Qianbo Chen1, Tianyong Hou, Fei Luo, Xuehui Wu, Zhao Xie, Jianzhong Xu. 1. National and Local United Engineering laboratory of Tissue Engineering, Department of Orthopaedics, Southwest Hospital, the Third Military Medical University, Chongqing, China.
Abstract
BACKGROUND AND AIMS: Osteomyelitis is a common manifestation of invasive Staphylococcus aureus infection characterized by bone loss and destruction. We investigated the role of toll-like receptor 2 (TLR2) in bacterial recognition and clearance in response to infection with an osteomyelitis isolate of S. aureus. METHODS: Apoptosis was assessed in the osteoblastic cell line MC3T3-E1 by Annexin V-FITC/PI staining and flow cytometry. The expression of TLR2 and apoptosis-related and mitogen-activated protein kinase pathway proteins was assessed by qRT-PCR and western blotting. Alkaline phosphatase (ALP) activity and calcium deposition were assessed by ALP activity assay and Alizarin red staining. RESULTS: S. aureus induced apoptosis, upregulated TLR2 expression, and activated mitogen-activated protein kinase pathways in a time dependent manner. Inhibition of the c-Jun N-terminal kinase (JNK) pathway downregulated TLR2 and suppressed the S. aureus induced activation of pro-apoptotic pathways. Short-hairpin RNA mediated silencing of TLR2 reversed S. aureus induced apoptosis and decrease in ALP activity and calcium deposition, and inhibition of JNK had a similar effect. CONCLUSION: We showed that osteoblast apoptosis and osteogenic differentiation in response to bacterial invasion are dependent on TLR2 expression and JNK activation, suggesting novel potential therapeutic targets for the treatment of osteomyelitis.
BACKGROUND AND AIMS: Osteomyelitis is a common manifestation of invasive Staphylococcus aureus infection characterized by bone loss and destruction. We investigated the role of toll-like receptor 2 (TLR2) in bacterial recognition and clearance in response to infection with an osteomyelitis isolate of S. aureus. METHODS: Apoptosis was assessed in the osteoblastic cell line MC3T3-E1 by Annexin V-FITC/PI staining and flow cytometry. The expression of TLR2 and apoptosis-related and mitogen-activated protein kinase pathway proteins was assessed by qRT-PCR and western blotting. Alkaline phosphatase (ALP) activity and calcium deposition were assessed by ALP activity assay and Alizarin red staining. RESULTS:S. aureus induced apoptosis, upregulated TLR2 expression, and activated mitogen-activated protein kinase pathways in a time dependent manner. Inhibition of the c-Jun N-terminal kinase (JNK) pathway downregulated TLR2 and suppressed the S. aureus induced activation of pro-apoptotic pathways. Short-hairpin RNA mediated silencing of TLR2 reversed S. aureus induced apoptosis and decrease in ALP activity and calcium deposition, and inhibition of JNK had a similar effect. CONCLUSION: We showed that osteoblast apoptosis and osteogenic differentiation in response to bacterial invasion are dependent on TLR2 expression and JNK activation, suggesting novel potential therapeutic targets for the treatment of osteomyelitis.
Authors: Johannes Maximilian Wagner; Yonca Steubing; Mehran Dadras; Christoph Wallner; Sebastian Lotzien; Julika Huber; Alexander Sogorski; Maxi Sacher; Felix Reinkemeier; Stephanie Dittfeld; Mustafa Becerikli; Marcus Lehnhardt; Björn Behr Journal: J Bone Miner Metab Date: 2021-09-25 Impact factor: 2.626