Department of Biomedicine, Aarhus University, 8000 Aarhus C, Denmark.
University Clinic for Plastic, Aesthetic and Hand Surgery, Otto-von-Guericke-University, 39120 Magdeburg, Germany.
Int J Mol Sci. 2017 Dec 3;18(12):2604. doi: 10.3390/ijms18122604.
To prepare the ESA (European Space Agency) spaceflight project "Wound healing and Sutures in Unloading Conditions", we studied mechanisms of apoptosis in wound healing models based on ex vivo skin tissue cultures, kept for 10 days alive in serum-free DMEM/F12 medium supplemented with bovine serum albumin, hydrocortisone, insulin, ascorbic acid and antibiotics at 32 °C. The overall goal is to test: (i) the viability of tissue specimens; (ii) the gene expression of activators and inhibitors of apoptosis and extracellular matrix components in wound and suture models; and (iii) to design analytical protocols for future tissue specimens after post-spaceflight download. Hematoxylin-Eosin and Elastica-van-Gieson staining showed a normal skin histology with no signs of necrosis in controls and showed a normal wound suture. TdT-mediated dUTP-biotin nick end labeling for detecting DNA fragmentation revealed no significant apoptosis. No activation of caspase-3 protein was detectable. , , , , , , , , , and mRNAs were not altered in epidermis and dermis samples with and without a wound compared to 0 day samples (specimens investigated directly post-surgery). , , and mRNAs were downregulated in epidermis/dermis samples with and/or without a wound compared to 0 day samples. , were upregulated in 10 day wound samples compared to 0 day samples in epidermis/dermis. mRNAs were elevated in 10 day wound and no wound samples compared to 0 day samples in dermis. In conclusion, we demonstrate that it is possible to maintain live skin tissue cultures for 10 days. The viability analysis showed no significant signs of cell death in wound and suture models. The gene expression analysis demonstrated the interplay of activators and inhibitors of apoptosis and extracellular matrix components, thereby describing important features in ex vivo sutured wound healing models. Collectively, the performed methods defining analytical protocols proved to be applicable for post-flight analyzes of tissue specimens after sample return.
为了准备欧洲航天局（ESA）的太空飞行项目“在减压条件下的伤口愈合和缝合”，我们研究了基于体外皮肤组织培养的伤口愈合模型中细胞凋亡的机制，这些组织在无血清 DMEM/F12 培养基中保持 10 天的存活，该培养基中添加了牛血清白蛋白、氢化可的松、胰岛素、抗坏血酸和抗生素，并在 32°C 下培养。总体目标是测试：（i）组织标本的活力；（ii）伤口和缝合模型中外源基质成分和凋亡激活剂和抑制剂的基因表达；以及（iii）为未来的组织标本设计分析方案，以便在太空飞行后下载。苏木精-伊红和弹性纤维-van-Gieson 染色显示，对照组无组织坏死迹象，且伤口和缝合均正常。末端转移酶介导的 dUTP-生物素缺口末端标记法检测 DNA 片段化显示无明显凋亡。不可检测到 caspase-3 蛋白的激活。与 0 天样本相比，在有和没有伤口的表皮和真皮样本中， 、 、 、 、 、 、 、 和 mRNAs 没有改变（直接手术后进行研究的标本）。与 0 天样本相比，在有和/或没有伤口的表皮/真皮样本中， 、 、 和 mRNAs 下调。与 0 天样本相比，在 10 天伤口样本中 上调。与 0 天样本相比，在 10 天伤口和无伤口样本中 mRNAs 升高。总之，我们证明了可以维持 10 天的活体皮肤组织培养。活力分析显示伤口和缝合模型中没有明显的细胞死亡迹象。基因表达分析表明了凋亡激活剂和抑制剂以及细胞外基质成分之间的相互作用，从而描述了体外缝合伤口愈合模型中的重要特征。总的来说，所采用的方法确定了适用于样本返回后组织标本飞行后分析的分析方案。
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