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4: Specific viscosity of high molecular weight (MW=1.4x10 6 Da) hyaluronic acid (HMW-HA) in water (Trendline a) and 0.1 M NaCl (Trendline b).
Source publication
Vibrational spectroscopic methods are minimally invasive, and are appropriate for use in clinical contexts. Methods were developed in this dissertation for evaluating joint damage and disease using Raman spectroscopy. Subtle changes in the molecular structure of joint tissue and synovial fluid precede morphological changes in the joint. The goal of...
Citations
... However, conventional arthroscopy relies on the surgeon's visual and tactile assessment of the tissue, which results in subjective and poorly reproducible outcomes [2]. Recently, several techniques including ultrasonography [3], high frequency [4] and intra-articular [5] ultrasound, optical coherence tomography [6,7], near-infrared (NIR) [8][9][10], mid-infrared [11,12], and RS [13][14][15][16] have been proposed as potential candidates for addressing the limitations of conventional arthroscopic approaches. ...
Objective
To differentiate healthy from artificially degraded articular cartilage and estimate its structural, compositional, and functional properties using Raman spectroscopy (RS).
Design
Visually normal bovine patellae ( n = 12) were used in this study. Osteochondral plugs ( n = 60) were prepared and artificially degraded either enzymatically (via Collagenase D or Trypsin) or mechanically (via impact loading or surface abrasion) to induce mild to severe cartilage damage; additionally, control plugs were prepared ( n = 12). Raman spectra were acquired from the samples before and after artificial degradation. Afterwards, reference biomechanical properties, proteoglycan (PG) content, collagen orientation, and zonal (%) thickness of the samples were measured. Machine learning models (classifiers and regressors) were then developed to discriminate healthy from degraded cartilage based on their Raman spectra and to predict the aforementioned reference properties.
Results
The classifiers accurately categorized healthy and degraded samples (accuracy = 86%), and successfully discerned moderate from severely degraded samples (accuracy = 90%). On the other hand, the regression models estimated cartilage biomechanical properties with reasonable error (≤ 24%), with the lowest error observed in the prediction of instantaneous modulus (12%). With zonal properties, the lowest prediction errors were observed in the deep zone, i.e., PG content (14%), collagen orientation (29%), and zonal thickness (9%).
Conclusion
RS is capable of discriminating between healthy and damaged cartilage, and can estimate tissue properties with reasonable errors. These findings demonstrate the clinical potential of RS.
... The mineral content of SB in horse's metacarpal joint increased, while its collagen composition decreased in the early stage of osteoarthritis [13]. However, the mineralization of SB was found to be reduced in a mouse model of early OA [14,15]. It can be seen that the biological composition of SB changed significantly during the progression of OA; however, the current results are not consistent. ...
... Nevertheless, these changes were not consistently significant in all related studies. It is important to note that although some previous studies on SP changes have reported similar findings, others have reported inconsistent observations [13,14,46,47]. The discrepancy may be due to the intraspecimen differences, the different stages of the disease, and variations in the region of interest (ROI) for the subchondral plate. ...
This study aims to investigate how biochemical composition in subchondral bone (SB) relates to the sulfated glycosaminoglycan (sGAG) content of articular cartilage (AC) in the knee joint of guinea pigs from the early to moderate osteoarthritis (OA). Male Dunkin Hartley strain guinea pigs were grouped according to age (1, 3, 6, and 9 months, with 10 guinea pigs in each group). The biochemical properties of the AC and SB in the tibial plateau of the guinea pigs were determined through histology and Raman spectroscopy, respectively. Furthermore, the microstructures of the SB were investigated using micro-computed tomography (micro-CT) and histology. Increased thickness and bone mineral density (BMD) and decreased porosity were observed in the subchondral plate (SP) with the progression of spontaneous OA, accompanied by a decreasing trend in sGAG integrated optical density (IOD) of AC. Compared with the changes in the microstructure of subchondral bone, the content of sGAG was more correlated to the changes in the mineral/matrix ratio of subchondral bone. The mineralization of the matrix was significantly correlated to the content of sGAG compared with crystallinity/maturity and Type B carbonate substitution. PO43− ν1/Amide III was more correlated to the content of sGAG than PO43− ν1/Amide I, PO43− ν1/CH2 wag during the progression of spontaneous osteoarthritis. This study demonstrated that the mineralization of subchondral bone plays a crucial role in the pathogenesis of OA. Future studies may access to the mineralization of subchondral bone in addition to its microstructure in the study for pathogenesis and early diagnosis of osteoarthritis.
... The analysis of the Raman spectroscopy bands is presented in the table 1. Amid III, СН2 oscillation of glycine and proline [16] 1445 (CH2)/(CH3) collagen and other proteins [11,17] 1587 Amid II (C -N, С-С strength) [11] 1660 Amid I, С-С valence oscillation [17] 1745 V(С=О) Lipids [18] After processing in Wolfram Mathematica, the spectra can still contain contours with an unresolved internal structure, which is caused by the overlapping of adjacent lines. In order to be able to analyze The results of the spectra deconvolution were used in further analysis. ...
