Single-Cell RNA Sequencing Reveals Transcriptional Changes in the Cartilage of Subchondral Insufficiency Fracture of the Knee

Wang Tang1 *, Zhen-Wei Li1 *, Gui-Qiang Miao2 *, Zhi-Peng Li,1 Tao Gui,1 Chong-Jie Wu,1 Zhen-Yan Li,1 Jie Yang,1 Xiao-Dong Zhao,2 Ning Liu,1 Zhen-Gang Zha,1 Lu-Tian Yao,3 Huan-Tian Zhang1 1Department of Bone and Joint Surgery, The First Affiliated Hospital of Jinan University, Guangzhou, 510630, People’s Republic of China; 2Department of Orthopedics, Foshan Fosun Chancheng Hospital, Foshan, 528010, People’s Republic of China; 3Department of Orthopedics, The First Hospital of China Medical University, Shenyang, 110001, People’s Republic of China*These authors contributed equally to this workCorrespondence: Lu-Tian Yao; Huan-Tian Zhang, Email ltyao@cmu.edu.cn; zhanghuantian@jnu.edu.cnPurpose: Subchondral insufficiency fracture of the knee (SIFK) is a common cause of knee joint pain that mainly afflicts the elderly. Until now, how a sudden insufficiency fracture of subchondral bone affects the transcriptomic profiles of cartilage in SIFK and OA patients are largely unknown.Methods: Single-cell RNA sequencing (scRNA-seq) was used to identify various cell subsets and evaluate transcriptomic differences in cartilage of SIFK and OA patients. In addition, the above findings were confirmed by histological evaluation and immunohistochemical (IHC) staining.Results: We found that the transcriptomic profiles of cartilage in the SIFK patient was completely different from those of normal and OA patients. Accordingly, several novel cell clusters with activation of hypoxia and endochondral ossification signaling were identified in the SIFK cartilage. Chondrocyte trajectories analysis and IHC staining revealed that transcription factors including TCF4 were found to be highly up-regulated during the occurrence of SIFK, which might drive the reactive formation of cartilage and fibrous tissue and the activation of endochondral ossification.Conclusion: This is the first report to elucidate the transcriptomic alterations and distinct cell type subpopulations in the cartilage of SIFK and OA by the use of scRNA-seq, which provides a new insight in the understanding of the initiation and progression of SIFK.Keywords: SIFK, scRNA, TCF4, cartilage, hypoxia