Noninvasive evaluation of tissue-engineered cartilage with time-resolved laser-induced fluorescence spectroscopy.
Regenerative medicine requires noninvasive evaluation. Our objective is to investigate the application of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) using a nano-second-pulsed laser for evaluation of tissue-engineered cartilage (TEC). To prepare scaffold-free TEC, articular chondrocytes from 4-week-old Japanese white rabbits were harvested, and were inoculated at a high density in a mold. Cells were cultured for 5 weeks by rotating culture (RC) or static culture (SC). The RC group and SC group at each week (n = 5), as well as normal articular cartilage and purified collagen type II (as controls), were analyzed by TR-LIFS. The peak wavelength was compared with those of type II collagen immunostaining and type II collagen quantification by enzyme-linked immunosorbent assay and tensile testing. The fluorescence peak wavelength of the TEC analyzed by this method shifted significantly in the RC group at 3 weeks, and in the SC group at 5 weeks (p < 0.01). These results correlated with changes in type II collagen (enzyme-linked immunosorbent assay) and changes in Young's modulus on tensile testing. The results were also supported by immunohistologic findings (type II collagen staining). Our findings show that TR-LIFS is useful for evaluating TEC.
Kutsuna
Toshiharu
T
Department of Orthopaedic Surgery, Tokai University School of Medicine, Kanagawa, Japan.
Sato
Masato
M
Ishihara
Miya
M
Furukawa
Katsuko S
KS
Nagai
Toshihiro
T
Kikuchi
Makoto
M
Ushida
Takashi
T
Mochida
Joji
J
D000818:Animals; D002358:Cartilage, Articular / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D002478:Cells, Cultured; D024043:Collagen Type II / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D007150:Immunohistochemistry; D011817:Rabbits; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013718:Tensile Strength; D023822:Tissue Engineering
D024043:Collagen Type II