Nanotechnological Strategies for Osteoarthritis Diagnosis, Monitoring, Clinical Management, and Regenerative Medicine: Recent Advances and Future Opportunities.

PURPOSE OF REVIEW: In this review article, we discuss the potential for employing nanotechnological strategies for the diagnosis, monitoring, and clinical management of osteoarthritis (OA) and explore how nanotechnology is being integrated rapidly into regenerative medicine for OA and related osteoarticular disorders. RECENT FINDINGS: We review recent advances in this rapidly emerging field and discuss future opportunities for innovations in enhanced diagnosis, prognosis, and treatment of OA and other osteoarticular disorders, the smart delivery of drugs and biological agents, and the development of biomimetic regenerative platforms to support cell and gene therapies for arresting OA and promoting cartilage and bone repair. Nanotubes, magnetic nanoparticles, and other nanotechnology-based drug and gene delivery systems may be used for targeting molecular pathways and pathogenic mechanisms involved in OA development. Nanocomposites are also being explored as potential tools for promoting cartilage repair. Nanotechnology platforms may be combined with cell, gene, and biological therapies for the development of a new generation of future OA therapeutics. Graphical Abstract.

Determination of the substrate repertoire of ADAMTS2, 3, and 14 significantly broadens their functions and identifies extracellular matrix organization and TGF-ß signaling as primary targets.

A disintegrin and metalloproteinase with thrombospondin type I motif (ADAMTS)2, 3, and 14 are collectively named procollagen N-proteinases (pNPs) because of their specific ability to cleave the aminopropeptide of fibrillar procollagens. Several reports also indicate that they could be involved in other biological processes, such as blood coagulation, development, and male fertility, but the potential substrates associated with these activities remain unknown. Using the recently described N-terminal amine isotopic labeling of substrate approach, we analyzed the secretomes of human fibroblasts and identified 8, 17, and 22 candidate substrates for ADAMTS2, 3, and 14, respectively. Among these newly identified substrates, many are components of the extracellular matrix and/or proteins related to cell signaling such as latent TGF-ß binding protein 1, TGF-ß RIII, and dickkopf-related protein 3. Candidate substrates for the 3 ADAMTS have been biochemically validated in different contexts, and the implication of ADAMTS2 in the control of TGF-ß activity has been further demonstrated in human fibroblasts. Finally, the cleavage site specificity was assessed showing a clear and unique preference for nonpolar or slightly hydrophobic amino acids. This work shows that the activities of the pNPs extend far beyond the classically reported processing of the aminopropeptide of fibrillar collagens and that they should now be considered as multilevel regulators of matrix deposition and remodeling.-Bekhouche, M., Leduc, C., Dupont, L., Janssen, L., Delolme, F., Vadon-Le Goff, S., Smargiasso, N., Baiwir, D., Mazzucchelli, G., Zanella-Cleon, I., Dubail, J., De Pauw, E., Nusgens, B., Hulmes, D. J. S., Moali, C., Colige, A. Determination of the substrate repertoire of ADAMTS2, 3, and 14 significantly broadens their functions and identifies extracellular matrix organization and TGF-ß signaling as primary targets.

ADAMTS3 activity is mandatory for embryonic lymphangiogenesis and regulates placental angiogenesis.

The only documented activity of a subclass of ADAMTS proteases comprising ADAMTS2, 3 and 14 is the cleavage of the aminopropeptide of fibrillar procollagens. A limited number of in vitro studies suggested that ADAMTS3 is mainly responsible for procollagen II processing in cartilage. Here, we created an ADAMTS3 knockout mouse (Adamts3(-/-)) model to determine in vivo the actual functions of ADAMTS3. Heterozygous Adamts3(+/-) mice were viable and fertile, but their intercrosses demonstrated lethality of Adamts3(-/-) embryos after 15 days of gestation. Procollagens I, II and III processing was unaffected in these embryos. However, a massive lymphedema caused by the lack of lymphatics development, an abnormal blood vessel structure in the placenta and a progressive liver destruction were observed. These phenotypes are most probably linked to dysregulation of the VEGF-C pathways. This study is the first demonstration that an aminoprocollagen peptidase is crucial for developmental processes independently of its primary role in collagen biology and has physiological functions potentially involved in several human diseases related to angiogenesis and lymphangiogenesis.

New prospects in the roles of the C-terminal domains of VEGF-A and their cooperation for ligand binding, cellular signaling and vessels formation.

VEGF-A is a crucial growth factor for blood vessel homeostasis and pathological angiogenesis. Due to alternative splicing of its pre-mRNA, VEGF-A is produced under several isoforms characterized by the combination of their C-terminal domains, which determines their respective structure, availability and affinity for co-receptors. As controversies still exist about the specific roles of these exon-encoded domains, we systematically compared the properties of eight natural and artificial variants containing the domains encoded by exons 1-4 and various combinations of the domains encoded by exons 5, 7 and 8a or 8b. All the variants (VEGF111a, VEGF111b, VEGF121a, VEGF121b, VEGF155a, VEGF155b, VEGF165a, VEGF165b) have a similar affinity for VEGF-R2, as determined by Surface plasmon resonance analyses. They strongly differ however in terms of binding to neuropilin-1 and heparin/heparan sulfate proteoglycans. Data indicate that the 6 amino acids encoded by exon 8a must be present and cooperate with those of exons 5 or 7 for efficient binding, which was confirmed in cell culture models. We further showed that VEGF165b has inhibitory effects in vitro, as previously reported, but that the shortest VEGF variant possessing also the 6 amino acids encoded by exon 8b (VEGF111b) is remarkably proangiogenic, demonstrating the critical importance of domain interactions for defining the VEGF properties. The number, size and localization of newly formed blood vessels in a model of tumour angiogenesis strongly depend also on the C-terminal domain composition, suggesting that association of several VEGF isoforms may be more efficient for treating ischemic diseases than the use of any single variant.

