The actions of methotrexate on endothelial cells are dependent on the shear stress-induced regulation of one carbon metabolism.
The actions of methotrexate on endothelial cells are dependent on the shear stress-induced regulation of one carbon metabolism.
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DOI：
10.3389/fimmu.2023.1209490
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发表时间：
2023
影响因子：
7.3
通讯作者：
中科院分区：
文献类型：
--
作者：
疾病缓解抗风湿药物甲氨蝶呤（MTX）被认为可以降低全身炎症性疾病患者的心血管风险。然而，这些心脏保护作用的分子基础仍然不完全清楚。本研究探讨了小剂量MTX对血管内皮的作用。在与炎性关节炎相关的体外条件下研究了人内皮细胞（EC）。这些包括在促炎微环境中培养和使用平行板模型暴露于流体剪切应力（FSS）。通过RNA测序和定量实时PCR分析分别处理的细胞的基因表达，通过免疫印迹分析蛋白质表达，通过磷酸激酶活性阵列，通过流式细胞术分析细胞周期，并通过质谱法评估叶酸代谢物水平。在静态条件下，MTX被EC有效地摄取，并引起细胞周期阻滞，同时调节细胞信号传导途径。这些反应被亚叶酸（FA）逆转，表明OCM是MTX的主要靶点。在FSS下，MTX不影响细胞增殖或促炎基因表达。暴露于FSS下调内皮细胞一碳代谢（OCM）的关键OCM基因和代谢产物的表达减少证明。我们发现，FSS显着下调OCM，从而使EC不太容易受到MTX治疗的影响。切应力对OCM的影响表明MTX不直接调节内皮功能。MTX的心脏保护作用可能反映了对炎症细胞的直接作用和对血管内皮的间接益处。
The disease-modifying anti-rheumatic drug methotrexate (MTX) is recognized to reduce cardiovascular risk in patients with systemic inflammatory diseases. However, the molecular basis for these cardioprotective effects remains incompletely understood. This study evaluated the actions of low-dose MTX on the vascular endothelium. Human endothelial cells (EC) were studied under in vitro conditions relevant to inflammatory arthritis. These included culture in a pro-inflammatory microenvironment and exposure to fluid shear stress (FSS) using a parallel plate model. Respectively treated cells were analyzed by RNA sequencing and quantitative real-time PCR for gene expression, by immunoblotting for protein expression, by phosphokinase activity arrays, by flow cytometry for cell cycle analyses and by mass spectrometry to assess folate metabolite levels. In static conditions, MTX was efficiently taken up by EC and caused cell cycle arrest concurrent with modulation of cell signaling pathways. These responses were reversed by folinic acid (FA), suggesting that OCM is a predominant target of MTX. Under FSS, MTX did not affect cell proliferation or pro-inflammatory gene expression. Exposure to FSS downregulated endothelial one carbon metabolism (OCM) as evidenced by decreased expression of key OCM genes and metabolites. We found that FSS significantly downregulated OCM and thereby rendered EC less susceptible to the effects of MTX treatment. The impact of shear stress on OCM suggested that MTX does not directly modulate endothelial function. The cardioprotective actions of MTX likely reflect direct actions on inflammatory cells and indirect benefit on the vascular endothelium.
影响因子：
5
作者：
Summers, Carolyn M.;Hammons, Andrea L.;Arora, Jasbir;Zhang, Suhong;Jochems, Jeanine;Blair, Ian A.;Whitehead, Alexander S.
通讯作者：
Whitehead, Alexander S.
影响因子：
4.9
作者：
Montesinos MC;Desai A;Cronstein BN
通讯作者：
Cronstein BN