SFR BioSciences, Skin Functional Integrity group. Laboratory for Tissue Biology and Therapeutics Engineering (LBTI) CNRS UMR5305 - University of Lyon, Lyon, France.
SFR BioSciences, UAR 3444, US8, ENS de Lyon, UCBL, AniRA ImmOs, Lyon, France.
Skin Res Technol. 2024 Mar;30(3):e13638. doi: 10.1111/srt.13638.
Skin wound healing is a complex mechanism which requires a lot of energy, mainly provided by mitochondrial respiration. However, little is known about the mitochondrial bioenergetics of mice skin. We sought to develop a microplate-based assay to directly measure oxygen consumption in whole mice skin with the goal of identifying mitochondrial dysfunction in diabetic skin using an extracellular flux.
Different parameters were optimized to efficiently measure the oxygen consumption rate (OCR). First, the most pertinent skin side of wild-type mice was first determined. Then, concentrations of mitochondrial inhibitors were then optimized to get the best efficacy. Finally, punch sizes were modulated to get the best OCR profile.
Dermis had the best metabolic activity side of the skin. Unlike the increased concentrations of carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) and rotenone/antimycin A, which showed no improvement of these drugs' effects, varying the skin punch size was successful. Finally, type II diabetic (T2D) skin produced less ATP through mitochondrial metabolism and had a greater non-mitochondrial oxygen consumption than wild-type or type I diabetic (T1D) skin.
Here we designed, for the first time, a reliable protocol to measure mitochondria function in whole mouse skin. Our optimized protocol was valuable in assessing alterations associated with diabetes and could be applied to future studies of pathological human skin metabolism.
皮肤伤口愈合是一个复杂的机制，需要大量的能量，主要由线粒体呼吸提供。然而，关于小鼠皮肤的线粒体生物能学知之甚少。我们试图开发一种基于微孔板的测定法，直接测量整个小鼠皮肤的耗氧量，目的是使用细胞外通量来鉴定糖尿病皮肤中的线粒体功能障碍。
优化了不同的参数，以有效地测量耗氧量（OCR）。首先，确定了野生型小鼠最相关的皮肤侧。然后，优化了线粒体抑制剂的浓度以获得最佳效果。最后，调节打孔器的尺寸以获得最佳的 OCR 谱。
真皮具有皮肤最佳的代谢活性侧。与增加羰基氰化物-对三氟甲氧基苯腙（FCCP）和鱼藤酮/抗霉素 A 的浓度不同，这些药物的效果没有改善，而改变皮肤打孔器的尺寸则是成功的。最后，2 型糖尿病（T2D）皮肤通过线粒体代谢产生的 ATP 较少，而非线粒体耗氧量大于野生型或 1 型糖尿病（T1D）皮肤。
我们首次设计了一种可靠的方案来测量整个小鼠皮肤的线粒体功能。我们优化的方案在评估与糖尿病相关的变化方面具有价值，并可应用于未来病理性人类皮肤代谢的研究。
