Poster abstracts

Poster number 75 submitted by Shahid Rameez

Liposome Encapsulated Hemoglobins as Red Blood Cell Substitutes.

Shahid Rameez (Chemical and Biomolecular Engineering), Andre F Palmer (Chemical and Biomolecular Engineering)

Abstract:
The limited supply of donor blood and its short shelf-life, coupled with the risks of infection and allergic reactions of donated blood, have led to the development of artificial red blood cell (RBC) substitutes (1, 2). Since hemoglobin (Hb) is responsible for oxygen (O2) storage and transport in the RBC, it is logical to use Hb for synthesis and formulation of RBC substitutes. Clinical trials of acellular RBC substitutes were largely unsuccessful due to the small size of these protein molecules, which allowed them to extravasate through pores in the blood vessel wall and scavenge endothelial-derived nitric oxide (NO), which resulted in vasoconstriction (reduction in blood vessel diameter), systemic hypertension (increase in mean arterial blood pressure) and oxidative tissue toxicity (3, 4). These side-effects can be attenuated, or even eliminated, by encapsulating Hb inside a particle that is larger than the size of pores lining the inner surface of blood vessels (5, 6).
During the last few decades, liposome encapsulated hemoglobin (LEH) dispersions have been investigated for use as cellular RBC substitutes. However, the process for formulating LEHs is cumbersome and the lipid mixture is often complex. This work investigates a simple approach for formulating LEHs from a simple lipid mixture composed of the high phase transition lipid distearoylphosphatidylcholine and cholesterol. In order to improve the circulation half-life and colloidal state of LEHs, the outer membrane of unmodified LEHs was conjugated with polyethylene glycol (PEG). PEG masks the LEH particle from being recognized by the immune system, hence increasing the particle circulation half-life (6). The results of this work show that PEG-LEH dispersions exhibited average diameters ranging from 166-195 nm that were colloidally stable for 4-5 months with internal Hb concentrations ranging from 50-75 g/dL.
We also measured the reactions of PEG-LEHs with physiologically important ligands such as O2, carbon monoxide (CO) and NO. PEG-LEHs and RBCs exhibited retarded gaseous ligand binding/release kinetics compared to acellular Hbs. This result provides important insight into the pivotal role the cellular membrane and intracellular diffusion barrier play in the transport of gases into and out of these structures.

References:
(1) E. Moore. J. Am. Coll. Surgeons. 196 (1):1-17, 2003.
(2) D. Zabarenko. www.abcnews.go.com, 2010.
(3) Poli De Figueiredo, M. Mathru, D. Solanki, V. W. Macdonald. G. C. Kramer and R. N. Garrison. Journal of Trauma-Injury Infection & Critical Care 42, 847-856, 1997
(4) H. Sakai, H. Hara, M. Yuasa, A. G. Tsai, S. Takeoka, E. Tsuchida, and M. Intaglietta. Am J Physiol Heart Circ Physiol 279, H908-915, 2000.
(5) H. Sakai, A. Sato, K. Masuda, S. Takeoka and E. Tsuchida. J. Biochem 283, 1508-1517, 2008.
(6) Sakai, H., Takeoka, S., Park, S. I., Kose, T., Nishide, H., Izumi, Y., Yoshizu, A., Kobayashi, K., and Tsuchida, E. Bioconjug Chem 8, 23-30, 1997.

Keywords: Liposome, RBC, vasoconstriction