Shandong Provincial Key Laboratory of Metabolic Diseases and Qingdao Key Laboratory of Gout, The Affiliated Hospital of Qingdao University, Qingdao, China.
Institute of Metabolic Diseases, Qingdao University, Qingdao, China.
Adv Rheumatol. 2023 Aug 8;63(1):39. doi: 10.1186/s42358-023-00307-1.
The deposition of monosodium urate (MSU) crystals within synovial joints and tissues is the initiating factor for gout arthritis. Thus, MSU crystals are a vital tool for studying gout's molecular mechanism in animal and cellular models. This study mainly compared the excellence and worseness of MSU crystals prepared by different processes and the degree of inflammation induced by MSU crystals.
MSU crystals were prepared using neutralization, alkali titration, and acid titration methods. The crystals' shape, length, quality, and uniformity were observed by polarized light microscopy and calculated by the software Image J. The foot pad and air pouch models were used to assess the different degrees of inflammation induced by the MSU crystals prepared by the three different methods at different time points. Paw swelling was evaluated by caliper. In air pouch lavage fluid, inflammatory cell recruitment was measured by hemocytometer, and the level of IL-1β, TNF-α, and IL-18 by ELISA. Inflammatory cell infiltration was assayed by immunohistochemistry of air pouch synovial slices.
For the preparation of MSU crystals with the same uric acid, the quantity acquired by the alkalization method was highest, followed by neutralization, with the acid titration method being the lowest. The crystals prepared by neutralization were the longest. The swelling index of the foot pad induced by MSU crystals prepared by acid titration was significantly lower than that of the other methods at 24 h. The inflammatory cell recruitment and level of IL-1β, TNF-α, and IL-18 in air pouch lavage fluid were lowest in animals with crystals prepared by acid titration. IL-1β secretion induced by MSU crystals prepared by acid titration was significantly lower than that of the other two groups, but there was no significant difference in IL-18 secretion between the three groups in THP-1 macrophages and BMDMs.
All three methods can successfully prepare MSU crystals, but the levels of inflammation induced by the crystals prepared by the three methods were not identical. The degree of inflammation induced by MSU crystals prepared by neutralization and alkalization is greater than by acid titration, but the quantity of MSU crystals obtained by the alkalization method is higher and less time-consuming. Apparently, the window of inflammation triggered by acid titration preparation is shorter compared to other forms of crystal preparation. Overall, MSU crystals prepared by the alkaline method should be recommended for studying the molecular mechanisms of gout in animal and cellular models.
单钠尿酸盐（MSU）晶体在滑膜关节和组织中的沉积是痛风关节炎的起始因素。因此，MSU 晶体是研究动物和细胞模型中痛风分子机制的重要工具。本研究主要比较了不同工艺制备的 MSU 晶体的优缺点以及 MSU 晶体诱导的炎症程度。
采用中和法、碱滴定法和酸滴定法制备 MSU 晶体。通过偏光显微镜观察晶体的形状、长度、质量和均匀性，并通过 Image J 软件进行计算。采用足垫和气囊模型评估三种不同方法在不同时间点制备的 MSU 晶体引起的不同程度的炎症。用卡尺评估爪肿胀。在气囊灌洗液中，用血细胞计数器测量炎性细胞募集情况，并通过 ELISA 测定 IL-1β、TNF-α 和 IL-18 的水平。通过气囊滑膜切片的免疫组织化学测定炎性细胞浸润。
对于用相同尿酸制备 MSU 晶体，碱化法获得的量最高，其次是中和法，酸化法最低。中和法制备的晶体最长。酸化法制备的 MSU 晶体诱导的足垫肿胀指数在 24 小时时明显低于其他方法。气囊灌洗液中炎性细胞募集和 IL-1β、TNF-α 和 IL-18 水平在酸化法制备晶体的动物中最低。酸化法制备的 MSU 晶体诱导的 IL-1β分泌明显低于其他两组，但在 THP-1 巨噬细胞和 BMDM 中三组之间的 IL-18 分泌无显著差异。
三种方法均可成功制备 MSU 晶体，但三种方法制备的晶体引起的炎症程度不同。中和法和碱化法制备的 MSU 晶体引起的炎症程度大于酸化法，但碱化法获得的 MSU 晶体量更高，耗时更短。显然，酸化法制备晶体引起的炎症窗口比其他形式的晶体制备更短。总的来说，碱性法制备的 MSU 晶体应推荐用于研究动物和细胞模型中痛风的分子机制。
