Hemocompatibility Evaluation of PEGylated Bovine Hemoglobin

Hemocompatibility is critical for intravenous hemoglobin-based oxygen carriers (HBOCs). We evaluated the hemocompatibility of polyethylene glycol-conjugated bovine hemoglobin (PEG-bHb) to facilitate its optimization and clinical translation. PEG-bHb was synthesized and characterized. In vitro hemocompatibility was assessed by incubating blood components with PEG-bHb (2.5-40 mg/mL), evaluating erythrocyte morphology/function, coagulation, complement activation, and leu kocyte phagocytosis. In vivo assessments in Wistar rats injecting PEG-bHb (50-100 mg/kg) included erythrocyte function, coagulation, complement activation, and histopathology. PEG-bHb exhibited increased hydrodynamic diameter, unaltered zeta potential, elevated colloidal osmotic pressure (COP) and viscosity, alongside a decreased P₅₀ versus bovine hemoglobin (bHb). In vitro, PEG-bHb preserved erythrocyte morphology without inducing eryptosis, and oxygen supply efficiency was unaffected. Moreover, it slightly disrupted the activated partial thromboplastin time (APTT), the thrombin time (TT), and the platelet adhesion, while platelet activation and thromboelastography (TEG) remained unchanged. PEG-bHb did not activate complement and only mildly enhanced phagocytosis at 2.5 mg/mL. In vivo, PEG-bHb did not affect eryptosis, oxygen supply efficiency, coagulation, complement activation, and no inflammatory infiltration was observed. PEG-bHb maintains erythrocyte morphology and function, slightly perturb coagulation without compromising immunocompatibility, and demonstrated excellent hemocompatibility.