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  • Title: Oral immunization against hepatitis B using bile salt stabilized vesicles (bilosomes).
    Author: Shukla A, Khatri K, Gupta PN, Goyal AK, Mehta A, Vyas SP.
    Journal: J Pharm Pharm Sci; 2008; 11(1):59-66. PubMed ID: 18445364.
    Abstract:
    PURPOSE: Most of the presently available vaccines including hepatitis B vaccines administered through parenteral route, fail to induce a mucosal antibody response. Therefore, oral immunization appears to be an effective and attractive alternative to parenteral immunization. However, the problem of degradation of antigen in the harsh and hostile environment of the gastrointestinal tract consequently requires larger doses and more frequent dosing of antigen. Furthermore, much larger doses can induce antigen tolerance. Therefore the purpose of the present study was firstly to overcome these problems by the use of bile salt stabilized vesicles (bilosomes) and HBsAg as the model antigen,which could provide both protection to the antigen as well as enable transmucosal uptake and subsequent immunization. Another purpose of this study was to determine the dose that could produce serum antibody titres against hepatitis B via the oral route compared to those following intramuscular immunization. METHODS: In the present study bilosomes containing recombinant hepatitis B surface antigen were prepared by a lipid cast film method. HBsAg loaded bilosomeswere characterized in vitro for their shape, size, percent antigen entrapment and stability. Fluorescence microscopy was carried out to confirm the uptake of bilosomes by gut associated lymphoid tissues (GALT). The in vivo part of the study comprised estimation of anti-HBsAg IgG response in serum and anti-HBsAg sIgA in various body secretions using specific ELISA techniques following oral immunization with low dose loaded bilosomes (B1, 10 microg), intermediate dose loaded bilosomes (B2, 20 microg) and high dose loaded bilosomes (B3, 50 microg) in BALB/c mice. RESULTS: Fluorescence microscopy suggested that there was an increase in fluorescence intensity following the uptake of bilosomes entrapped FITC-BSA in gut associated lymphoid tissues. The high dose HBsAg bilosomes (B3, 50 microg) produced comparable anti-HBsAg IgG levels in serum to those observed in the case of intramuscular administration of alum adsorbed HBsAg (10 microg). In addition, the bilosomal preparations elicited measurable sIgA in mucosal secretions, where the highest responses were observed with high dose HBsAg bilosomes (B3,50 microg) and as expected, intramuscular administered alum adsorbed HBsAg (10 microg) failed to elicit such responses. CONCLUSIONS: HBsAg loaded bilosomes produced both systemic as well as mucosal antibody responses upon oral administration. Furthermore, bilosomes with a five times higher dose upon oral administration produced comparable serum antibody titres to those obtained after intramuscular immunization without the induction of systemic tolerance.
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