Intra-operative Raman spectroscopy and ex vivo Raman mapping for assessment of cartilage degradation

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

Raman microspectroscopy demonstrates reduced mineralization of subchondral bone marrow lesions in knee osteoarthritis patients

Introduction: Bone marrow lesions (BMLs) are frequently identified by MRI in the subchondral bone in knee osteoarthritis (KOA). BMLs are known to be closely associated with joint pain, loss of the cartilage and structural changes in the subchondral trabecular bone (SCTB). Despite this, understanding of the nature of BMLs at the trabecular tissue level is incomplete. Thus, we used Raman microspectroscopy to examine the biochemical properties of SCTB from KOA patients with presence or absence of BMLs (OA-BML, OA No-BML; respectively), in comparison with age-matched cadaveric non-symptomatic controls (Non-OA CTL). Methods: Tibial plateau (TP) specimens were collected from 19 KOA arthroplasty patients (6-Male, 13-Female; aged 56–74 years). BMLs were identified ex-vivo by MRI, using PDFS- and T1-weighted sequences. The KOA specimens were then categorized into an OA-BML group (n = 12; containing a BML within the medial condyle only) and an OA No-BML group (n = 7; with no BMLs identified in the TP). The control (CTL) group consisted of Non-OA cadaveric TP samples with no BMLs and no macroscopic or microscopic evidence of OA-related changes (n = 8; 5-Male, 3-Female; aged 44–80 years). Confocal Raman microspectroscopy, with high spatial resolution, was used to quantify the biochemical properties of SCTB tissue of both the medial and the lateral condyle in each group. Results: The ratios of peak intensity and integrated area of bone matrix mineral (Phosphate (v1), Phosphate (v2) and Phosphate (v4)), to surrogates of the organic phase of bone matrix (Amide I, Proline and Amide III), were calculated. Within the medial compartment, the mineral:organic matrix ratios were significantly lower for OABML, compared to Non-OA CTL. These ratios were also significantly lower for the OA-BML medial compartment, compared to the OA-BML lateral compartment. There were no group or compartmental differences for Carbonate:Phosphate (v1, v2 and v4), Amide III (α-helix):Amide III (random-coil), Hydroxyproline:Proline, or Crystallinity. Conclusion: As measured by Raman microspectroscopy, SCTB tissue in BML zones in KOA is significantly less mineralized than the corresponding zones in individuals without OA. These data are consistent with those obtained using other methods (e.g. Fourier transform infrared spectroscopy; FTIR) and with the increased rate of bone remodeling observed in BML zones. Reduced mineralization may change the biomechanical properties of the trabecular bone in BMLs and the mechanical interaction between subchondral bone and its overlying cartilage, with potential implications for the development and progression of OA.Yea-Rin Lee, David M. Findlay, Dzenita Muratovic, Tiffany K. Gill, Julia S. Kuliwab

Variable bone fragility associated with an Amish COL1A2 variant and a knock-in mouse model

Osteogenesis imperfecta (OI) is a heritable form of bone fragility typically associated with a dominant COL1A1 or COL1A2 mutation. Variable phenotype for OI patients with identical collagen mutations is well established, but phenotype variability is described using the qualitative Sillence classification. Patterning a new OI mouse model on a specific collagen mutation therefore has been hindered by the absence of an appropriate kindred with extensive quantitative phenotype data. We benefited from the large sibships of the Old Order Amish (OOA) to define a wide range of OI phenotypes in 64 individuals with the identical COL1A2 mutation. Stratification of carrier spine (L1–4) areal bone mineral density (aBMD) Z -scores demonstrated that 73% had moderate to severe disease (less than −2), 23% had mild disease (−1 to −2), and 4% were in the unaffected range (greater than −1). A line of knock-in mice was patterned on the OOA mutation. Bone phenotype was evaluated in four F 1 lines of knock-in mice that each shared approximately 50% of their genetic background. Consistent with the human pedigree, these mice had reduced body mass, aBMD, and bone strength. Whole-bone fracture susceptibility was influenced by individual genomic factors that were reflected in size, shape, and possibly bone metabolic regulation. The results indicate that the G610C OI (Amish) knock-in mouse is a novel translational model to identify modifying genes that influence phenotype and for testing potential therapies for OI. © 2010 American Society for Bone and Mineral ResearchPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65040/1/90720_ftp.pd

