Objective assessment of endogenous collagen in vivo during tissue repair by laser induced fluorescence

Collagen, a triple helical protein with the primary role of mechanical function, provides tensile strength to the skin, and plays a pivotal task in tissue repair. During tissue regeneration, collagen level increases gradually and therefore, monitoring of such changes in vivo by laser induced fluores...

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Published in:	PloS one Vol. 9; no. 5; p. e98609
Main Authors:	Prabhu, Vijendra, Rao, Satish B S, Fernandes, Edward Mark, Rao, Anuradha C K, Prasad, Keerthana, Mahato, Krishna K
Format:	Journal Article
Language:	English
Published:	United States Public Library of Science 29-05-2014 Public Library of Science (PLoS)
Subjects:	Animal tissues Animals Biology and Life Sciences Biophysics Birefringence Collagen Collagen - metabolism Collagens Disease Models, Animal Female Fluorescence Fluorometry Granulation Granulation Tissue - metabolism Image analysis Image processing In vivo methods and tests Irradiation Laser induced fluorescence Lasers Life sciences Light microscopy Male Mechanical properties Medicine and Health Sciences Metabolism Mice Microscopy Microscopy, Polarization Monitoring Optical Imaging Physical Sciences Polarized light Principal components analysis Regeneration Repair Satellite radio Skin Skin - pathology Skin - radiation effects Spectrum analysis Studies Synthesis Tissue engineering Wound healing Wound Healing - physiology Wounding India
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Summary:	Collagen, a triple helical protein with the primary role of mechanical function, provides tensile strength to the skin, and plays a pivotal task in tissue repair. During tissue regeneration, collagen level increases gradually and therefore, monitoring of such changes in vivo by laser induced fluorescence was the main objective behind the present study. In order to accomplish this, 15 mm diameter excisional wounds were created on six to eight week old Swiss albino mice. The collagen deposition accelerated upon irradiation of single exposure of 2 J/cm2 He-Ne laser dose immediately after wounding was recorded by laser induced autofluorescence in vivo along with un-illuminated and un-wounded controls. Autofluorescence spectra were recorded for each animal of the experimental groups on 0, 5, 10, 30, 45 and 60 days post-wounding, by exciting the granulation tissue/skin with 325 nm He-Cd laser. The variations in the average collagen intensities from the granulation tissue/skin of mice were inspected as a function of age and gender. Further, the spectral findings of the collagen synthesis in wound granulation tissue/un-wounded skin tissues were validated by Picro-Sirius red- polarized light microscopy in a blinded manner through image analysis of the respective collagen birefringence. The in vivo autofluorescence studies have shown a significant increase in collagen synthesis in laser treated animals as compared to the un-illuminated controls. Image analysis of the collagen birefringence further authenticated the ability of autofluorescence in the objective monitoring of collagen in vivo. Our results clearly demonstrate the potential of laser induced autofluorescence in the monitoring of collegen synthesis during tissue regeneration, which may have clinical implications.
Bibliography:	Competing Interests: The authors have declared that no competing interests exist. Conceived and designed the experiments: KKM SBSR. Performed the experiments: VP. Analyzed the data: VP EMF ACKR KP. Wrote the paper: VP KKM SBSR.
ISSN:	1932-6203 1932-6203
DOI:	10.1371/journal.pone.0098609