Supplementary material
Supplementary material
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- 补充方法和结果部分
- 补充图S1-A)LPS剂量和时间轴研究:在腹腔内LPS(25μg,50μg,100μg)给药(每个)的4,24,48小时处处死小鼠(每次n = 4)。将在这些同一时间点处死的盐水灌注的小鼠被汇集为对照。LPS后48小时(100μg)时间点实验过早地终止,因为小鼠经历了突然死亡或瘫痪。基于Interllaque出血和血栓形成的组织学证据来定义滴漏稳定化。暴露于50μg和100μg的剂量肿瘤内LPS暴露于腹腔炎LPS的小鼠在4小时和24小时内显示出类似的动脉泡不稳定率。选择后LPS管理后24小时作为调查的主要时间点,因为LPS 24小时与盐水24小时表现出斑块脆弱性的显着变化(Fisher精确测试P <0.005)。b)斑块的不同形貌组分的代表性图:BCT的横截面中的血管内腔,斑块,坏死核和帽区。通过确定内皮层(内皮区域)包围的区域封闭的区域来确定斑块区域。坏死核被定义为不含任何基质材料(胶原或弹性蛋白)或细胞的斑块的组分。通过使用颜色分割量化该区域。 The percent (%) necrotic core was calculated by dividing the necrotic core area by the total plaque area; and c) the method used to determine cap-plaque thickness ratio using Image Pro Plus software. A plaque image was overlaid with a known pattern ray originating from the mid portion of inner elastic lamina (IEL) of the plaque. The cap-plaque thickness ratio was calculated by determining the distance from the endothelium to the necrotic core (yellow line) and then dividing this number by the distance from the endothelium to the spectrum origin (yellow line + blue line). An arithmetic mean value of five measurements was calculated and used for statistical analysis.
- 补充图S2- a)H&E染色小鼠留下肺部肺部肿块肿块肿瘤暴露在50μl无菌盐水(左)和100μg的LPS中溶解在50μl无菌盐水(右)。在暴露于LPS的小鼠肺中,中性粒细胞数量增加,中性粒细胞,上皮细胞,血液充血和水肿;b)显示从盐水和LPS暴露小鼠的支气管肺泡灌洗(BAL)流体中的中性粒细胞的定量。误差线表示均值的标准差。将BAL中的细胞轻轻地混合,然后将缩合到玻璃载玻片上,并在用H&E染色染色之前用甲醇固定。通过评估颗粒和多核的尺寸,形状和存在形态地鉴定PMN。单核细胞和肺泡巨噬细胞也通过形态学决定;c)来自LPS-CTRL组的支气管肺泡灌洗液(BAL)中嗜中性粒细胞的细胞螺旋藻图像的实例(注射对照抗体的LPS-暴露的小鼠;左)和LPS-Nd(LPS-暴露小鼠注入中性粒细胞特异性抗LY6G抗体,右侧);d)在BAL中的中性粒细胞计数的定量。误差线表示均值的标准差。 Neutrophil depletion in the LPS-exposed mice significantly attenuated the number of neutrophils in BAL.
- 补充图S3- Inflammatory cytokine concentration of a) keratinocyte chemoattractant (KC), b) monocyte chemotactic protein-1 (MCP-1), c) interleukin-6 (IL-6), and d) tumor necrosis factor alpha (TNF-α) in mouse plasma 24 hours after intratracheal exposure to saline (N=4) or LPS (N=4); error bars represent the standard errors.
- 补充图S4——急性斑块并发症由于有限合伙人肺博览会ure: a) The edge region of plaque in LPS-exposed mice displayed blood aggregated on the surface of plaque and mixed with plaque tissue (H&E). Loss of endothelium integrity was observed at the edge of plaque. b) The necrotic core region in the plaque of LPS treated mice showed intraplaque hemorrhage. c) Immunofluorescent staining of thrombin (green) in the plaque is shown. d) Both intraplaque hemorrhage and luminal thrombus were detected in the BCT of LPS-exposed mice by OPT (high relative intensity signals are demarcated by a blue line, vessel wall is demarked by a pink line, and the perimeter of the plaque is demarcated by a green line). e) 3-D OPT cross-sectional image of a plaque containing hemorrhage (Green: lower intensity blood signal, Yellow: medium intensity blood signal; Red: highest intensity blood signal). f) Immunfluorescent staining of MMP-9 (green) of plaque in LPS-exposed mice (left) and saline-exposed mice (right). There was MMP-9 in the sub-endothelial region and in areas where plaque had been perturbed; in contrast, there was no MMP-9 signal detected in saline treated plaque. g) Luminal blood clots (high intensity signal) were detected on plaque shoulders of BCT and extended into aortic arch in the LPS lung exposed animal.
