Haug Valentin, Peng Yifeng, Tchiloemba Bianief, Wang Alice T, Buerger Florian, Romfh Padraic, Kneser Ulrich, Polizzotti Brian D, Pomahac Bohdan
Division of Plastic Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Department of Hand, Plastic and Reconstructive Surgery, Microsurgery, Burn Trauma Center, BG Trauma Center Ludwigshafen, University of Heidelberg, 67071 Ludwigshafen, Germany.
J Clin Med. 2023 Oct 17;12(20):6568. doi: 10.3390/jcm12206568.
Normothermic ex situ perfusion of vascularized composite allografts (VCAs) necessitates high oxygen demand and, thus, increased metabolic activity, which, in turn, requires the use of blood-based perfusion solutions. However, blood-derived perfusates, in turn, constitute an antigenic load. To circumvent this immunogenic problem, we used a perfusate enriched with acellular dextrane oxygen microcarriers to perfuse rat hindlimbs.
Rat hindlimbs (n = 11) were perfused with either (non-), oxygenated dextrane-enriched Phoxilium, or Phoxilium enriched with dextrane oxygen microcarriers (MO) for 12 h at 37 °C or stored on ice. Oxygenation of the skeletal muscle was assessed with Raman spectroscopy, tissue pO-probes, and analysis of the perfusate. Transmission electronic microscopy was utilized to assess the ultrastructure of mitochondria of the skeletal muscle.
For all evaluated conditions, ischemia time until perfusion was comparable (22.91 ± 1.64 min; = 0.1559). After 12 h, limb weight increased significantly by at least 81%, up to 124% in the perfusion groups, and by 27% in the static cold storage (SCS) group. Raman spectroscopy signals of skeletal muscle did not differ substantially among the groups during either perfusion or static cold storage across the duration of the experiment. While the total number of skeletal muscle mitochondria decreased significantly compared to baseline, mitochondrial diameter increased in the perfusion groups and the static cold storage group.
The use of oxygen microcarriers in ex situ perfusion of VCA with acellular perfusates under normothermic conditions for 12 h facilitates the maintenance of mitochondrial structure, as well as a subsequent recovery of mitochondrial redox status over time, while markers of muscle injury were lower compared to conventional oxygenated acellular perfusates.
血管化复合异体移植（VCA）的常温体外灌注需要高氧需求，从而增加代谢活性，这反过来又需要使用基于血液的灌注溶液。然而，源自血液的灌注液又构成了抗原负荷。为了规避这个免疫原性问题，我们使用富含脱细胞葡聚糖氧微载体的灌注液灌注大鼠后肢。
11只大鼠后肢分别用（非）氧合富含葡聚糖的Phoxilium或富含葡聚糖氧微载体（MO）的Phoxilium在37℃下灌注12小时或置于冰上保存。用拉曼光谱、组织pO探针和灌注液分析评估骨骼肌的氧合情况。利用透射电子显微镜评估骨骼肌线粒体的超微结构。
在所有评估条件下，直至灌注的缺血时间相当（22.91±1.64分钟；P = 0.1559）。12小时后，灌注组肢体重量显著增加至少81%，最高达124%，静态冷藏（SCS）组增加27%。在实验期间的灌注或静态冷藏过程中，各组骨骼肌的拉曼光谱信号没有显著差异。与基线相比，骨骼肌线粒体总数显著减少，而灌注组和静态冷藏组的线粒体直径增加。
在常温条件下用脱细胞灌注液对VCA进行12小时的体外灌注时使用氧微载体，有助于维持线粒体结构，以及随后线粒体氧化还原状态随时间的恢复，同时与传统的氧合脱细胞灌注液相比，肌肉损伤标志物较低。
