Supplementary Material for: Interleukin-11 Is Elevated in Patients with Atrial Fibrillation, Correlates with Serum Fibrosis Markers, and Represents a Therapeutic Target for Atrial Fibrosis

<b><i>Introduction:</i></b> Inflammatory cytokines are closely associated with developing cardiac fibrosis. This research aimed to explore the significant role of IL-11 in atrial fibrosis progression and potential therapeutic targets. <b><i>Methods:</i></b> 207 AF patients and 160 healthy subjects were included in the case-control study. Blood samples were analyzed for the level of IL-11 by enzyme-linked immunosorbent assay (ELISA). Angiotensin II (Ang II)-treated fibrosis mouse models were generated, and expression of IL-11 mRNA and protein was detected by RT-qPCR and Western blot. IL-11 antagonist was used to evaluating atrial fibrosis-related markers. <b><i>Results:</i></b> The persistent atrial fibrillation patients (<i>n</i> = 76) had significantly larger left atrial size, higher serum levels of hypertrophic protein BNP, proinflammatory cytokine high-sensitivity C-reactive protein (hs-CRP), and interleukin-6 (IL-6) compared to paroxysmal atrial fibrillation patients (<i>n</i> = 131), and healthy subjects (all <i>p</i> &lt; 0.05). Pearson correlation analysis revealed significant positive correlation between serum IL-11 and cardiac fibrosis markers BNP (<i>r</i> = 0.394, <i>p</i> &lt; 0.001), CTX-I (<i>r</i> = 0.418, <i>p</i> &lt; 0.001), PICP (<i>r</i> = 0.306, <i>p</i> &lt; 0.001), PIIINP (<i>r</i> = 0.335, <i>p</i> &lt; 0.001), and TGF-β1 (<i>r</i> = 0.273, <i>p</i> &lt; 0.001). In the fibrosis mouse model, Ang II infusion significantly upregulated IL-11 mRNA and protein expression in the left atrium of mice (<i>p</i> &lt; 0.05), as well as staining intensity of Masson trichrome, the intensity of α-SMA, and it increased mRNA expression of collagen I and III in atrial tissue. IL-11 antagonist treatment significantly attenuated Masson trichrome, number of α-SMA-positive myofibroblasts in atrial tissue. Also, it significantly reduced the p-ERK1/2 in atrial tissue of mice infused with Ang II (<i>p</i> &lt; 0.05). <b><i>Conclusions:</i></b> IL-11 is upregulated in the serum of AF patients, and IL-11 inhibitor significantly inhibited Ang II-induced atrial fibrosis, a key pathological feature of AF. Therefore, IL-11 could be a potential therapeutic target for AF.

<b><i>研究背景：</i></b> 炎症细胞因子与心脏纤维化的发生发展密切相关。本研究旨在探讨白细胞介素-11（IL-11）在心房纤维化进展中的关键作用及潜在治疗靶点。
<b><i>研究方法：</i></b> 本研究为病例对照研究，共纳入207例心房颤动（AF）患者与160名健康受试者。采用酶联免疫吸附试验（ELISA）检测血液样本中IL-11的水平。构建血管紧张素II（Ang II）诱导的纤维化小鼠模型，通过实时定量聚合酶链反应（RT-qPCR）与蛋白质印迹（Western blot）检测小鼠左心房组织中IL-11的mRNA与蛋白表达水平。使用IL-11拮抗剂评估心房纤维化相关标志物的表达变化。
<b><i>研究结果：</i></b> 与阵发性心房颤动患者（<i>n</i> = 131）及健康受试者相比，持续性心房颤动患者（<i>n</i> = 76）的左心房内径显著更大，血清肥厚标志物B型利钠肽（BNP）、促炎细胞因子高敏C反应蛋白（hs-CRP）及白细胞介素-6（IL-6）水平均显著升高（所有<i>p</i> &lt; 0.05）。Pearson相关分析显示，血清IL-11与心脏纤维化标志物BNP（<i>r</i> = 0.394, <i>p</i> &lt; 0.001）、I型胶原C端肽（CTX-I）（<i>r</i> = 0.418, <i>p</i> &lt; 0.001）、I型前胶原C端肽（PICP）（<i>r</i> = 0.306, <i>p</i> &lt; 0.001）、III型前胶原N端肽（PIIINP）（<i>r</i> = 0.335, <i>p</i> &lt; 0.001）及转化生长因子-β1（TGF-β1）（<i>r</i> = 0.273, <i>p</i> &lt; 0.001）均呈显著正相关。在纤维化小鼠模型中，Ang II灌注可显著上调小鼠左心房组织中IL-11的mRNA与蛋白表达水平（<i>p</i> &lt; 0.05），同时增强马松三色染色（Masson trichrome）的染色强度与α-平滑肌肌动蛋白（α-SMA）的表达强度，并升高心房组织中I型与III型胶原的mRNA表达水平。IL-11拮抗剂治疗可显著减弱马松三色染色的阳性信号，减少心房组织中α-SMA阳性肌成纤维细胞的数量，此外该治疗还可显著降低Ang II灌注小鼠心房组织中的磷酸化细胞外信号调节激酶1/2（p-ERK1/2）水平（<i>p</i> &lt; 0.05）。
<b><i>研究结论：</i></b> 本研究结果显示，心房颤动患者血清中IL-11表达上调，IL-11抑制剂可显著抑制Ang II诱导的心房纤维化——心房颤动的关键病理特征。因此，IL-11有望成为心房颤动的潜在治疗靶点。