Mimetic Membrane System to Carry Multiple Antigenic Proteins from Leishmania amazonensis

Abstract

Liposomes have long been used as models for lipid membranes and for the reconstitution of a single or multiple proteins. Also, liposomes have adjuvant activity in vaccines against several protozoan or bacterial organisms. Thus, the main objective of the present study was to obtain a crude extract of detergent-solubilized proteins of Leishmania amazonensis amastigotes and reconstitute them into liposomes. Neutral and zwiterionic detergents were less efficient than an ionic detergent. In order to obtain efficient solubilization using only sodium dodecyl sulfate (SDS), the effects of detergent and protein concentration and incubation time were studied. The maximum of solubilized proteins was obtained instantaneously using a ratio of 0.5 mg/ml of protein to 0.1% (w/v) detergent at 4 degrees C. Dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylserine (DPPS) and cholesterol in a weight ratio of 5:1:4 were used for protein reconstitution into liposomes using the cosolubilization method, yielding 60% of incorporation. The incorporation of multiple parasite proteins results in a vesicular diameter of proteoliposomes of about 140 nm, presenting a final lipid weight ratio for DPPC, DPPS and cholesterol of 1:1:5, with high stability. The detergent-solubilized proteins of L. amazonensis amastigotes present in the proteoliposome, when analyzed by SDS-polyacrylamide gel electrophoresis, include a wide range of parasite-incorporated proteins. BALB/c mice inoculated with these proteoliposomes were able to produce antibodies against the proteins reconstituted in DPPC:DPPS:cholesterol liposomes and were partially resistant to infection with L. amazonensis promastigotes. These results indicate that this system can be used as a possible vaccine against L. amazonensis.

Proteoliposomes in nanobiotechnology

“…Several publications have demonstrated how liposomes are being used as appropriate antigen carriers for the generation of antibody response in vivo or in vitro (Daghastanli et al 2004;Santos et al 2006a;Migliaccio et al 2008) and as effective vehicles for peptides and proteins, thus enhancing their immunogenicity in leishmaniasis infection models (Afrin et al 2002;Jaafari et al 2006;Badiee et al 2007;Santos et al 2006a;Migliaccio et al 2008). Depending on the nature of the antigen, however, the procedure for the preparation of the liposomes may vary considerably.…”

“…Reconstitution of membrane proteins into liposomes has proved a viable alternative for the preparation of antigenic components to induce immunogenicity and humoral protection in animal models (Leserman 2004). In addition to preserving the native structure of proteins, liposomes have the advantage of giving adjuvant activity in vaccines against various pathogens (Daghastanli et al 2004;Santos et al 2006a;Migliaccio et al 2008;Colhone et al 2009). …”

Liposomal systems as carriers for bioactive compounds

“…Such systems can be used as biomimetic models for studies of interaction with various hydrophobic drugs and membrane proteins, with potential applications in the field of immunology (vaccines and drugs targeting specific cell membranes). Liposomes associated with proteins can be used to study the activity of photosensitive dyes applied during photodynamic therapy (Bolfarini et al 2012;Longo et al 2012;Rocha et al 2012;Bicalho et al 2013;Faria et al 2015), as well for studies related to the ability of proteins to prevent, treat and/or act as nanosensors for various diseases (Daghastanli et al 2004;Santos et al 2006;Zucolotto et al 2007;Migliaccio et al 2008;Colhone et al 2009Colhone et al , 2015Calderon et al 2009a, b;Perinoto et al 2010;Paulovich et al 2011;Barbosa et al 2011;Salay et al 2011;Barros et al 2013;Fotoran et al 2015).…”

Use of proteoliposome as a vaccine against Trypanosoma cruzi in mice

“…The reconstitution of membrane proteins into liposomes, lipid vesicles constituted by one or more lipid layers, has been shown to be a useful tool to prepare antigenic components to induce immunity in experimental models (Kita et al, 2005;Santos et al, 2006). Liposomes have been shown to serve as appropriate antigen carriers for the generation of antibody response in vivo and effective vehicles for peptides and proteins, thus enhancing their immunogenicity (Brewer et al, 1998;Ignatius et al, 2000).…”