MP4, a Human Hemoglobin Modified with Maleimide-Peg, Reduces Sickling of Red Blood Cells in an In Vitro Model.

Human hemoglobin surface-modified with maleimide polyethylene glycol (MP4) has been shown to be effective in transporting O₂ and maintaining tissue perfusion in several animal models. MP4 appears to be free of vasoconstriction, and clinical trials have shown MP4 to be free of the side-effects commonly associated with hemoglobin-based products. Since MP4 is effective in delivering O₂ to capillaries in the hamster microcirculation, our hypothesis is that it would be of therapeutic benefit in Sickle Cell Disease. In order to test this hypothesis, we measured pressure that develops as dilute sickle cells pass through a filter with and without MP4. Cells from patients with sickling disorders were washed with phosphate-buffered saline (PBS) containing 1% bovine serum albumin (BSA) and suspended at 1% hematocrit in the presence and absence of MP4 (2 g/dL). The suspension was then pumped through a hollow fiber oxygenator (Living Systems Instrumentation, Model Number OXR) using an array of 162 polypropylene fibers, 300 mm OD, 240 mm ID, surface area 0.0115 m². After emerging from the oxygenator the suspension passed through a filter (Millipore Isopore, pore size 5.0 mm, 13 mm diameter) at a pump rate of 1 mL/min. All experiments were performed at 21^oC. After achieving a steady flow rate, the oxygenator gas was abruptly changed to N₂, and the PO₂ and pressure of the prefilter suspension was continuously monitored. As PO₂ fell, pressure began to rise in all experiments, but in the presence of MP4, the pressure rise was offset, being lower by about 10 mm Hg, indicating a favorable effect on sickling at constant PO₂. If MP4 was saturated with CO, sickling was inhibited, even in the absence of O₂. These results indicate that MP4 has a significant effect on sickling, apart from any effect it might have in promoting capillary perfusion. Furthermore, the results confirm that despite its low P50 (~5 mm Hg), MP4 does not “steal” O₂ from sickle cells. Supported by grants HL64395 and HL076163 from the NHLBI