RNA interference targeting p110β reduces tumor necrosis factor-alpha production in cellular response to wear particles in vitro and osteolysis in vivo

摘要

Aseptic joint loosening is a key factor that reduces the life span of arthroplasty. There are currently few effective treatments for joint loosening except surgical revision. We explored the inhibitory effects of p110β-targeted small interfering RNA (siRNA) and lentivirus on particle-induced inflammatory cytokine expression in the murine macrophage cell line, RAW264.7. 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 efficiency of siRNA transfection and lentivirus infection were 74.2 ± 4.2 and 92.3 ± 2.6 %, respectively. 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). Similarly, protein levels of TNF-alpha in RNAi-treated groups were significantly decreased after transfection or infection (P < 0.05). It showed that Phosphor-AKT (Ser473) activation was significantly reduced by RNAi through western blot. As assessed by CT, micro-CT and histological analysis, particle implantation induced a significant osteolysis in mice calvaria, which was limited by p110β lentivirus addition. These results suggested that p110β subtype of PI3K, followed by activation of Ser473, may possibly participate in the regulation of macrophages activity by wear particles, ultimately resulting in the TNF-α secretion and osteolysis.