L. Utomo

Preparation and characterization of a decellularized cartilage scaffold for ear cartilage reconstruction

Scaffolds are widely used to reconstruct cartilage. Yet, the fabrication of a scaffold with a highly organized microenvironment that closely resembles native cartilage remains a major challenge. Scaffolds derived from acellular extracellular matrices are able to provide such a microenvironment. Currently, no report specifically on decellularization of full thickness ear cartilage has been published. In this study, decellularized ear cartilage scaffolds were prepared and extensively characterized. Cartilage decellularization was optimized to remove cells and cell remnants from elastic cartilage. Following removal of nuclear material, the obtained scaffolds retained their native collagen and elastin contents as well as their architecture and shape. High magnification scanning electron microscopy showed no obvious difference in matrix density after decellularization. However, glycosaminoglycan content was significantly reduced, resulting in a loss of viscoelastic properties. Additionally, in contact with the scaffolds, human bone-marrow-derived mesenchymal stem cells remained viable and are able to differentiate toward the chondrogenic lineage when cultured in vitro. These results, including the ability to decellularize whole human ears, highlight the clinical potential of decellularization as an improved cartilage reconstruction strategy.

Cartilage inflammation and degeneration is enhanced by pro-inflammatory (M1) macrophages in vitro, but not inhibited directly by anti-inflammatory (M2) macrophages

OBJECTIVE Macrophages play a crucial role in the progression of osteoarthritis (OA). Their phenotype may range from pro-inflammatory to anti-inflammatory. The aim of this study was to evaluate the direct effects of macrophage subtypes on cartilage by culturing macrophage conditioned medium (MCM) on human articular cartilage. DESIGN Human OA cartilage explants were cultured with MCM of pro-inflammatory M(IFNγ+TNFα), or anti-inflammatory M(IL-4) or M(IL-10) human monocyte-derived macrophages. To assess effects of anti-inflammatory macrophages, the cartilage was cultured with a combination of MCM phenotypes as well as pre-stimulated with IFNγ+TNFα cartilage before culture with MCM. The reactions of the explants were assessed by gene expression, nitric oxide (NO) production and release of glycosaminoglycans (GAGs). RESULTS M(IFNγ+TNFα) MCM affected OA cartilage by upregulation of IL1B (Interleukin 1β), IL6, MMP13 (Matrix Metalloproteinase-13) and ADAMTS5 (A Disintegrin And Metalloproteinase with Thrombospondin Motifs-5), while inhibiting ACAN (aggrecan) and COL2A1 (collagen type II). M(IL-10) upregulated IL1B and Suppressor of cytokine signaling 1 (SOCS1). NO production and GAG release by the cartilage was increased when cultured with M(IFNγ+TNFα) MCM. M(IL-4) and M(IL-10) did not inhibit the effects of M(IFNγ+TNFα) MCM of neither phenotype affected IFNγ+TNFα pre-stimulated cartilage, in which an inflammatory gene response was deliberately induced. CONCLUSION M(IFNγ+TNFα) macrophages have a prominent direct effect on OA cartilage, while M(IL-4) and M(IL-10) do not inhibit the effects of M(IFNγ+TNFα), or IFNγ+TNFα induced inflammation of the cartilage. Therapies aiming at inhibiting cartilage degeneration may take this into account by directing suppression of pro-inflammatory macrophages or stimulation of anti-inflammatory macrophages.

Tissue composition regulates distinct viscoelastic responses in auricular and articular cartilage.

It is well-accepted that articular (ART) cartilage composition and tissue architecture are intimately related to mechanical properties. On the other hand, very little information about other cartilage tissues is available, such as elastin-rich auricular (AUR) cartilage. While thorough investigation of ART cartilage has enhanced osteoarthritis research, ear cartilage reconstruction and tissue engineering (TE) could benefit in a similar way from in-depth analysis of AUR cartilage properties. This study aims to explore the constituent-function relationships of AUR cartilage, and how elastin influences mechanical behavior. Stress-relaxation indentation and tensile tests were performed on bovine ART and AUR cartilage. Elastase incubation was performed to simultaneously deplete elastin and sulfated glycosaminoglycans (sGAG), while hyaluronidase incubation was used to deplete sGAG-only, in order to systematically investigate matrix components in material behavior. ART and AUR cartilages showed different viscoelastic behaviors, with AUR cartilage exhibiting a more elastic behavior. Higher equilibrium properties and limited viscous dissipation of strain energy were observed in AUR cartilage, while ART cartilage exhibited a rapid viscous response and high resistance to instantaneous loading. In conclusion, loss of sGAG had no effect on auricular mechanics in contrast to articular cartilage where GAG loss clearly correlated with mechanical properties. Auricular cartilage without elastin lost all compressive mechanical integrity, whereas in articular cartilage this was provided by collagen. This work shows for the first time the involvement of elastin in the mechanical behavior of ear cartilage. In future, this data can be used in AUR cartilage TE efforts to support reproduction of tissue-specific mechanical properties.

Meniscal extrusion and degeneration during the course of osteoarthritis in the Murine collagenase-induced osteoarthritis model: MENISCAL DEGENERATION DURING EXPERIMENTAL OSTEOARTHRITIS

Meniscal damage is, despite its major role in knee osteoarthritis (OA), often neglected in OA animal models. We evaluated structural meniscal degeneration during the course of OA in the murine collagenase‐induced OA (CIOA) model. To investigate this, OA was induced in the knee joints of 33 male C57BL/6 mice by an intra‐articular injection of 10U collagenase. The mice were sacrificed after 1, 3, 7, 14, 28, and 56 days, and the knees were harvested and processed for histological analysis. As control, six knees were obtained from 16‐week‐old mice in which no OA was induced. Meniscal damage, meniscal extrusion, and articular cartilage damage were evaluated on thionin‐stained sections. Associations between parameters of interest were evaluated with Spearman rho correlation tests. When compared to non‐OA knees, meniscal extrusion was visible from day 1 onwards and meniscal degeneration had a tendency to increase over time. The meniscus damage appeared around the same time as articular cartilage damage (day 14–28) and was statistically significantly more pronounced anterior than posterior, and no differences were seen between medial and lateral menisci. Meniscus and articular cartilage damage were moderately associated in the CIOA knees (ρ = 0.57; 95%CI [0.23–0.78]). Our findings suggest that the CIOA model is a valuable model to study the role of meniscal damage during OA progression and can support the development of future preventative treatment strategies.