title: Low pulse energy Nd:YAG laser irradiation exerts a biostimulative effect on different cells of the oral microenvironment: "an in vitro study".
abstract: OBJECTIVE
  Dental lasers represent a promising therapeutic tool in the treatment of periodontal and peri-implant diseases. However, their clinical application remains still limited. Here, we investigated the potential biostimulatory effect of low pulse energy neodymium:yttrium-aluminum-garnet (Nd:YAG) laser irradiation on different cells representative of the oral microenvironment and elucidated the underlying molecular mechanisms.
  
  
  METHODS
  Saos-2 osteoblasts, H-end endothelial cells, and NIH/3T3 fibroblasts pre-treated or not with photosensitizing dye methylene blue (MB), were irradiated with low pulse energy (20 mJ) and high repetition rate (50-70 Hz) Nd:YAG laser, and evaluated for cell viability and proliferation as well as for the expression of specific differentiation markers by confocal immunofluorescence and real-time RT-PCR. Changes in intracellular Ca(2+) levels after laser exposure were also evaluated in living osteoblasts.
  
  
  RESULTS
  Nd:YAG laser irradiation did not affect cell viability in all the tested cell types, even when combined with pre-treatment with MB, and efficiently stimulated cell growth in the non-sensitized osteoblasts. Moreover, a significant induction in the expression of osteopontin, ALP, and Runx2 in osteoblasts, type I collagen in fibroblasts, and vinculin in endothelial cells could be observed in the irradiated cells. Pre-treatment with MB negatively affected cell differentiation in the unstimulated and laser-stimulated cells. Notably, laser irradiation also caused an increase in the intracellular Ca(2+) in osteoblasts through the activation of TRPC1 ion channels. Moreover, the pharmacologic or genetic inhibition of these channels strongly attenuated laser-induced osteopontin expression, suggesting a role for the laser-mediated Ca(2+) influx in regulating osteoblast differentiation.
  
  
  CONCLUSIONS
  Low pulse energy and high repetition rate Nd:YAG laser irradiation may exert a biostimulative effect on different cells representative of the oral microenvironment, particularly osteoblasts. Pre-treatment with MB prior to irradiation hampers this effect and limits the potential clinical application of photosensitizing dyes in dental practice.
authors.affiliation: Department of Anatomy, Histology and Forensic Medicine, University of Florence, Florence, Italy. f _ chellini@hotmail.com
authors.forename: Flaminia
authors.initials: F
authors.lastname: Chellini
authors.forename: Chiara
authors.initials: C
authors.lastname: Sassoli
authors.forename: Daniele
authors.initials: D
authors.lastname: Nosi
authors.forename: Cristiana
authors.initials: C
authors.lastname: Deledda
authors.forename: Paolo
authors.initials: P
authors.lastname: Tonelli
authors.forename: Sandra
authors.initials: S
authors.lastname: Zecchi-Orlandini
authors.forename: Lucia
authors.initials: L
authors.lastname: Formigli
authors.forename: Marco
authors.initials: M
authors.lastname: Giannelli
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D002118:Calcium / Q000378:metabolism; D049109:Cell Proliferation / Q000528:radiation effects; D002470:Cell Survival / Q000528:radiation effects; D002478:Cells, Cultured; D024042:Collagen Type I / Q000378:metabolism; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism / Q000528:radiation effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism / Q000528:radiation effects*; D005455:Fluorescent Antibody Technique; D006801:Humans; D053844:Lasers, Solid-State; D028022:Low-Level Light Therapy; D008751:Methylene Blue / Q000494:pharmacology; D051379:Mice; D009061:Mouth Mucosa / Q000166:cytology*; D010006:Osteoblasts / Q000378:metabolism / Q000528:radiation effects*; D053495:Osteopontin / Q000378:metabolism; D016133:Polymerase Chain Reaction; D050052:TRPC Cation Channels / Q000502:physiology; D016596:Vinculin / Q000378:metabolism
chemical_list: D024042:Collagen Type I; D050659:Core Binding Factor Alpha 1 Subunit; D004791:Enzyme Inhibitors; D050052:TRPC Cation Channels; C096351:transient receptor potential cation channel, subfamily C, member 1; D053495:Osteopontin; D016596:Vinculin; D000469:Alkaline Phosphatase; D002118:Calcium; D008751:Methylene Blue