2021 Volume 30 Issue 4 Pages 333-338
Osteoclast differentiation is one of the key steps that regulate bone mass and involves RANKL-induced changes in the intracellular Ca2+ levels. Previously, we reported the function of a lysosomal Ca2+ channel, two-pore channel (TPC) subtype 2 (TPC2), using TPC2-knockout RAW264.7 cell line (RAW) during osteoclastogenesis. However, the effects of overexpressed TPC2 have not been examined because of the relatively low lipid-based efficiency transfection in RAW. In this study, TPC2 was transfected in RAW and RAW-derived mature osteoclast-like cells using an improved lipid-based transfection method. TPC2 was predominantly localized in the ruffled border-like structure of the osteoclasts. Moreover, overexpression of TPC2 promoted osteoclast differentiation. In addition, expression levels of TPC1 were measured, and TPC1-expressing vectors were transfected into RAW to investigate the role of TPC1 in osteoclastogenesis. Osteoclast differentiation was promoted in TPC1-transfected RAW. Notably, TPC2 expression was not influenced by TPC1 overexpression. Furthermore, TPC1 was knocked down by siRNA, resulting in reduced TRAP activity. TRAP is a biochemical indicator of osteoclastogenesis. Our findings suggest that TPC1 and TPC2 have independent essential roles in the differentiation of osteoclasts.