Powered by TCPDF (www.tcpdf.org) This material is protected by copyright and other intellectual property rights, and duplication or sale of all or part of any of the repository collections is not permitted, except that material may be duplicated by you for your research use or educational purposes in electronic or print form. You must obtain permission for any other use. Electronic or print copies may not be offered, whether for sale or otherwise to anyone who is not an authorised user. Kauppinen, S.; Karhula, S. S.; Thevenot, J.; Ylitalo, T.; Rieppo, L.; Kestilä, I.; Haapea, M.; Hadjab, I.; Finnilä, M. A.; Quenneville, E.; Garon, M.; Gahunia, H. K.; Pritzker, K. P.H.; Buschmann, M. D.; Nieminen, Heikki

Powered by TCPDF (www.tcpdf.org) This material is protected by copyright and other intellectual property rights, and duplication or sale of all or part of any of the repository collections is not permitted, except that material may be duplicated by you for your research use or educational purposes in electronic or print form. You must obtain permission for any other use. Electronic or print copies may not be offered, whether for sale or otherwise to anyone who is not an authorised user. Kestilä, I.; Thevenot, J.; Finnilä, M. A.; Karhula, S. S.; Hadjab, I.; Kauppinen, S.; Garon, M.; Quenneville, E.; Haapea, M.; Rieppo, L.; Pritzker, K. P.; Buschmann, M. D.; Nieminen, Heikki; Saarakkala, Seppo

Quantitative assessments of articular cartilage function are needed to aid clinical decision making. Our objectives were to develop a new electromechanical grade to assess quantitatively cartilage quality and test its reliability. Electromechanical properties were measured using a hand-held electromechanical probe on 200 human articular surfaces from cadaveric donors and osteoarthritic patients. These data were used to create a reference electromechanical property database and to compare with visual arthroscopic International Cartilage Repair Society (ICRS) grading of cartilage degradation. The effect of patient-specific and location-specific characteristics on electromechanical properties was investigated to construct a continuous and quantitative electromechanical grade analogous to ICRS grade. The reliability of this novel grade was assessed by comparing it with ICRS grades on 37 human articular surfaces. Electromechanical properties were not affected by patient-specific characteristics for each ICRS grade, but were significantly different across the articular surface. Electromechanical properties varied linearly with ICRS grade, leading to a simple linear transformation from one scale to the other. The electromechanical grade correlated strongly with ICRS grade (r = 0.92, p < 0.0001). Additionally, the electromechanical grade detected lesions that were not found visually. This novel grade can assist the surgeon in assessing human knee cartilage by providing a quantitative and reliable grading system.

Purpose: Calcified cartilage (CC) has an important role in solute transportation and biomechanics between the subchondral bone and articular cartilage. The morphological changes of CC, including thickening of CC, tidemark duplication and tidemark roughness, have all been associated with cartilage degeneration during osteoarthritis (OA). Current methods to visualize these changes are mainly based on the use of 2D histological sections. Micro-computed tomography (μCT) is a volumetric imaging technique useful for characterizing calcified tissues. Here we present a novel method to volumetrically analyze the roughness of tidemark and the vessel perforations through CC from μCT image stacks. Furthermore, we investigated these changes at different stages of OA. Methods: Samples were harvested from six patients (age 49–67) undergoing total knee replacement surgery and two asymptomatic cadavers (age 26 and 49). Osteochondral cores (n = 15, Ø = 4mm) were drilled from the weight bearing area of lateral tibial plateaus. Samples were fixed in buffered 4% formaldehyde and subsequently imaged with μCT (Skyscan 1272, Brüker microCT, Kontich, Belgium: voltage 50 kV, current 200 μA, exposure 3200 ms, frame averaging 3, projection images 1200, isotropic voxel size 2.8 μm). Projections were reconstructed with Nrecon software (v1.6.9.8). After μCT imaging, samples were subjected to conventional histological sectioning and stained with safranin O for histopathological OARSI grading. Furthermore, the number of tidemarks (TM.N) and CC thickness (CC.Th) were analyzed from the stained histological sections. The μCT data analysis was conducted with Matlab software (v8.5), except for calcified tissue and vessel segmentations that were done with a custom-made C++ algorithm and CTAn software (v1.14.4.1), respectively. Volumes of interest (VOIs) of 600×400×Z (Z proportional to CC thickness) were fitted in the center of the CC surface. The calculated parameters were: 1) vessel area fraction (VAF, ratio of flat vessel surface area to VOI surface