Evaluation of the oxidative stress of psoriatic fibroblasts based on spectral two-photon fluorescence lifetime imaging
Authors: Kapsokalyvas D., Barygina V., Cicchi R., Fiorillo C., Pavone FS.
Autors Affiliation: European Laboratory for Non-linear Spectroscopy (LENS), University of Florence, Via Nello Carrara 1, 50019, Sesto Fiorentino, Italy
Department of Biochemical Sciences, University of Florence, Viale GB Morgagni 50, 50134, Florence, Italy
National Institute of Optics, National Research Council, Largo E. Fermi 6, 50125, Florence, Italy
Abstract: Psoriasis is an autoimmune disease of the skin characterized by hyperkeratosis, hyperproliferation of the epidermis, inflammatory cell accumulation and increased dilatation of dermal papillary blood vessels. Metabolic activity is increased in the epidermis and the dermis. Oxidative stress is high mainly due to reactive oxygen species (ROS) originating from the skin environment and cellular metabolism. We employed a custom multiphoton microscope coupled with a FLIM setup to image primary culture fibroblast cells from perilesional and lesional psoriatic skin in-vitro. Twophoton excited fluorescence images revealed the morphological differences between healthy and psoriatic fibroblasts. Based on the spectral analysis of the NADH and FAD components the oxidative stress was assessed and found to be higher in psoriatic cells. Furthermore the fluorescence lifetime properties were investigated with a TCSPC FLIM module. Mean fluorescence lifetime was found to be longer in psoriatic lesional cells. Analysis of the fast (?1) and slow (?2) decay lifetimes revealed a decrease of the ratio of the contribution of the fast (a1) parameter to the contribution of the slow (a2) parameter. The fluorescence in the examined part of the spectrum is attributed mainly to NADH. The decrease of the ratio (a1)/ (a2) is believed to correlate strongly with the anti-oxidant properties of NADH which can lead to the variation of its population in high ROS environment. This methodology could serve as an index of the oxidative status in cells and furthermore could be used to probe the oxidative stress of tissues in-vivo.
Conference title: Photonics West – BIOS 2013
Place: San Francisco, US
More Information: The research leading to these results has received funding from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement n 228334 and n 284464 (Bioptichal) and from the Italian Ministry for education, University and Research in the framework of the Flagship Project NANOMAX. Financial support by the Ente Cassa di Risparmio di Firenze (private foundation) is acknowledged.KeyWords: Autofluorescence; FLIM; Fluorescence lifetimes; Multi-photon microscopy; NADH; Psoriasis; Redox ratio, Blood vessels; Cell culture; Dermatology; Fibroblasts; Oxidative stress; Spectrum analysis; Tissue, FluorescenceDOI: 10.1117/12.2001508Citations: 2data from “WEB OF SCIENCE” (of Thomson Reuters) are update at: 2019-12-01References taken from IsiWeb of Knowledge: (subscribers only)