Tissue oxygenation after exchange transfusion with ultrahigh-molecular-weight tense- and relaxed-state polymerized bovine hemoglobins

Abstract

Hemoglobin (Hb)-based O(2) carriers (HBOCs) constitute a class of therapeutic agents designed to correct the O(2) deficit under conditions of anemia and traumatic blood loss. The O(2) transport capacity of ultrahigh-molecular-weight bovine Hb polymers (PolybHb), polymerized in the tense (T) state and relaxed (R) state, were investigated in the hamster chamber window model using microvascular measurements to determine O(2) delivery during extreme anemia. The anemic state was induced by hemodilution with a plasma expander (70-kDa dextran). After an initial moderate hemodilution to 18% hematocrit, animals were randomly assigned to exchange transfusion groups based on the type of PolybHb solution used (namely, T-state PolybHb and R-state PolybHb groups). Measurements of systemic parameters, microvascular hemodynamics, capillary perfusion, and intravascular and tissue O(2) levels were performed at 11% hematocrit. Both PolybHbs were infused at 10 g/dl, and their viscosities were higher than nondiluted blood. Restitution of the O(2) carrying capacity with T-state PolybHb exhibited lower arterial pressure and higher functional capillary density compared with R-state PolybHb. Central arterial O(2) tensions increased significantly for R-state PolybHb compared with T-state PolybHb; conversely, microvascular O(2) tensions were higher for T-state PolybHb compared with R-state PolybHb. The increased tissue Po(2) attained with T-state PolybHb results from the larger amount of O(2) released from the PolybHb and maintenance of macrovascular and microvascular hemodynamics compared with R-state PolybHb. These results suggest that the extreme high O(2) affinity of R-state PolybHb prevented O(2) bound to PolybHb from been used by the tissues. The results presented here show that T-state PolybHb, a high-viscosity O(2) carrier, is a quintessential example of an appropriately engineered O(2) carrying solution, which preserves vascular mechanical stimuli (shear stress) lost during anemic conditions and reinstates oxygenation, without the hypertensive or vasoconstriction responses observed in previous generations of HBOCs.

Biophysical Properties and Oxygenation Potential of High-Molecular-Weight Glutaraldehyde-Polymerized Human Hemoglobins Maintained in the Tense and Relaxed Quaternary States

“…This pattern is consistent with PolybHb (the P 50 is 41 mm Hg for 50:1 T-state PolybHb and 0.66 mm Hg for 40:1 R-state PolybHb). 21,22 Both PolyhHbs show no cooperativity. Polymerization of hHb freezes the structure of hHb in a well-defined quaternary state.…”

“…[18][19][20] PolyhHb synthesis T-state PolyhHb was synthesized as previously described in the literature. 9,[21][22][23] hHb was diluted with 20 mM phosphate buffer (PB; pH 8.0) to yield 1.5 L of 0.3 mM hHb that was stored in a 2 L glass bottle submerged in an ice bath. The contents of the glass bottle were then subjected to vacuum for 1 min, followed by argon purging for 1 min.…”

“…[21][22][23] About 1.5 L of 0.3 mM hHb in 20 mM PB was purged with pure O 2 for 2 h in an ice bath, and its pO 2 was continuously monitored with a RapidLab 248 Blood Gas Analyzer. After the pO 2 was out of range (>749 mm Hg), glutaraldehyde was added at the following glutaraldehyde to hHb molar ratios-30:1 and 20:1.…”

Simulation of NO and O2 transport facilitated by polymerized hemoglobin solutions in an arteriole that takes into account wall shear stress-induced NO production

“…The arteriole diameter, average blood velocity and discharge hct ( H D ) for these simulations were taken from experimental data reported in reference [39] and personal communications from Dr. Cabrales (Bioengineering Department) at the University of California, San Diego (UCSD): the H D was set to 11% to mimic animal studies conducted using 50:1 L-PolyHb and 40:1 H-PolyHb solutions, while the H D was set to 18% to mimic animal studies conducted using 20:1 and 30:1 L- and H-PolyHb solutions. The radius of the RBC-rich region ( r 1 ) changes proportionally with the H D [58, 59], and the r 1 value used in this work was extrapolated from values of r 1 measured at hct s of 20%, 30% and 40% that were obtained from the literature [59].…”

“…In the physiological range of observed blood vessel wall shear stresses ranging from 6–25 dyne/cm 2 , the rate of release of NO was found to be linearly dependent on the shear stress acting on the blood vessel wall [34, 35]. To further emphasize the importance of shear stress-induced NO production, Cabrales et al reported that transfusion of high viscosity >500 kDa fraction of ultrahigh molecular weight (MW) glutaraldehyde polymerized bHb (PolyHb) solutions synthesized by our group [36, 37] elicited little to no vasoconstriction in a hamster top-load model [38] and in an exchange transfusion model [39], while the commercial low MW polymerized HBOC Oxyglobin ® (average MW ~ 200 kDa) elicited severe hypertension [40], although these various PolyHbs possessed NO dioxygenation rate constants that were similar to that of cell-free Hb. Compared to Oxyglobin ® (1.8 cP at a total Hb concentration of 13 g/dL) [39], the viscosities of PolyHbs synthesized by our group are much higher than that of Oxyglobin ® (3.6 – 14 cP at a total Hb concentration of 10 g/dL).…”