Zone Of Articular Cartilage Research Articles

Cartilage progenitor cells derived extracellular vesicles-based cell-free strategy for osteoarthritis treatment by efficient inflammation inhibition and extracellular matrix homeostasis restoration

Osteoarthritis (OA) is a common degenerative joint disease which currently lacks of effective agents. It is therefore urgent and necessary to seek an effective approach that can inhibit inflammation and promote cartilage matrix homeostasis. Cartilage progenitor cells (CPCs) are identified as a cell population of superficial zone in articular cartilage which possess strong migration ability, proliferative capacity, and chondrogenic potential. Recently, the application of CPCs may represent a novel cell therapy strategy for OA treatment. There is growing evidence that extracellular vesicles (EVs) are primary mediators of the benefits of stem cell-based therapy. In this study, we explored the protective effects of CPCs-derived EVs (CPCs-EVs) on IL-1β-induced chondrocytes. We found CPCs-EVs exhibited chondro-protective effects in vitro. Furthermore, our study demonstrated that CPCs-EVs promoted matrix anabolism and inhibited inflammatory response at least partially via blocking STAT3 activation. In addition, liquid chromatography-tandem mass spectrometry analysis identified 991 proteins encapsulated in CPCs-EVs. By bioinformatics analysis, we showed that STAT3 regulatory proteins were enriched in CPCs-EVs and could be transported to chondrocytes. To promoting the protective function of CPCs-EVs in vivo, CPCs-EVs were modified with cationic peptide ε-polylysine-polyethylene-distearyl phosphatidylethanolamine (PPD) for surface charge reverse. In posttraumatic OA mice, our results showed PPD modified CPCs-EVs (PPD-EVs) effectively inhibited extracellular matrix catabolism and attenuated cartilage degeneration. Moreover, PPD-EVs down-regulated inflammatory factors expressions and reduced OA-related pain in OA mice. In ex-vivo cultured OA cartilage explants, PPD-EVs successfully promoted matrix anabolism and inhibited inflammation. Collectively, CPCs-EVs-based cell-free therapy is a promising strategy for OA treatment.

Prg4-Expressing Chondroprogenitor Cells in the Superficial Zone of Articular Cartilage.

Joint-resident chondrogenic precursor cells have become a significant therapeutic option due to the lack of regenerative capacity in articular cartilage. Progenitor cells are located in the superficial zone of the articular cartilage, producing lubricin/Prg4 to decrease friction of cartilage surfaces during joint movement. Prg4-positive progenitors are crucial in maintaining the joint's structure and functionality. The disappearance of progenitor cells leads to changes in articular hyaline cartilage over time, subchondral bone abnormalities, and the formation of ectopic ossification. Genetic labeling cell technology has been the main tool used to characterize Prg4-expressing progenitor cells of articular cartilage in vivo through drug injection at different time points. This technology allows for the determination of the origin of progenitor cells and the tracking of their progeny during joint development and cartilage damage. We endeavored to highlight the currently known information about the Prg4-producing cell population in the joint to underline the significance of the role of these cells in the development of articular cartilage and its homeostasis. This review focuses on superficial progenitors in the joint, how they contribute to postnatal articular cartilage formation, their capacity for regeneration, and the consequences of Prg4 deficiency in these cells. We have accumulated information about the Prg4+ cell population of articular cartilage obtained through various elegantly designed experiments using transgenic technologies to identify potential opportunities for further research.

Endogenus chondrocytes immobilized by G-CSF in nanoporous gels enable repair of critical-size osteochondral defects

