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Osteocytic cells exposed to titanium particles increase sclerostin expression and inhibit osteoblastic cell differentiation mostly via direct cell-to-cell contact.

ABSTRACT: The mechanism underlying induction of periprosthetic osteolysis by wear particles remains unclear. In this study, cultured MLO-Y4 osteocytic cells were exposed to different concentrations of titanium (Ti) particles. The results showed that Ti particles increased expression of the osteocytic marker SOST/sclerostin in a dose-dependent manner, accelerated apoptosis of MLO-Y4 cells, increased the expression of IL-6, TNF-α and connexin 43. SOST silence alleviated the increase of MLO-Y4 cells apoptosis, decreased the expression of IL-6, TNF-α and connexin 43 caused by Ti particles. The different co-culture systems of MLO-Y4 cells with MC3T3-E1 osteoblastic cells were further used to observe the effects of osteocytic cells' changes induced by Ti particles on osteoblastic cells. MLO-Y4 cells treated with Ti particles inhibited dramatically differentiation of MC3T3-E1 cells mostly through direct cell-to-cell contact. SOST silence attenuated the inhibition effects of Ti-induced MLO-Y4 on MC3T3-E1 osteoblastic differentiation, which ALP level and mineralization of MC3T3-E1 cells increased and the expression of ALP, OCN and Runx2 increased compared to the Ti-treated group. Taken together, Ti particles had negative effects on MLO-Y4 cells and the impact of Ti particles on osteocytic cells was extensive, which may further inhibit osteoblastic differentiation mostly through intercellular contact directly. SOST/sclerostin plays an important role in the process of mutual cell interaction. These findings may help to understand the effect of osteocytes in wear particle-induced osteolysis.

SUBMITTER: Chai H

PROVIDER: S-EPMC9345295 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Osteocytic cells exposed to titanium particles increase sclerostin expression and inhibit osteoblastic cell differentiation mostly via direct cell-to-cell contact.

Chai Hao H Zhang Zai Hang ZH Fang Jing Yi JY She Chang C Geng De Chun C Xu Wei W

Journal of cellular and molecular medicine 20220628 15

The mechanism underlying induction of periprosthetic osteolysis by wear particles remains unclear. In this study, cultured MLO-Y4 osteocytic cells were exposed to different concentrations of titanium (Ti) particles. The results showed that Ti particles increased expression of the osteocytic marker SOST/sclerostin in a dose-dependent manner, accelerated apoptosis of MLO-Y4 cells, increased the expression of IL-6, TNF-α and connexin 43. SOST silence alleviated the increase of MLO-Y4 cells apoptosi ...[more]

PMID: 35762300

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Project description:Periprosthetic osteolysis (PPO) induced by wear particles is the most severe complication of total joint replacement; however, the mechanism behind PPO remains elusive. Previous studies have shown that osteocytes play important roles in wear-particle-induced osteolysis. In this study, we investigated the effects of connexin 43 (Cx43) on the regulation of osteocyte-to-osteoblast differentiation. We established an in vivo murine model of calvarial osteolysis induced by titanium (Ti) particles. The osteolysis characteristic and osteogenesis markers in the osteocyte-selective Cx43 (CKO)-deficient and wild-type (WT) mice were observed. The calvarial osteolysis induced by Ti particles was partially attenuated in CKO mice. The expression of β-catenin and osteogenesis markers increased significantly in CKO mice. In vitro, the osteocytic cell line MLO-Y4 was treated with Ti particles. The co-culturing of MLO-Y4 cells with MC3T3-E1 osteoblastic cells was used to observe the effects of Ti-treated osteocytes on osteoblast differentiation. When Cx43 of MLO-Y4 cells was silenced or overexpressed, β-catenin was detected. Additionally, co-immunoprecipitation detection of Cx43 and β-catenin binding in MLO-Y4 cells and MC3T3-E1 cells was performed. Finally, β-catenin expression in MC3T3-E1 cells and osteoblast differentiation were evaluated after 18α-glycyrrhetinic acid (18α-GA) was used to block the intercellular communication of Cx43 between MLO-Y4 and MC3T3-E1 cells. Ti particles increased Cx43 expression and decreased β-catenin expression in MLO-Y4 cells. The silencing of Cx43 increased the β-catenin expression, and the over-expression of Cx43 decreased the β-catenin expression. In the co-culture model, Ti treatment of MLO-Y4 cells inhibited the osteoblastic differentiation of MC3T3-E1 cells and Cx43 silencing in MLO-Y4 cells attenuated the inhibitory effects on osteoblastic differentiation. With Cx43 silencing in the MLO-Y4 cells, the MC3T3-E1 cells, co-cultured alongside MLO-Y4, displayed decreased Cx43 expression, increased β-catenin expression, activation of Runx2, and promotion of osteoblastic differentiation in vitro co-culture. Finally, Cx43 expression was found to be negatively correlated to the activity of the Wnt signaling pathway, mostly through the Cx43 binding of β-catenin from its translocation to the nucleus. The results of our study suggest that Ti particles increased Cx43 expression in osteocytes and that osteocytes may participate in the regulation of osteoblast function via the Cx43 during PPO.

