Medicinal Plants of Jiangsu Province, Xuzhou Normal University, People’s Republic of China. Rabbit anti-mouse IgG horseradish peroxidase (HRP)-conjugated antibody was purchased from Promega, USA.