Reconstructive surgery transfers viable tissue to cover defects and to restore aesthetic and functional properties. Failure rates after free flap surgery range from 3 to 7%. Co‐morbidities such as diabetes mellitus or peripheral vascular disease increase the risk of flap failure up to 4.5‐fold. Experimental therapeutic concepts commonly use a monocausal approach by applying single growth factors. The secretome of γ‐irradiated, stressed peripheral blood mononuclear cells (PBMCsec) resembles the physiological environment necessary for tissue regeneration. Its application led to improved wound healing rates and a two‐fold increase in blood vessel counts in previous animal models. We hypothesized that PBMCsec has beneficial effects on the survival of compromised flap tissue by reducing the necrosis rate and increasing angiogenesis. Surgery was performed on 39 male Sprague–Dawley rats (control, N = 13; fibrin sealant, N = 14; PBMCsec, N = 12). PBMCsec was produced according to good manufacturing practices (GMP) guidelines and 2 ml were administered intraoperatively at a concentration of 2.5 × 10⁷ cells/ml using fibrin sealant as carrier substance. Flap perfusion and necrosis (as percentage of the total flap area) were analyzed using Laser Doppler Imaging and digital image planimetry on postoperative days 3 and 7. Immunohistochemical stainings for von Willebrand factor (vWF) and Vascular Endothelial Growth Factor‐receptor‐3 (Flt‐4) were performed on postoperative day 7 to evaluate formation of blood vessels and lymphatic vessels. Seroma formation was quantified using a syringe and flap adhesion and tissue edema were evaluated clinically through a cranial incision by a blinded observer according to previously described criteria on postoperative day 7. We found a significantly reduced tissue necrosis rate (control: 27.8% ± 8.6; fibrin: 22.0% ± 6.2; 20.9% reduction, p = .053 vs. control; PBMCsec: 19.1% ± 7.2; 31.1% reduction, p = .012 vs. control; 12.9% reduction, 0.293 vs. fibrin) together with increased vWF+ vessel counts (control: 70.3 ± 16.3 vessels/4 fields at 200× magnification; fibrin: 67.8 ± 12.1; 3.6% reduction, p = .651, vs. control; PBMCsec: 85.9 ± 20.4; 22.2% increase, p = .045 vs. control; 26.7% increase, p = .010 vs. fibrin) on postoperative day 7 after treatment with PBMCsec. Seroma formation was decreased after treatment with fibrin sealant with or without the addition of PBMCsec. (control: 11.9 ± 9.7 ml; fibrin: 1.7 ± 5.3, 86.0% reduction, 0.004 vs. control; PBMCsec: 0.6 ± 2.0; 94.8% reduction, p = .001 vs. control; 62.8% reduction, p = .523 vs. fibrin). We describe the beneficial effects of a secretome derived from γ‐irradiated PBMCs on tissue survival, angiogenesis, and clinical parameters after flap surgery in a rodent epigastric flap model.

中文翻译：

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应激外周血单核细胞的分泌组可提高啮齿动物上腹皮瓣模型中的组织存活率


重建手术转移活组织来覆盖缺陷并恢复美观和功能特性。游离皮瓣手术后的失败率为 3% 至 7%。糖尿病或周围血管疾病等合并症可使皮瓣失败的风险增加至 4.5 倍。实验性治疗概念通常使用单因素方法，即应用单一生长因子。 γ 辐射、应激的外周血单核细胞 (PBMCsec) 的分泌组类似于组织再生所需的生理环境。它的应用提高了伤口愈合率，并使之前的动物模型中的血管数量增加了一倍。我们假设 PBMCsec 通过降低坏死率和增加血管生成对受损皮瓣组织的存活产生有益影响。对 39 只雄性 Sprague-Dawley 大鼠进行了手术（对照， N = 13；纤维蛋白密封剂， N = 14；PBMCsec， N = 12）。 PBMCsec是根据良好生产规范(GMP)指南生产的，并且使用纤维蛋白密封剂作为载体物质以2.5×10 ^{7 个}细胞/ml的浓度在术中施用2ml。术后第 3 天和第 7 天，使用激光多普勒成像和数字图像面积测定法分析皮瓣灌注和坏死（占皮瓣总面积的百分比）。对血管性血友病因子 (vWF) 和血管内皮生长因子受体 3 (Flt) 进行免疫组织化学染色‐4) 在术后第 7 天进行，以评估血管和淋巴管的形成。 使用注射器对血清形成进行定量，并由盲人观察者根据先前描述的标准在术后第 7 天通过颅骨切口对皮瓣粘连和组织水肿进行临床评估。我们发现组织坏死率显着降低（对照：27.8％±8.6；对照：27.8％±8.6）。纤维蛋白：22.0% ± 6.2；与对照相比减少 20.9%；与对照相比，PBMCsec：19.1% ± 7.2；减少 31.1% ；与纤维蛋白相比减少 12.9%；0.293 vWF+ 血管计数（对照：70.3 ± 16.3 血管/4 个视野，200 倍放大倍数；纤维蛋白：67.8 ± 12.1；与对照相比，减少 3.6%， p = .651；PBMCsec：85.9 ± 20.4；增加 22.2%， p = . 045 与对照相比；在术后第 7 天用 PBMCsec 治疗后增加 26.7%， p = .010。在添加或不添加 PBMCsec 的纤维蛋白密封剂治疗后，血清形成减少。 （对照：11.9 ± 9.7 ml；纤维蛋白：1.7 ± 5.3，减少 86.0%，与对照相比减少 0.004；PBMCsec：0.6 ± 2.0；减少 94.8%，与对照相比p = .001；减少 62.8%，与对照相比p = .523纤维蛋白）。我们描述了来自 γ 照射的 PBMC 的分泌蛋白组对啮齿动物上腹皮瓣模型皮瓣手术后组织存活、血管生成和临床参数的有益影响。