Procalcitonin Impairs Endothelial Cell Function and Viability
Procalcitonin Impairs Endothelial Cell Function and Viability
复制标题
降钙素原损害内皮细胞功能和活力
DOI：
10.1213/ane.0000000000001574
复制
发表时间：
2017-03-01
影响因子：
5.7
通讯作者：
Roesner, Jan Patrick
中科院分区：
文献类型：
--
作者：
Wagner, Nana-Maria;Van Aken, Caroline;Roesner, Jan Patrick
背景：降钙素原被用作脓毒症患者识别和风险分层的诊断工具。降钙素原血浆浓度与正在进行的炎症反应的严重程度密切相关，可高达10,000倍。脓毒症时血管内皮细胞功能受损在低血压和器官灌流障碍的发病机制中起重要作用。方法：将人内皮细胞暴露于0.01~100 ng/mL的降钙素原中，观察内皮细胞的通透性、划痕实验中的迁移和细胞外基质上新生毛细血管的形成。肿瘤坏死因子-a和血管内皮生长因子作为阳性对照。在单侧股动脉结扎的小鼠模型上，研究了降钙素原对内皮细胞对缺血反应的影响。用免疫印迹法检测降钙素原暴露的内皮细胞中血管内皮细胞钙粘附素的表达和血管生成信号通路。用流式细胞仪检测细胞的炎症活性和活性，并进行基因组分析。结果：肿瘤坏死因子-α和0.1 ng/mL原降钙素诱导内皮细胞单层孵育6小时后内皮屏障破坏(分别为-2.53[-4.16~-0.89]，P=.0008和-2.09[-3.73~-0.45]，P-ADJUST=0.0064)。与血管内皮生长因子的促血管生成作用相比，降钙素原在浓度为0.02 ng/mL时可减少内皮细胞的迁移(0.26[0.06至0.47]，P-ADJUST=0.0069)和体外新生毛细血管形成(0.47[0.28至0.66]，P-ADJUST<；0.0001)。在缺血诱导后第1、3和5天，小鼠左室注射降钙素原可损害新的毛细血管形成和体内后肢血流灌注的恢复(赋形剂治疗组和降钙素治疗组小鼠缺血腿的毛细血管数/mm(2)，852.6[383.4-1322]，P-ADJUST=.0002)。与降钙素原孵育24小时后，血管内皮细胞钙粘附素的表达减少(0.39[0.06~0.71]，P-ADJET=0.0167)，并诱导内皮细胞死亡(-5.4[-10.67~-0.13]，P-ADJET=0.0431)。细胞间黏附分子-1、血管细胞黏附分子-1或细胞外信号调节激酶1/2的表达及AKT信号转导通路未见改变。基因组分析揭示了参与炎症、血管生成和细胞生长的各种基因的调节。结论：本研究发现降钙素原本身就损害了内皮细胞功能的几个方面。降钙素原诱导的内皮屏障功能丧失可能导致脓毒症时毛细血管渗漏和治疗难治性低血压。低浓度的降钙素原的抗血管生成特性也可以确定降钙素原是与代谢综合征相关的血管疾病的媒介。未来的研究需要进一步测试降钙素原作为一种潜在的治疗靶点，在急性和慢性炎症性疾病期间保留血管功能障碍。
BACKGROUND: Procalcitonin is used as a diagnostic tool for the identification and risk stratification of septic patients. Procalcitonin plasma concentrations tightly correlate with the severity of the ongoing inflammatory reaction and can rise up to 10,000-fold. Impairment of endothelial cell function plays an important role in the pathogenesis of hypotension and disturbed organ perfusion during sepsis. We investigated the possible effects of procalcitonin itself on endothelial cell function and viability.METHODS: Human endothelial cells were exposed to 0.01 to 100 ng/mL procalcitonin and investigated for endothelial permeability using transwells, migration in a scratch wound assay and new capillary formation on extracellular matrix in vitro. Tumor necrosis factor-a and vascular endothelial growth factor served as positive controls. Procalcitonin's impact on the response of endothelial cells toward ischemia was investigated in vivo in the murine model of unilateral femoral artery ligation. Procalcitonin-exposed endothelial cells were subjected to immunoblot for the investigation of vascular endothelial-cadherin expression and angiogenic signaling pathways. Flow cytometry was used for the detection of inflammatory activation and viability, and genomic analysis was performed. Data are presented as difference in means and 95% confidence intervals; statistical analyses were performed using analysis of variance/Bonferroni, and P values are reported as adjusted for multiple comparisons (P-adjust).RESULTS: Tumor necrosis factor-alpha and 0.1 ng/mL procalcitonin induced endothelial barrier disruption after incubation of endothelial monolayers for 6 hours (-2.53 [-4.16 to -0.89], P = .0008 and -2.09 [-3.73 to -0.45], P-adjust = .0064 compared with vehicle-treated control, respectively). Procalcitonin beginning at concentrations of 0.02 ng/mL reduced endothelial cell migration (0.26 [0.06 to 0.47], P-adjust = .0069) and new capillary formation in vitro (0.47 [0.28 to 0.66], P-adjust < .0001) contrasting the proangiogenic action of vascular endothelial growth factor. Left ventricular injection of procalcitonin in mice on postoperative day 1, 3, and 5 after induction of ischemia impaired new capillary formation and recovery of hindlimb perfusion in vivo (number of capillaries/mm(2) in the ischemic leg of vehicle-treated versus procalcitonin-treated mice, 852.6 [383.4-1322], P-adjust = .0002). Twenty-four-hour incubation with procalcitonin reduced the expression of vascular endothelial-cadherin at 100 ng/mL (0.39 [0.06-0.71], P-adjust = .0167) and induced endothelial cell death (apoptosis, -5.4 [-10.67 to -0.13], P-adjust = .0431). No alteration in the expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1 or extracellular signal-regulated kinase 1/2, and AKT signaling pathways was observed. Genomic analysis revealed regulation of a variety of genes involved in inflammation, angiogenesis, and cell growth.CONCLUSIONS: This study found that procalcitonin itself impaired several aspects of endothelial cell function. Procalcitonin-induced loss of endothelial barrier function may contribute to capillary leakage and therapy-refractory hypotension during sepsis. Anti-angiogenic properties of procalcitonin at low concentrations could also identify procalcitonin as a mediator of vascular disease associated with the metabolic syndrome. Future studies are needed to further test procalcitonin as a potential therapeutic target for preserving vascular dysfunction during acute and chronic inflammatory disorders.