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.
The aging process is often associated with impaired wound healing, but the cellular and molecular mechanisms implicated are not completely understood. Accordingly, we have investigated the response of human fibroblasts from donors of various ages to platelet‐derived and autocrine growth factors, in terms of mitogenicity as well as extracellular matrix synthesis and degradation. Our data indicate that fibroblast responses persist during aging, suggesting the involvement of systemic factors in the regulation of the healing process. In this context, we have found that neutral endopeptidase‐24.11, a metalloproteinase controlling the action of neuroendocrine peptides and also of immunocyte chemotaxis, is overexpressed during aging. Finally, the connection between these data and those from in vitro aging studies is discussed.
Oxidative stress has been implicated in several biological and pathological aspects. Reactive oxygen species (ROS) have been proposed to act as signal transduction molecules activating reactions leading to cell rescue or to cell apoptosis/necrosis. In the present study, oxidative stress was induced by photosensitization of zinc phthalocyanine (ZnPc) in human fibroblasts using a photodynamic dose that did not lead to apoptosis or necrosis. The induction of oxidative stress was performed at the microscope stage in preassigned time. The cascade of phenomena evoked was studied in real time and at the single cell level using confocal laser scanning microscopy. Using specific vital fluorescent probes, alterations induced by oxidative stress in mitochondria membrane potential, in intracellular pH and in calcium concentration were recorded. Image processing and analysis techniques were used to quantify the observed changes. Subcellular localization of the photosensitizer was studied in order to determine the primary and immediate ROS target. It was found that ZnPc is mainly localized in the mitochondria region. Downloaded From: http://proceedings.spiedigitallibrary.org/ on 06/20/2016 Terms of Use: http://spiedigitallibrary.org/ss/TermsOfUse.aspx Proc. of SPIE Vol. 5142 3 Downloaded From: http://proceedings.spiedigitallibrary.org/ on 06/20/2016 Terms of Use: http://spiedigitallibrary.org/ss/TermsOfUse.aspx Proc. of SPIE Vol. 5142 7 Downloaded From: http://proceedings.spiedigitallibrary.org/ on 06/20/2016 Terms of Use: http://spiedigitallibrary.org/ss/TermsOfUse.aspx
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.