Merigo Elisabetta, Chevalier Marlène, Conti Stefania, Ciociola Tecla, Fornaini Carlo, Manfredi Maddalena, Vescovi Paolo, Doglio Alain
Department of Medicine and Surgery, University of Parma, Italy.
Micoralis Laboratory, Faculty of Dentistry, University of Nice Sophia Antipolis, Nice, France.
Laser Ther. 2019 Sep 30;28(3):180-186. doi: 10.5978/islsm.28_19-OR-14.
The aim of this study was to test the application of different laser wavelengths at a low fluence in combination or not with proper photosensitizing dyes on biofilm with or without a synthetic killer decapeptide (KP). SC5314 was grown on Sabouraud dextrose agar plates at 37°C for 24 h. Cells were suspended in RPMI 1640 buffered with MOPS and cultured directly on the flat bottom of 96-wells plates. The previously described killer decapeptide KP was used in this study. Three different combinations of wavelengths and dyes were applied, laser irradiation has been performed at a fluence of 10 J/cm. The effect on biofilm was evaluated by the XTT assay. Microscopic observations were realized by fluorescence optic microscopy with calcofluor white and propidium iodide. Compared with control, no inhibition of biofilm viability was obtained with application of red, blue and green lasers alone or with any combination of red diode laser, toluidine blue and KP. The combined application of blue diode laser with curcumin and/or KP showed always a very significant inhibition, as curcumin alone and the combination of curcumin and KP did, while combination of blue diode laser and KP gave a less significant inhibition, the same obtained with KP alone. The combined application of green diode laser with erythrosine and/or KP showed always a very significant inhibition, as the combination of erythrosine and KP did, but no difference was observed with respect to the treatment with erythrosine alone. Again, combination of green diode laser and KP gave a significant inhibition, although paradoxically lower than the one obtained with KP alone. Treatment with KP alone, while reducing biofilm viability did not cause death in the adopted experimental conditions. On the contrary, combined treatment with blue laser, curcumin and KP, as well as green laser, erythrosine and KP led to death most cells. The combination of laser light at a fluence of 10 J/cm and the appropriate photosensitizing agent, together with the use of KP, proved to exert differential effects on biofilm.
本研究的目的是测试不同激光波长在低能量密度下，与或不与合适的光敏染料联合应用于有或没有合成杀伤十肽（KP）的生物膜上的效果。将白色念珠菌SC5314在37°C的沙氏葡萄糖琼脂平板上培养24小时。细胞悬浮于用MOPS缓冲的RPMI 1640中，并直接在96孔板的平底上培养。本研究使用了先前描述的杀伤十肽KP。应用了三种不同的波长和染料组合，激光照射能量密度为10 J/cm²。通过XTT法评估对生物膜的影响。通过用荧光增白剂和碘化丙啶的荧光光学显微镜进行微观观察。与对照组相比，单独应用红色、蓝色和绿色激光，或红色二极管激光、甲苯胺蓝和KP的任何组合，均未对生物膜活力产生抑制作用。蓝色二极管激光与姜黄素和/或KP的联合应用始终显示出非常显著的抑制作用，单独使用姜黄素以及姜黄素与KP的组合也是如此，而蓝色二极管激光与KP的组合产生的抑制作用较小，单独使用KP时也是如此。绿色二极管激光与赤藓红和/或KP的联合应用始终显示出非常显著的抑制作用，赤藓红与KP的组合也是如此，但与单独使用赤藓红处理相比未观察到差异。同样，绿色二极管激光与KP的组合产生了显著的抑制作用，尽管自相矛盾的是，其低于单独使用KP时的抑制作用。单独使用KP处理，虽然降低了生物膜活力，但在所采用的实验条件下并未导致细胞死亡。相反，蓝色激光、姜黄素和KP以及绿色激光、赤藓红和KP的联合处理导致大多数细胞死亡。能量密度为10 J/cm²的激光与合适的光敏剂联合使用，再加上使用KP，对生物膜产生了不同的影响。
