Type-2 cannabinoid receptor regulates proliferation, apoptosis, differentiation, and OPG/RANKL ratio of MC3T3-E1 cells exposed to Titanium particles

Abstract

The type-2 cannabinoid receptor (CB2) is expressed in osteoblasts and plays a role in bone metabolism through regulation on bone mass and bone turnover, but the functional importance of CB2 in osteoblasts under Titanium (Ti) stimulation is incompletely understood. This study aimed to investigate the CB2 expression in osteoblasts under Ti stimulation and the effects of CB2 activation on proliferation, apoptosis, differentiation, mineralization, OPG, and RANKL expression of MC3T3-E1 cells exposed to Ti particles. MC3T3-E1 cells were incubated in the presence of Ti particles with or without CB2-specific agonist HU-308 and antagonist SR144528. Ti particles treatment obviously induced the CB2 expression in MC3T3-E1 cells, and reduced the cell survival in a dose- and time-dependent manner (p < 0.05). Addition of HU-308 could dose-dependently alleviate the Ti-induced decrease of cell survival (p < 0.05). The flow cytometry assay showed that comparing with the control group, the apoptosis rate and caspase-3 activity in the Ti group were significantly elevated (p < 0.05), which could be alleviated by HU-308. Moreover, HU-308 effectively attenuated the decrease of cell mineralization capability, alkaline phosphates (ALP) and osteocalcin activity, and increase of OPG/RANKL ratio induced by Ti particles treatment (p < 0.05). These effects were partially counteracted by combined treatment of CB2 antagonist SR144528 (p < 0.05). In conclusion, CB2 activation has a favorable inhibitory effect on Ti-induced reactions in MC3T3-E1 cell through modulating proliferation, apoptosis, differentiation, and RANKL expression. These findings suggest that activation of CB2 might be an effective therapeutic strategy to promote bone formation and reduce bone dissolution.

Parthenolide reduces empty lacunae and osteoclastic bone surface resorption induced by polyethylene particles in a murine calvarial model of peri‐implant osteolysis

“…Osteocyte apoptosis generates signals that increase bone loss and direct it toward the area of bone containing the dead or dying cells . Metal particles have been shown to induce osteocyte apoptosis and inflammatory cytokine production in vitro . This study determines the effects of PE particles on osteocyte death in a murine model of PE‐induced osteolysis.…”

Gene Expression in Osteolysis: Review on the Identification of Altered Molecular Pathways in Preclinical and Clinical Studies

“…Despite the great variability found in particle size, most Ti and PE particles used in vitro had diameters of 4–4.5 µm [31,33,38,42,49] and 1.7 µm [20,43,48], respectively, with dimensions never exceeding 10 µm. In regards to in vivo models, Ti ranged from 0.1 to 20 µm [55,57,59,60], while PE particles dimensions were usually about 7 µm [61,62,63].…”

“…Mouse macrophage cell lines (RAW264.7) were the most used [4,6,27,28,29,30,31,32,33,34,35,36,37], followed by OB precursors derived from mouse calvaria (MC3T3-E1) [27,28,38,39,40,41] and human (Saos-2 and MG-63) [1,42,43] or rat (ROS 17/2.8) [44] osteosarcoma and monocitic (THP-1) [20,45,46] cell lines. In addition, other studies also used primary cells: mouse [28,47] or human [42,48] OBs, mouse or rat bone-marrow-derived macrophages (BMM) [28,49,50,51], human MSCs [39,52], mouse [47] or human [53] FBs and mouse peritoneal macrophages [54].…”

“…In one study, mouse calvaria [4] were cultured in toto. Regarding the different wear particles used, Ti [4,6,27,28,29,30,31,32,33,38,39,42,45,50,52,53] was the most employed, followed by PE [20,34,37,43,47,48], PMMA [36,41,51] and Co-Cr alloy [1,40,46]. Three studies compared Ti effects with PMMA [49], zirconia (ZrO 2 ) [54] and aluminia ceramic (CE) [34] and one compared PE and hydroxyapatite (HA) particles [44].…”

“…In cells of the osteoblastic lineage, Ti particles induced an increase in the pro-inflammatory genes and a decrease in Osteoprotegerin (OPG) gene observed in macrophagic cells and in CB2 [27,28,38,39,42]. In addition, one study analyzed osteogenesis and apoptotic pathways with microarray in hMSCs, showing an up-regulation of genes of pro-apoptotic proteins (Bcl-2-associated death promoter (BAD), Cluster of Differentiation 70 (CD70), B-cell lymphoma 2 (BCL2) and a down-regulation of anti-apoptotic and osteogenesis genes, encoding for Cartilage oligomeric matrix protein (COMP), Fibroblast Growth Factor Receptor 2 (FGFR2), Insulin-like growth factor 1 (IGF-1), Bone morphogenetic protein 6 (BMP6), Collagens (COLLs), (sex determining region Y)-box 9 (SOX9) and OPG [52].…”