Sox9 is a transcription factor that regulates chondrogenesis, but its role in the chondrogenic differentiation of mesenchymal stem cells (MSCs) triggered by materials is poorly understood. In this study, we investigated the effect of Sox9 interference on collagen-induced chondrogenesis and further collagen-based therapies for cartilage defects. In this paper, MSCs were infected with a vector carrying the Sox9 promoter and related markers were detected. A lentivirus-mediated vector targeting the silencing of the Sox9 gene was used in bone marrow-derived MSCs prior to being encapsulated in a collagen hydrogel. The collagen hydrogel as a sole inducer was also compared with transforming growth factor-β1 (TGF-β1). Before being implanted into the articular cartilage defect in rats, the cell-hydrogel pellets were cultured in vitro for 14 days. The effect of Sox9 transfection on cell proliferation was evaluated by measuring the total DNA content. Safranin-O staining and a biochemistry assay were performed to assess the synthesis and secretion of glycosaminoglycan (GAG) of MSCs. The real-time fluorescent quantitative polymerase chain reaction (RT-PCR) was performed to detect the gene expression levels of Col1a1, Col2a1, Acan and Sox9. The protein expression of collagen type II and collagen type I was analyzed by immunohistochemical analysis. Collagen alone significantly increased the luciferase activity of the Sox9 promoter, which was in parallel with the upregulation of cartilage specific markers. In vitro, the chondrogenic differentiation ability of MSCs was greatly inhibited after Sox9 interference, both in the collagen and TGF-β1-induced groups. In vivo, a further study showed that cartilage regeneration was arrested by using transfected MSCs with an injectable collagen gel or induced by TGF-β1. The results indicated that collagen may mediate Sox9 expression by providing a biomimetic microenvironment favoring cell condensation prior to chondrogenesis. The role of Sox9 regulation by materials is similar to that by growth factors, suggesting that well-designed scaffolds may replace growth factors in chondrogenesis. Thus, interventions targeting Sox9 may help improve articular cartilage repair.
Collagen hydrogel has been widely used for osteochondral repair, but its mechanical properties cannot meet the requirements of clinical application. Previous studies have shown that the addition of either polysaccharide or inorganic particles could reinforce the polymer matrix. However, their synergic effects on collagen-based hydrogel have seldom been studied, and the potential application of triple-phased composite gel in osteochondral regeneration has not been reported. In this study, nano-hydroxyapatite (nano-HA) reinforced collagen-alginate hydrogel (nHCA) was prepared by the in situ synthesis of nano-HA in collagen gel followed by the addition of alginate and Ca(2+). The properties of triple-phased nHCA hydrogel were studied and compared with pure collagen and biphasic gels, and the triple-phased composite of collagen-alginate-HA gels showed a superiority in not only mechanical but also biological features, as evidenced by the enhanced tensile and compressive modulus, higher cell viability, faster cell proliferation and upregulated hyaline cartilage markers. In addition, it was found that the synthesis process could also affect the properties of the triple-phased composite, compared to blend-mixing HCA. The in situ-synthesized nHCA hydrogel showed an enhanced tensile modulus, as well as enhanced biological features compared with HCA. Our study demonstrated that the nHCA composite hydrogel holds promise in osteochondral regeneration. The addition of alginate and nano-HA contribute to the increase in both mechanical and biological properties. This study may provide a valuable reference for the design of an appropriate composite scaffold for osteochondral tissue engineering.
The three-phase CCH hydrogel, which is closer to natural cartilage matrix and is stiffer than collagen, may replace collagen as the "gold standard" for cartilage tissue engineering. This study may provide a new insight for cartilage repair using ectopic cartilage reconstructed from functional materials and allogeneic cells.
Intra-articular (IA) injection is an efficient treatment for osteoarthritis, which will minimize systemic side effects. However, the joint experiences rapid clearance of therapeutics after intra-articular injection. Delivering system modified through active targeting strategies to facilitate localization within specific joint tissues such as cartilage is hopeful to increase the therapeutic effects. In this study, we designed a nanoscaled amphiphilic and cartilage-targeting polymer-drug delivery system by using formononetin (FMN)-poly(ethylene glycol) (PEG) (denoted as PCFMN), which was prepared by PEGylation of FMN followed by coupling with cartilage-targeting peptide (CollBP). Our results showed that PCFMN was approximately regular spherical with an average diameter about 218 nm. The in vitro test using IL-1β stimulated chondrocytes indicated that PCFMN was biocompatible and upregulated anabolic genes while simultaneously downregulated catabolic genes of the articular cartilage. The therapeutic effects in vivo indicated that PCFMN could effectively attenuate the progression of OA as evidenced by immunohistochemical staining and histological analysis. In addition, PCFMN showed higher intention time in joints and better anti-inflammatory effects than FMN, indicating the efficacy of cartilage targeting nanodrug on OA. This study may provide a reference for clinical OA therapy.
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