Figueroa Brian A, Said Sayf A, Ordenana Carlos, Rezaei Majid, Orfahli Lynn M, Dubé Gregory P, Papay Francis, Brunengraber Henri, Dasarathy Srinivasan, Rampazzo Antonio, Gharb Bahar Bassiri
From the Department of Plastic Surgery (B.A.F., S.A.S., C.O., M.R., L.M.O., F.P., A.R., B.B.G.), Cleveland Clinic; Department of Nutrition (H.B.), School of Medicine, Case Western Reserve University; Department of Gastroenterology (S.D.), Cleveland Clinic, Cleveland, Ohio; and Hemoglobin Oxygen Therapeutics, LLC (G.P.D.), Souderton, Pennsylvania.
J Trauma Acute Care Surg. 2022 Feb 1;92(2):388-397. doi: 10.1097/TA.0000000000003395.
Ex vivo normothermic limb perfusion (EVNLP) preserves amputated limbs under near-physiologic conditions. Perfusates containing red blood cells (RBCs) have shown to improve outcomes during ex vivo normothermic organ perfusion, when compared with acellular perfusates. To avoid limitations associated with the use of blood-based products, we evaluated the feasibility of EVNLP using a polymerized hemoglobin-based oxygen carrier-201 (HBOC-201).
Twenty-four porcine forelimbs were procured from Yorkshire pigs. Six forelimbs underwent EVNLP with an HBOC-201-based perfusate, six with an RBC-based perfusate, and 12 served as static cold storage (SCS) controls. Ex vivo normothermic limb perfusion was terminated in the presence of systolic arterial pressure of 115 mm Hg or greater, fullness of compartments, or drop of tissue oxygen saturation by 20%. Limb contractility, weight change, compartment pressure, tissue oxygen saturation, oxygen uptake rates (OURs) were assessed. Perfusate fluid-dynamics, gases, electrolytes, metabolites, methemoglobin, creatine kinase, and myoglobin concentration were measured. Uniformity of skin perfusion was assessed with indocyanine green angiography and infrared thermography.
Warm ischemia time before EVNLP was 35.50 ± 8.62 minutes (HBOC-201), 30.17 ± 8.03 minutes (RBC) and 37.82 ± 10.45 (SCS) (p = 0.09). Ex vivo normothermic limb perfusion duration was 22.5 ± 1.7 hours (HBOC-201) and 28.2 ± 7.3 hours (RBC) (p = 0.04). Vascular flow (325 ± 25 mL·min-1 vs. 444.7 ± 50.6 mL·min-1; p = 0.39), OUR (2.0 ± 1.45 mL O2·min-1·g-1 vs. 1.3 ± 0.92 mL O2·min-1·g-1 of tissue; p = 0.80), lactate (14.66 ± 4.26 mmol·L-1 vs. 13.11 ± 6.68 mmol·L-1; p = 0.32), perfusate pH (7.53 ± 0.25 HBOC-201; 7.50 ± 0.23 RBC; p = 0.82), flexor (28.3 ± 22.0 vs. 27.5 ± 10.6; p = 0.99), and extensor (31.5 ± 22.9 vs. 28.8 ± 14.5; p = 0.82) compartment pressures, and weight changes (23.1 ± 3.0% vs. 13.2 ± 22.7; p = 0.07) were not significantly different between HBOC-201 and RBC groups, respectively. In HBOC-201 perfused limbs, methemoglobin levels increased, reaching 47.8 ± 12.1% at endpoint. Methemoglobin saturation did not affect OUR (ρ = -0.15, r2 = 0.022; p = 0.45). A significantly greater number of necrotic myocytes was found in the SCS group at endpoint (SCS, 127 ± 17 cells; HBOC-201, 72 ± 30 cells; RBC-based, 56 ± 40 cells; vs. p = 0.003).
HBOC-201- and RBC-based perfusates similarly support isolated limb physiology, metabolism, and function.
体外常温肢体灌注（EVNLP）可在接近生理条件下保存截肢肢体。与无细胞灌注液相比，含红细胞（RBC）的灌注液已显示可改善体外常温器官灌注期间的结果。为避免与使用基于血液的产品相关的局限性，我们评估了使用聚合血红蛋白基氧载体-201（HBOC-201）进行EVNLP的可行性。
从约克夏猪获取24个猪前肢。6个前肢用基于HBOC-201的灌注液进行EVNLP，6个用基于RBC的灌注液进行EVNLP，12个作为静态冷藏（SCS）对照。当收缩压达到或高于115 mmHg、组织腔饱满或组织氧饱和度下降20%时，终止体外常温肢体灌注。评估肢体收缩性、重量变化、组织腔压力、组织氧饱和度、氧摄取率（OURs）。测量灌注液的流体动力学、气体、电解质、代谢物、高铁血红蛋白、肌酸激酶和肌红蛋白浓度。用吲哚菁绿血管造影和红外热成像评估皮肤灌注的均匀性。
EVNLP前的热缺血时间为35.50±8.62分钟（HBOC-201）、30.17±8.03分钟（RBC）和37.82±10.45分钟（SCS）（p = 0.09）。体外常温肢体灌注持续时间为22.5±1.7小时（HBOC-201）和28.2±7.3小时（RBC）（p = 0.04）。血管流量（325±25 mL·min-1对444.7±50.6 mL·min-1；p = 0.39）、OUR（2.0±1.45 mL O2·min-1·g-1组织对1.3±0.92 mL O2·min-1·g-1组织；p = 0.80）、乳酸（14.66±4.26 mmol·L-1对13.11±6.68 mmol·L-1；p = 0.32）、灌注液pH（HBOC-201为7.53±0.25；RBC为7.50±0.23；p = 0.82）、屈肌（28.3±22.0对27.5±10.6；p = 0.99）和伸肌（31.5±22.9对28.8±14.5；p = 0.82）组织腔压力以及重量变化（23.1±3.0%对13.2±22.7；p = 0.07）在HBOC-201组和RBC组之间分别无显著差异。在HBOC-201灌注的肢体中，高铁血红蛋白水平升高，在终点时达到47.8±（此处原文可能有误，推测为12.1）%。高铁血红蛋白饱和度不影响OUR（ρ = -0.15，r2 = 0.022；p = 0.45）。在终点时，SCS组发现坏死肌细胞数量显著更多（SCS为127±17个细胞；HBOC-201为72±30个细胞；基于RBC的为56±40个细胞；p = 0.003）。
基于HBOC-201和RBC的灌注液同样支持离体肢体的生理、代谢和功能。
