Multiphoton imaging and quantitative analysis of collagen production by chondrogenic human mesenchymal stem cells cultured in chitosan scaffold.
We used the combined imaging modality of multiphoton autofluorescence and second-harmonic generation microscopy to investigate the chondrogenic process of human mesenchymal stem cells cultured in chitosan scaffold. Isolated human mesenchymal stem cells seeded onto chitosan scaffold were induced to undergo chondrogenesis by addition of the transforming growth factor-&#x03B2;3. After continuous culturing, the engineered tissues at the same scaffold location were imaged at different time points for up to 49 days. Using the acquired images of the chondrogenic process, we quantify tissue morphogenesis by monitoring the changes in multiphoton autofluorescence and second-harmonic generation signals from the engineered tissues. We found that the extracellular matrix generation can be modeled by an exponential function during the initial growth stage and that saturation occurs between days 11 and 14. Further, the growth rate of the extracellular matrix was found to increase toward the surface of the chitosan scaffold. Our work demonstrates the use of multiphoton microscopy for performing long-term monitoring and quantification of the tissue engineering process.
Chen
Wei-Liang
WL
Department of Physics, National Taiwan University, Taipei, Taiwan.
Huang
Chi-Hsiu
CH
Chiou
Ling-Ling
LL
Chen
Te-Hsuen
TH
Huang
Yi-You
YY
Jiang
Ching-Chuan
CC
Lee
Hsuan-Shu
HS
Dong
Chen-Yuan
CY
D002478:Cells, Cultured; D048271:Chitosan; D003094:Collagen / Q000096:biosynthesis*; D003470:Culture Media; D005109:Extracellular Matrix / Q000378:metabolism; D005453:Fluorescence; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D023822:Tissue Engineering
D003470:Culture Media; D003094:Collagen; D048271:Chitosan