Monitoring Wound Healing Of Elastic Cartilage By Using Multiphoton Microscopy

Multiphoton microscopy (MPM) based on second harmonic generation (SHG) and two-photon excited fluorescence (TPEF), with the advantages of reduced specimen photo-bleaching, enhanced penetration depth and high resolution, has the ability to image cellular and subcellular microstructure of tissues without additional exogenous fluorophores and provide the particular emission spectra information from biological tissue. The objective of this study was to demonstrate the ability of MPM for monitoring morphological characterization and wound healing monitoring of elastic cartilage. Firstly, the morphology and distribution of main components in elastic cartilage including chondrocytes, collagen and elastic fibers were clearly observed from the high-resolution MPM images. The spectral characteristics from elastic cartilage were also obtained and analyzed by using MPM. And then, MPM was employed to image and quantify the elastic cartilage regeneration tissue in a rabbit ear model. And the morphological images of normal elastic cartilage and elastic cartilage regeneration tissue were compared. Morphology of cells and collagen matrix were analyzed, showing significant difference between regenerated and intact cartilage in cellular size and collagen distribution. At last, MPM was employed to provide high-resolution images of elastic cartilage at varying healing time points. Comparisons of the images of specimens at different healing stages showed obvious cell growth and matrix deposition. The results are consistent with the histological results. Moreover, quantitative analysis results show significant alteration in the collagen cavity size or collagen orientation index during wound healing of elastic cartilage, indicating the possibility to act as indicators for monitoring wound healing. Our results suggested that MPM has the ability to monitor the wound healing progression of elastic cartilage. With additional development, this technique may be developed into an effective clinical diagnostic and monitoring tool for establishment of novel and effective therapies in plastic surgery for accelerating cartilage repair.