TY - JOUR
T1 - In vitro study of extracellular vesicles migration in cartilage-derived osteoarthritis samples using real-time quantitative multimodal nonlinear optics imaging
AU - Mortati, Leonardo
AU - de Girolamo, Laura
AU - Orfei, Carlotta Perucca
AU - Viganò, Marco
AU - Brayda-Bruno, Marco
AU - Ragni, Enrico
AU - Colombini, Alessandra
N1 - Funding Information:
Funding: This research was funded by the Italian Ministry of Health, “Ricerca Corrente”.
Publisher Copyright:
© 2020 by the authors. Licensee MDPI, Basel, Switzerland.
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/8
Y1 - 2020/8
N2 - Mesenchymal stromal cells (MSCs)-derived extracellular vesicles (EVs) are promising therapeutic nano-carriers for the treatment of osteoarthritis (OA). The assessment of their uptake in tissues is mandatory but, to date, available technology does not allow to track and quantify incorporation in real-time. To fill this knowledge gap, the present study was intended to develop an innovative technology to determine kinetics of fluorescent MSC-EV uptake by means of time-lapse quantitative microscopy techniques. Adipose-derived mesenchymal stromal cells (ASCs)-EVs were fluorescently labeled and tracked during their uptake into chondrocytes micromasses or cartilage explants, both derived from OA patients. Immunofluorescence and time-lapse coherent anti-Stokes Raman scattering, second harmonic generation and two-photon excited fluorescence were used to follow and quantify incorporation. EVs penetration appeared quickly after few minutes and reached 30–40 µm depth after 5 h in both explants and micromasses. In explants, uptake was slightly faster, with EVs signal overlapping both extracellular matrix and chondrocytes, whereas in micromasses a more homogenous diffusion was observed. The finding of this study demonstrates that this innovative technology is a powerful tool to monitor EVs migration in tissues characterized by a complex extracellular network, and to obtain data resembling in vivo conditions.
AB - Mesenchymal stromal cells (MSCs)-derived extracellular vesicles (EVs) are promising therapeutic nano-carriers for the treatment of osteoarthritis (OA). The assessment of their uptake in tissues is mandatory but, to date, available technology does not allow to track and quantify incorporation in real-time. To fill this knowledge gap, the present study was intended to develop an innovative technology to determine kinetics of fluorescent MSC-EV uptake by means of time-lapse quantitative microscopy techniques. Adipose-derived mesenchymal stromal cells (ASCs)-EVs were fluorescently labeled and tracked during their uptake into chondrocytes micromasses or cartilage explants, both derived from OA patients. Immunofluorescence and time-lapse coherent anti-Stokes Raman scattering, second harmonic generation and two-photon excited fluorescence were used to follow and quantify incorporation. EVs penetration appeared quickly after few minutes and reached 30–40 µm depth after 5 h in both explants and micromasses. In explants, uptake was slightly faster, with EVs signal overlapping both extracellular matrix and chondrocytes, whereas in micromasses a more homogenous diffusion was observed. The finding of this study demonstrates that this innovative technology is a powerful tool to monitor EVs migration in tissues characterized by a complex extracellular network, and to obtain data resembling in vivo conditions.
KW - Cartilage
KW - Coherent anti-stokes raman scattering
KW - Extracellular vesicles
KW - Mesenchymal stem cells
KW - Microscopy
KW - Osteoarthritis
KW - Second harmonic generation
KW - Time-lapse
KW - Two-photon excitation fluorescence
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U2 - 10.3390/pharmaceutics12080734
DO - 10.3390/pharmaceutics12080734
M3 - Article
AN - SCOPUS:85090682997
VL - 12
SP - 1
EP - 18
JO - Pharmaceutics
JF - Pharmaceutics
SN - 1999-4923
IS - 8
M1 - 734
ER -