Citrullinated histone H3: a novel target for the treatment of sepsis.
Citrullinated histone H3: a novel target for the treatment of sepsis.
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DOI：
10.1016/j.surg.2014.04.009
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发表时间：
2014-08
期刊：
影响因子：
3.8
通讯作者：
Alam, Hasan B.
中科院分区：
文献类型：
--
作者：
Li, Yongqing;Liu, Zhengcai;Liu, Baoling;Zhao, Ting;Chong, Wei;Wang, Yanming;Alam, Hasan B.
我们最近证明，在脂多糖 (LPS) 诱导的休克啮齿动物模型中，循环瓜氨酸组蛋白 H3 (Cit H3) 的增加与败血症的致死率相关，而用组蛋白脱乙酰酶 (HDAC) 抑制剂 (HDACI) 辛二酰苯胺异羟肟酸 (SAHA) 治疗可显着提高生存率。然而，Cit H3 在脓毒症发病机制和治疗中的作用尚不清楚。本研究旨在测试 HDACI 治疗是否可以抑制细胞 Cit H3 的产生，以及用 Cl-脒（PAD 抑制剂）抑制肽酰精氨酸脱亚胺酶（PAD，一种产生 Cit H3 的酶）或用抗 Cit H3 抗体中和血液 Cit H3 是否可以提高盲肠结扎穿刺 (CLP) 诱导的败血性休克临床相关小鼠模型的存活率。进行了三个实验。在实验I中，在存在或不存在SAHA（5μmol）的情况下，用LPS（100ng/ml）处理盖玻片上生长的HL-60中性粒细胞3小时，并用抗Cit H3抗体进行免疫染色，以在荧光显微镜下评估SAHA对Cit H3产生的影响。 Cit H3 阳性细胞的比例计算为平均值±SD (n=3)。实验II中，雄性C57BL/6J小鼠接受CLP，1小时后随机分为三组腹腔注射：（1）二甲基亚砜（DMSO），（2）SAHA（50 mg/kg）DMSO溶液，（3）Cl-脒（80 mg/kg）DMSO溶液（n=10/组）。实验III中，将雄性C57BL/6J小鼠分为对照组和治疗组，并进行CLP。两小时后，对照组和治疗组分别注射免疫球蛋白（IgG）和Cit H3抗体（20 mg/kg iv；n=5/组）。对存活率进行长达 10 天的监测。在实验I中，LPS诱导HL-60细胞中Cit H3的产生，而SAHA处理显着抑制H3瓜氨酸化（p<0.05）。在实验 II 中，所有注射媒介物的小鼠均在 3 天内死亡，循环 Cit H3 水平增加，而 HDACI 或 Cl-脒治疗显着改善了长期存活率 (p < 0.01)。在实验 III 中，给予 IgG 并没有提高生存率，但使用 Cit H3 特异性抗体进行单次治疗可显着提高生存率 (p<0.014)。抑制 HDAC 或 PAD 可显着抑制体外 Cit H3 的产生，并提高体内存活率。 Cit H3 的中和可显着提高脓毒症小鼠的存活率。总的来说，我们的研究结果首次表明 Cit H3 不仅可以作为潜在的生物标志物，而且可以作为脓毒症的新治疗靶点。
We have recently demonstrated that in a rodent model of lipopolysaccharide (LPS)-induced shock, an increase in circulating citrullinated histone H3 (Cit H3) is associated with lethality of sepsis, and treatment with suberoylanilide hydroxamic acid (SAHA), a histone deacetylase (HDAC) inhibitor (HDACI), significantly improves survival. However, the role of Cit H3 in pathogenesis and therapeutics of sepsis are largely unknown. The present study was designed to test whether treatment with HDACI could inhibit cellular Cit H3 production, and inhibition of peptidyalarginine deiminase (PAD, an enzyme producing Cit H3) with Cl-amidine (PAD inhibitor) or neutralization of blood Cit H3 with anti-Cit H3 antibody could improve survival in a clinically relevant mouse model of cecal ligation and puncture (CLP) induced septic shock. Three experiments were carried out. In experiment I, HL-60 neutrophilic cells grown on a coverslip were treated with LPS (100 ng/ml) in the presence or absence of SAHA (5 µmol) for 3 h, and subjected to immuno-staining with anti-Cit H3 antibody to assess effect of SAHA on Cit H3 production under a fluorescence microscope. The ratio of Cit H3 positive cells was calculated as mean ± SD (n=3). In experiment II, male C57BL/6J mice were subjected to CLP, and 1 hour later randomly divided into three groups for intraperitoneal injection as follows: (1) dimethyl sulfoxide (DMSO), (2) SAHA (50 mg/kg) in DMSO, and (3) Cl-amidine (80 mg/kg) in DMSO (n=10/group). In experiment III, male C57BL/6J mice were divided into control and treatment groups, and subjected to CLP. Two hours later, immunoglobulin (IgG) and Cit H3 antibody (20 mg/kg iv; n=5/group) were injected into the control and treatment groups, respectively. Survival was monitored for up to 10 days. In experiment I, LPS induced Cit H3 production in the HL-60 cells, while SAHA treatment inhibited H3 citrullination significantly (p<0.05). In experiment II, all vehicle injected mice died within 3 days with increased circulating Cit H3 levels, whereas treatment with HDACI or Cl-amidine notably improved long-term survival (p< 0.01). In experiment III, administration of IgG did not improve survival, but a single treatment with Cit H3 specific antibody significantly improved survival (p<0.014). Inhibition of HDAC or PAD significantly suppresses Cit H3 production in vitro, and improves survival in vivo. Neutralization of Cit H3 significantly improves survival in septic mice. Collectively, our findings indicate for the first time that Cit H3 could not only serve as a potential biomarker but also a novel therapeutic target in sepsis.
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