Application of polyethyleneglycol (PEG)-modified liposomes for oral vaccine: effect of lipid dose on systemic and mucosal immunity

Abstract

To examine the systemic and mucosal immunity towards a liposomal antigen in an oral vaccine, we prepared ovalbumin (OVA)-encapsulating polyethyleneglycol (PEG)-modified liposomes and unmodified ones, and orally administered two different concentrations of them to mice. Unmodified liposomes tended to induce a stronger systemic immune response than the PEG-modified ones especially at the higher concentration of liposomes. Whereas at the lower liposome concentration the mucosal immune response was stronger for the PEG-modified liposomes than for the unmodified ones but nearly the same at the higher concentration. The relative amount of immunoglobulin G (IgG) against OVA in the plasma was 1.7-fold higher for a 12.5 μmol phospholipid dose of PEG-liposomes encapsulating OVA than for a 5.0 μmol one encapsulating the same amount of OVA. On the contrary, the relative amount of IgA in the intestinal wash was 2.6-fold higher for the 5.0 μmol phospholipid dose than for the 12.5 μmol one. These results indicate that OVA encapsulated in a small number of liposomes, especially the PEG-modified ones, is favorable for inducing a mucosal immune response and that the same amount of OVA in a large number of liposomes tends to improve the systemic immune response. A possible explanation for this tendency is the differential release rate of OVA from the liposomes at the intestinal mucosa. Our present study suggests that the dose of liposomes containing antigen is an important factor for controlling the response of systemic and mucosal immune systems.

Introduction

Vaccine is one of the best preventive strategies against infectious diseases. Infusion vaccines chiefly used are not fully beneficial, since they require trained persons to administer them. In comparison, oral vaccines have the advantages of self-administration at ease, low risk of contamination, and a reduced cost. Therefore, the development of oral vaccines has become noticed. Oral immunization acts mainly in the gut-associated lymphoid tissue (GALT), forms a network with other mucosal sites such as respiratory and urogenital mucosa, and represents the first line of defense against colonization by viral and bacterial pathogens. A certain foreign substance is taken up into Peyer’s patches, which are epithelia enriched with phagocytic microfold (M) cells. Then the substance is transmited to macrophages and lymphocytes following secretion of immunoglobulin A (IgA) from antigen-specific B cells at general mucosal sites. IgA can neutralize biologically active antigens to prevent pathogens from invading the body [1], [2].

Oral immunization, however, has a problem of low bioavailability: antigens are degraded by gastric acidity and proteolytic enzymes in the intestinal lumen and therefore extremely large doses are required to achieve an adequate immune response. In recent years, the use of microparticles such as liposomes as a carrier of antigens for inducing immunity has been explored; and microparticles were found to prevent the degradation of antigens in the gut [3]. Oral immunization with antigen incorporated in microparticles was also reported to induce not only mucosal but also systemic immunity [4], [5]. Therefore, the development of optimum microparticle formulations enabling the prevention of infections by activating the systemic and mucosal immune systems has been attempted. Among the microparticles evaluated, liposomes are notable, since they can be easily controlled in terms of size, charge, membrane fluidity, etc. The change in liposomal properties such as particle size, surface charge, saturation degree of structural lipids, and lipid doses is known to affect the pharmacokinetics of the encapsulated drugs after systemic administration [6], [7]. Furthermore, particle size, surface charge and lipid composition were reported to affect liposomal uptake by Peyer’s patches [8], [9]. However, the effect of liposomal lipid doses on the induction of immunity has not yet been fully examined.

In the present study, the effect of two liposomal lipid doses on systemic and mucosal immunity following oral administration was examined by use of ovalbumin-containing unmodified or polyethyleneglycol-modified liposomes.