Department of Medicine, University of California, San Diego, La Jolla, CA, United States.
Department of Orthopedic Surgery, University of California, San Diego, La Jolla, CA, United States.
Front Immunol. 2023 Jul 17;14:1103231. doi: 10.3389/fimmu.2023.1103231. eCollection 2023.
Glucose metabolism, specifically, hexokinase 2 (HK2), has a critical role in rheumatoid arthritis (RA) fibroblast-like synoviocyte (FLS) phenotype. HK2 localizes not only in the cytosol but also in the mitochondria, where it protects mitochondria against stress. We hypothesize that mitochondria-bound HK2 is a key regulator of RA FLS phenotype.
HK2 localization was evaluated by confocal microscopy after FLS stimulation. RA FLSs were infected with Green fluorescent protein (GFP), full-length (FL)-HK2, or HK2 lacking its mitochondrial binding motif (HK2ΔN) expressing adenovirus (Ad). RA FLS was also incubated with methyl jasmonate (MJ; 2.5 mM), tofacitinib (1 µM), or methotrexate (1 µM). RA FLS was tested for migration and invasion and gene expression. Gene associations with HK2 expression were identified by examining single-cell RNA sequencing (scRNA-seq) data from murine models of arthritis. Mice were injected with K/BxN serum and given MJ. Ad-FLHK2 or Ad-HK2ΔN was injected into the knee of wild-type mice.
Cobalt chloride (CoCl) and platelet-derived growth factor (PDGF) stimulation induced HK2 mitochondrial translocation. Overexpression of the HK2 mutant and MJ incubation reversed the invasive and migrative phenotype induced by FL-HK2 after PDGF stimulation, and MJ also decreased the expression of C-X-C Motif Chemokine Ligand 1 (CXCL1) and Collagen Type I Alpha 1 Chain (COL1A1). Of interest, tofacitinib but not methotrexate had an effect on HK2 dissociation from the mitochondria. In murine models, MJ treatment significantly decreased arthritis severity, whereas HK2FL was able to induce synovial hypertrophy as opposed to HK2ΔN.
Our results suggest that mitochondrial HK2 regulates the aggressive phenotype of RA FLS. New therapeutic approaches to dissociate HK2 from mitochondria offer a safer approach than global glycolysis inhibition.
葡萄糖代谢，特别是己糖激酶 2（HK2），在类风湿关节炎（RA）成纤维样滑膜细胞（FLS）表型中起着关键作用。HK2 不仅位于细胞质中，还位于线粒体中，在那里它可以保护线粒体免受应激。我们假设线粒体结合的 HK2 是 RA FLS 表型的关键调节因子。
通过共聚焦显微镜评估 FLS 刺激后 HK2 的定位。用绿色荧光蛋白（GFP）、全长（FL）-HK2 或缺乏其线粒体结合基序的 HK2（HK2ΔN）表达的腺病毒（Ad）感染 RA FLS。RA FLS 还与茉莉酸甲酯（MJ；2.5 mM）、托法替尼（1 μM）或甲氨蝶呤（1 μM）孵育。测试 RA FLS 的迁移和侵袭以及基因表达。通过检查关节炎小鼠模型的单细胞 RNA 测序（scRNA-seq）数据，确定与 HK2 表达相关的基因。给 K/BxN 血清注射的小鼠给予 MJ。将 Ad-FLHK2 或 Ad-HK2ΔN 注射到野生型小鼠的膝关节中。
氯化钴（CoCl）和血小板衍生生长因子（PDGF）刺激诱导 HK2 线粒体易位。HK2 突变体的过表达和 MJ 孵育逆转了 PDGF 刺激后 FL-HK2 诱导的侵袭和迁移表型，MJ 还降低了 C-X-C 基序趋化因子配体 1（CXCL1）和 I 型胶原α 1 链（COL1A1）的表达。有趣的是，与甲氨蝶呤相比，托法替尼对 HK2 从线粒体解离有影响。在小鼠模型中，MJ 治疗显著降低关节炎严重程度，而 HK2FL 能够诱导滑膜肥大，而不是 HK2ΔN。
我们的结果表明，线粒体 HK2 调节 RA FLS 的侵袭表型。将 HK2 从线粒体解离的新治疗方法提供了一种比全局糖酵解抑制更安全的方法。