Injured articular cartilage is a leading cause for osteoarthritis. We recently discovered that endogenous stem/progenitor cells not only reside in the superficial zone of mouse articular cartilage, but also regenerated heterotopic bone and cartilage in vivo. However, whether critical-size osteochondral defects can be repaired by pure induced chemotatic cell homing of these endogenous stem/progenitor cells remains elusive. Here, we first found that cells in the superficial zone of articular cartilage surrounding surgically created 3 × 1 mm defects in explant culture of adult goat and rabbit knee joints migrated into defect-filled fibrin/hylaro1nate gel, and this migration was significantly more robust upon delivery of exogenous granulocyte-colony stimulating factor (G-CSF). Remarkably, G–CSF–recruited chondrogenic progenitor cells (CPCs) showed significantly stronger migration ability than donor-matched chondrocytes and osteoblasts. G–CSF–recruited CPCs robustly differentiated into chondrocytes, modestly into osteoblasts, and barely into adipocytes. In vivo, critical-size osteochondral defects were repaired by G–CSF–recruited endogenous cells postoperatively at 6 and 12 weeks in comparison to poor healing by gel-only group or defect-only group. ICRS and O'Driscoll scores of articular cartilage were significantly higher for both 6- and 12-week G-CSF samples than corresponding gel-only and defect-only groups. Thus, endogenous stem/progenitor cells may be activated by G-CSF, a Food and Drug Administration (FDA)-cleared bone-marrow stimulating factor, to repair osteochondral defects.

Broadband scattering properties of articular cartilage zones and their relationship with the heterogenous structure of articular cartilage extracellular matrix.

Articular cartilage exhibits a zonal architecture, comprising three distinct zones: superficial, middle, and deep. Collagen fibers, being the main solid constituent of articular cartilage, exhibit unique angular and size distribution in articular cartilage zones. There is a gap in knowledge on how the unique properties of collagen fibers across articular cartilage zones affect the scattering properties of the tissue. This study hypothesizes that the structural properties of articular cartilage zones affect its scattering parameters. We provide scattering coefficient and scattering anisotropy factor of articular cartilage zones in the spectral band of 400 to 1400nm. We enumerate the differences and similarities of the scattering properties of articular cartilage zones and provide reasoning for these observations. We utilized collimated transmittance and integrating sphere measurements to estimate the scattering coefficients of bovine articular cartilage zones and bulk tissue. We used the relationship between the scattering coefficients to estimate the scattering anisotropy factor. Polarized light microscopy was applied to estimate the depth-wise angular distribution of collagen fibers in bovine articular cartilage. We report that the Rayleigh scatterers contribution to the scattering coefficients, the intensity of the light scattered by the Rayleigh and Mie scatterers, and the angular distribution of collagen fibers across tissue depth are the key parameters that affect the scattering properties of articular cartilage zones and bulk tissue. Our results indicate that in the short visible region, the superficial and middle zones of articular cartilage affect the scattering properties of the tissue, whereas in the far visible and near-infrared regions, the articular cartilage deep zone determines articular cartilage scattering properties. This study provides scattering properties of articular cartilage zones. Such findings support future research to utilize optical simulation to estimate the penetration depth, depth-origin, and pathlength of light in articular cartilage for optical diagnosis of the tissue.

Induced regeneration of articular cartilage: Identification of a dormant regeneration program for a non-regenerative tissue.

An organoid culture model was developed to regenerate articular cartilage by sequential treatment with BMP2 and BMP9 that parallels induced joint regeneration at digit amputation wounds in vivo. BMP9 induced chondrogenesis was used to identify clonal cell lines of articular chondrocyte and hypertrophic chondrocyte progenitor cells from digit fibroblasts. A protocol that includes cell aggregation enhanced by BMP2 followed by BMP9 induced chondrogenesis resulted in the differentiation of organized layers of articular chondrocytes similar to the organization of middle and deep zones of articular cartilage in situ and retained a differentiated phenotype following transplantation. In addition, the differentiation of a non-chondrogenic connective tissue layer containing articular chondrocyte progenitor cells demonstrates that progenitor cell sequestration is coupled with articular cartilage differentiation at a clonal level. The studies identify a dormant endogenous regenerative program for a non-regenerative tissue in which fibroblast-derived progenitor cells can be induced to initiate morphogenetic and differentiative programs that include progenitor cell sequestration. The identification of dormant regenerative programs in non-regenerative tissues such as articular cartilage represents a novel strategy that integrates regeneration biology with regenerative medicine.

Effect of Moderate Exercise on the Superficial Zone of Articular Cartilage in Age-Related Osteoarthritis.