Bioplastics from sweet potatoes

As oil runs out and the use of fossil fuels becomes expensive, the need for replacement source of raw material for the manufacture of plastics becomes essential. Bioplastics are essential as an alternative of commercial plastics from fossil fuels. Bioplastics are eco-friendly and biodegradable hence provide an effective way to replace the commercial plastics. Producing bioplastics from three types of sweet potatoes which are white, orange and purple in colour is the main goal of the study. Extraction techniques were applied to obtain starch from the sweet potatoes. The starch was then mixed with chemical solution such as glycerine, vinegar and distilled water to form bioplastic. The bioplastics were tested for biodegradability, stretch and water adsorption tests. The results show that the bioplastics from white potatoes degrade faster than the other types of sweet potatoes while commercial plastics cannot be degraded at all. White sweet potatoes have less absorption of water which it is the best criteria for bioplastics. Stretchable of white sweet potatoes is more compared to the other types of sweet potatoes. Bioplastics from white sweet potatoes have a good potential as a replacement of commercial plastics

Optical Biomarkers for the Diagnosis of Osteoarthritis through Raman Spectroscopy: Radiological and Biochemical Validation Using Ex Vivo Human Cartilage Samples

[Abstract] Osteoarthritis (OA) is the most common rheumatic disease, characterized by progressive articular cartilage degradation. Raman spectroscopy (RS) has been recently proposed as a label-free tool to detect molecular changes in musculoskeletal tissues. We used cartilage samples derived from human femoral heads to perform an ex vivo study of different Raman signals and ratios, related to major and minor molecular components of articular cartilage, hereby proposed as candidate optical biomarkers for OA. Validation was performed against the radiological Kellgren–Lawrence (K-L) grading system, as a gold standard, and cross-validated against sulfated glycosaminoglycans (sGAGs) and total collagens (Hyp) biochemical contents. Our results showed a significant decrease in sGAGs (SGAGs, A1063 cm−1/A1004 cm−1) and proteoglycans (PGs, A1375 cm−1/A1004 cm−1) and a significant increase in collagen disorganization (ColD/F, A1245 cm−1/A1270 cm−1), with OA severity. These were correlated with sGAGs or Hyp contents, respectively. Moreover, the SGAGs/HA ratio (A1063 cm−1/A960 cm−1), representing a functional matrix, rich in proteoglycans, to a mineralized matrix-hydroxyapatite (HA), was significantly lower in OA cartilage (K-L I vs. III–IV, p < 0.05), whilst the mineralized to collagenous matrix ratio (HA/Col, A960 cm−1/A920 cm−1) increased, being correlated with K-L. OA samples showed signs of tissue mineralization, supported by the presence of calcium crystals-related signals, such as phosphate, carbonate, and calcium pyrophosphate dihydrate (MGP, A960 cm−1/A1004 cm−1, MGC, A1070 cm−1/A1004 cm−1 and A1050 cm−1/A1004 cm−1). Finally, we observed an increase in lipids ratio (IL, A1450 cm−1/A1670 cm−1) with OA severity. As a conclusion, we have described the molecular fingerprint of hip cartilage, validating a panel of optical biomarkers and the potential of RS as a complementary diagnostic tool for OA.Xunta de Galicia; ED431E 2018/03Xunta de Galicia; IN607A2017/1

Raman Spectroscopy identifies differences in ochronotic and non-ochronotic cartilage:a potential novel technique for monitoring ochronosis

