Department of Oncology and Diagnostic Sciences, School of Dentistry, University of Maryland, 650 W Baltimore Street, Baltimore, MD, 21201, USA.
Preventive Dental Sciences Department, School of Dentistry, Imam Abdulrahman Bin Faisal University, Dammam, Saudi Arabia.
BMC Immunol. 2021 Mar 25;22(1):23. doi: 10.1186/s12865-021-00409-9.
Lipopolysaccharide (LPS) is an endotoxin and a vital component of gram-negative bacteria's outer membrane. During gram-negative bacterial sepsis, LPS regulates osteoclast differentiation and activity, in addition to increasing inflammation. This study aimed to investigate how LPS regulates osteoclast differentiation of RAW 264.7 cells in vitro.
Herein, we revealed that RAW cells failed to differentiate into mature osteoclasts in vitro in the presence of LPS. However, differentiation occurred in cells primed with receptor activator of nuclear factor-kappa-Β ligand (RANKL) for 24 h and then treated with LPS for 48 h (henceforth, denoted as LPS-treated cells). In cells treated with either RANKL or LPS, an increase in membrane levels of toll-like receptor 4 (TLR4) receptor was observed. Mechanistically, an inhibitor of TLR4 (TAK-242) reduced the number of osteoclasts as well as the secretion of tumor necrosis factor (TNF)-α in LPS-treated cells. RANKL-induced RAW cells secreted a very basal level TNF-α. TAK-242 did not affect RANKL-induced osteoclastogenesis. Increased osteoclast differentiation in LPS-treated osteoclasts was not associated with the RANKL/RANK/OPG axis but connected with the LPS/TLR4/TNF-α tumor necrosis factor receptor (TNFR)-2 axis. We postulate that this is because TAK-242 and a TNF-α antibody suppress osteoclast differentiation. Furthermore, an antibody against TNF-α reduced membrane levels of TNFR-2. Secreted TNF-α appears to function as an autocrine/ paracrine factor in the induction of osteoclastogenesis independent of RANKL.
TNF-α secreted via LPS/TLR4 signaling regulates osteoclastogenesis in macrophages primed with RANKL and then treated with LPS. Our findings suggest that TLR4/TNF-α might be a potential target to suppress bone loss associated with inflammatory bone diseases, including periodontitis, rheumatoid arthritis, and osteoporosis.
脂多糖（LPS）是革兰氏阴性菌外膜的内毒素和重要组成部分。在革兰氏阴性菌败血症中，LPS 不仅能增加炎症，还能调节破骨细胞分化和活性。本研究旨在探讨 LPS 如何在体外调节 RAW264.7 细胞的破骨细胞分化。
在此，我们发现 LPS 存在时，RAW 细胞无法在体外分化为成熟破骨细胞。但是，在经过核因子-κB 受体激活剂配体（RANKL）预刺激 24 小时然后用 LPS 处理 48 小时的细胞（以下简称 LPS 处理细胞）中发生了分化。在用 RANKL 或 LPS 处理的细胞中，观察到 TLR4 受体的膜水平增加。从机制上讲，TLR4 抑制剂（TAK-242）减少了 LPS 处理细胞中的破骨细胞数量和肿瘤坏死因子（TNF）-α的分泌。RANKL 诱导的 RAW 细胞分泌出非常基础水平的 TNF-α。TAK-242 不影响 RANKL 诱导的破骨细胞生成。在 LPS 处理的破骨细胞中，破骨细胞分化的增加与 RANKL/RANK/OPG 轴无关，而与 LPS/TLR4/TNF-α肿瘤坏死因子受体（TNFR）-2 轴有关。我们假设这是因为 TAK-242 和 TNF-α 抗体抑制了破骨细胞分化。此外，TNF-α 抗体降低了 TNFR-2 的膜水平。分泌的 TNF-α似乎作为一种自分泌/旁分泌因子在独立于 RANKL 的破骨细胞生成诱导中发挥作用。
通过 LPS/TLR4 信号传导分泌的 TNF-α调节经 RANKL 预处理然后用 LPS 处理的巨噬细胞中的破骨细胞生成。我们的研究结果表明，TLR4/TNF-α 可能是抑制与炎症性骨病相关的骨丢失的潜在靶点，包括牙周炎、类风湿关节炎和骨质疏松症。
