Abstract
Adipose-derived mesenchymal stem cell (ADSC) with a high capacity of chondrogenic differentiation was a promising candidate for cartilage defect treatment. This study's objective is to study the roles of integrin β1 (ITGB1) in regulating ADSC chondrogenic differentiations as well as the underlying mechanisms. The identity of ADSC was confirmed by flow cytometry. ITGB1 gene was overexpressed in human ADSC (hADSC) by transfection with LV003-recombinant plasmids. Gene mRNA and protein levels were examined using quantitative RT-PCR and western blotting, respectively. Differentially expressed mRNAs and proteins were characterized by next-generation RNA sequencing and label-free quantitative proteomics, respectively. ERK signaling and AKT signaling in hADSCs were inhibited by treating with SCH772984 and GSK690693, respectively. ITGB1 gene overexpression substantially increased collagen type II alpha 1 chain (COL2A1), aggrecan (ACAN), and SRY-box transcription factor 9 (SOX9) expression but suppressed collagen type I alpha 1 chain (COL1A1) expression in hADSCs. Next-generation RNA sequencing identified a total of 246 genes differentially expressed in hADSCs by ITGB1 overexpression, such as 183 upregulated and 63 downregulated genes. Label-free proteomics characterized 34 proteins differentially expressed in ITGB1-overexpressing hADSCs. Differentially expressed genes and proteins were enriched by different biological processes such as cell adhesion and differentiation and numerous signaling pathways such as the ERK signaling pathway. ERK inhibitor treatment caused substantially enhanced chondrogenic differentiation in ITGB1-overexpressing hADSCs. ITGB1 promoted the chondrogenic differentiation of human ADSCs via the activation of the ERK signaling pathway.