Chondrocyte Migration Affects Tissue-Engineered Cartilage Integration by Activating the Signal Transduction Pathways Involving Src, PLCγ1, and ERK1/2

Abstract

To determine the signal transduction pathways involved in chondrocyte migration and their effects on cartilage integration in autologous chondrocyte implantation. Articular chondrocytes were divided into three inhibitor groups pretreated with different inhibitors to Src, phospholipase Cγ1 (PLCγ1), and extracellular signal-regulated kinase (ERK)1/2 signaling pathways and one control group pretreated with vehicle. The effect of these pathways on chondrocyte migration was first explored by Boyden chamber assay, and then by an in vitro cell/ring integration model. Chondrocyte migration was visualized and quantified by cell tracking, and the activity of Src, PLCγ1, and ERK1/2 was determined by Western blotting. The effect of these pathways on cartilage integration was evaluated histologically, biochemically, and biomechanically. Boyden chamber assay revealed that the number of migrated cells was significantly increased in the control group without inhibitors. In an in vitro integration model, the implanted chondrocytes were observed to migrate through the interface and infiltrate into the native cartilage. Additionally, chondrocyte migration could be improved in the absence of inhibitors After 4 weeks of culture, the control group demonstrated a significantly higher cellularity, larger amount of chemical content deposition, stronger extracellular matrix staining in the integration zone, and higher integrative strength as compared to the inhibitor groups. Western blotting demonstrated that the Src-PLCγ1-ERK1/2 signaling pathway was promoted in the integration process. This study is the first to show that the Src-PLCγ1-ERK1/2 signaling transduction pathway is involved in cartilage tissue integration by affecting chondrocyte migration. Our results raise the importance of the chondrocyte migration enhancement therapy or the development of new agents specifically targeting the pathways to ensure long-term functionality of the restored joint surface.