强直性脊柱炎患者细胞微粒对人脐静脉内皮细胞的影响
发布时间：2019-05-05 13:17
【摘要】：目的:1、研究细胞微粒(Microparticles,MPs)血小板微粒(Platelets microparticles,PMPs)、CD41a+CD62P+PMPs、粒细胞微粒(Granulocyte microparticles,GMPs)、CD4+T细胞微粒(CD4 positive T cells microparticles,CD4+T-LMPs)、CD8+T细胞微粒(CD8 positive T cells microparticles,CD8+T-LMPs)和B细胞微粒(B-cell microparticles,B-LMPs)在强直性脊柱炎(Ankylosing Spondylitis,AS)患者和正常人中的变化,观察AS患者MPs、PMP、CD41a+CD62P+PMPs、GMPs、CD4+T-LMPs、CD8+T-LMPs和B-LMPs水平与临床炎症指标的关系。2、探讨AS患者MPs对人脐静脉内皮细胞(human umbilical vein endothelial cell,HUVEC)的细胞存活率影响。进一步分析AS患者血清刺激形成的PMPs对HUVEC细胞间粘附分子-1(Intercellular adhesion molecule-1,ICAM-1)、血管细胞粘附分子-1(Vascular cell adhesion molecule-1,VCAM-1)和内皮型一氧化氮合酶(Endothelial nitric oxide synthase,e NOS)基因表达的影响。方法:收集47例AS患者外周血,所有患者均符合1984年修订的AS纽约标准,同时收集15例来院体检的健康人外周血作为对照组。(1)收集47例AS患者的空腹静脉血并检测血沉(ESR)、C反应蛋白(CRP)水平、评定BASDAI评分。(2)采用流式细胞术(Flow cytometry,FCM)检测AS患者和HC组MPs、PMPs、GMPs、B-LMPs、CD4+T-LMPs、CD8+T-LMPs和CD41a+CD62P+PMPs表达水平并分析AS患者MPs、PMPs、GMPs、B-LMPs、CD4+T-LMPs、CD8+T-LMPs和CD41a+CD62P+PMPs数量与临床炎症指标的相关性。(3)CCK8法检测HUVEC与MPs共孵育后的细胞存活率。(4)RT-PCR检测HUVEC与AS患者和HC刺激形成PMPs共孵育24 h后ICAM-1、VCAM-1和e NOSm RNA水平表达。结果:(1)AS患者MPs、PMPs、CD41a+CD62P+PMPs和GMPs数量显著高于健康对照组(p0.05)。AS患者B-LMPs,CD4+T-LMPs、CD8+T-LMPs数量与健康对照组相比,差异无统计学意义(p0.05)。(2)AS患者MPs、PMPs、CD41a+CD62P+PMPs、GMPs、B-LMPs、CD4+T-LMPs和CD8+T-LMPs数量与ESR,CRP无相关性。(3)AS患者MPs与BASDAI呈正相关(r=0.5548,P=0.0004)、PMPs与BASDAI呈正相关(r=0.5423,P=0.0006)、CD41a+CD62P+PMPs、GMPs与BASDAI无相关性(p0.05)。(4)AS患者与HC组MPs与HUVEC共培养24小时后,AS患者MPs能显著降低HUVEC细胞存活率(p0.01)。(5)AS与HC组血清刺激形成的PMPs与HUVEC细胞共培养24小时后,AS组、HC组和空白组之间e NOS、VCAM-1m RNA表达无明显统计学差异(p0.05)。而AS组ICAM-1m RNA表达显著高于空白组和HC组,差异具有统计学意义(p0.01)。结论:AS患者MPs、PMPs、GMPs和CD41a+CD62P+PMP表达增加。并且AS患者MPs,PMPs数量与BASDAI呈正相关,提示MPs,PMPs可能参与AS发病。此外AS患者MPs能够明显降低HUVEC细胞存活率,AS患者MPs可能损伤内皮细胞而参与心血管功能调控。此外,AS患者血清刺激产生的PMPs能够上调ICAM-1m RNA表达水平,提示MPs中的PMPs可能通过调节内皮细胞相关功能基因损伤内皮细胞,这可能在一定程度上导致AS患者心血管风险增加。
[Abstract]:Objective: 1. To study the cellular microparticles (Microparticles,MPs), platelet microparticles (Platelets microparticles,PMPs), CD41a CD62P PMPs, granulocyte particles (Granulocyte microparticles,GMPs), CD4 T cell particles (CD4 positive T cells microparticles,CD4 T-LMPs), CD8 T cell particles (CD8 positive T cells microparticles,). The changes of CD8 T-LMPs and B cell microparticles (B-cell microparticles,B-LMPs) in patients with ankylosing spondylitis (Ankylosing Spondylitis,AS) and normal controls were observed. To investigate the effect of MPs on the survival rate of human umbilical vein endothelial cells (human umbilical vein endothelial cell,HUVEC) in patients with AS. 2. The relationship between the levels of CD8 T-LMPs and B-LMPs and clinical inflammatory indexes. Further analysis of HUVEC intercellular adhesion molecule-1 (Intercellular adhesion molecule-1,ICAM-1 (ICAM-1 (Intercellular adhesion molecule-1,ICAM-1), vascular cell adhesion molecule-1 (Vascular cell adhesion molecule-1,VCAM-1 (VCAM-1 (Vascular cell adhesion molecule-1,VCAM-1) and endothelial nitric oxide synthase (Endothelial nitric oxide synthase, (Enos) induced by serum-stimulated PMPs in AS patients was carried out. The effect of e NOS) gene expression. Methods: peripheral blood samples were collected from 47 patients with AS, all of whom met the AS New York standard revised in 1984. At the same time, the peripheral blood of 15 healthy people were collected as control group. (1) fasting venous blood was collected from 47 patients with AS and erythrocyte sedimentation rate (ESR) (ESR), C reactive protein (CRP) level was measured. (2) flow cytometry (Flow cytometry,FCM) was used to