Abstract
Stem cell therapy and tissue engineering represent a promising approach for cartilage regeneration. However, they present limits in terms of mechanical properties and premature de-differentiation of engineered cartilage. Cycloastragenol (CAG), a triterpenoid saponin compound and a hydrolysis product of the main ingredient in Astragalus membranaceous, has been explored for cartilage regeneration. The aim of this study was to investigate CAG’s ability to promote cell proliferation, maintain cells in their stable active phenotype, and support the production of cartilaginous extracellular matrix (ECM) in human adipose-derived mesenchymal stem cells (hAMSCs) in up to 28 days of three-dimensional (3D) chondrogenic culture. The hAMSC pellets were cultured in chondrogenic medium (CM) and in CM supplemented with CAG (CAG–CM) for 7, 14, 21, and 28 days. At each time-point, the pellets were harvested for histological (hematoxylin and eosin (H&E)), histochemical (Alcian-Blue) and immunohistochemical analysis (Type I, II, and X collagen, aggrecan, SOX9, lubricin). After excluding CAG’s cytotoxicity (MTT Assay), improved cell condensation, higher glycosaminoglycans (sGAG) content, and increased cell proliferation have been detected in CAG–CM pellets until 28 days of culture. Overall, CAG improved the chondrogenic differentiation of hAMSCs, maintaining stable the active chondrocyte phenotype in up to 28 days of 3D in vitro chondrogenic culture. It is proposed that CAG might have a beneficial impact on cartilage regeneration approaches.
Highlights
Articular cartilage shows poor regenerative properties, due to its alymphatic, avascular, aneural nature and the limited cellularity of the tissue, composed of chondrocytes sparsely embedded within the collagen and proteoglycan-based extracellular matrix (ECM) [1]
It has been shown that one of the most important age-related epigenetic factors associated with OA is accelerated telomere shortening [3]. The latter implies that OA chondrocytes lose their ability to synthesize matrix components and begin to synthesize proteins, which contribute to ECM degradation [4]
The present study aimed to investigate, for the first time in literature, the effects of CAG on chondrogenic differentiation of human adipose-derived Mesenchymal stem cells (MSCs) cultured as self-assembling 3D spherical aggregates, in terms of maintenance of stable chondrocyte phenotype, anti-hypertrophic property, and ECM component deposition
Summary
Articular cartilage shows poor regenerative properties, due to its alymphatic, avascular, aneural nature and the limited cellularity of the tissue, composed of chondrocytes sparsely embedded within the collagen and proteoglycan-based extracellular matrix (ECM) [1]. Changes in biomechanical and metabolic features of articular cartilage, related to aging or injury, lead to matrix degradation, resulting in progressive tissue degeneration and causing severe pain and disability of the joint. An inadequate tissue repair, followed by a progressive loss of cartilage and remodeling of the underlying subchondral bone, leads to osteoarthritis (OA), a very common form of severe degenerative articular cartilage disease [2]. It has been shown that one of the most important age-related epigenetic factors associated with OA is accelerated telomere shortening [3]. The latter implies that OA chondrocytes lose their ability to synthesize matrix components and begin to synthesize proteins, which contribute to ECM degradation [4]
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