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For the past thirty years, hemoglobin-based oxygen carriers (HBOCs) have been under development as a red blood cell substitute. Side-effects such as vasoconstriction, oxidative injury, and cardiac toxicity have prevented clinical approval of HBOCs. Recently, high molecular weight (MW) polymerized human hemoglobin (PolyhHb) has shown positive results in rats. Studies have demonstrated that high MW PolyhHb increased O2 delivery, with minimal effects on blood pressure, without vasoconstriction, and devoid of toxicity. In this study, we used guinea pigs to evaluate the efficacy and safety of high MW PolyhHb, since like humans guinea pigs cannot produce endogenous ascorbic acid, which limits the capacity of both species to deal with oxidative stress. Hence, this study evaluated the efficacy and safety of resuscitation from severe hemorrhagic shock with high MW PolyhHb, fresh blood, and blood stored for 2 weeks. Animals were randomly assigned to each experimental group, and hemorrhage was induced by the withdrawal of 40% of the blood volume (BV, estimated as 7.5% of body weight) from the carotid artery catheter. Hypovolemic shock was maintained for 50 min. Resuscitation was implemented by infusing 25% of the animal's BV with the different treatments. Hemodynamics, blood gases, total hemoglobin, and lactate were not different before hemorrhage and during shock between groups. The hematocrit was lower for the PolyhHb group compared to the fresh and stored blood groups after resuscitation. Resuscitation with stored blood had lower blood pressure compared to fresh blood at 2 h. There was no difference in mean arterial pressure between groups at 24 h. Resuscitation with PolyhHb was not different from fresh blood for most parameters. Resuscitation with PolyhHb did not show any remarkable change in liver injury, inflammation, or cardiac damage. Resuscitation with stored blood showed changes in liver function and inflammation, but no kidney injury or systemic inflammation. Resuscitation with stored blood after 24 h displayed sympathetic hyper-activation and signs of cardiac injury. These results suggest that PolyhHb is an effective resuscitation alternative to blood. The decreased toxicities in terms of cardiac injury markers, vital organ function, and inflammation following PolyhHb resuscitation in guinea pigs indicate a favorable safety profile. These results are promising and support future studies with this new generation of PolyhHb as alternative to blood when blood is unavailable.

Fluid resuscitation following severe inflammation-induced hypoperfusion is critical for the restoration of hemodynamics and the prevention of multiorgan dysfunction syndrome during septic shock. Fluid resuscitation with commercially available crystalloid and colloid solutions only provides transient benefits, followed by fluid extravasation and tissue edema through the inflamed endothelium. The increased molecular weight (M.W.) of polymerized human serum albumin (PolyHSA) can limit fluid extravasation, leading to restoration of hemodynamics. In this prospective study, we evaluated how fluid resuscitation with PolyHSA impacts the hemodynamic and immune response in a lipopolysaccharide (LPS) induced endotoxemia mouse model. Additionally, we evaluated fluid resuscitation with PolyHSA in a model of polymicrobial sepsis induced by cecal ligation and puncture (CLP). Resuscitation with PolyHSA attenuated the immune response and improved the maintenance of systemic hemodynamics and restoration of microcirculatory hemodynamics. This decrease in inflammatory immune response and maintenance of vascular wall shear stress likely contributes to the maintenance of vascular integrity following fluid resuscitation with PolyHSA. The sustained restoration of perfusion, decrease in pro-inflammatory immune response, and improved vascular integrity that results from the high M.W. of PolyHSA indicates that a PolyHSA based solution is a potential resuscitation fluid for endotoxic and septic shock.

Traumatic brain injury (TBI) is often accompanied by hemorrhage, and treatment of hemorrhagic shock (HS) after TBI is particularly challenging because the two therapeutic treatment strategies for TBI and HS often conflict. Ischemia/reperfusion injury from HS resuscitation can be exaggerated by TBI-induced loss of autoregulation. In HS resuscitation, the goal is to restore lost blood volume, while in the treatment of TBI the priority is focused on maintenance of adequate cerebral perfusion pressure and avoidance of secondary bleeding. In this study, we investigate the responses to resuscitation from severe HS after TBI in rats, using fresh blood, polymerized human hemoglobin (PolyhHb), and lactated Ringer's (LR). Rats were subjected to TBI by pneumatic controlled cortical impact. Shortly after TBI, HS was induced by blood withdrawal to reduce mean arterial pressure (MAP) to 35-40 mmHg for 90 min before resuscitation. Resuscitation fluids were delivered to restore MAP to ~ 65 mmHg and animals were monitored for 120 min. Increased systolic blood pressure variability (SBPV) confirmed TBI-induced loss of autoregulation. MAP after resuscitation was significantly higher in the blood and PolyhHb groups compared to the LR group. Furthermore, blood and PolyhHb restored diastolic pressure, while this remained depressed for the LR group, indicating a loss of vascular tone. Lactate increased in all groups during HS, and only returned to baseline level in the blood reperfused group. The PolyhHb group possessed lower SBPV compared to LR and blood groups. Finally, sympathetic nervous system (SNS) modulation was higher for the LR group and lower for the PolyhHb group compared to the blood group after reperfusion. In conclusion, our results suggest that PolyhHb could be an alternative to blood for resuscitation from HS after TBI when blood is not available, assuming additional testing demonstrate similar favorable results. PolyhHb restored hemodynamics and oxygen delivery, without the logistical constraints of refrigerated blood.

Administration of hemoglobin-based oxygen carriers (HBOCs) into the systemic circulation is a potential strategy to relieve solid tumor hypoxia in order to increase the effectiveness of chemotherapeutics. Previous computational analysis indicated that the oxygen (O2) status of the tumor and HBOC O2 affinity may play a role in increased O2 delivery to the tumor. However, no study has experimentally investigated how low- and high-affinity HBOCs would perform in normoxic and hypoxic tumors. In this study, we examined how the HBOC, polymerized human hemoglobin (PolyhHb), in the relaxed (R) or tense (T) quaternary state modulates O2 delivery to hypoxic (FME) and normoxic (LOX) human melanoma xenografts in a murine window chamber model. We examined microcirculatory fluid flow via video shearing optical microscopy, and O2 distributions via phosphorescence quenching microscopy. Additionally, we examined how weekly infusion of a 20% top-load dose of PolyhHb influences growth rate, vascularization, and regional blood flow in the FME and LOX tumor xenografts. Infusion of low-affinity T-state PolyhHb led to increased tissue oxygenation, decreased blood flow, decreased tumor growth, and decreased vascularization in hypoxic tumors. However, infusion of both T-state and R-state PolyhHbs led to worse outcomes in normoxic tumors. Of particular concern was the high-affinity R-state PolyhHb, which led to no improvement in hypoxic tumors and significantly worsened outcomes in normoxic tumors. Taken together, the results of this study indicate that the tumor O2 status is a primary determinant of the potency and outcomes of infused PolyhHb.