detect the expression of MPs,PMPs,GMPs,B-LMPs,CD4 T cells, CD 8 T-LMPs and CD41a CD62P PMPs in patients with AS and HC and to analyze MPs,PMPs,GMPs,B-LMPs, in patients with AS. (2) BASDAI scores were evaluated by flow cytometry (Flow cytometry,FCM). CD4 T\ x {e16c} LMPs, The relationship between the number of CD8 T-LMPs and CD41a CD62P PMPs and clinical inflammatory markers. (3) the survival rate of HUVEC-MPs co-incubated with HUVEC was detected by CCK8. (4) ICAM-1, was detected by RT-PCR after PMPs was co-incubated with AS and HC for 24 h. VCAM-1 and e NOSm RNA were expressed at the level. Results: (1) the number of MPs,PMPs,CD41a CD62P PMPs and GMPs in AS patients was significantly higher than that in healthy controls (p0.05). There was no significant difference (p0.05). (2) between the number of MPs,PMPs,CD41a CD62P PMPs,GMPs,B-LMPs,CD4 T-LMPs and CD8 T-LMPs and ESR,CRP in AS patients. (3) there was a positive correlation between MPs and BASDAI in AS patients (r = 0.555 8, P < 0.0004). There was a positive correlation between PMPs and BASDAI (r = 0.5423, P = 0.0006), but there was no correlation between CD41a CD62P PMPs,GMPs and BASDAI (p0.05). (4). After 24 hours of co-culture of MPs and HUVEC between AS patients and HC group, MPs and HUVEC were co-cultured for 24 hours. The survival rate of HUVEC cells was significantly decreased by MPs in AS patients (p0.01). (5). After 24 hours of co-culture of PMPs and HUVEC cells induced by AS and serum stimulation in HC group, e NOS, was observed between AS group, HC group and blank group. There was no significant difference in the expression of VCAM-1m RNA (p0.05). The expression of ICAM-1m RNA in AS group was significantly higher than that in blank group and HC group (p0.01). Conclusion: the expression of MPs,PMPs,GMPs and CD41a CD62P PMP increased in AS patients. There was a positive correlation between the number of MPs,PMPs and BASDAI in patients with AS, suggesting that MPs,PMPs may be involved in the pathogenesis of AS. In addition, MPs can significantly reduce the survival rate of HUVEC cells in AS patients, and MPs in AS patients may damage endothelial cells and participate in cardiovascular function regulation. In addition, PMPs produced by serum stimulation in patients with AS can up-regulate the expression of ICAM-1m RNA, suggesting that PMPs in MPs may damage endothelial cells by regulating endothelial cell-related functional genes. This may, to some extent, lead to increased cardiovascular risk in patients with AS.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.23
本文编号：2469594
[Abstract]:Objective: 1. To study the cellular microparticles (Microparticles,MPs), platelet microparticles (Platelets microparticles,PMPs), CD41a CD62P PMPs, granulocyte particles (Granulocyte microparticles,GMPs), CD4 T cell particles (CD4 positive T cells microparticles,CD4 T-LMPs), CD8 T cell particles (CD8 positive T cells microparticles,). The changes of CD8 T-LMPs and B cell microparticles (B-cell microparticles,B-LMPs) in patients with ankylosing spondylitis (Ankylosing Spondylitis,AS) and normal controls were observed. To investigate the effect of MPs on the survival rate of human umbilical vein endothelial cells (human umbilical vein endothelial cell,HUVEC) in patients with AS. 2. The relationship between the levels of CD8 T-LMPs and B-LMPs and clinical inflammatory indexes. Further analysis of HUVEC intercellular adhesion molecule-1 (Intercellular adhesion molecule-1,ICAM-1 (ICAM-1 (Intercellular adhesion molecule-1,ICAM-1), vascular cell