X. Zhu, Y. Tang, J. Chen, S. Xiong, S. Zhuo, J. Chen

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Presentation transcript:

Monitoring wound healing of elastic cartilage using multiphoton microscopy X. Zhu, Y. Tang, J. Chen, S. Xiong, S. Zhuo, J. Chen Osteoarthritis and Cartilage Volume 21, Issue 11, Pages 1799-1806 (November 2013) DOI: 10.1016/j.joca.2013.08.016 Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 1 Schematic illustration of a wound model of elastic cartilage in the rabbit external ear. The region marked with dot lines indicates the ROI. Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 2 (a) Schematic of elastic cartilage structure in the rabbit external ear, showing chondrocytes and the sparse amount of intercellular substance mainly including collagen and elastic fibers. (b) Representative SHG image of collagen and (c) TPEF image from an intact elastic cartilage showing elastic fibers (white arrows) and pericellular matrix (yellow arrows). The dark regions in (b) and (c) represent the locations of the chondrocytes, labeled by * for an example. (d) 12 sequential SHG/TPEF overlapping images at intervals of 2 μm, providing the 3D shape of chondrocytes and the morphology of the surrounding collagen and elastic fibers. The arrow points to a chondrocyte to visualize the shape varied with depth. Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 3 High-resolution large-area SHG/TPEF images obtained by MPM from wound margins on (a) 1-day, (b) 1-, (c) 4-, and (d) 20-week repair time points, respectively. SHG images of collagen (green) are extracted from 398 to 409 nm, while TPEF images of elastic and cells (red) are extracted from 430 to 697 nm. The excited wavelength is 810 nm. The images are obtained at 3.2 μm per pixel. Scale bar is 100 μm. Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 4 TPEF images (a–d) from elastin and cells, SHG images (e–h) from collagen, and overlapped SHG/TPEF images (230.8 × 230.8 μm with 512 × 512 pixels spatial resolution) (i–l) from 1-, 4-, 20-week repair and intact samples are listed for analysis. White circles in SHG images are drawed to roughly measure the cavity sizes, where I and A represent the length and area of the circle, respectively. The SHG/TPEF images are obtained at 6.4 μm per pixel. Scale bar is 50 μm. In addition, the corresponding H&E stained images (×20) (m–p) are presented for comparison. Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 5 (a) A binary image of the 2D SHG image of center chondrocytes in Fig. 4(h), separating the collagen from the cavities. (b) Collagen cavity sizes from 1-, 4-, 20-week wound healing and intact samples are determined for quantification, showing increasing tendency of collagen cavity size during wound healing. It shows significant difference between any two groups (P < 0.05). Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions

Fig. 6 (a) Schematic of the collagen orientation index evaluation (1 − W/L), representatively presenting the FFT result of Fig. 4(g) fitted with a 95% confidence ellipse (red circle) combined with its width (W) and length (L) (red arrows). (b) Collagen orientation indexes from 1-, 4-, 20-week wound healing and intact samples are determined for quantification, showing decreasing tendency of collage orientation during wound healing. It shows significant difference between any two groups (P < 0.05). Osteoarthritis and Cartilage 2013 21, 1799-1806DOI: (10.1016/j.joca.2013.08.016) Copyright © 2013 Osteoarthritis Research Society International Terms and Conditions