17 June 2002 Identification of chondrocyte cell division in intact rabbit cartilage grafts following photothermal heating using fluorescence-activated cell counting
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Proceedings Volume 4609, Lasers in Surgery: Advanced Characterization, Therapeutics, and Systems XII; (2002) https://doi.org/10.1117/12.441077
Event: International Symposium on Biomedical Optics, 2002, San Jose, CA, United States
Abstract
The lack of adequate cartilage for aesthetic and reconstructive head and neck surgery has prompted intense research in developing methods to stimulate chondrocyte proliferation in cartilage, a terminally differentiated tissue. In this study, we propose that laser heating can stimulate chondrocyte proliferation in rabbit nasal septal cartilage. Ex-vivo rabbit nasal septal cartilage grafts were irradiated using a CO2 laser (?=10.6 um, 5, 10, or 20 W, 0.1 or 0.5 seconds) and then placed in tissue culture for 7-21 days with 10 _M of 5-bromo-2’-deoxyuridine (BrdU), a thymidine analog that is incorporated into DNA during the S phase of the cell cycle. Chondrocytes were then isolated using enzymatic digestion, incubated with fluorescein conjugated anti-BrdU monoclonal antibodies (Roche Diagnostic, Basel, Switzerland) and counterstained with propidium iodide (PI), a DNA intercalater.? Fluorescent-activated cell counting was performed by flow cytometry (excitation wavelength ?=488 nm, Model BD LSR Flow Cytometer, Becton Dickson, Franklin Lakes, NJ) identifying cells with emissions measured at 515 nm (fluorescein labeled anti-BrdU) and 620 nm (PI).? Cellular debris and cell aggregates were gated out of the signal.?Cartilage grafts irradiated approximately 24 times with the CO2 laser were found to have a significant proliferating population by flow cytometry. Chondrocytes incorporating BrdU into their DNA were clearly identified and the signal intensity increased in integral multiples from baseline values indicating these cells had undergone cell division.? This provides further evidence that chondroctyes have a proliferative response to photothermal stimulation, which may be a response to injury to thermal heating by laser irradiation.
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Mai Thy Truong, Kenneth K. H. Chao, Nidhi S. Pandoh, Fang-Yao Hou, David M. Gardiner, Brian Jet-Fei Wong, "Identification of chondrocyte cell division in intact rabbit cartilage grafts following photothermal heating using fluorescence-activated cell counting", Proc. SPIE 4609, Lasers in Surgery: Advanced Characterization, Therapeutics, and Systems XII, (17 June 2002); doi: 10.1117/12.441077; https://doi.org/10.1117/12.441077
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