Magnetic-Responsive Carbon Nanotubes Composite Scaffolds for Chondrogenic Tissue Engineering.

The demand for engineered scaffolds capable of delivering multiple cues to cells continues to grow as the interplay between cell fate with microenvironmental and external cues is revealed. Emphasis has been given to develop stimuli-responsive scaffolds. These scaffolds are designed to sense an external stimulus triggering a specific response (e.g., change in the microenvironment, release therapeutics, etc.) and then initiate/modulate a desired biofunction. Here, magnetic-responsive carboxylated multi-walled carbon nanotubes (cMWCNTs) are integrated into 3D collagen/polylactic acid (PLA) scaffold via a reproducible filtration-based method. The integrity and biomechanical performance of the collagen/PLA scaffolds are preserved after cMWCNT integration. In vitro safety assessment of cMWCNT/collagen/PLA scaffolds shows neither cytotoxicity effects nor macrophage pro-inflammatory response, supporting further in vitro studies. The cMWCNT/collagen/PLA scaffolds enhance chondrocytes metabolic activity while maintaining high cell viability and extracellular matrix (i.e., type II collagen and aggrecan) production. Comprehensive in vitro study applying static and pulsed magnetic field on seeded scaffolds shows no specific cell response in dependence with the applied field. This result is independent of the presence or absence of cMWCNT into the collagen/PLA scaffolds. Taken together, these findings provide additional evidence of the benefits to exploit the CNTs outstanding properties in the design of stimuli-responsive scaffolds.

Effects of platelet-rich plasma (PRP) on the healing of Achilles tendons of rats.

Platelet-rich plasma (PRP) contains growth factors involved in the tissular healing process. The aim of the study was to determine if an injection of PRP could improve the healing of sectioned Achilles tendons of rats. After surgery, rats received an injection of PRP (n = 60) or a physiological solution (n = 60) in situ. After 5, 15, and 30 days, 20 rats of both groups were euthanized and 15 collected tendons were submitted to a biomechanical test using cryo-jaws before performing transcriptomic analyses. Histological and biochemical analyses were performed on the five remaining tendons in each group. Tendons in the PRP group were more resistant to rupture at 15 and 30 days. The mechanical stress was significantly increased in tendons of the PRP group at day 30. Histological analysis showed a precocious deposition of fibrillar collagen at day 5 confirmed by a biochemical measurement. The expression of tenomodulin was significantly higher at day 5. The messenger RNA levels of type III collagen, matrix metalloproteinases 2, 3, and 9, were similar in the two groups at all time points, whereas type I collagen was significantly increased at day 30 in the PRP group. In conclusion, an injection of PRP in sectioned rat Achilles tendon influences the early phase of tendon healing and results in an ultimately stronger mechanical resistance.

Histochemical quantification of collagen content in articular cartilage.

BACKGROUND: Articular cartilage (AC) is mainly composed of water, type II collagen, proteoglycans (PGs) and chondrocytes. The amount of PGs in AC is routinely quantified with digital densitometry (DD) from Safranin O-stained sections, but it is unclear whether similar method could be used for collagens. OBJECTIVE: The aim of this study was to clarify whether collagens can be quantified from histological AC sections using DD. MATERIAL AND METHODS: Sixteen human AC samples were stained with Masson's trichrome or Picrosirius red. Optical densities of histological stains were compared to two commonly used collagen parameters (amide I and collagen CH2 side chain peak at 1338cm-1) measured using Fourier Transform Infrared (FTIR) spectroscopic imaging. RESULTS: Optical density of Modified Masson's trichrome staining, which included enzymatic removal of PGs before staining, correlated significantly with FTIR-derived collagen parameters at almost all depths of cartilage. The other studied staining protocols displayed significant correlations with the reference parameters at only few depth layers. CONCLUSIONS: Based on our findings, modified Masson's trichrome staining protocol is suitable for quantification of AC collagen content. Enzymatic removal of PGs prior to staining is critical as us allows better staining of the collagen. Further optimization of staining protocols may improve the results in the future studies.

Vascular Endothelial Growth Factor-111 (VEGF-111) and tendon healing: preliminary results in a rat model of tendon injury.

Tendon lesions are among the most frequent musculoskeletal pathologies. Vascular endothelial growth factor (VEGF) is known to regulate angiogenesis. VEGF-111, a biologically active and proteolysis-resistant splice variant of this family, was recently identified. This study aimed at evaluating whether VEGF-111 could have a therapeutic interest in tendon pathologies. Surgical section of one Achilles tendon of rats was performed before a local injection of either saline or VEGF-111. After 5, 15 and 30 days, the Achilles tendons of 10 rats of both groups were sampled and submitted to a biomechanical tensile test. The force necessary to induce tendon rupture was greater for tendons of the VEGF-111 group (p<0.05) while the section areas of the tendons were similar. The mechanical stress was similar at 5 and 15 days in the both groups but was improved for the VEGF-111 group at day 30 (p <0.001). No difference was observed in the mRNA expression of collagen III, tenomodulin and MMP-9. In conclusion, we observed that a local injection of VEGF-111 improves the early phases of the healing process of rat tendons after a surgical section. Further confirmatory experimentations are needed to consolidate our results.