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Development of recombinant hemoglobin-based oxygen carriers.

ABSTRACT: The worldwide blood shortage has generated a significant demand for alternatives to whole blood and packed red blood cells for use in transfusion therapy. One such alternative involves the use of acellular recombinant hemoglobin (Hb) as an oxygen carrier.Large amounts of recombinant human Hb can be expressed and purified from transgenic Escherichia coli. The physiological suitability of this material can be enhanced using protein-engineering strategies to address specific efficacy and toxicity issues. Mutagenesis of Hb can (i) adjust dioxygen affinity over a 100-fold range, (ii) reduce nitric oxide (NO) scavenging over 30-fold without compromising dioxygen binding, (iii) slow the rate of autooxidation, (iv) slow the rate of hemin loss, (v) impede subunit dissociation, and (vi) diminish irreversible subunit denaturation. Recombinant Hb production is potentially unlimited and readily subjected to current good manufacturing practices, but may be restricted by cost. Acellular Hb-based O(2) carriers have superior shelf-life compared to red blood cells, are universally compatible, and provide an alternative for patients for whom no other alternative blood products are available or acceptable.Remaining objectives include increasing Hb stability, mitigating iron-catalyzed and iron-centered oxidative reactivity, lowering the rate of hemin loss, and lowering the costs of expression and purification. Although many mutations and chemical modifications have been proposed to address these issues, the precise ensemble of mutations has not yet been identified.Future studies are aimed at selecting various combinations of mutations that can reduce NO scavenging, autooxidation, oxidative degradation, and denaturation without compromising O(2) delivery, and then investigating their suitability and safety in vivo.

SUBMITTER: Varnado CL

PROVIDER: S-EPMC3638513 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature

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Development of recombinant hemoglobin-based oxygen carriers.

Varnado Cornelius L CL Mollan Todd L TL Birukou Ivan I Smith Bryan J Z BJ Henderson Douglas P DP Olson John S JS

Antioxidants & redox signaling 20121116 17

<h4>Significance</h4>The worldwide blood shortage has generated a significant demand for alternatives to whole blood and packed red blood cells for use in transfusion therapy. One such alternative involves the use of acellular recombinant hemoglobin (Hb) as an oxygen carrier.<h4>Recent advances</h4>Large amounts of recombinant human Hb can be expressed and purified from transgenic Escherichia coli. The physiological suitability of this material can be enhanced using protein-engineering strategie ...[more]

PMID: 23025383

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Project description:BackgroundHyperfibrinolysis plays an integral role in the genesis of trauma-induced coagulopathy. Recent data demonstrate that red blood cell lysis promotes fibrinolysis; however, the mechanism is unclear. Hemoglobin-based oxygen carriers (HBOCs) have been developed for resuscitation and have been associated with coagulopathy. We hypothesize that replacement of whole blood (WB) using an HBOC results in a coagulopathy because of the presence of free hemoglobin.Materials and methodsWB was sampled from healthy donors (n = 6). The clotting profile of each citrated sample was evaluated using native thromboelastography. Serial titrations were performed using both HBOC (PolyHeme) and normal saline (NS; 5%, 25%, and 50%) and evaluated both with and without a 75-ng/mL tissue plasminogen activator (tPA) challenge. Tranexamic acid (TXA) was added to inhibit plasmin-dependent fibrinolysis. Fibrinolysis was measured and recorded as lysis at 30 min (LY30), the percentage of clot LY30 after maximal clot strength. Dilution of WB with NS or HBOC was correlated using LY30 via Spearman rho coefficients. Groups were also compared using a Friedman test and post hoc analysis with a Bonferroni adjustment.ResultstPA-provoked fibrinolysis was enhanced by both HBOC (median LY30 at 5%, 25%, and 50% titrations: 11%, 21%, and 44%, respectively; Spearman = 0.94; P < 0.001) and NS (11%, 28%, and 58%, respectively; Spearman = 0.790; P < 0.001). However, HBOC also enhanced fibrinolysis without the addition of tPA (1%, 4%, 5%; Spearman = 0.735; P = 0.001) and NS did not (1%, 2%, 1%; r = 0.300; P = 0.186. Moreover, addition of TXA did not alter or inhibit this fibrinolysis (WB versus 50% HBOC: 1.8% versus 5.7%, P = 0.04). There was no significant difference in fibrinolysis of HBOC with or without TXA (50% HBOC versus 50% HBOC + TXA: 5.6% versus 5.7%, P = 0.92). In addition, the increased fibrinolysis seen with NS was reversed when TXA was present (WB versus 50% NS: 1.8% versus 1.7%, P = 1.0).ConclusionsHBOCs enhance fibrinolysis both with and without addition of tPA; moreover, this mechanism is independent of plasmin as the phenomenon persists in the presence of TXA. Our findings indicate the hemoglobin molecule or its components stimulate fibrinolysis by both tPA-dependent and innate mechanisms.

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Development of recombinant hemoglobin-based oxygen carriers.

Publications

Development of recombinant hemoglobin-based oxygen carriers.

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