Objective Alkaptonuria (AKU) is a rare, inherited disorder of tyrosine metabolism, where patients are unable to breakdown homogentisic acid (HGA), which increases systemically over time. It presents with a clinical triad of features; HGA in urine, ochronosis of collagenous tissues, and the subsequent ochronotic arthritis of these tissues. In recent years the advance in the understanding of the disease and the potential treatment of the disorder looks promising with the data on the efficacy of nitisinone. However, there are limited methods for the detection and monitoring of ochronosis in vivo, or for treatment monitoring. The study aim was to test the hypothesis that Raman spectra would identify a distinct chemical fingerprint for the non-ochronotic, compared to ochronotic cartilage. Design: Ochronotic and non-ochronotic cartilage from human hips and ears were analysed using Raman spectroscopy. Results: Non-ochronotic cartilage spectra were similar and reproducible and typical of normal articular cartilage. Conversely, the ochronotic cartilage samples were highly fluorescent and displayed limited or no discernible Raman peaks in the spectra, in stark contrast to their non-ochronotic pairs. Interestingly, a novel peak was observed associated with the polymer of HGA in the ochronotic cartilage that was confirmed by analysis of pigment derived from synthetic HGA. Conclusion: This technique reveals novel data on the chemical differences in ochronotic compared with non-ochronotic cartilage, these differences are detectable by a technique that is already generating in vivo data and demonstrates the first possible procedure to monitor the progression of ochronosis in tissues of patients with AKU

Molecular analysis of the destruction of articular joint tissues by Raman spectroscopy

Review[Abstract] Introduction: Osteoarthritis (OA) is a highly heterogenous disease influenced by different molecular, anatomic, and physiologic imbalances. Some of the bottlenecks for enhanced diagnosis and therapeutic assessment are the lack of validated biomarkers and early diagnosis tools. In this narrative review, we analyze the potential of Raman spectroscopy (RS) as a label-free optical tool for the characterization of articular joint tissues and its application as a diagnosis tool for OA. Areas covered: Raman spectra produce a unique 'molecular fingerprint' providing rotational and vibrational molecular information, allowing the identification and follow-up of molecular changes associated with OA pathological mechanisms. Focusing on multiple joint tissues (cartilage, synovium, bone, tendons, ligaments, and meniscus) and their contribution in disease incidence and progression, this review highlights the current knowledge on the application of RS in the characterization of organic and inorganic molecules present at these tissues and alterations that occur in the onset of OA. Expert opinion: Vibrational spectroscopy techniques, such as RS, are low cost, rapid and minimally invasive approaches that offer high specificity in the assessment of the molecular composition of complex tissues. Combined with multivariate statistical methods, RS offers great potential for optical biomarkers discovery or disease diagnosis applications, and we hereby discuss clinical translational progresses on the field

Fatty acids in seed oil of wild and cultivated rosehip (Rosa canina L.) from different locations in Serbia

Rosehip (Rosa canina L.) seeds are rich in bioactive compounds and nutrients and hence with a great potential to be employed in production of functional foods. This work aimed to evaluate the fatty acid composition of seed oil from wild and cultivated rosehip collected at different locations in the Republic of Serbia. Unsaturated fatty acids were dominant in majority of seed oil samples, with linoleic (LA), α-linolenic (ALA) and oleic (OA) acids (24.53–46.68 %, 4.73–12.39 % and 3.89–13.82 %, respectively) as the most abundant ones. Based on the analyses of most dominant bands in Raman spectra of seeds (∼1265 and ∼1660 cm-1) characteristic for unsaturated fatty acids, ANOVA revealed significantly higher content in two seed samples (5SW and 10SC). Ratios of UFAs/ SFAs, ω-6/ω-3 and LA/ALA and desirable fatty acids (DFA) indicated that most studied rosehip seed oils showed good quality. Factors such as genetic characteristics and agro-ecological conditions most likely affected FAs composition of seed oils. © 2022 The Author