TGF-β1 regulates chondrocyte proliferation and extracellular matrix synthesis via circPhf21a-Vegfa axis in osteoarthritis

Abstract

Background The transforming growth factor-beta (TGF-β) signaling pathway is an important pathway associated with the pathogenesis of osteoarthritis (OA). This study was to investigate the involvement of circRNAs in the TGF-β signaling pathway. Methods Cell Counting Kit-8 (CCK-8) assay and 5-ethynyl-2′-deoxyuridine (EdU) assay were used to detect the proliferation of primary mouse chondrocytes (PMCs). RNA-sequencing together with bioinformatics analysis were used to systematically clarify TGF-β1 induced alternations of circRNAs in PMCs. The regulatory and functional role of circPhf21a was examined in PMCs. Downstream targets of circPhf21a were explored by RNA-sequencing after overexpression of circPhf21a and verified by RT-qPCR in PMCs. Finally, the role and mechanism of circPhf21a in OA were explored in mouse models. Results We found that TGF-β1 promoted the proliferation of PMCs. Meanwhile, RT-qPCR and western blotting indicated that TGF-β1 promoted extracellular matrix (ECM) anabolism. RNA-sequencing revealed that a total of 36 circRNAs were differentially expressed between PMCs treated with and without TGF-β1. Of these, circPhf21a was significantly decreased by TGF-β1. Furthermore, circPhf21a knockdown promoted the proliferation and ECM synthesis of PMCs, whereas overexpression of circPhf21a showed the opposite effects. Mechanically, the expression profiles of the mRNAs revealed that Vegfa may be the target of circPhf21a. Additionally, we found that circPhf21a was significantly upregulated in the mouse OA model, and inhibition of circPhf21a significantly relieved the progression of OA. Conclusions Our results found that TGF-β1 promoted the proliferation and ECM synthesis of PMCs via the circPhf21a-Vegfa axis, which may provide novel therapeutic targets for OA treatment.

circRNA-ZCCHC14 affects the chondrogenic differentiation ability of peripheral blood-derived mesenchymal stem cells by regulating GREM1 through miR-181a

“…For example, circ_0000423 regulates cartilage ECM synthesis through the miRNA-27b-3p/MMP-13 axis 29 . TGF-β1 regulates chondrocyte proliferation and extracellular matrix synthesis in osteoarthritis through the circPhf21a-Vegfa axis 30 . circCDK14 ameliorated IL-1β-induced osteoarthritis chondrocyte injury through the miR-1183/KLF5 pathway 31 .…”

Co-aggregation of MSC/chondrocyte in a dynamic 3D culture elevates the therapeutic effect of secreted extracellular vesicles on osteoarthritis in a rat model

“…Another study by Kubosch et al has shown that the co-culture of chondrocytes with human synovial MSCs (SMSCs) leads to self-organization, chondrogenic differentiation of SMSC, and TGFβ secretion 43 . In addition, researchers demonstrated that TGFβ could stimulate chondrogenesis by provoking the expression of Sox9 and Col II, and TGFβ signaling could lead to further stimulation of chondrocyte proliferation and inhibition of chondrocyte hypertrophy 44 , 45 . These data justify our results about the increasing diameter of micromass by Co-ag-EV and MSC-ag-EV and their chondrogenic potential on MSC micromass culture, but more research is still required, especially about the existence of TGFβ in Cho-ag-EV.…”

Sirtuin 4 (Sirt4) downregulation contributes to chondrocyte senescence and osteoarthritis via mediating mitochondrial dysfunction

“…The extraction of total protein from chondrocytes was performed using RIPA lysis buffer (Solarbio, China) supplemented with 1% PMSF protease inhibitor (Solarbio, China). Western blotting was performed following established protocols as described in our previous work 32 . Firstly, the protein concentrations were determined using the Enhanced Bicinchoninic Acid (BCA) protein assay kit (Beyotime, China).…”