This study aimed to evaluate the effect of exercise on the superficial zone of the osteoarticular cartilage during osteoarthritis progression. Three-month-old, nine-month-old, and eighteen-month-old Sprague Dawley rats were randomly divided into two groups, moderate exercise and no exercise, for 10 weeks. Histological staining, immunostaining, and nanoindentation measurements were conducted to detect changes in the superficial zone. X-ray and micro-CT were quantitated to detect alterations in the microarchitecture of the tibial subchondral bone. Cells were extracted from the superficial zone of the cartilage under fluid-flow shear stress conditions to further verify changes in vitro. The number of cells and proteoglycan content in the superficial zone increased more in the exercise group than in the control group. Exercise can change the content and distribution of collagen types I and III in the superficial layer. In addition, TGFβ/pSmad2/3 and Prg4 expression levels increased under the intervention of exercise on the superficial zone. Exercise can improve the Young's modulus of the cartilage and reduce the abnormal subchondral bone remodeling which occurs after superficial zone changes. Moderate exercise delays the degeneration of the articular cartilage by its effect on the superficial zone, and the TGFβ/pSmad2/3 signaling pathways and Prg4 play an important role.

Development of a computational-experimental framework for enhanced mechanical characterization and cross-species comparison of the articular cartilage superficial zone

To provide a better understanding of the contribution of specific constituents (i.e. proteoglycan, collagen, fluid) to the mechanical behavior of the superficial zone of articular cartilage, a complex biological tissue with several time-dependent properties, a finite element model was developed. Optimization was then used to fit the model to microindentation experiments. We used this model to compare superficial zone material properties of mature human vs. immature bovine articular cartilage. Non-linearity and stiffness of the fiber-reinforced component of the model differed between human and bovine tissue. This may be due to the more complex collagen architecture in mature tissue and is of interest to investigate in future work.

Optimizing Bioink Composition for Human Chondrocyte Expression of Lubricin.

The surface zone of articular cartilage is the first area impacted by cartilage defects, commonly resulting in osteoarthritis. Chondrocytes in the surface zone of articular cartilage synthesize and secrete lubricin, a proteoglycan that functions as a lubricant protecting the deeper layers from shear stress. Notably, 3D bioprinting is a tissue engineering technique that uses cells encapsulated in biomaterials to fabricate 3D constructs. Gelatin methacrylate (GelMA) is a frequently used biomaterial for 3D bioprinting cartilage. Oxidized methacrylated alginate (OMA) is a chemically modified alginate designed for its tunable degradation rate and mechanical properties. To determine an optimal combination of GelMA and OMA for lubricin expression, we used our novel high-throughput human articular chondrocyte reporter system. Primary human chondrocytes were transduced with PRG4 (lubricin) promoter-driven Gaussia luciferase, allowing for temporal assessment of lubricin expression. A lubricin expression-driven Design of Experiment screen and subsequent validation identified 14% GelMA/2% OMA for further study. Therefore, DoE optimized 14% GelMA/2% OMA, 14% GelMA control, and 16% GelMA (total solid content control) were 3D bioprinted. The combination of lubricin protein expression and shape retention over the 22 days in culture, successfully determined the 14% GelMA/2%OMA to be the optimal formulation for lubricin secretion. This strategy allows for rapid analysis of the role(s) of biomaterial composition, stiffness or other cell manipulations on lubricin expression by chondrocytes, which may improve therapeutic strategies for cartilage regeneration.

Early Degenerative Changes in a Spontaneous Osteoarthritis Model Assessed by Nanoindentation.

Understanding early mechanical changes in articular cartilage (AC) and subchondral bone (SB) is crucial for improved treatment of osteoarthritis (OA). The aim of this study was to develop a method for nanoindentation of fresh, unfixed osteochondral tissue to assess the early changes in the mechanical properties of AC and SB. Nanoindentation was performed throughout the depth of AC and SB in the proximal tibia of Dunkin Hartley guinea pigs at 2 months, 3 months, and 2 years of age. The contralateral tibias were either histologically graded for OA or analyzed using immunohistochemistry. The results showed an increase in the reduced modulus (Er) in the deep zone of AC during early-stage OA (6.0 ± 1.75 MPa) compared to values at 2 months (4.04 ± 1.25 MPa) (*** p < 0.001). In severe OA (2-year) specimens, there was a significant reduction in Er throughout the superficial and middle AC zones, which correlated to increased ADAMTS 4 and 5 staining, and proteoglycan loss in these regions. In the subchondral bone, a 35.0% reduction in stiffness was observed between 2-month and 3-month specimens (*** p < 0.001). The severe OA age group had significantly increased SB stiffness of 36.2% and 109.6% compared to 2-month and 3-month-old specimens respectively (*** p < 0.001). In conclusion, this study provides useful information about the changes in the mechanical properties of both AC and SB during both early- and late-stage OA and indicates that an initial reduction in stiffness of the SB and an increase in stiffness in the deep zone of AC may precede early-stage cartilage degeneration.

