Fabrication of Tissue-Engineered Cartilage Using Decellularized Scaffolds and Chondrocytes.
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ABSTRACT:
SUBMITTER: Lu L
PROVIDER: S-EPMC9316171 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:PURPOSE:The significant shortcomings associated with current autologous reconstructive options for auricular deformities have inspired great interest in a tissue engineering solution. A major obstacle in the engineering of human auricular cartilage is the availability of sufficient autologous human chondrocytes. A clinically obtainable amount of auricular cartilage tissue (ie, 1 g) only yields approximately 10 million cells, where 25 times this amount is needed for the fabrication of a full-scale pediatric ear. It is thought that repeated passaging of chondrocytes leads to dedifferentiation and loss of the chondrogenic potential. However, little to no data exist regarding the ideal number of times that human auricular chondrocytes (HAuCs) can be passaged in a manner that maximizes the cellular expansion while minimizing dedifferentiation. METHODS:Human auricular chondrocytes were isolated from discarded otoplasty specimens. The HAuCs were then expanded, and cells from passages 3, 4, and 5 were encapsulated into discs 8 mm in diameter made from type I collagen hydrogels with a cell density of 25 million cells/mL. The constructs were implanted subcutaneously in the dorsa of nude mice and harvested after 1 and 3 months for analysis. RESULTS:Constructs containing passages 3, 4, and 5 chondrocytes all maintained their original cylindrical geometry. After 3 months in vivo, the diameters of the P3, P4, and P5 discs were 69 ± 9%, 67 ± 10%, and 73 ± 15% of their initial diameter, respectively. Regardless of the passage number, all constructs developed a glossy white cartilaginous appearance, similar to native auricular cartilage. Histologic analysis demonstrated development of an organized perichondrium composed of collagen, a rich proteoglycan matrix, cellular lacunae, and a dense elastin fibrin network by Safranin-O and Verhoeff stain. Biochemical analysis confirmed similar amounts of proteoglycan and hydroxyproline content in late passage constructs when compared with native auricular cartilage. CONCLUSIONS:These data indicate that late passage HAuCs (up to passage 5) form elastic cartilage that is histologically, biochemically, and biomechanically similar to native human elastic cartilage and have the potential to be used for auricular cartilage engineering.
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Project description:Articular cartilage defects, and subsequent degeneration, are prevalent and account for the poor quality of life of most elderly persons; they are also one of the main predisposing factors to osteoarthritis. Articular cartilage is an avascular tissue and, thus, has limited capacity for healing and self-repair. Damage to the articular cartilage by trauma or pathological causes is irreversible. Many approaches to repair cartilage have been attempted with some potential; however, there is no consensus on any ideal therapy. Tissue engineering holds promise as an approach to regenerate damaged cartilage. Since cell adhesion is a critical step in tissue engineering, providing a 3D microenvironment that recapitulates the cartilage tissue is vital to inducing cartilage regeneration. Decellularized materials have emerged as promising scaffolds for tissue engineering, since this procedure produces scaffolds from native tissues that possess structural and chemical natures that are mimetic of the extracellular matrix (ECM) of the native tissue. In this work, we present, for the first time, a study of decellularized scaffolds, produced from avian articular cartilage (extracted from Gallus Gallus domesticus), reseeded with human chondrocytes, and we demonstrate for the first time that human chondrocytes survived, proliferated and interacted with the scaffolds. Morphological studies of the decellularized scaffolds revealed an interconnected, porous architecture, ideal for cell growth. Mechanical characterization showed that the decellularized scaffolds registered stiffness comparable to the native cartilage tissues. Cell growth inhibition and immunocytochemical analyses showed that the decellularized scaffolds are suitable for cartilage regeneration.
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Project description:The scaffold is a key element in cartilage tissue engineering. The components of Wharton's jelly are similar to those of articular cartilage and it also contains some chondrogenic growth factors, such as insulin-like growth factor I and transforming growth factor-?. We fabricated a tissue-engineered cartilage scaffold derived from Wharton's jelly extracellular matrix (WJECM) and compared it with a scaffold derived from articular cartilage ECM (ACECM) using freeze-drying. The results demonstrated that both WJECM and ACECM scaffolds possessed favorable pore sizes and porosities; moreover, they showed good water uptake ratios and compressive moduli. Histological staining confirmed that the WJECM and ACECM scaffolds contained similar ECM. Moreover, both scaffolds showed good cellular adherence, bioactivity, and biocompatibility. MTT and DNA content assessments confirmed that the ACECM scaffold tended to be more beneficial for improving cell proliferation than the WJECM scaffold. However, RT-qPCR results demonstrated that the WJECM scaffold was more favorable to enhance cellular chondrogenesis than the ACECM scaffold, showing more collagen II and aggrecan mRNA expression. These results were confirmed indirectly by glycosaminoglycan and collagen content assessments and partially confirmed by histology and immunofluorescent staining. In conclusion, these results suggest that a WJECM scaffold may be favorable for future cartilage tissue engineering.
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