The Impact of Cellular Debris on Pseudomonas aeruginosa Adherence to Silicone Hydrogel Contact Lenses and Contact Lens Storage Cases
The Impact of Cellular Debris on Pseudomonas aeruginosa Adherence to Silicone Hydrogel Contact Lenses and Contact Lens Storage Cases
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DOI：
10.1097/icl.0b013e31823bad0e
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发表时间：
2012-01-01
影响因子：
2.3
通讯作者：
Robertson, Danielle M.
中科院分区：
文献类型：
--
作者：
Burnham, Geoffrey W.;Cavanagh, Dwight;Robertson, Danielle M.
目的：评价硅胶隐形眼镜上中性粒细胞增强的铜绿假单胞菌(PA)生物膜的形成情况，以及角膜上皮细胞生物碎片对PA黏附的影响。未佩戴的洛曲菲康A隐形眼镜在37℃下以不同比例的PA与中性粒细胞(PMN)孵育24小时，用激光扫描共聚焦显微镜评价晶状体相关PA，并用碘化丙啶显示无活性PA。通过菌落形成单位(CFU)分析进行活菌计数。在急性上皮细胞研究中，PA活性在与冻融上皮细胞裂解物在96孔聚苯乙烯平板中共同孵育后测定。结果：激光扫描共聚焦显微镜显示，与PMA刺激的中性粒细胞共同处理后，PA在24小时内增加了PA与晶状体表面的粘附性，并显著改变了PA的结构。碘化丙啶染色显示粘附菌由活性PA和非活性PA组成；CFU证实活性PA与中性粒细胞相关增加(PA：PMN0.1：1，P=0.025；PA：PMN1：1，P=0.005)。急性上皮细胞碎片研究显示，96孔板中活性PA显著增加(PA：碎片1：1，P=0.002；PA：碎片100：1，P=0.002)。结晶紫染色显示所有时间点都有残留的细胞碎片，这与微生物污染无关；所有使用9个月或更长时间的镜盒均与高水平的活菌群一致相关。结论：这些结果表明，当面临延长配戴的PA挑战时，长期的角膜炎症和PMN的存在可能会刺激硅胶水凝胶隐形眼镜上生物膜的形成。这些发现进一步表明，晶状体储存情况下细胞外碎片的持续堆积可能是这些表面报告的厚重生物膜的原因之一。
Objectives: To evaluate neutrophil-enhanced Pseudomonas aeruginosa (PA) biofilm formation on silicone hydrogel contact lenses and to determine the effect of epithelial biodebris on PA adherence in contact lens storage cases.Methods: A fully invasive PA corneal isolate stably conjugated to green fluorescent protein was used. Unworn lotrafilcon A contact lenses were incubated at various ratios of PA to polymorphonuclear neutrophil (PMN) for 24 hours at 37 degrees C. Lens-associated PA was evaluated using laser scanning confocal microscopy and nonviable PA were visualized using propidium iodide. Viable bacteria were enumerated by colony-forming unit (CFU) analysis. For acute epithelial cell studies, PA viability was determined after coincubation with freeze-thaw epithelial cell lysates in 96-well polystyrene plates. Levels of residual cellular debris and bacterial viability were further assessed in used contact lens storage cases.Results: Laser scanning confocal microscopy demonstrated that cotreatment with PMA-stimulated neutrophils increased PA adherence over 24 hours to lens surfaces with a striking alteration of PA architecture. Propidium iodide staining showed that the adherent bacteria consisted of a mixture of viable and nonviable PA; a PMN-associated increase in viable PA was confirmed by CFU (PA: PMN 0.1:1, P = 0.025; PA:PMN 1: 1, P = 0.005). Acute epithelial cell debris studies revealed a significant increase in viable PA in 96-well plates in the presence of epithelial freeze-thaw lysates (PA: debris 1:1, P = 0.002; PA:debris 100:1, P = 0.002). Crystal violet staining of used lens storage cases revealed residual cellular debris at all time points, which was independent of microbial contamination; all lens cases used for periods of 9 months or more were uniformly associated with high levels of viable microorganisms.Conclusion: These results demonstrate that prolonged corneal inflammation with the presence of PMNs when confronted with simultaneous PA challenge in extended contact lens wear has the potential to stimulate biofilm formation on silicone hydrogel contact lenses. These findings further suggest that a persistent buildup of extracellular debris in lens storage cases may contribute to the heavy biofilms reported on these surfaces.