Biomechanical Impact of Phosphate Wasting on Articular Cartilage Using the Murine Hyp Model of X-linked hypophosphatemia.

Degenerative osteoarthritis (OA) is recognized as an early-onset comorbidity of X-linked hypophosphatemia (XLH), contributing to pain and stiffness and limiting range of motion and activities of daily living. Here, we extend prior findings describing biochemical and cellular changes of articular cartilage (AC) in the phosphate-wasting environment of XLH to determine the impact of these changes on the biomechanical properties of AC in compression and potential role in the etiology of OA. We hypothesize that despite increased proteoglycan biosynthesis, disruption of the mineralized zone of AC impacts the mechanical properties of cartilage that function to accommodate loads and that therapeutic restoration of this zone will improve the mechanical properties of AC. Data were compared between three groups: wild type (WT), Hyp, and Hyp mice treated with calcitriol and oral phosphate. EPIC microCT confirmed AC mineral deficits and responsiveness to therapy. MicroCT of the Hyp subchondral bone plate revealed that treatment improved trabecular bone volume (BV/TV) but remained significantly lower than WT mice in other trabecular microstructures (p < 0.05). Microindentation AC studies revealed that, compared with WT mice, the mean stiffness of tibial AC was significantly lower in untreated Hyp mice (2.65 ± 0.95 versus 0.87 ± 0.33 N/mm, p < 0.001) and improved with therapy (2.15 + 0.38 N/mm) to within WT values. Stress relaxation of AC under compressive loading displayed similar biphasic relaxation time constants (Taufast and Tauslow) between controls and Hyp mice, although Tauslow trended toward slowed relaxation times. In addition, Taufast and Tauslow times correlated with peak load in WT mice (r = 0.80; r = 0.78, respectively), whereas correlation coefficient values for Hyp mice (r = 0.46; r = 0.21) improved with treatment (r = 0.71; r = 0.56). These data provide rationale for therapies that both preserve AC stiffness and recovery from compression. The Hyp mouse also provides unique insight into determinants of structural stiffness and the viscoelastic properties of AC in the progression of OA. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Single-cell transcriptomic analysis identifies a highly replicating Cd168+ skeletal stem/progenitor cell population in mouse long bones

Skeletal stem/progenitor cells (SSPCs) are tissue-specific stem/progenitor cells localized within skeletons and contribute to bone development, homeostasis, and regeneration. However, the heterogeneity of SSPC populations in mouse long bones and their respective regenerative capacity remain to be further clarified. In this study, we perform integrated analysis using single-cell RNA sequencing (scRNA-seq) datasets of mouse hindlimb buds, postnatal long bones, and fractured long bones. Our analyses reveal the heterogeneity of osteochondrogenic lineage cells and recapitulate the developmental trajectories during mouse long bone growth. In addition, we identify a novel Cd168+ SSPC population with highly replicating capacity and osteochondrogenic potential in embryonic and postnatal long bones. Moreover, the Cd168+ SSPCs can contribute to newly formed skeletal tissues during fracture healing. Furthermore, the results of multicolor immunofluorescence show that Cd168+ SSPCs reside in the superficial zone of articular cartilage as well as in growth plates of postnatal mouse long bones. In summary, we identify a novel Cd168+ SSPC population with regenerative potential in mouse long bones, which adds to the knowledge of the tissue-specific stem cells in skeletons.

Recent Advances in "Functional Engineering of Articular Cartilage Zones by Polymeric Biomaterials Mediated with Physical, Mechanical, and Biological/Chemical Cues".

