Abstract
Summary
RNAi targeting p110β reduces TNF-alpha production and osteolysis in response to wear particles.
Introduction
Aseptic joint loosening is a key factor that reduces the life span of joint prosthesis. Prosthetic wear particles are thought to play a central role in the initiation and development of periprosthetic osteolysis, leading to aseptic loosening of prostheses. This study aims to explore the effect of p110β-targeted small interfering RNA (siRNA) and lentivirus on particle-induced inflammatory cytokine expression in murine macrophage.
Methods
siRNA and lentivirus targeting p110β were transfected and infected prior to particle stimulation, respectively. Ceramic and titanium particles of different sizes were prepared to stimulate macrophages. Fluorescence microscopy showed that the siRNA transfection and lentivirus infection efficiency were 74.2 ± 4.2% and 92.3 ± 2.6%, respectively.
Results
Real-time polymerase chain reaction (PCR) showed that the levels of tumor necrosis factor-alpha (TNF-alpha) mRNA in the particle stimulation plus RNA interference (RNAi) groups were significantly lower compared with the particle stimulation-only groups (P<0.05), respectively. Similarly, enzyme-linked immunosorbent assay (ELISA) showed that protein levels of TNF-alpha in RNAi-treated groups were significantly decreased after transfection or infection (P<0.05), respectively. Western Blot showed that Phospho-Akt activation was significantly reduced by RNAi. As assessed by CT and micro-CT, particle implantation induced a significant osteolysis effect in mice calvaria, which was limited by p110β-lentivirus addition.
Conclusions
p110β subtype of PI3K, followed by activation of phosphor-AKT (Ser473), may possibly participate in the regulation of activating macrophages by wear particles, ultimately resulting in the secretion of TNF-α and osteolysis.
