James Isaac, Bourne Debra, Silva Mayara, Havis Emmanuelle, Albright Kassandra, Zhang Liyong, Kostereva Nataliya, Wang Sheri, DiBernardo Gabriella, Guest Rachel, Lei Jenny, Almadori Aurora, Satish Latha, Marra Kacey, Rubin J Peter
Department of Plastic Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania.
Department of Plastic Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania; Discipline of Plastic Surgery, Department of Surgery, Federal University of São Paulo, São Paulo, Brazil.
J Surg Res. 2018 Sep;229:243-253. doi: 10.1016/j.jss.2018.03.068. Epub 2018 May 3.
Adipose-derived stem cells (ASCs) are capable of secreting regenerative growth factors and replacing multiple tissue types. Although current literature suggests that ASCs accelerate wound healing and reduce scarring, the dose-response relationship has not been adequately investigated in large animals. We sought to establish a porcine model to optimize dose and delivery.
Four-centimeter circular, full thickness excisional wounds were created on the backs of Yorkshire pigs. Fluorescently labeled allogeneic porcine ASCs were injected into the superficial wound bed and around the wound perimeter at high (3.0 × 10 cells/cm; n = 8), medium (1.0 × 10 cells/cm; n = 8), and low (0.3 × 10 cells/cm; n = 8) doses. Control wounds received saline injections (n = 8) or no treatment (n = 8). Dressings were changed twice per week, and wound closure was tracked by surface area tracing. Animals were sacrificed at 1 and 2 wk. Wounds were harvested for real-time quantitative reverse transcriptase polymerase chain reaction, immunohistochemistry, and ASC tracking.
Labeled ASCs integrated into treated wounds by 1 wk in a dose-dependent fashion. Epithelial coverage was achieved by 14 d in all wounds. Wounds receiving high-dose ASCs exhibited thicker granulating neodermis at 7 d and greater wound contraction at 14 d. real-time quantitative reverse transcriptase polymerase chain reaction revealed improved collagen 1:collagen 3 (Col1:Col3) ratio in the medium-dose group and enhanced α-smooth muscle actin in the high-dose group at 14 d. Western blot demonstrated increased cluster of differentiation 31 protein at 2 wk in wounds receiving >10 cells/cm.
Doses up to 3.0 × 10 cells/cm were well-tolerated. High-dose ASCs accelerate wound contraction, enhance neovascularization, and may improve scar quality in excisional wounds healing by secondary intention. Doses greater than those previously used may be necessary to achieve desired effects.
脂肪来源干细胞（ASC）能够分泌再生生长因子并替代多种组织类型。尽管当前文献表明ASC可加速伤口愈合并减少瘢痕形成，但在大型动物中尚未充分研究剂量反应关系。我们试图建立一个猪模型来优化剂量和给药方式。
在约克郡猪的背部制作4厘米圆形全层切除伤口。将荧光标记的同种异体猪ASC以高剂量（3.0×10⁶细胞/cm²；n = 8）、中剂量（1.0×10⁶细胞/cm²；n = 8）和低剂量（0.3×10⁶细胞/cm²；n = 8）注入浅表伤口床和伤口周边。对照伤口接受盐水注射（n = 8）或不进行治疗（n = 8）。每周更换两次敷料，并通过表面积追踪来跟踪伤口闭合情况。在1周和2周时处死动物。采集伤口用于实时定量逆转录聚合酶链反应、免疫组织化学和ASC追踪。
标记的ASC在1周时以剂量依赖方式整合到治疗的伤口中。所有伤口在14天时实现上皮覆盖。接受高剂量ASC的伤口在7天时表现出更厚的肉芽新真皮，在14天时伤口收缩更大。实时定量逆转录聚合酶链反应显示，中剂量组在14天时胶原1:胶原3（Col1:Col3）比例改善，高剂量组在14天时α平滑肌肌动蛋白增强。蛋白质印迹法显示，在接受>10⁶细胞/cm²的伤口中，2周时分化簇31蛋白增加。
高达3.0×10⁶细胞/cm²的剂量耐受性良好。高剂量ASC可加速伤口收缩，增强新血管形成，并可能改善二期愈合的切除伤口的瘢痕质量。可能需要大于先前使用的剂量才能达到预期效果。
