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Journal of Biomedical Optics

Mechanistic study of apoptosis induced by high-fluence low-power laser irradiation using fluorescence imaging techniques
Author(s): Shengnan Wu; Xing Da; Fang Wang; Tongsheng Chen; Wei R. Chen
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Paper Abstract

Low-power laser irradiation (LPLI) can cause cell proliferation, differentiation, or death; however, the cellular mechanisms of these effects of LPLI, at high or low fluences, are not well known. To investigate the mechanism of high-fluence LPLI-induced apoptosis, both human lung adenocarcinoma cells (ASTC-a-1) and African green monkey SV40-transformed kidney fibroblast cells (COS-7) were irradiated with a He-Ne laser for 10 min under a fluence of 120 J/cm2 and 80 J/cm2, respectively. The dynamics of reactive oxygen species (ROS) generation was determined by measuring changes in fluorescence resulting from oxidation of intracellular dichlorodihydrofluorescein diacetate (H2DCFDA) to (DCF). The changes of mitochondrial membrane potential, ΔΨm, were studied by measuring the reduction of cellular fluorescence of Rhodamine 123 dyes using confocal laser scanning microscopy. The activation of caspase-3 in cells transfected by [SCAT3] reporters was observed using fluorescence resonance energy transfer (FRET) imaging. The activity of caspase-8 during high-fluence LPLI-induced apoptosis was studied by monitoring the cellular distribution of [Bid-CFP] reporters using fluorescence imaging. The following temporal sequence of cellular events was observed during apoptosis induced by high-fluence LPLI (120 J/cm2, ASTC-a-1 cells): (1) immediate generation of mitochondrial ROS following laser irradiation, reaching a maximum level 60 min after irradiation; (2) onset of ΔΨm decrease 15 min after laser irradiation, reaching a minimum level 50 min after irradiation; and (3) activation of caspase-3 between 30 min and 180 min after laser irradiation. Our results also show that the high-fluence LPLI does not activate caspase-8, indicating that the induced apoptosis was initiated directly from mitochondrial ROS generation andΔΨm decrease, independent of the caspase-8 activation.

Paper Details

Date Published: 1 November 2007
PDF: 10 pages
J. Biomed. Opt. 12(6) 064015 doi: 10.1117/1.2804923
Published in: Journal of Biomedical Optics Volume 12, Issue 6
Show Author Affiliations
Shengnan Wu, South China Normal Univ. (China)
Xing Da, South China Normal Univ. (China)
Fang Wang, South China Normal Univ. (China)
Tongsheng Chen, South China Normal Univ. (China)
Wei R. Chen, Univ. of Central Oklahoma (United States)

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