Articular cartilage (AC) plays an unquestionable role in joint movements but unfortunately the healing capacity is restricted due to its avascular and acellular nature. While cartilage tissue engineering has been lifesaving, it is very challenging to remodel the complex cartilage composition and architecture with gradient physio-mechanical properties vital to proper tissue functions. To address these issues, a better understanding of the intrinsic AC properties and how cells respond to stimuli from the external microenvironment must be better understood. This is essential in order to take one step closer to producing functional cartilaginous constructs for clinical use. Recently, biopolymers have aroused much attention due to their versatility, processability, and flexibility because the properties can be tailored to match the requirements of AC. This review highlights polymeric scaffolds developed in the past decade for reconstruction of zonal AC layers including the superficial zone, middle zone, and deep zone by means of exogenous stimuli such as physical, mechanical, and biological/chemical signals. The mimicked properties are reviewed in terms of the biochemical composition and organization, cell fate (morphology, orientation, and differentiation), as well as mechanical properties and finally, the challenges and potential ways to tackle them are discussed.

Early patellofemoral cartilage and bone pathology in a rat model of noninvasive anterior cruciate ligament rupture

ABSTRACT Objective Anterior cruciate ligament rupture (ACLR) is a risk factor for the development of post-traumatic osteoarthritis (PTOA). While PTOA in the tibiofemoral joint compartment is well-characterized, very little is known about pathology in the patellofemoral compartment after ACL injury. Here, we evaluated the extent to which ACLR induces early patellofemoral joint damage in a rat model. Methods Adult female Lewis rats were randomized to noninvasive ACLR or Sham. Two weeks post-injury, contrast-enhanced micro-computed tomography (µCT) of femoral and patellar cartilage was performed using 20% v/v ioxaglate. Morphometric parameters of femoral trochlear and patellar cartilage, subchondral bone, and trabecular bone were derived from µCT. Sagittal Safranin-O/Fast-Green-stained histologic sections were graded using the OARSI score in a blinded fashion. Results Cartilage and bone remodelling consistent with an early PTOA phenotype were observed in both femoral trochleas and patellae. ACLR caused osteophyte formation of the patella and pathology in the superficial zone of articular cartilage, including surface fibrillation, fissures, increased cellularity, and abnormal chondrocyte clustering. There were significant increases in thickness of patellar and trochlear cartilage. Loss of subchondral bone thickness, bone volume fraction, and tissue mineral density, as well as changes to patellar and trochlear trabecular microarchitecture, were indicative of catabolic bone remodelling. Several injury-induced changes, including increased cartilage thickness and trabecular spacing and decreased trabecular number were more severe in the patella compared to the trochlea. Conclusion The patellofemoral joint develops mild but evident pathology in the early period following ACL rupture, extending the utility of this model to the study of patellofemoral PTOA.

Tissue Engineering Cartilage with Deep Zone Cytoarchitecture by High-Resolution Acoustic Cell Patterning.

The ultimate objective of tissue engineering is to fabricate artificial living constructs with a structural organization and function that faithfully resembles their native tissue counterparts. For example, the deep zone of articular cartilage possesses a distinctive anisotropic architecture with chondrocytes organized in aligned arrays ≈1-2 cells wide, features that are oriented parallel to surrounding extracellular matrix fibers and orthogonal to the underlying subchondral bone. Although there are major advances in fabricating custom tissue architectures, it remains a significant technical challenge to precisely recreate such fine cellular features in vitro. Here, it is shown that ultrasound standing waves can be used to remotely organize living chondrocytes into high-resolution anisotropic arrays, distributed throughout the full volume of agarose hydrogels. It is demonstrated that this cytoarchitecture is maintained throughout a five-week course of in vitro tissue engineering, producing hyaline cartilage with cellular and extracellular matrix organization analogous to the deep zone of native articular cartilage. It is anticipated that this acoustic cell patterning method will provide unprecedented opportunities to interrogate in vitro the contribution of chondrocyte organization to the development of aligned extracellular matrix fibers, and ultimately, the design of new mechanically anisotropic tissue grafts for articular cartilage regeneration.

Regulatory Mechanisms of Prg4 and Gdf5 Expression in Articular Cartilage and Functions in Osteoarthritis.