area [600x400]), 2) number of vessels per mm2 (NV/A), and 3) tidemark roughness (TMR, mean ratio of tidemark length vs. fitted line along the tidemark from all slices). Furthermore, a local binary pattern (LBP) -based analysis was applied to the segmented tidemark in 3D. Briefly, the LBP method defines a specific pattern for each studied voxel based on its neighborhood, as a measure of local volumetric orientation. From the LBP analysis, the amount of different patterns (ADP) as well as the entropy of patterns (EP) were calculated to describe the variance in local CC surface orientation. Figure 1 shows the full analysis protocol. Spearman's correlations were calculated to associate analyzed features with OARSI grading and TM.N. Pearson's correlations were used in all other comparisons. Results: Volumetric representation of two VOIs are shown in Figure 2. VAF correlated with ADP (rp = 0.833, p < 0.0001), and TMR (rp = 0.923, p < 0.0001). EP showed a similar trend with VAF, although not statistically significant (rp = 0.369, p = 0.176). NV/A correlated with ADP (rp = 0.633, p = 0.011), CC.Th (rp = −0.615, p = 0.015) and the TM.N (rs = −0.531, p = 0.042). OARSI grade correlated with EP (rs = −0.578, p = 0.024) and ADP (rs = −0.600, p = 0.018). Scatter plots from μCT volume analyses are shown in Figure 3. Conclusions: Tidemark roughness and vessel perforations through CC were analyzed volumetrically from μCT images using the novel method. The parameters describing local morphology of the tidemark were significantly associated with OA progression. In contrast, no association between the vessel perforations through CC and OA progression was found. On the other hand, increase in vessel perforations was associated with a thinner CC and fewer tidemark duplications. Increased vessel perforations were also linked to the increase in the tidemark roughness and complexity, and to some extent with the randomness of the tidemark morphology. These results suggest an interaction between CC internal and external structure and vessel perforations through CC. Furthermore, these findings support that OA progression may change the local morphology of the tidemark. This volumetric analysis method provides means for further investigation of the tidemark morphology and the structural changes of CC with OA progression.Figure 2: 3D visualization of the 600×400×Z VOIs used in the analyzes. Vessels that perforate to the tidemark surface are shown in red and the calcified tissue is shown in cyan.View Large Image Figure ViewerDownload Hi-res image Download (PPT)Figure 3: Scatter plots of Vessel Area Fraction (VAF) and Number of vessels per mm2 (NV/A) against Tidemark roughness (TMR), Entropy of patterns (EP) and Amount of different patterns (ADP). Colors indicate different OARSI grade groups. Positive trend is observed between vessel perforation and parameters that define roughness (TMR) and local orientation variances (ADP, EP) of the tidemark. ADP is the amount of different patterns per average surface area evaluated.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Purpose: The collagen network has a unique organization in articular cartilage. The most common ways to characterize the collagen network orientation are polarized light microscopy (PLM) and electron microscopy techniques. However, these techniques are limited to the study of thin sections or tissue surfaces. Here, we present an analysis method for micro-computed tomography (micro-CT) data to evaluate the microstructural orientation of dehydrated articular cartilage samples in 3D using structure tensor analysis. Methods: Cylindrical osteochondral samples (n = 6, diameter 4 mm) were prepared from tibial plateaus of human cadavers (N = 3) and of patients who underwent total knee replacement surgery (N = 3). The samples were split in half, and then fixed in formaldehyde. One half was subjected for standard histological sectioning protocol, while the second half was dehydrated in ascending ethanol series and treated with hexamethyldisilazane (HMDS). Subsequently, the second half was dried in room temperature overnight and imaged using desktop μCT (SkyScan 1272, Bruker microCT, Kontich, Belgium; 40kV, 250 μA, 3600 projections, 5 frames/projection, 1815 ms/frame, isotropic voxel size 1.6 μm, no additional filtration). The data was reconstructed using NRecon software (v 1.6.10.4). Structure tensor analysis was applied to the micro-CT data (volume-of-interest: 500 μm x 500 μm x cartilage depth) in 2D (i.e. x-z plane) and in 3D to determine the extracellular matrix orientation in articular cartilage. Eigen analysis was applied to the structure tensors in each voxel, and the eigen-vector with the smallest eigenvalue (i.e. the smallest gradient) was set as the direction of the extracellular matrix. Depth-dependent elevation angle (elevation from the x-y plane) profiles were calculated by averaging the elevation angles of the direction vectors in x-y plane. The histological