Abstract
The development of a label-free, non-destructive and safe analytical method such as Raman spectroscopy for assessing cartilage degradation is highly desirable. Compared to non-optical imaging modalities, Raman mapping offers a more sensitive means of directly assessing the chemical composition of cartilage in three-dimensional space and the potential to monitor cartilage degeneration to inform intervention and treatment strategies. Herein, we report the application of Raman spectroscopic methods ex vivo and at arthroscopy to identify molecular alterations in cartilage specimens containing minor focal lesions characteristic of the early disease phase. Our initial ex vivo analysis, obtained by single-point Raman spectroscopy of cartilage samples, supports previous findings based on S-O stretching vibration bands associated with sulphated glycosaminoglycans (sGAGs). We extended the analyses to the high-wavenumber region where we observed that vibrational bands assigned to C-H and O-H stretching modes discriminated early cartilage alterations from healthy cartilage samples. Furthermore, we performed a proof-of-concept in-clinic study using a custom-built optical probe to acquire Raman spectral measurements for the first time in patients undergoing arthroscopy of knee joints. Spectra were obtained with adequate signal-to-noise ratios that similarly discriminated between lesion and adjacent cartilage sites and identified reductions in sGAGs in apparently healthy cartilage. Building on this, we present initial results from Raman mapping to spatially resolve the molecular constituents of cartilage through its depth and across a lesion. Mapping revealed a non-uniform and reduced sGAG distribution within the lesion and peripheral cartilage that was otherwise visually normal, similar to the in-clinic observations, showing that the degradative influence of the lesion extended beyond its border. This was accompanied by a decreased fluorescence signal intensity, which suggests that fluorescence may provide valuable information as an adjunct to the Raman signal in discriminating normal and degenerating cartilage. This work demonstrates the value of Raman mapping over single-point Raman measurements for the analysis of the anisotropy of articular cartilage and highlights the potential of the technology for in vivo articular joint arthroscopy applications.
Highlights
Diarthrodial joints are dependent on a thin layer of hyaline cartilage at their bony surfaces that enables near-frictionless movement and dissipation of the mechanical loads experienced during locomotion
These consisted of a series of bands in the fingerprint region, at approximately 400–1800 cm–1, characteristic of the collagen, aggrecan and other components of cartilage [44]
Our initial investigation focused on the 1063 cm–1 S-O stretching band of sulphated glycos aminoglycan (sGAG), which is an abundant moiety of aggrecan that is known to be lost in the early phases of cartilage degradation [10,11]
Summary
Diarthrodial joints are dependent on a thin layer of hyaline cartilage at their bony surfaces that enables near-frictionless movement and dissipation of the mechanical loads experienced during locomotion. Articular cartilage is a highly specialised tissue and structural lesions (fibrillations) that occur at its surface do not readily heal [1,2]. Such fibrillar lesions are considered to progress to osteoarthritis (OA), a dis ease characterised by the progressive loss of a smooth articulating sur face, pain, remodelling of joint tissues and eventual biomechanical failure of the joint [3,4].
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