Nondestructive Assessment of Engineered Cartilage Constructs Using Near-Infrared Spectroscopy

Abstract

Noninvasive assessment of engineered cartilage properties would enable better control of the developing tissue towards the desired structural and compositional endpoints through optimization of the biochemical environment in real time. The objective of this study is to assess the matrix constituents of cartilage using near-infrared spectroscopy (NIRS), a technique that permits full-depth assessment of developing engineered tissue constructs. Mid-infrared (mid-IR) and NIR data were acquired from full-thickness cartilage constructs that were grown up to 4 weeks with and without mechanical stimulation. Correlations were assessed between established mid-IR peak areas that reflect the relative amount of collagen (amide I, amide II, and 1338 cm−1) and proteoglycan (PG), (850 cm−1), and the integrated area of the NIR water absorbance at 5190 cm−1. This analysis was performed to evaluate whether simple assessment of the NIR water absorbance could yield information about matrix development. It was found that an increase in the mid-IR PG absorbance at 850 cm−1 correlated with the area of the NIR water peak (Spearman’s rho = 0.95, p < 0.0001). In the second analysis, a partial least squares method (PLS1) was used to assess whether an extended NIR spectral range (5400–3800 cm−1) could be utilized to predict collagen and proteoglycan content of the constructs based on mid-IR absorbances. A subset of spectra was randomly selected as an independent prediction set in this analysis. Average of the normalized root mean square errors of prediction of first-derivative NIR spectral models were 7% for 850 cm−1 (PG), 11% for 1338 cm−1 (collagen), 8% for amide II (collagen), and 8% for amide I (collagen). These results demonstrate the ability of NIRS to monitor macromolecular content of cartilage constructs and is the first step towards employing NIR to assess engineered cartilage in situ.

Analysis of collagen preservation in bones recovered in archaeological contexts using NIR Hyperspectral Imaging

“…As collagen is the most abundant protein in bones, there would appear to be a direct link between these wavelengths and the presence of collagen. In addition, the 2030 and 2150 nm peaks are also consistent with the spectra presented by Baykal et al [17]. In addition, the possibility of 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 interference of water absorbed by the samples can be rejected since all samples are at the same low humidity level.…”

“…Comparison between the data presented in Baykal et al [17] and spectra from the COL+ class and the medieval cow appears to show similar characteristic wavelengths (vertical lines in Figure 4). Further, spectral comparison between COL+ and COL-samples seems to indicate the potential for segregation.…”

“…The prediction scores of the spectra from a single sample were then pooled to compute the mean prediction score of the sample and its estimated standard deviation (confidence level of 95%). Figure 4 In their paper, Baykal et al [17] discuss characteristic wavelengths related to the presence of collagen in samples analyzed in the NIR range. Figure 4 presents the mean spectra of samples from COL+ and COL-classes used for the model construction.…”

“…Other studies have investigated collagen in blood vessels or muscles using NIR spectroscopy [17][18]. For hyperspectral imaging, very few applications have been attempted in archaeology (see e.g., [19][20], [12]), but not at the scale of assemblage or artifact and none on bone or collagen.…”

Near infrared spectroscopic evaluation of water in hyaline cartilage

“…In a recent study from our group, water content in engineered cartilage, based on the 5200 cm −1 absorbance band, was correlated to matrix composition 8 . However, to date, NIR spectroscopic assessment of water in cartilage has not been shown to correlate to the gold standard, gravimetric water determination, and therefore quantitative data on changes in water content have not been obtained.…”