sections (thickness 5 μm) were imaged with PLM (Abrio PLM system, CRi, Inc., Woburn, MA, USA) to obtain a reference for the collagen network orientation. The micro-CT-based and PLM depth-dependent profiles were rescaled to 200 pixels and compared to each other with Pearson's correlation analysis. Data analyses were conducted using MATLAB (v 8.5, Natick, MA, USA). Results: Figure 1 shows representative images of single micro-CT slice, and the elevation angles obtained using structure tensor methods and PLM. When the structure tensor analysis was conducted in 2D, the average elevation angle in the surface layer (1–10% of the thickness) was 41 ± 4 degrees (with respect to the cartilage surface) and gradually changed to 61 ± 5 degrees in the deep layer (40–100% of the thickness) (Figure 2A). The mean correlation (± standard deviation) between the depth-dependent profiles obtained using 2D structure tensor analysis and PLM was r = 0.82 ± 0.08. The elevation angles obtained using 3D structure tensor analysis are presented in Figure 2B. The average elevation angle obtained using 3D structure tensor analysis was 29 ± 4 degrees at the surface layer and gradually changed to 47 ± 3 degrees in the deep layer. The mean correlation between the depth-dependent profiles obtained using 3D structure tensor analysis and PLM was r = 0.87 ± 0.05. Conclusions: The natural contrast of dehydrated articular cartilage obtained using micro-CT imaging was utilized to reveal 3D structural information of extracellular matrix. The presented micro-CT analysis approach is fully based on image texture analysis, which evaluates the orientations of structures from the grayscale differences between the neighboring voxels. On the other hand, PLM gives direct information on the collagen network orientation. Therefore, it is not surprising that the techniques are not in absolute agreement. Nevertheless, the shapes of the depth-dependent elevation angle profiles obtained using micro-CT analysis and PLM are very similar, as indicated by the high correlation coefficients (r > 0.8) between the profiles of adjacent blocks. The main advantage of the presented method over PLM is its ability to study larger tissue volumes in 3D, while PLM is limited to the study of thin tissue sections in 2D and collagen orientation only in that plane. Based on the findings of this study, we suggest that micro-CT imaging of HMDS-dehydrated articular cartilage samples can be used to obtain information on the orientation of extracellular matrix.Figure 2. Depth-dependent average elevation angles as a function of normalized cartilage thickness obtained using PLM in 2D (red) and using structure tensor analysis A) in 2D (black) and B) in 3D (black). The shaded areas represent the standard deviations in the sample set.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

Objective: To evaluate cross-correlations of ex vivo electromechanical properties with cartilage and subchondral bone plate thickness, as well as their sensitivity and specificity regarding early cartilage degeneration in human tibial plateau. Method: Six pairs of tibial plateaus were assessed ex vivo using an electromechanical probe (Arthro-BST) which measures a quantitative parameter (QP) reflecting articular cartilage compression-induced streaming potentials. Cartilage thickness was then measured with an automated thickness mapping technique using Mach-1 multiaxial mechanical tester. Subsequently, a visual assessment was performed by an experienced orthopedic surgeon using the International Cartilage Repair Society (ICRS) grading system. Each tibial plateau was finally evaluated with mCT scanner to determine the subchondral-bone plate thickness over the entire surface. Results: Cross-correlations between assessments decreased with increasing degeneration level. Moreover, electromechanical QP and subchondral-bone plate thickness increased strongly with ICRS grade (p = 0.86 and p = 0.54 respectively), while cartilage thickness slightly increased (p = 0.27). Sensitivity and specificity analysis revealed that the electromechanical QP is the most performant to distinguish between different early degeneration stages, followed by subchondral-bone plate thickness and then cartilage thickness. Lastly, effect sizes of cartilage and subchondral-bone properties were established to evaluate whether cartilage or bone showed the most noticeable changes between normal (ICRS 0) and each early degenerative stage. Thus, the effect sizes of cartilage electromechanical QP were almost twice those of the subchondral-bone plate thickness, indicating greater sensitivity of electromechanical measurements to detect early osteoarthritis. Conclusion: The potential of electromechanical properties for the diagnosis of early human cartilage degeneration was highlighted and supported by cartilage thickness and mCT assessments. (c) 2017 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Purpose: Imaging the articular cartilage (AC) chondrons