Sobral Christiane S, Gragnani Alfredo, Cao Xudong, Morgan Jeffrey R, Ferreira Lydia Masako
Skin Cell Culture Laboratory, Division of Plastic Surgery, Federal University of Sao Paulo (UNIFESP), Sao Paulo, Brazil.
J Burns Wounds. 2007 Oct 30;7:e6.
In experimental models in vivo, it is difficult to characterize the effect of thermal burns on epidermal keratinocytes. Since the response to thermal injury involves several systemic mechanisms, especially because of the stimulus to coagulation and inflammatory cascades, it becomes hard to evaluate the specific effect of thermal burns on keratinocytes. The aim of this study is to propose the use of human keratinocytes cultured on collagen matrix as an in vitro experimental burn model.
Human keratinocytes derived from neonatal foreskins were isolated and cultured following standard methods. All experiments used the same keratinocyte lineage and were carried out in triplicate. Initially, gels of collagen and Matrigel were prepared. For each gel, 2 x 10(6) keratinocytes were seeded and cultured to form stratified epithelia. Following, burn wounds were induced at 170 degrees C.
Keratinocytes were cultured on collagen-coated Millicell membranes. Stratified epithelia were formed and burned on the seventh day after the cultures were raised to the air-liquid interface. The burn procedure is reproducible and can be easily executed.
The proposed model can be used to study the effects of induced burn wounds on keratinocytes in a specific way.
在体内实验模型中，很难描述热烧伤对表皮角质形成细胞的影响。由于对热损伤的反应涉及多种全身机制，特别是由于对凝血和炎症级联反应的刺激，因此很难评估热烧伤对角质形成细胞的具体影响。本研究的目的是提出使用在胶原基质上培养的人角质形成细胞作为体外实验性烧伤模型。
按照标准方法分离和培养源自新生儿包皮的人角质形成细胞。所有实验均使用相同的角质形成细胞谱系，并重复进行三次。最初，制备胶原蛋白和基质胶凝胶。对于每种凝胶，接种2×10⁶个角质形成细胞并培养以形成分层上皮。随后，在170℃诱导烧伤创面。
角质形成细胞在胶原包被的Millicell膜上培养。在培养物升至气液界面后第7天形成分层上皮并进行烧伤。烧伤程序具有可重复性且易于执行。
所提出的模型可用于以特定方式研究诱导烧伤创面对角质形成细胞的影响。
