Integrative Proteomics and Phosphoproteomics Profiling of Symptomatic Accessory Navicular Bone Based on Tandem Mass Tag Technology

Bin Liu,* Ran Wei,* Yuqing Wang, Zishen Cheng, Liangliang Jiang, Xiaopeng Pu, Yaxing Zhang, Yantao Wang, Qiangjun Kang Department of Orthopaedics, Bethune International Peace Hospital, Shijiazhuang, 050082, China*These authors contributed equally to this workCorrespondence: Qiangjun Kang, Department of Orthopaedics, Bethune International Peace Hospital, No. 398, Zhongshan Road, Qiaoxi District, Shijiazhuang, Hebei, 050082, China, Email [email protected]: The accessory navicular bone (ANB) is a common accessory bone in the foot. Certain ANBs significantly impair patients’ feet normal walking function. Foot injury is associated with ANB after athletic training. However, the molecular mechanism of foot injury with ANB after athletic training remains unclear. This study aims to investigate the proteomics and phosphoproteomics profile of foot injury with the ANB after athletic training.Patients and Methods: We collected ANB tissues and normal bone (NB) tissues from 5 foot injury patients with ANB after 3 months of athletic training to perform proteome sequencing by tandem mass tag (TMT) technology. Then, the differentially expressed proteins (DEPs) and phosphorylation proteins (DPPs) were identified between the ANB and NB groups. Furthermore, the potential functions of DEPs and DPPs were annotated, respectively. Besides, the protein-protein interaction (PPI) network was constructed for DEPs.Results: A total of 147 DEPs (129 upregulated and 18 downregulated) were detected. Functional enrichment suggested that they were involved in extracellular matrix (ECM)-receptor interaction and cell adhesion. PPI network showed that COL4A1 and COL4A2 had the highest interaction score, followed by RBBP4 and RBBP7. In addition, phosphoproteomics analysis identified 4 upregulated and 1 downregulated DPPs, and they were primarily enriched in regulating lipolysis in adipocytes.Conclusion: Our study found that foot injury with ANB after exercise training may be associated with proteins related to inflammation and immunity (such as MRC1, UBE2N, CYCS), bone repair and regeneration (such as Emilin2, COL4A1, COL4A2, and ITGA9), and bone microstructure homeostasis (such as GCA and ANXA3). This provides new insights into understanding its pathogenesis and guiding treatment strategies.Keywords: accessory navicular bone, athletic training, proteomics, phosphoproteomics