in 3D has shown to be feasible using a hexamethyldisilazane-based (HMDS) sample dehydration followed by micro-computed tomography (μCT). However, manual selection and segmentation of chondrons is challenging, highly user-dependent and time-consuming. In this study, we present an automatic algorithm to select and volumetrically segment chondrons from HMDS μCT image stacks. Using the developed method, we further compared chondrons in intact (OARSI 0–1.0) and degenerated (OARSI 3.0–3.5) human AC at different depths. According to previous literature, it was hypothesized that chondron volumes are larger in the degenerated AC due to chondrocyte hypertrophy in osteoarthritis (OA). Methods: Osteochondral samples from three human cadaver tibiae and seven tibiae from patients who underwent total knee replacement surgery (n = 12, Ø = 4 mm) were prepared and cut in half; one part for histology and the other for μCT. After preparation, samples were fixed in 4% saline-buffered formaldehyde, dehydrated in ascending ethanol series, treated with HMDS and dried in room temperature overnight. The samples were then imaged with desktop μCT (SkyScan 1272, Bruker microCT, Kontich, Belgium; scanning parameters: 40 kV, 250 μA, 1.6 μm voxel side length, 1815 ms, no additional filtration) and reconstructed using NRecon software (v1.6.10.4). Subsequently, volumes of interest (VOIs) with the size of 300 × 300 × Z (Z = the height of the AC) voxels were chosen for analysis. A custom-made algorithm developed in Matlab (v8.5) was then applied to automatically select and segment the chondrons. A second algorithm was used for manual verification by the superposition of the original image and the segmented mask in three orthogonal views from the center of chondron. Finally, the user validated the accuracy of segmentation and differentiated chondrons containing a single cell from the ones containing a cluster (Fig. 1, bottom left). For all correctly segmented chondrons, their volume (μm3) and depth (%) from the AC surface were calculated. The average chondron volumes were divided into three AC depth zones (zone 1: 0–10%; zone 2: 10–40%; zone 3: 40–100%) and grouped by the OARSI histological grades (OARSI 0–1.0, n = 6; OARSI 3.0–3.5, n = 6). The groups were statistically compared with Mann-Whitney U-test. Results: A total of approximately 6 000 chondrons were automatically segmented from all the 12 samples, and 25% of them were approved and further analyzed. The user input time with this new algorithm was roughly 48 hours, decreasing significantly the estimated time of 600 hours required by the previous manual method. The average chondron volumes in the zones 2 and 3 were significantly larger (zone 2: p = 0.004; zone 3: p = 0.004) in the OARSI 3.0–3.5 group compared to OARSI 0–1.0 group (Fig. 1, top left). Similar results were observed for chondrons containing only a single cell (zone 2: p = 0.028; zone 3: p = 0.016) and the ones with clusters only (zone 2: p = 0.004; zone 3: p = 0.006) (Fig. 1, top middle-right). Volumetric models of the segmented chondrons for representative samples from both OARSI groups can be seen in Fig. 1 (bottom right). Conclusions: In this study, we present an automated 3D selection and segmentation algorithm for AC chondrons imaged with HMDS μCT. The main advantage of this method is that it allows fast 3D analyses for multiple chondrons. Furthermore, results obtained with this approach concur with the previous studies; the chondrons in OA samples were significantly larger in the zones 2 and 3 than in the respective zones of intact samples. The observed hypertrophic morphological changes could be explained by their upregulated metabolism induced by OA activity. In zone 1, statistically significant differences were not observed either due to a lack of segmented chondrons or simply because hypertrophy is less common in the AC superficial layer. The superficial layer still remains challenging for chondron segmentation, mainly due to the shrinkage of the cartilage surface during sample processing. Verification could be further improved with an evaluation from the full 3D volume instead of the current approach from three orthogonal 2D views. Consequently, with the current approach some details may not be detected during the verification process. However, the presented protocol for imaging and segmenting chondrons in 3D volumes provides a new approach to spatially evaluate chondron/chondrocyte properties during different phases of OA.

INTRODUCTION: We published a recent study showing superior sensitivity of electromechanical and indentation (instantaneous response) assessments versus well-established techniques, including histological Mankin score, to characterize cartilage degeneration [1]. This study aims to determine whether the combination of instantaneous, relaxation and equilibrium mechanical properties and friction measurements (surface integrity) could increase sensitivity to detect cartilage degeneration.