- 补充图S5- a)代表自体荧光强度map of mouse BCT from LPS-Ctrl group demonstrating intraplaque high intensity signal consistent with intraplaque hemorrhage (left) and that from LPS-ND group demonstrating lower signal intensity consistent with a stable plaque (right). b) A representative OPT cross-sectional image of mouse BCT from LPS-Ctrl group that shows high intensity signal at the shoulder region consistent with intraplaque hemorrhage (left) and that from LPS-ND group that shows a stable phenotype (right). c) An OPT cross-sectional image from the proximal end of BCT from a LPS-Ctrl mouse that shows a luminal blood clot at the edge of a plaque. d) OPT cross-sectional images from the proximal end of BCT of a mouse injected with control antibodies (Ctrl, left) and of a mouse injected with neutrophil-specific antibody (ND, right). Both images show a stable plaque phenotype with no features of vulnerability. e) Immunofluorescent staining of MPO (green) and CD68 (red) in cross-sections of BCT from LPS-exposed mice that shows MPO+ polymorphonuclear cells and CD68+ macrophages localized at the edges of a plaque, and f) these MPO+ polymorphonuclear cells (green) also observed in the plaques of LPS-Ctrl mice (mouse treated with control antibodies) g) An confocal image of representative neutrophil stained with MPO (green) found in an area of plaque rupture.
- 补充图S6- Inflammatory cell counts from the bronchoalveolar lavage (BAL); Error bars represent standard error of the mean: a) BAL neutrophil counts from saline group, Ctrl group (with injections of control IgG antibodies), and ND group (injections of neutrophil-specific antibodies) are shown. Injections of antibodies into mice in vivo did not induce neutrophil lung infiltration. b) Quantitation of macrophage in the BAL of mice in the saline group, Ctrl group, ND group, LPS group, LPS-Ctrl group (LPS group with injections of control IgG antibodies), and LPS-ND group (LPS group with injections of neutrophil specific antibodies. c) Comparisons of BAL neutrophil counts between IT (intratracheal) route and IP (intraperitoneal) route at the 8 hour time point. LPS exposure via IT route induced neutrophil lung inflammation, but not LPS exposure via IP route. d) There were no significant changes in the macrophage counts between these groups. Eosinophil and basophil counts were consistently rare (less than 1% of the total cell counts); thus, they were not included in these data.
- 补充图S7- Circulating white blood cell counts from IT (intratracheal) and IP (intraperitoneal) groups are shown. Error bars represent the standard errors: a) Total white blood cells, b) neutrophils, c) lymphocytes, and d) monocytes in mouse blood 8 hours after saline and LPS administration demonstrated a similar pattern change. Eosinophil and basophil counts were consistently rare (less than 1% of the total cell counts); thus, they were not included in these data.
- Supplementary video S1- 从LPS暴露的鼠标含有Brococephalic Trunk(BCT)的小鼠主动脉弓的3D介绍:血液(红色)是比血管壁(蓝色)高10倍的自动荧光。LPS小鼠中的血凝块与BCT中的破裂斑块的肩部相关。
- Supplementary video S2- 从盐水暴露的鼠标中的3D呈小鼠主动脉弓和肱骨躯干(BCT):在BCT的动脉瘤中没有观察到血液凝块或肿瘤发血。
- Supplementary video S3- 3D presentation of aortic arch containing brachiocephalic trunk (BCT) from another LPS-exposed mouse that displayed intraplaque hemorrhage (red color) inside atheroma.
- Supplementary video S4- 3D presentation of aortic arch containing brachiocephalic trunk (BCT) from a different LPS-exposed mouse: Blood clots (color red) localized in the plaque shoulders at the proximal end of BCT and extended into the aortic arch.
- Supplementary video S5- 3D presentation of brachiocephalic trunk (BCT) from LPS-exposed mice treated with neutrophil-specific antibodies (LPS-ND, left) and control antibodies (LPS-Ctrl, right): Blood clots (color red) localized at the plaque shoulders of BCT in the LPS-Ctrl group but not in the LPS-ND group.