Department of Medicine, Faculty of Medical and Health Sciences, University of Auckland, 85 Park Rd, Grafton, Auckland, New Zealand.
Department of Surgery, University of Auckland, Auckland, New Zealand.
Arthritis Res Ther. 2022 Sep 5;24(1):212. doi: 10.1186/s13075-022-02900-z.
Tophi are lesions commonly present at sites of bone erosion in gout-affected joints. The tophus comprises a core of monosodium urate (MSU) crystals surrounded by soft tissue that contains macrophages and other immune cells. Previous studies found that MSU crystals directly reduce osteoblast viability and function. The aim of the current study was to determine the indirect, macrophage-mediated effects of MSU crystals on osteoblasts.
Conditioned medium from the RAW264.7 mouse macrophage cell line cultured with MSU crystals was added to the MC3T3-E1 mouse osteoblastic cell line. Conditioned medium from the THP-1 human monocytic cell line cultured with MSU crystals was added to primary human osteoblasts (HOBs). Matrix mineralization was assessed by von Kossa staining. Gene expression was determined by real-time PCR, and concentrations of secreted factors were determined by enzyme-linked immunosorbent assay.
In MC3T3-E1 cells cultured for 13 days in an osteogenic medium, the expression of the osteoblast marker genes Col1a1, Runx2, Sp7, Bglap, Ibsp, and Dmp1 was inhibited by a conditioned medium from MSU crystal-stimulated RAW264.7 macrophages. Mineral staining of MC3T3-E1 cultures on day 21 confirmed the inhibition of osteoblast differentiation. In HOB cultures, the effect of 20 h incubation with a conditioned medium from MSU crystal-stimulated THP-1 monocytes on osteoblast gene expression was less consistent. Expression of the genes encoding cyclooxygenase-2 and IL-6 and secretion of the proinflammatory mediators PGE and IL-6 were induced in MC3T3-E1 and HOBs incubated with conditioned medium from MSU crystal-stimulated macrophages/monocytes. However, inhibition of cyclooxygenase-2 activity and PGE secretion from HOBs indicated that this pathway does not play a major role in mediating the indirect effects of MSU crystals in HOBs.
Factors secreted from macrophages stimulated by MSU crystals attenuate osteoblast differentiation and induce the expression and secretion of proinflammatory mediators from osteoblasts. We suggest that bone erosion in joints affected by gout results from a combination of direct and indirect effects of MSU crystals.
痛风患者关节中骨质侵蚀部位常出现痛风石。痛风石由尿酸单钠（MSU）晶体核心组成，周围是含有巨噬细胞和其他免疫细胞的软组织。先前的研究发现，MSU 晶体直接降低成骨细胞的活力和功能。本研究旨在确定 MSU 晶体对成骨细胞的间接、巨噬细胞介导的影响。
用 MSU 晶体培养 RAW264.7 小鼠巨噬细胞系的条件培养基添加到 MC3T3-E1 小鼠成骨细胞系中。用 MSU 晶体培养 THP-1 人单核细胞系的条件培养基添加到原代人成骨细胞（HOB）中。通过 von Kossa 染色评估基质矿化。通过实时 PCR 确定基因表达，通过酶联免疫吸附测定法确定分泌因子的浓度。
在矿化培养基中培养 13 天的 MC3T3-E1 细胞中，MSU 晶体刺激的 RAW264.7 巨噬细胞的条件培养基抑制成骨细胞标记基因 Col1a1、Runx2、Sp7、Bglap、Ibsp 和 Dmp1 的表达。第 21 天 MC3T3-E1 培养物的矿化染色证实了成骨细胞分化的抑制。在 HOB 培养物中，MSU 晶体刺激的 THP-1 单核细胞的 20 小时孵育条件培养基对成骨细胞基因表达的影响不太一致。在 MC3T3-E1 和 HOB 中，用 MSU 晶体刺激的巨噬细胞/单核细胞的条件培养基孵育诱导编码环氧化酶-2 和 IL-6 的基因表达和促炎介质 PGE 和 IL-6 的分泌。然而，HOB 中环氧化酶-2 活性和 PGE 分泌的抑制表明该途径在介导 HOB 中 MSU 晶体的间接作用中不起主要作用。
MSU 晶体刺激的巨噬细胞分泌的因子减弱成骨细胞分化，并诱导成骨细胞中促炎介质的表达和分泌。我们认为痛风影响的关节中的骨质侵蚀是由 MSU 晶体的直接和间接作用共同作用的结果。
