Effects of Parathyroid Hormone on Osteoporotic Fracture Healing in Mice via Non-Phospholipases C-Dependent Protein Kinase C Signaling Pathway

Abstract

Objectives: This study was aimed to explore the effects of parathyroid hormone (PTH) on osteoporotic fracture healing in mice and the underlying mechanisms. Methods: Microarray analysis was conducted to analyze the gene expression level in MC3T3-E1 cells. Carboxyfluorescein succinimidyl ester (CFSE) staining and flow cytometry was adopted to analyze the proliferation and apopto-sis of MC3T3-E1 cells. qRT-PCR was used to analyze the mRNA expression level. Fluorescence resonance energy transfer (FRET) assay was conducted to detect PKC activity. The bone mineral density (BMD) and bone volume (BV)/total volume (TV) were determined via enzyme-linked immunosorbent assay (ELISA) and microscopic computed tomography (micro-CT). Results: ERK1/2 was abnormally expressed in MC3T3-E1 cells after GlylArg19hPTH (1-34) + KT5720 treatment. GlylArg19hPTH (1-34)+ KT5720 treatment promoted cell proliferation, inhibited cell apoptosis, and upregulatedthe expression of osteogenesis-related genes (ALP, OPN, Runx2 and OPG) in MC3T3-E1 cells, which were due to the activation of the non-PLC-dependent PKC signaling pathway and can be blocked by PKC inhibitor Go6983 or ERK1/2 inhibitor BVD-523. Moreover, the activity of PKC in MC3T3-E1 cells treated with GlylArg19hPTH (1-34) + KT5720 + Go6983 was alleviated by ERK1/2 inhibitor BVD-523. In vivo, specific activation of the non-PLC-dependent PKC signaling pathway increased the serum levels of APL and OPG in mice with osteoporotic fracture, which were reversed by PKC inhibitor Go6983 and ERK1/2 inhibitor BVD-523. Moreover, PKC inhibitor Go6983 and ERK1/2 inhibitor BVD-523 suppressed the elevation of BV/TV and BMD induced by specific activation of the non-PKC-dependent signaling pathway. Conclusions: Taken together, PTH stimulates osteoporotic fracture healing in mice through the non-PLC-dependent PKC signaling pathway in which ERK1/2 exerts a vital role.