Cellular Hemoglobin-Based Oxygen Carriers as Potential Artificial Blood Substitutes

The objective of this study was to develop an effective and reliable cellular hemoglobin-based blood substitute (HBOC). Cellular HBOCs were created by encapsulating bovine hemoglobin (Hb) into the aqueous cores of liposomes, poly(ethylene glycol) (PEG) conjugated liposomes and polymersomes. In order to evaluate the potential of the dispersions as cellular HBOCs, the following physical properties were measured: vesicle size distribution, Hb encapsulation efficiency, oxygen binding property (as indicated by P50 and cooperativity coefficient), and encapsulated methemoglobin level.

The oxygen binding properties of unmodified liposomes encapsulated Hb (ULEHs), PEGylated LEHs (PEG-LEHs) and polymersomes encapsulated Hb (PEHs) were comparable to human RBCs, indicating that these vesicles displayed good potential as cellular HBOCs. The physical integrity of ULEH dispersions in phosphate buffered saline at physiological pH and temperature was unstable post-production, due to osmosis of water into and out of the liposome core, which implies that ULEHs will be osmotically fragile in the blood stream. This potential problem was solved by grafting PEG molecules onto the liposome surface, thereby strengthening the liposome bilayers. However, the Hb encapsulation efficiency of PEG-LEH dispersions was low. The intravascular circulation, biocompatibility and colloidal state of PEG-LEH dispersions were limited due to the limited PEG surface coverage and molecular weight that can be stably conjugated onto the liposome surface.

In contrast, PEH dispersions possessed higher Hb encapsulation efficiencies compared to ULEH, PEG-LEH, PEGylated actin-containing LEH, lipogel particle and nanoscale hydrogel particle dispersions loaded with Hb. Unlike LEH dispersions, encapsulation of Hb into polymersomes did not enhance Hb oxidation. Polymersomes possessed superior PEG shielding ability compared to PEG-liposomes, due to 100% PEG surface coverage with longer PEG brushes. Since polymersomes can be synthesized with thicker hydrophobic membranes compared to liposomes, polymersomes are mechanically stronger than liposomes. Simulation of in-vivo oxygen transport in a capillary and to surrounding tissues, demonstrated that PEH dispersions could be engineered for applications from routine surgery to treatment of trauma. We concluded that PEH dispersions were readily prepared and exhibited good potential as a cellular HBOC, while offering superior physical properties, which may alleviate the limitations encountered with current designs of cellular HBOCs.