The skin barrier poses an ongoing challenge for the cosmetics industry. Its penetration, by non-invasive means, can readily be achieved with currents and ultrasound or radiofrequency devices through electroporation, sonophoresis, iontophoresis or cavitation. When several types of energy are applied simultaneously, we expect the effects to be magnified and all the more effective. Although the mechanism of action of each technology on the skin is not entirely controlled, and is even less so when multiple technologies are applied concurrently, some studies demonstrate that nitric oxide (NO) plays a pivotal role in skin wound-healing and regeneration. With regard to wound healing, one of the key functions of NO appears to be its permissive effect on keratinocyte and fibroblast proliferation, which helps promote wound re-epithelialization. The objective of the actual research is to gain an in-depth understanding of the mechanisms generated by NO through the application of a specific combination of technologies.

Currently, there are no established treatments to prevent, stop, or even retard the degeneration of articular cartilage in osteoarthritis (OA). Biological repair of the degenerating articular cartilage would be preferable to surgery. There is no benign site where autologous chondrocytes can be harvested and used as a cell source for cartilage repair, leaving mesenchymal stem cells (MSCs) as an attractive option. However, MSCs from OA patients have been shown to constitutively express collagen type X (COL-X), a marker of late-stage chondrocyte hypertrophy. We recently found that naproxen (Npx), but not other nonsteroidal anti-inflammatory drugs, can induce collagen type X alpha 1 (COL10A1) gene expression in bone marrow-derived MSCs from healthy and OA donors. In this study, we determined the effect of Npx on COL10A1 expression and investigated the intracellular signaling pathways that mediate such effect in normal human MSCs during chondrogenesis. MSCs were cultured in standard chondrogenic differentiation media supplemented with or without Npx. Our results show that Npx can regulate chondrogenic differentiation by affecting the gene expression of both Indian hedgehog and parathyroid hormone/parathyroid hormone-related protein signaling pathways in a time-dependent manner, suggesting a complex interaction of different signaling pathways during the process.

Material & Methods: Electromechanical ex vivo mappings of articular surfaces were obtained in distal condyles from 5 sheep (8 – 9 y-o, 9 months post-surgery, bone marrow stimulation model) and 2 control sheep (8 years old) with the Arthro-BST (Biomomentum, Laval) which has a hemispherical indenter (r=3.175mm) to measure electromechanical properties by the quantitative parameter (QP), followed by automated indentation mappings to 50 μm with the Mach-1 (Biomomentum, Laval) with a spherical indenter (r=0.5mm) at the same positions to measure structural stiffness (load/indentation depth). Selected test sites were analyzed histologically and by unconfined compression.

Introduction Intervertebral disc (IVD) structure changes throughout life ultimately resulting in disc degeneration and back pain. Most of the IVD extracellular matrix alterations are related this degeneration. Fourier transform infrared (FTIR) spectroscopy has been shown to be a powerful tool in the study of molecular changes associated with matrix structure (collagen denaturation, cross-links, proteoglycans [PGs], and calcification). FTIR analysis is based on monitoring vibrations that originate from molecular components in tissues. Accordingly, evaluation of changes in molecular structure, concentration, and spatial distribution of the tissue components can be performed by direct analysis of spectral maps. Once the spectral maps are calculated, the spatial quantitative and qualitative information on the composition and organization of the tissue compounds can be obtained. FTIR has been successfully utilized in bone and cartilage research in the differentiation between normal and diseased tissues. Our study aims to investigate spatial changes in human IVD composition and microstructural organization in relation to increasing grades of degeneration. Materials and Methods Sample Preparation Human IVDs were obtained from donor lumbar spines of Thompson grades 2 to 5 through organ donations within 24 hours after death. Two IVDs per grade of degeneration were used for FTIR procedure. All tissues were fixed in Accustain (Sigma-Aldrich, St. Louis, Missouri, United States), paraffin embedded and sectioned into 20-mm-thick sagittal sections. Consecutive slices were assessed for chemical