adhesion molecule-1 (Vascular cell adhesion molecule-1,VCAM-1 (VCAM-1 (Vascular cell adhesion molecule-1,VCAM-1) and endothelial nitric oxide synthase (Endothelial nitric oxide synthase, (Enos) induced by serum-stimulated PMPs in AS patients was carried out. The effect of e NOS) gene expression. Methods: peripheral blood samples were collected from 47 patients with AS, all of whom met the AS New York standard revised in 1984. At the same time, the peripheral blood of 15 healthy people were collected as control group. (1) fasting venous blood was collected from 47 patients with AS and erythrocyte sedimentation rate (ESR) (ESR), C reactive protein (CRP) level was measured. (2) flow cytometry (Flow cytometry,FCM) was used to detect the expression of MPs,PMPs,GMPs,B-LMPs,CD4 T cells, CD 8 T-LMPs and CD41a CD62P PMPs in patients with AS and HC and to analyze MPs,PMPs,GMPs,B-LMPs, in patients with AS. (2) BASDAI scores were evaluated by flow cytometry (Flow cytometry,FCM). CD4 T\ x {e16c} LMPs, The relationship between the number of CD8 T-LMPs and CD41a CD62P PMPs and clinical inflammatory markers. (3) the survival rate of HUVEC-MPs co-incubated with HUVEC was detected by CCK8. (4) ICAM-1, was detected by RT-PCR after PMPs was co-incubated with AS and HC for 24 h. VCAM-1 and e NOSm RNA were expressed at the level. Results: (1) the number of MPs,PMPs,CD41a CD62P PMPs and GMPs in AS patients was significantly higher than that in healthy controls (p0.05). There was no significant difference (p0.05). (2) between the number of MPs,PMPs,CD41a CD62P PMPs,GMPs,B-LMPs,CD4 T-LMPs and CD8 T-LMPs and ESR,CRP in AS patients. (3) there was a positive correlation between MPs and BASDAI in AS patients (r = 0.555 8, P < 0.0004). There was a positive correlation between PMPs and BASDAI (r = 0.5423, P = 0.0006), but there was no correlation between CD41a CD62P PMPs,GMPs and BASDAI (p0.05). (4). After 24 hours of co-culture of MPs and HUVEC between AS patients and HC group, MPs and HUVEC were co-cultured for 24 hours. The survival rate of HUVEC cells was significantly decreased by MPs in AS patients (p0.01). (5). After 24 hours of co-culture of PMPs and HUVEC cells induced by AS and serum stimulation in HC group, e NOS, was observed between AS group, HC group and blank group. There was no significant difference in the expression of VCAM-1m RNA (p0.05). The expression of ICAM-1m RNA in AS group was significantly higher than that in blank group and HC group (p0.01). Conclusion: the expression of MPs,PMPs,GMPs and CD41a CD62P PMP increased in AS patients. There was a positive correlation between the number of MPs,PMPs and BASDAI in patients with AS, suggesting that MPs,PMPs may be involved in the pathogenesis of AS. In addition, MPs can significantly reduce the survival rate of HUVEC cells in AS patients, and MPs in AS patients may damage endothelial cells and participate in cardiovascular function regulation. In addition, PMPs produced by serum stimulation in patients with AS can up-regulate the expression of ICAM-1m RNA, suggesting that PMPs in MPs may damage endothelial cells by regulating endothelial cell-related functional genes. This may, to some extent, lead to increased cardiovascular risk in patients with AS.
【学位授予单位】：西南医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.23
【参考文献】
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1 仲悦娇;陈宝安;黄成垠;李翠萍;高峰;费菲;裴孝平;高冲;丁家华;孙耘玉;程坚;王骏;赵刚;马燕;;血小板衍生膜微粒对人脐静脉内皮细胞增殖和凋亡的影响[J];中国实验血液学杂志;2007年04期
，本文编号：2469594
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