... The analysis of the Raman spectroscopy bands is presented in the table 1. Amid III, СН2 oscillation of glycine and proline [16] 1445 (CH2)/(CH3) collagen and other proteins [11,17] 1587 Amid II (C -N, С-С strength) [11] 1660 Amid I, С-С valence oscillation [17] 1745 V(С=О) Lipids [18] After processing in Wolfram Mathematica, the spectra can still contain contours with an unresolved internal structure, which is caused by the overlapping of adjacent lines. In order to be able to analyze The results of the spectra deconvolution were used in further analysis. ...
This paper presents the results of expanded analysis after the experimental researches with the samples femoral bones of rabbits in distal epiphysis area using the Raman spectroscopy method. It has been found that values of the optical ratios characterized by reduction of Raman bands during transition from the intact cartilage zone to the PFP plasty region on the wave numbers 956 cm ⁻¹ (PO 3 ⁻⁴ (symmetric stretching phosphate oscillation)), 1069 cm ⁻¹ (CO 3 ²⁻ (C-O planar stretching vibration)) and relatively constant coefficients on the wavenumbers 852 cm ⁻¹ (Proline), 1250 cm ⁻¹ (Amid III), 1587 cm ⁻¹ (Amid II) и 1660 cm ⁻¹ (Amid I ), and also on the 1745 cm ⁻¹ ((C=O) Lipids).
Le cartilage articulaire est un tissu conjonctif, non innervé et non vascularisé, qui recouvre les extrémités des os au sein des articulations. Les fortes contraintes auxquelles est soumis le cartilage provoquent souvent l’altération de la matrice articulaire. Les défauts du cartilage articulaire sont généralement associés à la douleur et à une perte de la mobilité articulaire, et ont un impact sur la qualité de la vie, y compris le bien-être physique, social et économique. En raison du défaut de régénération ou de cicatrisation spontanée du cartilage, le recours à des techniques d’ingénierie tissulaire semble une voie prometteuse pour répondre aux besoins clinques des patients souffrant d’arthrose et de troubles de l’articulation temporo-mandibulaire (TATM). Pour optimiser l’ingénierie du tissu cartilagineux, des techniques (nanométriques) non invasives, non destructives et à haute résolution sont nécessaires, afin d’évaluer la qualité et l’arrangement structurel du cartilage fabriqué in vitro. L’objectif de notre étude était de suivre la maturation du cartilage artificiel sous charges mécaniques avec des méthodes innovantes de microscopies, dont la microscopie confocale Raman, une technique d’analyse sans contact, non destructive et sans aucune préparation de l’échantillon. Nous avons, à partir des spectres et des images hyperspectrales Raman, caractériser les différences morphologiques des chondrons suivant les différentes zones de la matrice du cartilage et établir une cartographie représentative de la distribution spatiale des composants biochimiques de ladite matrice sur toute sa profondeur. Le calcul de l’épaisseur de la matrice péricellulaire a mise en évidence des variations croissantes dépendantes de la profondeur (zone superficielle est = 2,7±0,8, zone intermédiaire = 3,6±1,1et la zone profonde = 4,9±1,8, p<0,001). Sur la base des données spectrales Raman, il a été possible de suivre en temps réel les mécanismes sous-jacents au développement de tissus cartilagineux par ingénierie tissulaire, notamment la dédifférenciation des chondrocytes et les cellules souches mésenchymateuses (CSM) différenciées en chondrocytes. Les résultats ont été comparés à des tests immunohistologiques et de réaction en chaine polymérase (RT-qPCR). En comparant le cartilage dégradé versus le cartilage normal, il a été possible de caractériser les modifications subtiles de la structure moléculaire du tissu cartilagineux précédant les modifications morphologiques de la matrice dans l’arthrose. Ces résultats montrent à la fois la capacité et la sensibilité de la microscopie confocale Raman a caractérisé la structure du cartilage articulaire et fournit des informations précieuses pouvant servir de base dans la conception de tissus cartilagineux par ingénierie tissulaire et un outil diagnostic pour détecter de façon précoce les modifications subcellulaires liées à l’arthrose.
A home-made small-sized Raman spectrometer combined with machine learning algorithms was used to study and identify healthy and multi-period osteoarthritis (OA) canine knee joints. Nine canines were equally divided into three groups according to the post-operative (OA modeling) time of 2-month, 3-month and 7-month. Other two normal canines were used as control. It was found that the degeneration degree of cartilage was positively correlated with post-operative time by doing anatomical analysis. The mixed Raman spectra of cartilage and subchondral bone were collected and analyzed, which reveals subchondral bone demineralization and carbonate ion substituting into the apatite mineral during OA. Raman spectra combined with principal component analysis (PCA) further disclosed that collagen matrix became unordered, both content ratios of amide I/matrix and phenylalanine/matrix in OA cartilage and subchondral bone increased. Based on the PCA getting five principal components, all groups were effectively discriminated by Fisher discriminant analysis (FDA) with high accuracy of 91.07% for the validation set, as well as 95.45% for the test set. It suggests that Raman spectroscopy combined with machine learning is capable to become an effective tool to achieve in situ identification of multi-period OA with high accuracy and preclinical significance.