Owing to the rapid aging of society, the numbers of patients with joint disease continue to increase. Accordingly, a large number of patients require appropriate treatment for osteoarthritis (OA), the most frequent bone and joint disease. Thought to be caused by the degeneration and destruction of articular cartilage following persistent and excessive mechanical stimulation of the joints, OA can significantly impair patient quality of life with symptoms such as knee pain, lower limb muscle weakness, or difficulty walking. Because articular cartilage has a low self-repair ability and an extremely low proliferative capacity, healing of damaged articular cartilage has not been achieved to date. The current pharmaceutical treatment of OA is limited to the slight alleviation of symptoms (e.g., local injection of hyaluronic acid or non-steroidal anti-inflammatory drugs); hence, the development of effective drugs and regenerative therapies for OA is highly desirable. This review article summarizes findings indicating that proteoglycan 4 (Prg4)/lubricin, which is specifically expressed in the superficial zone of articular cartilage and synovium, functions in a protective manner against OA, and covers the transcriptional regulation of Prg4 in articular chondrocytes. We also focused on growth differentiation factor 5 (Gdf5), which is specifically expressed on the surface layer of articular cartilage, particularly in the developmental stage, describing its regulatory mechanisms and functions in joint formation and OA pathogenesis. Because several genetic studies in humans and mice indicate the involvement of these genes in the maintenance of articular cartilage homeostasis and the presentation of OA, molecular targeting of Prg4 and Gdf5 is expected to provide new insights into the aetiology, pathogenesis, and potential treatment of OA.

Raman spectroscopy-based water measurements identify the origin of MRI T2 signal in human articular cartilage zones and predict histopathologic score.

We investigated for the first time zonal-dependent water distribution in articular cartilage by Raman spectroscopy (RS). We further investigated the association of histopathologic score with RS- and magnetic resonance imaging (MRI)-based water measurements. Cadaveric human cartilage plugs (N=16) with different osteoarthritis (OA) severity were used. Water content distribution in cartilage zones was probed using RS- and MRI-based techniques. Histopathologic scoring was performed by two independent observers blindly. Moderate associations existed between RS- and MRI-based water measurements across all cartilage zones. RS-based analysis of different water compartments helped assign the origin of the T2 signal collected from the various cartilage zones. RS-based water parameters significantly correlated with OA-severity score, whereas MRI-based water measurements did not. RS can probe different water compartments in cartilage zones and predict up to 66% of the variation observed in the histopathologic score. RS-based water measurement could be developed further to assess cartilage quality in the clinic.

Early Efficacy of Type I Collagen-Based Matrix-Assisted Autologous Chondrocyte Transplantation for the Treatment of Articular Cartilage Lesions.

Background: Articular cartilage is a complex structure that allows for low frictional gliding and effective shock absorption. Various sports injuries and inflammatory conditions can lead to lesions in the articular cartilage, which has limited regenerative potential. Type I collagen combined with autologous chondrocytes in a three-dimensional culture were used to induce the regeneration of single-layer autologous expanded chondrocytes without chondrogenic differentiation. Purpose: To assess the clinical, radiological, and histological changes following collagen-based autologous chondrocyte transplantation (MACT) for chondral knee lesions. Methods: The study prospectively enrolled 20 patients with symptomatic knee chondral lesions (mean size lesion was 2.41 ± 0.43 cm2, range: 2.0–3.4 cm2) in the lateral femoral condyle and femoral groove who underwent type I collagen-based MACT between July 2017 and July 2019. knee injury and osteoarthritis outcome score (KOOS) was assessed before the procedure, and periodic clinical follow-up was conducted every 3 months for a maximum of 12 months following the procedure and at 1-year intervals thereafter. Magnetic resonance imaging (MRI) T2 mapping of repaired cartilage was also used for the quantitative analysis of regeneration. In one patient, second-look arthroscopy was performed to assess cartilage regeneration characteristics, and a portion of regenerated cartilage was harvested for histological evaluation 12 months after implantation. Results: At pre-operation and at three, six, 12, and 24 months after the operation, KOOS pain, symptoms, daily life activities, sports and recreation, as well as the quality of life were significantly improved between every two time points. Hematoxylin and eosin (HE) staining indicated that the newly formed cartilage was comprised of naive chondrocytes. Safranin O-fast (S-O) green staining of the regenerated tissue revealed fibroblast-like cells surrounded by glycosaminoglycans. Immunohistochemistry (IHC) analysis indicated that collagen type II was uniformly distributed at the deep zone of articular cartilage and type I collagen mainly depositing in the superficial cartilage layer. The T2 values for repaired tissue gradually decreased, eventually approaching near-average values. Conclusion: The present study demonstrated that type I collagen-based MACT is a clinically effective treatment for improving functionality and pain levels. Histological evidence confirmed hyaline cartilage induction and showed that repaired cartilage tended to emerge from the deep to the superficial layer. The quantitative MRI T2 mapping test indicated that there still was a difference between the transplanted cartilage and the surrounding hyaline cartilage. Taken together, the current method represents an efficient approach for the restoration of knee cartilage lesions.

