Lone Abdul G, Atci Erhan, Renslow Ryan, Beyenal Haluk, Noh Susan, Fransson Boel, Abu-Lail Nehal, Park Jeong-Jin, Gang David R, Call Douglas R
Paul G. Allen School for Global Animal Health, Washington State University, Pullman, Washington, USA.
School of Chemical Engineering & Bioengineering, Washington State University, Pullman, Washington, USA.
Infect Immun. 2015 Aug;83(8):3026-34. doi: 10.1128/IAI.00175-15. Epub 2015 May 18.
A partial-thickness epidermal explant model was colonized with green fluorescent protein (GFP)-expressing Staphylococcus aureus, and the pattern of S. aureus biofilm growth was characterized using electron and confocal laser scanning microscopy. The oxygen concentration in explants was quantified using microelectrodes. The relative effective diffusivity and porosity of the epidermis were determined using magnetic resonance imaging, while hydrogen peroxide (H2O2) concentration in explant media was measured by using microelectrodes. Secreted proteins were identified and quantified using elevated-energy mass spectrometry (MS(E)). S. aureus biofilm grows predominantly in lipid-rich areas around hair follicles and associated skin folds. Dissolved oxygen was selectively depleted (2- to 3-fold) in these locations, but the relative effective diffusivity and porosity did not change between colonized and control epidermis. Histological analysis revealed keratinocyte damage across all the layers of colonized epidermis after 4 days of culture. The colonized explants released significantly (P < 0.01) more antioxidant proteins of both epidermal and S. aureus origin, consistent with elevated H2O2 concentrations found in the media from the colonized explants (P< 0.001). Caspase-14 was also elevated significantly in the media from the colonized explants. While H2O2 induces primary keratinocyte differentiation, caspase-14 is required for terminal keratinocyte differentiation and desquamation. These results are consistent with a localized biological impact from S. aureus in response to colonization of the skin surface.
用表达绿色荧光蛋白（GFP）的金黄色葡萄球菌定殖部分厚度的表皮外植体模型，并用电子显微镜和共聚焦激光扫描显微镜表征金黄色葡萄球菌生物膜的生长模式。使用微电极对外植体中的氧浓度进行定量。利用磁共振成像确定表皮的相对有效扩散率和孔隙率，同时使用微电极测量外植体培养基中的过氧化氢（H2O2）浓度。使用高能质谱（MS(E)）鉴定和定量分泌蛋白。金黄色葡萄球菌生物膜主要生长在毛囊周围富含脂质的区域以及相关的皮肤褶皱处。这些部位的溶解氧被选择性消耗（2至3倍），但定殖表皮和对照表皮之间的相对有效扩散率和孔隙率没有变化。组织学分析显示，培养4天后，定殖表皮的所有层均出现角质形成细胞损伤。定殖的外植体释放出显著更多（P < 0.01）的源自表皮和金黄色葡萄球菌的抗氧化蛋白，这与定殖外植体培养基中发现的H2O2浓度升高一致（P < 0.001）。定殖外植体的培养基中半胱天冬酶-14也显著升高。虽然H2O2诱导初级角质形成细胞分化，但半胱天冬酶-14是终末角质形成细胞分化和脱屑所必需的。这些结果与金黄色葡萄球菌对皮肤表面定殖的局部生物学影响一致。