Project description:Type 1 diabetes with periodontitis shows elevated TNF-α expression. Tumor necrosis factor (TNF)-α stimulates the expression of receptor activator of nuclear factor-κB ligand (RANKL) and sclerostin. The objective of this study was to determine the effect of TNF-α expression of osteocytic RANKL and sclerostin in type 1 diabetes rats with periodontitis using infliximab (IFX), a TNF-α antagonist. Rats were divided into two timepoint groups: day 3 and day 20. Each timepoint group was then divided into four subgroups: 1) control (C, n = 6 for each time point); 2) periodontitis (P, n = 6 for each time point); 3) diabetes with periodontitis (DP, n = 8 for each time point); and 4) diabetes with periodontitis treated with IFX (DP+IFX, n = 8 for each time point). To induce type 1 diabetes, rats were injected with streptozotocin (50 mg/kg dissolved in 0.1 M citrate buffer). Periodontitis was then induced by ligature of the mandibular first molars at day 7 after STZ injection (day 0). IFX was administered once for the 3 day group (on day 0) and twice for the 20 day group (on days 7 and 14). The DP group showed greater alveolar bone loss than the P group on day 20 (P = 0.020). On day 3, higher osteoclast formation and RANKL-positive osteocytes in P group (P = 0.000 and P = 0.011, respectively) and DP group (P = 0.006 and P = 0.017, respectively) than those in C group were observed. However, there was no significant difference in osteoclast formation or RANKL-positive osteocytes between P and DP groups. The DP+IFX group exhibited lower alveolar bone loss (P = 0.041), osteoclast formation (P = 0.019), and RANKL-positive osteocytes (P = 0.009) than that of the DP group. On day 20, DP group showed a lower osteoid area (P = 0.001) and more sclerostin-positive osteocytes (P = 0.000) than P group. On days 3 and 20, the DP+IFX group showed more osteoid area (P = 0.048 and 0.040, respectively) but lower sclerostin-positive osteocytes (both P = 0.000) than DP group. Taken together, these results suggest that TNF-α antagonist can diminish osteocytic RANKL/sclerostin expression and osteoclast formation, eventually recovering osteoid formation. Therefore, TNF-α might mediate alveolar bone loss via inducing expression of osteocytic RANKL and sclerostin in type 1 diabetes rats with periodontitis.

Project description:Osteoblastic cells in the bone marrow produce the bone matrix, participate in mineralization of bone and regulate the balance of calcium and phosphate ions in the developing bone. In addition, the radiosensitive hematopoietic stem/progenitor cells form a niche with osteoblastic cells to maintain immature state and self-renewal in the bone marrow microenvironment. In the field of radiotherapy, internal radiotherapy using radioactive isotopes that emit alpha particles is performed, and antitumor effect is expected by radioisotopes (such as 223-Ra) when the primary cancer cells metastasize to bone marrow. A major problem in this treatment is that the efficacy and side effects form show a high degree of individual variability, and the response of a patient is very difficult to predict. The mechanisms of the variability are not known, but it is possible that some contributing factors are the alpha radiation-induced modulation of osteoblastic cell activity. However, the characteristics of micro-RNA expressions in alpha-irradiated osteoblastic cells remain unclear. The purpose of this study is to clarify the pattern of miRNA expressions in osteoblast derived from freshly bone marrow cells exposed to alpha particle radiation. Murine bone marrow cells from both femurs were differentiated to osteoblastic cells using cell culture medium and exposed to 0-1 Gy of alpha radiation. The 24 miRNAs were significantly up- or down-regulated: mmu-let-7a-5p, mmu-let-7b-5p, mmu-let-7c-5p, mmu-miR-1224-5p, mmu-miR-125b-5p, mmu-miR-296-5p, mmu-miR-331-3p, mmu-miR-3473b, mmu-miR-3960, mmu-miR-3963, mmu-miR-466i-5p, mmu-miR-494-3p, mmu-miR-5126, mmu-miR-6360, mmu-miR-6366, mmu-miR-6378, mmu-miR-6769b-5p, mmu-miR-6937-5p, mmu-miR-7005-5p, mmu-miR-7042-5p, mmu-miR-7047-5p, mmu-miR-709, mmu-miR-8110, mmu-miR-8117). These results indicate that exposure of osteoblastic cells to alpha particles leads to unique expression of miRNAs and it may give the cell radiosensitive environment.

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Osteocytic cells exposed to titanium particles increase sclerostin expression and inhibit osteoblastic cell differentiation mostly via direct cell-to-cell contact.

Publications

Osteocytic cells exposed to titanium particles increase sclerostin expression and inhibit osteoblastic cell differentiation mostly via direct cell-to-cell contact.

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