Poly (Lactic- co-Glycolic Acid) Nanoparticles and Nanoliposomes for Protein Delivery in Targeted Therapy: A Comparative In Vitro Study

Abstract

Over the previous years, the design, development, and potential application of nanocarriers in the medical field have been intensively studied for their ability to preserve drug properties, especially their pharmacological activity, and to improve their bioavailability. This work is a comparative study between two different types of nanocarriers, poly (lactic-co-glycolic acid)-based nanoparticles and phosphatidylcholine-based nanoliposomes, both prepared for the encapsulation of bovine serum albumin as a model protein. Polymeric nanoparticles were produced using the double emulsion water-oil-water evaporation method, whereas nanoliposomes were obtained by the thin-film hydration method. Both nanocarriers were characterized by morphological analysis, particle mean size, particle size distribution, and protein entrapment efficiency. Invitro release studies were performed for 12 days at 37 °C. In order to explore a possible application of these nanocarriers for a targeted therapy in the cardiovascular field, hemolytic activity and biocompatibility, in terms of cell viability, were performed by using human red blood cells and EA.hy926 human endothelial cell line, respectively.

Keywords: biocompatibility; cardiovascular diseases; hemolysis; lipid-based nanosystem; nanocarriers; phosphatidylcholine; poly (lactic-co-glycolic acid); polymer-based nanosystem; protein drug delivery.

Figure 1

Representative particle size distribution of (A) PNPs and (B) NLPs. PNPs: polymeric nanoparticles, NLPs: nanoliposomes.

Figure 2

Representative SEM images of (A) PNP_E and (B) NLP_E. SEM: scanning electron microscopy, PNP_E: empty polymeric nanoparticles, NLP_E: empty nanoliposomes.

Figure 3

Release profile of BSA from (A) PNPs: PNP_{BSA (30 mg/mL)}, PNP_{BSA (35 mg/mL)}, PNP_{BSA (40 mg/mL)} and from (B) NLPs: NLP_{BSA (3 mg/mL)}, NLP_{BSA (4 mg/mL)}, NLP_{BSA (6 mg/mL)}. Data are expressed as mean of three measurements. Error bars indicate SD. BSA: bovine serum albumin, PNPs: polymeric nanoparticles, NLPs: nanoliposomes.

Figure 4

Hemolysis percentage of RBCs after contact with different concentrations of (A) PNPs: PNP_E, PNP_{BSA (30 mg/mL)}, PNP_{BSA (35 mg/mL)}, PNP_{BSA (40 mg/mL)} and of (B) NLPs: NLP_E, NLP_{BSA (3 mg/mL)}, NLP_{BSA (4 mg/mL)}, NLP_{BSA (6 mg/mL)}. RBCs: red blood cells, PNP_E: empty polymeric nanoparticles, PNP_BSA: BSA-loaded polymeric nanoparticles, NLP_E: empty nanoliposomes, NLP_BSA: BSA-loaded nanoliposomes. Data are expressed as mean of three measurements. Error bars indicate SD. Different symbols refer to statistically significant differences among results (p < 0.05, ANOVA with Tukey’s HSD post hoc multiple comparison test). ^a: statistically different to empty nanocarriers (PNP_E or NLP_E), ^aa: statistically different to PNP_{BSA (30 mg/mL)} or NLP_{BSA (3 mg/mL)}, ^aaa: statistically different to PNP_{BSA (35 mg/mL)} or NLP_{BSA (4 mg/mL)}. The dotted line refers to 5% of hemolysis.

Figure 4

Hemolysis percentage of RBCs after contact with different concentrations of (A) PNPs: PNP_E, PNP_{BSA (30 mg/mL)}, PNP_{BSA (35 mg/mL)}, PNP_{BSA (40 mg/mL)} and of (B) NLPs: NLP_E, NLP_{BSA (3 mg/mL)}, NLP_{BSA (4 mg/mL)}, NLP_{BSA (6 mg/mL)}. RBCs: red blood cells, PNP_E: empty polymeric nanoparticles, PNP_BSA: BSA-loaded polymeric nanoparticles, NLP_E: empty nanoliposomes, NLP_BSA: BSA-loaded nanoliposomes. Data are expressed as mean of three measurements. Error bars indicate SD. Different symbols refer to statistically significant differences among results (p < 0.05, ANOVA with Tukey’s HSD post hoc multiple comparison test). ^a: statistically different to empty nanocarriers (PNP_E or NLP_E), ^aa: statistically different to PNP_{BSA (30 mg/mL)} or NLP_{BSA (3 mg/mL)}, ^aaa: statistically different to PNP_{BSA (35 mg/mL)} or NLP_{BSA (4 mg/mL)}. The dotted line refers to 5% of hemolysis.

Figure 5

Cell viability of PNPs and NLPs by MTS assay after (A,B) 24, (C,D) 48, and (E,F) 72 h. control (without particles), PNP_E (A,C,E), PNP_{BSA (40mg/mL)} (A,C,E), NLP_E (B,D,F), NLP_{BSA (3 mg/mL)} (B,D,F). PNP_E: empty polymeric nanoparticles, PNP_BSA: BSA-loaded polymeric nanoparticles, NLP_E: empty nanoliposomes, NLP_BSA: BSA-loaded nanoliposomes. Data are expressed as mean of three measurements. Error bars indicate SD. Different symbols refer to statistically significant differences among results (p < 0.05, ANOVA with Tukey’s HSD post hoc multiple comparison test). ^a: statistically different to control, ^aa: statistically different to empty carrier.