Human fibroblast alterations induced by low power laser irradiation at the single cell level using confocal microscopy.
Low power laser irradiation is regarded to have a significant role in triggering cellular proliferation and in treating diseases of diverse etiologies. The present work contributes to the understanding of the mechanisms of action by studying low power laser effects in human fibroblasts. Confocal laser scanning microscopy is used for irradiation and observation of the same area of interest allowing the imaging of laser effects at the single cell level and in real time. Coverslip cultures were placed in a small incubation chamber for in vivo microscopic observation. Laser stimulation of the cells was performed using the 647 nm line of the confocal laser through the objective lens of the microscope. Mitochondrial membrane potential (delta psi(m)), intracellular pH, calcium alterations and generation of reactive oxygen species (ROS) were monitored using specific fluorescent vital probes. The induced effects were quantified using digital image processing techniques. After laser irradiation, a gradual alkalinization of the cytosolic pH and an increase in mitochondrial membrane potential were observed. Recurrent spikes of intracellular calcium concentration were also triggered by laser. Reactive oxygen species were generated as a result of biostimulation. No such effects were monitored in microscopic fields other than the irradiated ones.
Alexandratou
Eleni
E
National Technical University of Athens, Department of Electrical Engineering & Computing, Applied Biophysics and Biomedical Optics Laboratory, 157 73 Zografou Campus, Athens, Greece. ealexan@central.ntua.gr
Yova
Dido
D
Handris
Panagiotis
P
Kletsas
Dimitris
D
Loukas
Spyros
S
D002478:Cells, Cultured; D004307:Dose-Response Relationship, Radiation; D005347:Fibroblasts / Q000378:metabolism / Q000502:physiology / Q000528:radiation effects*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000528:radiation effects; D007834:Lasers; D008564:Membrane Potentials / Q000528:radiation effects; D018613:Microscopy, Confocal; D017382:Reactive Oxygen Species
D017382:Reactive Oxygen Species