Periplaneta Americana extract inhibits osteoclastic differentiation in vitro.

Abstract

Periplaneta americana extract (PAE) is proven to be promising in treating fever, wound healing, liver fibrosis, and cardiovascular disease. However, the role of PAE in skeletal disorders remains unclear. This study investigated whether PAE regulates osteoclastic differentiation in vitro via the culture using RAW264.7 cells and bone marrow derived macrophages (BMDMs). RAW264.7 cells and BMDMs were cultured and induced for osteoclastic differentiation supplementing with different concentrations of PAE (0, 0.1, 1, and 10mg/mL). Cell counting kit-8 (CCK-8) assay was used to detect the cytotoxicity and cell proliferation. TRAP staining, actin ring staining, real-time quantitative PCR (RT-qPCR), and bone resorption activity test were performed to detect osteoclastic differentiation. RT-qPCR and enzyme-linked immunosorbent assay (ELISA) were conducted to assay the expression and secretion of inflammatory factors. RNA sequencing (RNA-seq) and western blot analysis were carried out to uncover the underlying mechanism. CCK-8 results showed that 10mg/mL and a lower concentration of PAE did not affect cell proliferation. RT-qPCR analysis verified that PAE down-regulated the osteoclastic genes Nfatc1, Ctsk, and Acp5 in macrophages. Moreover, PAE restrained the differentiation, formation, and function of osteoclasts. Besides, RT-qPCR and ELISA assays showed that PAE decreased inflammatory genes expression and reduced the secretion of inflammatory factors, including IL1β, IL6, and TNFα. Subsequent RNA-seq analysis identified possible genes and signaling pathways of PAE-mediated osteoclastogenesis suppression. Our study indicates that PAE has inhibitive effects on osteoclastogenesis and may be a potential therapeutic alternative for bone diseases.