Zirconia is the most common ceramic used in dental implants. Even if it possesses both mechanical strength and esthetics, its intrinsic bio-inertness often results in a lack of biological integration. On the other hand, Si3N4 has been proved to be bio-active and to be able to easily osteointegrate. In this work, a Si3N4 powder-based laser-cladding process has been developed in order to improve the biological response to biomedical zirconia. The process resulted in the formation of a composite coating with Si3N4 particles dispersed in nano-crystalline/amorphous silicon. Microscopic observation showed that the layer is adherent to the substrate. The application of the laser cladding treatment resulted in an increase of the roughness, further enhancing the probability of interaction with biological tissues and consequently the bioactivity. Testing with human osteosarcoma cell lines resulted in the formation of bone tissue with high collagen maturity, high carbonate to phosphate ratios and good tissue mineralization, but lower cell proliferation when compared to stoichiometric Si3N4. The bone tissue quality parameters, as measured by Raman spectroscopy, resulted to be comparable to healthy human bone tissue and suggest that laser-cladded Si3N4 treatments might be able to improve the stability of zirconia implants.
The results of the study of synovial fluid (SF) of patients with knee joint osteoarthritis (OA). The optical analysis of the SF samples, harvested during exploratory punctures of knee joints of patients suffering the osteoarthrosis in different stages using the standard method, was made. A certain component composition of surface of SF samples that differs for healthy people and the patients, having knee joint osteoarthrosis in different stages, is possible to identify as a result of analysis with the use of Raman spectroscopy method. The introduced optical coefficients help to estimate the spectral composition of surface of SF samples of the patients, suffering the early and late stages of ОА.
The onset of osteoarthritis (OA)in articular cartilage is characterized by degradation of extracellular matrix (ECM). Specifically, breakage of cross-links between collagen fibrils in the articular cartilage leads to loss of structural integrity of the bulk tissue. Since there are no broadly accepted, non-invasive, label-free tools for diagnosing OA at its early stage, Raman spectroscopyis therefore proposed in this work as a novel, non-destructive diagnostic tool. In this study, collagen thin films were employed to act as a simplified model system of the cartilage collagen extracellular matrix. Cross-link formation was controlled via exposure to glutaraldehyde (GA), by varying exposure time and concentration levels, and Raman spectral information was collected to quantitatively characterize the cross-link assignments imparted to the collagen thin films during treatment. A novel, quantitative method was developed to analyze the Raman signal obtained from collagen thin films. Segments of Raman signal were decomposed and modeled as the sum of individual bands, providing an optimization function for subsequent curve fitting against experimental findings. Relative changes in the concentration of the GA-induced pyridinium cross-links were extracted from the model, as a function of the exposure to GA. Spatially resolved characterization enabled construction of spectral maps of the collagen thin films, which provided detailed information about the variation of cross-link formation at various locations on the specimen. Results showed that Raman spectral data correlate with glutaraldehyde treatment and therefore may be used as a proxy by which to measure loss of collagen cross-links in vivo. This study proposes a promising system of identifying onset of OA and may enable early intervention treatments that may serve to slow or prevent osteoarthritis progression.
Biomolecular changes in the cartilage matrix during the early stage of osteoarthritis may be detected by Raman spectroscopy. The objective of this investigation was to determine vibrational spectral differences among different grades (grades I, II, and III) of osteoarthritis in human osteoarthritic cartilage, which was classified according to the International Cartilage Repair Society (ICRS) grading system. Degenerative articular cartilage samples were collected during total joint replacement surgery and were classified according to the ICRS grading system for osteoarthritis. Twelve cartilage sections (4 sections of each ICRS grades I, II, and III) were selected for Raman spectroscopic analysis. Safranin-O/Fast green was used for histological staining and assignment of the Osteoarthritis Research Society International (OARSI) grade. Multivariate principal component analysis (PCA) was used for data analysis. Spectral analysis indicates that the content of disordered coil collagen increases significantly during the early progression of osteoarthritis. However, the increase was not statistically significant during later stages of the disease. A decrease in the content of proteoglycan was observed only during advanced stages of osteoarthritis. Our investigation shows that Raman spectroscopy can classify the different stage of osteoarthritic cartilage and can provide details on biochemical changes. This proof-of-concept study encourages further investigation of fresh cartilage on a larger population using fiber-based miniaturized Raman probe for the development of in vivo Raman arthroscopy as a potential diagnostic tool for osteoarthritis.