Fourier Transform Infrared Microspectroscopy Combined with Principal Component Analysis and Artificial Neural Networks for the Study of the Effect of β-Hydroxy-β-Methylbutyrate (HMB) Supplementation on Articular Cartilage.

The potential of Fourier Transform infrared microspectroscopy (FTIR microspectroscopy) and multivariate analyses were applied for the classification of the frequency ranges responsible for the distribution changes of the main components of articular cartilage (AC) that occur during dietary β-hydroxy-β-methyl butyrate (HMB) supplementation. The FTIR imaging analysis of histological AC sections originating from 35-day old male piglets showed the change in the collagen and proteoglycan contents of the HMB-supplemented group compared to the control. The relative amount of collagen content in the superficial zone increased by more than 23% and in the middle zone by about 17%, while no changes in the deep zone were observed compared to the control group. Considering proteoglycans content, a significant increase was registered in the middle and deep zones, respectively; 62% and 52% compared to the control. AFM nanoindentation measurements collected from animals administered with HMB displayed an increase in AC tissue stiffness by detecting a higher value of Young’s modulus in all investigated AC zones. We demonstrated that principal component analysis and artificial neural networks could be trained with spectral information to distinguish AC histological sections and the group under study accurately. This work may support the use and effectiveness of FTIR imaging combined with multivariate analyses as a quantitative alternative to traditional collagenous tissue-related histology.

Role of WNT Agonists, BMP and VEGF Antagonists in Rescuing Osteoarthritic Knee Cartilage in a Rat Model.

The superficial zone of articular cartilage (AC) is vital for its function and biomechanics. The damaged AC gets vascularized and undergoes hypertrophy and ossification. Studies have highlighted these two as the major causative factors in osteoarthritis (OA). We aimed at preventing the OA progression in a rat knee instability model by inhibiting the vascular ingrowth and ossification using VEGF and BMP antagonist. A WNT agonist was also used to promote AC regeneration because of its protective effect on the superficial layer. Rat knee OA was created by surgical excision of the medial meniscus and medial collateral ligament. Forty rats were divided into two groups of twenty each for surgical control and tests (surgery + intra-articular injection of drugs every two weeks). Ten animals from each group were sacrificed at four and eight weeks. Histology was mainly used to evaluate the outcome. A surgical OA model was successfully created with higher histological scores for operated knees, both in short- (P = 0.0001) and long-term (P = 0.001). Modified Mankin score was lesser in the test animals as compared to control (P = 0.17) in the short-term, but the trend was reversed in the long-term (P = 0.13). Subgroup analysis revealed that repeated injections in the anterolateral compartment contributed to higher scores in the lateral (P = 0.03) and anterior (P = 0.03) compartment of the knee in the long-term. The combinatorial approach was effective in controlling the OA in short-term. Further studies are needed to test the sustained drug delivery system to improve the outcome.

Creb5 establishes the competence for Prg4 expression in articular cartilage

A hallmark of cells comprising the superficial zone of articular cartilage is their expression of lubricin, encoded by the Prg4 gene, that lubricates the joint and protects against the development of arthritis. Here, we identify Creb5 as a transcription factor that is specifically expressed in superficial zone articular chondrocytes and is required for TGF-β and EGFR signaling to induce Prg4 expression. Notably, forced expression of Creb5 in chondrocytes derived from the deep zone of the articular cartilage confers the competence for TGF-β and EGFR signals to induce Prg4 expression. Chromatin-IP and ATAC-Seq analyses have revealed that Creb5 directly binds to two Prg4 promoter-proximal regulatory elements, that display an open chromatin conformation specifically in superficial zone articular chondrocytes; and which work in combination with a more distal regulatory element to drive induction of Prg4 by TGF-β. Our results indicate that Creb5 is a critical regulator of Prg4/lubricin expression in the articular cartilage.