Human amniotic membrane modulates Wnt/β-catenin and NF-κβ signaling pathways in articular chondrocytes in vitro

• Published in: Osteoarthritis and cartilage open Vol. 3; no. 4; p. 100211
• Main Authors: Chung, Connie, Massee, Michelle, Koob, Thomas J.
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 01.12.2021; Elsevier
• Subjects: Amniotic membrane; Chondrocyte; ECM catabolism; NF-κβ; ORIGINAL PAPER; Osteoarthritis; Wnt; NF-κβ; Amniotic membrane; ECM catabolism; Chondrocyte; Wnt; Osteoarthritis
• ISSN: 2665-9131; 2665-9131
• DOI: 10.1016/j.ocarto.2021.100211

Inflammation, catabolism, and hypertrophy in chondrocytes play a central role in osteoarthritis (OA). The Wnt/β-catenin and NF-κβ pathways contribute to these degradative processes. This in vitro study evaluates the inhibitory effect of a novel therapeutic, micronized dehydrated human amnion/chorion membrane (μdHACM), as a potential treatment to offset elevated Wnt/β-catenin and NF-κβ signaling. Three-dimensional human articular chondrocyte pellets were stimulated with an inflammatory cocktail to induce a degenerative phenotype. Treatments included varying doses of μdHACM. Protein and gene expression were analyzed using qRT-PCR, immunoblotting, and immunofluorescence to assess changes in the major constituents of Wnt/β-catenin and NF-κβ signaling. Regulation of catabolic activity was evaluated using enzymatic assays that detect MMP-13 and aggrecanase-mediated degradation products in conditioned media. Confirmation of the model was established through the expression of specific markers and extracellular matrix genes, verifying a chondrogenic phenotype was maintained. Inflammatory stimulation elicited a change in the chondrocyte proteome and secretome, elevating Wnt/β-catenin and NF-κβ signaling and downstream expression of inflammatory, proteolytic, and hypertrophic markers, while decreasing primary cartilage matrix components, ACAN and COL2A1. μdHACM reversed these inflammatory-induced changes, suppressing phospho-GSK-3β, β-catenin expression/nuclear localization of the Wnt signaling axis and inhibiting IKKβ, phospho-IκBα, and phospho-p65 in the NF-κβ signaling cascade. Additionally, μdHACM altered expression of direct downstream targets, namely MCP1, MMP3, MMP13, ADAMTS4, ADAMTS5, RUNX2 and COL10A1. Moreover, μdHACM reduced MMP-13 and aggrecanase-mediated substrate degradation. μdHACM ameloriated the effects of inflammatory-induced degeneration in chondrocytes through Wnt/β-catenin and NF-κβ inhibition, subsequently downregulating key inflammatory, hypertrophic and catabolic mediators in vitro.