K.E.M. Benders

Extracellular matrix scaffolds for cartilage and bone regeneration.

Regenerative medicine approaches based on decellularized extracellular matrix (ECM) scaffolds and tissues are rapidly expanding. The rationale for using ECM as a natural biomaterial is the presence of bioactive molecules that drive tissue homeostasis and regeneration. Moreover, appropriately prepared ECM is biodegradable and does not elicit adverse immune responses. Successful clinical application of decellularized tissues has been reported in cardiovascular, gastrointestinal, and breast reconstructive surgery. At present, the use of ECM for osteochondral tissue engineering is attracting interest. Recent data underscore the great promise for future application of decellularized ECM for osteochondral repair. This review describes the rationale for using ECM-based approaches for different regenerative purposes and details the application of ECM for cartilage or osteochondral repair.

Endochondral bone formation in gelatin methacrylamide hydrogel with embedded cartilage-derived matrix particles

The natural process of endochondral bone formation in the growing skeletal system is increasingly inspiring the field of bone tissue engineering. However, in order to create relevant-size bone grafts, a cell carrier is required that ensures a high diffusion rate and facilitates matrix formation, balanced by its degradation. Therefore, we set out to engineer endochondral bone in gelatin methacrylamide (GelMA) hydrogels with embedded multipotent stromal cells (MSCs) and cartilage-derived matrix (CDM) particles. CDM particles were found to stimulate the formation of a cartilage template by MSCs in the GelMA hydrogel in vitro. In a subcutaneous rat model, this template was subsequently remodeled into mineralized bone tissue, including bone-marrow cavities. The GelMA was almost fully degraded during this process. There was no significant difference in the degree of calcification in GelMA with or without CDM particles: 42.5 ± 2.5% vs. 39.5 ± 8.3% (mean ± standard deviation), respectively. Interestingly, in an osteochondral setting, the presence of chondrocytes in one half of the constructs fully impeded bone formation in the other half by MSCs. This work offers a new avenue for the engineering of relevant-size bone grafts, by the formation of endochondral bone within a degradable hydrogel.

Of Mice, Men and Elephants: The Relation between Articular Cartilage Thickness and Body Mass

Mammalian articular cartilage serves diverse functions, including shock absorption, force transmission and enabling low-friction joint motion. These challenging requirements are met by the tissue’s thickness combined with its highly specific extracellular matrix, consisting of a glycosaminoglycan-interspersed collagen fiber network that provides a unique combination of resilience and high compressive and shear resistance. It is unknown how this critical tissue deals with the challenges posed by increases in body mass. For this study, osteochondral cores were harvested post-mortem from the central sites of both medial and lateral femoral condyles of 58 different mammalian species ranging from 25 g (mouse) to 4000 kg (African elephant). Joint size and cartilage thickness were measured and biochemical composition (glycosaminoclycan, collagen and DNA content) and collagen cross-links densities were analyzed. Here, we show that cartilage thickness at the femoral condyle in the mammalian species investigated varies between 90 µm and 3000 µm and bears a negative allometric relationship to body mass, unlike the isometric scaling of the skeleton. Cellular density (as determined by DNA content) decreases with increasing body mass, but gross biochemical composition is remarkably constant. This however need not affect life-long performance of the tissue in heavier mammals, due to relatively constant static compressive stresses, the zonal organization of the tissue and additional compensation by joint congruence, posture and activity pattern of larger mammals. These findings provide insight in the scaling of articular cartilage thickness with body weight, as well as in cartilage biochemical composition and cellularity across mammalian species. They underscore the need for the use of appropriate in vivo models in translational research aiming at human applications.

Multipotent Stromal Cells Outperform Chondrocytes on Cartilage-Derived Matrix Scaffolds

Objective Although extracellular matrix (ECM)–derived scaffolds have been extensively studied and applied in a number of clinical applications, the use of ECM as a biomaterial for (osteo)chondral regeneration is less extensively explored. This study aimed at evaluating the chondrogenic potential of cells seeded on cartilage-derived matrix (CDM) scaffolds in vitro. Design Scaffolds were generated from decellularized equine articular cartilage and seeded with either chondrocytes or multipotent stromal cells (MSCs). After 2, 4, and 6 weeks of in vitro culture, CDM constructs were analyzed both histologically (hematoxylin and eosin, Safranin-O, collagen types I and II) and biochemically (glycosaminoglycan [GAG] and DNA content). Results After 4 weeks, both cell types demonstrated chondrogenic differentiation; however, the MSCs significantly outperformed chondrocytes in producing new GAG-containing cartilaginous matrix. Conclusion These promising in vitro results underscore the potency of CDM scaffolds in (osteo)chondral defect repair.

Formalin fixation affects equilibrium partitioning of an ionic contrast agent-microcomputed tomography (EPIC-μCT) imaging of osteochondral samples

OBJECTIVE Equilibrium Partitioning of an Ionic Contrast agent with microcomputed tomography (EPIC-μCT) is a non-invasive technique to quantify and visualize the three-dimensional distribution of glycosaminoglycans (GAGs) in fresh cartilage tissue. However, it is unclear whether this technique is applicable to already fixed tissues. Therefore, this study aimed at investigating whether formalin fixation of bovine cartilage affects X-ray attenuation, and thus the interpretation of EPIC-μCT data. DESIGN Osteochondral samples (n=24) were incubated with ioxaglate, an ionic contrast agent, for 22h prior to μCT scanning. The samples were scanned in both formalin-fixed and fresh conditions. GAG content was measured using a biochemical assay and normalized to wet weight, dry weight, and water content to determine potential reasons for differences in X-ray attenuation. RESULTS The expected zonal distribution of contrast agent/GAGs was observed for both fixed and fresh cartilage specimens. However, despite no significant differences in GAG concentrations or physical properties between fixed and fresh samples, the average attenuation levels of formalin-fixed cartilage were 14.3% lower than in fresh samples. CONCLUSIONS EPIC-μCT is useful for three-dimensional visualization of GAGs in formalin-fixed cartilage. However, a significant reduction in X-ray attenuation for fixed (compared to fresh) cartilage must be taken into account and adjusted for accordingly when quantifying GAG concentrations using EPIC-μCT.