Biostimulation Effects Of Low Power Laser At Wavelengths Of 592nm And 630nm Irradiation On Human Skin Fibroblasts

Objects:To detect the biostimulation effects of Low Power Laser at wavelengths of 590 nm and 630 nm on proliferation and procollagen gene expression of human skin fibroblasts, as well as SMADs gene expression, so as to provide theoretical evidence and possible parameters for clinical improvement of Low Power Laser therapy in aging akin at wavelengths of 590 nm and 630 nm.Methods:1. Select cultured adult and aging skin fibroblasts as cell model. To test the stimulation effects on cell proliferation, cultured fibroblasts were treated in a controlled manner during three consecutive days, either with a yellow laser or with a red laser source emitting two wavelengths(592 nm,630 nm) and two level power density outputs(3.5mw/cm2,10.6mw/cm2).Treatment duration including 2min,5min and 10min varied in relation to varying surface power densities (radiant exposures). To test gene expression, cultured fibroblasts were treated in a controlled manner during three consecutive days, either with a yellow laser or with a red laser source emitting two wavelengths(592nm,630nm) and two level power density outputs(3.5 mw/cm2,10.6 mw/cm2). Treatment duration including 5min and 10min varied in relation to varying surface power densities (radiant exposures).2. Each time before and after 24 h of irradiation, cells were photographed using a phase-contrast microscope, to observe the variation of cell morphous and viabilities.3. Using MTT method to test cell proliferation status.4. Utilizing RT-PCR technology to detect gene expressions of procollagen typeⅠ-α1,Ⅰ-α2, typeⅢ-α2 and SMAD2,3. Measure the optical density values with an analysis software Bandscan5.0 of each PCR product after electrophoresis.5. Utilizing SPSS11.5 statistical analysis software to analyze data. Results:1.Statistical analysis revealed a higher rate of proliferation (p<0.005) in all irradiated cultures in comparison with the controls. Low power density of both 592 nm and 630 nm (3.5mw/cm2) lasers yielded a significantly higher number of cells, than high power density (10.6 mw/cm2) lasers (p<0.005).2. Statistical analysis revealed a higher rate of proliferation (p<0.005) in all irradiated cultures in comparison with the controls.5 minutes' irradiation of both 592 nm and 630 nm lasers yielded a significantly higher number of cells, than 2 minutes' irradiation and 10 minutes'by turns.3. Statistical analysis revealed a higher rate of gene express (p<0.005) in those irradiated cultures at low power density (3.5mw/cm2) in comparison with the controls. Both 592 nm and 630 nm lasers yielded a significantly higher expression of procollagen typeⅠ-α1,Ⅰ-α2, typeⅢ-α2 and SMAD2,3, than high power density(10.6mw/cm2) lasers.5 minutes' irradiation of both 592 nm and 630 nm (3.5 mw/cm2) lasers yielded a significantly higher gene expression thanlO minutes' irradiation.4. Statistical analysis revealed a higher rate of gene expression (p<0.005) in those irradiated cultures under high power density (10.6 mw/cm2) in comparison with the controls. High power density (10.6mw/cm2) of both 592 nm and 630 nm lasers yielded a significantly higher expression of gene after irradiation 5 min than 10 min. There is no statistical significance (p>0.05) of gene expression increase in comparison with the controls when irradiated for 10 min at high power density (10.6mw/cm2) lasers with wavelengths of 592 nm and 630 nm.Conclusions:1.590 nm and 630 nm low power lasers irradiation resulted in an increased human fibroblast proliferation in vitro, and the trend varied between different power densities and irradiation duration times. Low power density (3.5mw/cm2) lasers yielded higher proliferation rate than high power density (10.6mw/cm2) lasers. Shorter irradiation duration time (5min) lead to higher proliferation rate than longer irradiation duration time (10min).2. The increase of gene expression of SMADs suggesting that the secretions of TGF-βby fibroblasts were increased.3. Base on the evidence of stimulated proliferation and gene expression and their penetration depth of each wavelength at 592 nm and 630 nm, this study therefore postulates possible improvements on wrinkle in vivo at the applied dosimetric parameters.4. Since the fibroblasts irradiated by 630nm laser were derived from aging skin, there is greater clinical significance based on the result of this study.