Articular cartilage in the tibiofemoral joint contains unique tissue microstructures that serve specific functions, including reduction of friction and distributing the dynamic and static cyclic loading at the ends of diarthrodial joints. A proficient understanding of these microstructures can lead to significant clinical advances in diagnosing orthopedic diseases such as osteoarthritis and improving cartilage repairs. The surface of tibiofemoral condyles can be roughly separated into loadbearing and meniscus-covered areas. Due to the difference in mechanical loading between the two regions, we hypothesize that their microstructures differ. To test this hypothesis, we used cartilage punches harvested from the tibial condyle of porcine knee joints as an example tissue and a custom nonlinear optical microscope for performing a dye-free imaging study. The custom nonlinear optical microscope could simultaneously acquire Two-Photon excitation Auto-Fluorescence (TPAF) and Second Harmonic Generation (SHG) images. Through the TPAF channel, elastin fibers are visible along with chondrocytes. The SHG channel was utilized for observing the vast collagen network and its evident orientation throughout the tibial condyle. Images were analyzed by ImageJ to reveal alignment angles of the collagen network and elastin fibers. The load-bearing region exhibits a denser uniform collagen network with minimum elastin fibers. In contrast, the meniscuscovered areas have a distinctive collagen orientation with a greater magnitude of co-localized elastin fibers. The biological differences are likely derived from their different biomechanical environments in the tibiofemoral joint.
Visualization of collagen fibers in cardiac tissues is essential for clinical diagnosis and pathological analysis of cardiac fibrosis. Selecting a proper imaging method is still challenging for researchers and clinicians who want to determine specific information about the collagen network in cardiac tissues. We examined fibrillar collagen network from mouse ventricular myocardium by commonly available light microscopy techniques using our home-built multimodal microscope. Myocardial slices were unstained or stained with either Picrosirius red or collagen type I antibody/dye conjugation, then imaged by polarized light, confocal fluorescence, second harmonic generation (SHG), two-photon excited fluorescence (TPEF), and stimulated emission depletion (STED) microscopy techniques. This study is intended to serve as a reference for basic research and clinical evaluation of fibrillar collage network in cardiac tissues.
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