properties. FTIR Spectra Acquisition Human IVD sections were placed on barium fluoride infrared transparent windows and infrared absorbance spectra were acquired using a classical FTIR spectrometer FTS 7000 series’ (DGILAB) coupled to UMA 600 microscope. For FTIR spectra acquisition, the system was used in point mode (aperture of 250 ×250 µm) with a 4.0/cm resolution and using 128 scans in transmittance mode, and spectra acquisitions were performed under complete N2 purge of the analytical system. Four repeated scans were performed on the spectral region of approximately 900 to 2,000/cm in each sample in the annulus fibrosus (AF) and nucleus pulposus (NP) regions. Analysis of FTIR Absorption Spectrum of IVD Spectral data were analyzed using Wire 3.0 software. All spectra were baselined and the absorbance of the collagen (COL), elastin, and PG were monitored in the 1,690 to 1,660; 1,595 to 1,500; and 1,140 to 985/cm spectral regions. A univariate analysis was used to evaluate the collagen maturity as the ratio of the integrated area under the amide subpeaks (1,660:1,690/cm). Results The elastin and COL content associated with the stretching vibrations of carbonyl (C=O), C-N, and N-H in the amide II spectral region (1,595-1,500/cm) indicated a significant decrease according to degeneration grade (in AF, from 29.73 ± 0.83 in grade 2 to 20.19 ± 0.65 in grade 5, and in NP from 24.92 ± 0.71 in grade 2 to 8.76 ± 0.56 in grade 5). A significant decrease was also found for the PG content, which is reflected by the stretching vibrations of C-O, C-OH, and as C-C in the carbohydrate chemical group. PG content significantly decreased from 7.95 ± 0.89 in grade 2 to 2.77 ± 0.11 in grade 5 ( p < 0.05) in AF, and from 9.2 ± 1.02 in grade 2 to 5.67 ± 0.72 in grade 5 ( p < 0.03) in NP. Moreover, the ratio of the integrated area of the amide subpeaks (1,660:1690/cm) accompanying collagen maturity, doubled in value with increasing degeneration grades. Conclusion Our studies indicate that FTIR spectral imaging, coupled with univariate data processing techniques, can be used to image the spatial distribution of matrix constituents in human discs with different grades of degeneration. It can therefore provide unique quantitative information on how human disc degeneration can affect the functional state of the disc. Disclosure of Interest None declared

Introduction We previously showed that type X collagen, a marker of late stage chondrocyte hypertrophy (associated with endochondral ossification), is constitutively expressed by mesenchymal stem cells (MSCs) from osteoarthritis patients and this may be related to Naproxen (Npx), a nonsteroidal anti-inflammatory drug used for therapy. Hedgehog (HH) signaling plays an important role during the development of bone. We tested the hypothesis that Npx affected osteogenic differentiation of human MSCs through the expression of Indian hedgehog ( IHH ), Patched-1 ( PTC1 ) and GLI family members GLI1 , GLI2, GLI3 in vitro . Methods MSCs were cultured in osteogenic differentiation medium without (control) or with 0.5 μM Npx. The expression of collagen type X, alpha 1 ( COL10A1 ), alkaline phosphatase ( ALP ), osteopontin ( OPN ), osteocalcin ( OC ), collagen type I, alpha 1 ( COL1A1 ) was analyzed with real-time reverse transcription (RT) PCR, and the ALP activity was measured. The osteogenesis of MSCs was monitored by mineral staining and quantification with alizarin red S. To examine whether Npx affects osteogenic differentiation through HH signaling, the effect of Npx on the expression of IHH , GLI1 , GLI2 , GLI3 and PTC1 was analyzed with real-time RT PCR. The effect of cyclopamine (Cpn), a HH signaling inhibitor, on the expression of COL10A1 , ALP , OC and COL1A1 was also determined. Results When MSCs were cultured in osteogenic differentiation medium, Npx supplementation led to a significant decrease in ALP gene expression as well as its activity, and had a tendency to decrease mineral deposition. It also decreased the expression of COL1A1 significantly. In contrast, the gene expression of COL10A1 and OPN were upregulated significantly by Npx. No significant effect was found on OC expression. The expression of IHH , PTC1 , GLI1, and GLI2 was increased by Npx, while no significant difference was observed on GLI3 expression. Cpn reversed the effect of Npx on the expression of COL10A1 , ALP , OPN and COL1A1 . Conclusions These results indicate that Npx can affect gene expression during osteogenic differentiation of MSCs, and downregulate mineral deposition in the extracellular matrix through IHH signaling. Therefore, Npx could affect MSC-mediated repair of subchondral bone in OA patients.