儿茶酚胺调节人单核细胞来源树突状细胞迁移功能及其机制研究

发布时间：2018-11-13 17:45

【摘要】： 目的：本研究观察慢性应激的主要神经递质儿茶酚胺(Catecholamines, CAs)对人外周血单核细胞来源的树突状细胞(Monocyte-derived dendritic cells, MoDCs)迁移功能以及趋化因子受体(Chemokine Receptor-7, CCR7)表达的影响,并进一步研究肾上腺素受体(adrenergic receptors, AR)-丝裂原活化蛋白激酶(Mitogen-Activated Protein Kinase, MAPK)-核因子-κB(nuclear factor-κB, NF-κB)通路在其中起的作用。方法：磁珠分选法纯化人外周血单核细胞,经rhGM-CSF和rh-IL4诱导分化为不成熟DCs。LPS诱导成熟的基础上给予不同浓度肾上腺素或去甲肾上腺素刺激,采用transwell小室检测DCs向MIP-3迁移能力,流式细胞仪及荧光定量逆转录多聚酶链反应检测CCR7基因和蛋白的表达,ELISA检测细胞因子IL10分泌的变化,Western-blot观察p38 MAPK及NF-κB通路的蛋白表达变化情况。进一步通过给予α、β1、β2、β等不同肾上腺素受体阻滞剂,以及MAPK、NF-κB等通路抑制剂,研究肾上腺素受体和信号通路在其中的作用。结果：肾上腺素和去甲肾上腺素不影响DCs的凋亡和成熟(CD86,HLA-DR),但均可抑制LPS诱导成熟的DCs向MIP-3β迁移(p0.05)。肾上腺素下调成熟DCs表达CCR7,而去甲肾上腺素上调成熟DCs表达CCR7并促进DC分泌IL10。D2受体阻滞剂对肾上腺素作用的抑制最明显,β1受体阻滞剂能抑制甲肾上腺素对IL10分泌的影响。p38 MAPK抑制剂、NF-κB抑制剂可以抑制LPS诱导的DCs迁移能力增强和CCR7表达增加(p0.01)；肾上腺素能抑制LPS刺激的磷酸化p38-MAPK、NF-κB表达。结论：肾上腺素可能通过p受体-p38 MAPK-NF-κB通路下调成熟DCs表面CCR7的表达,从而抑制成熟DCs的迁移功能。慢性应激时分泌增多的儿茶酚胺类神经递质,可能是动脉粥样硬化(Atherosclerosis, AS)条件下调节DCs迁移功能的因素之一,这可部分解释慢性应激在AS发病中的作用机制。
[Abstract]:Aim: to investigate the effects of catecholamine (Catecholamines, CAs), a major neurotransmitter of chronic stress, on the migration of dendritic cells (Monocyte-derived dendritic cells, MoDCs) derived from human peripheral blood monocytes and the chemokine receptor (Chemokine Receptor-7,). To investigate the role of Mitogen-Activated Protein Kinase, MAPK)-nuclear factor-魏 B (nuclear factor- 魏 B (NF- 魏 B) pathway in the expression of epinephrine receptor (adrenergic receptors, AR)-mitogen-activated protein kinase (NF- 魏 B). Methods: human peripheral blood monocytes were purified by magnetic bead sorting and differentiated into immature DCs.LPS by rhGM-CSF and rh-IL4. Different concentrations of epinephrine or norepinephrine were stimulated on the basis of differentiation into immature DCs.LPS. The migration ability of DCs to MIP-3 was detected by transwell chamber, the expression of CCR7 gene and protein was detected by flow cytometry and fluorescence quantitative reverse transcriptase polymerase chain reaction (FQ-PCR), and the secretion of cytokine IL10 was detected by ELISA. The protein expression of p38 MAPK and NF- 魏 B pathway was observed by Western-blot. The role of adrenergic receptor and signal pathway was studied by administration of different adrenergic receptor blockers, such as 伪, 尾 _ 1, 尾 _ 2, 尾 _ 2 and 尾, as well as MAPK,NF- 魏 B pathway inhibitors. Results: epinephrine and norepinephrine did not affect the apoptosis and maturation (CD86,HLA-DR) of DCs, but could inhibit the migration of mature DCs to MIP-3 尾 induced by LPS (p0.05). Epinephrine down-regulated the expression of CCR7, in mature DCs, and norepinephrine up-regulated the expression of CCR7 in mature DCs and promoted the secretion of IL10.D2 receptor blockers by DC. 尾 1 receptor blocker inhibited the effect of epinephrine on IL10 secretion. P38 MAPK inhibitor and NF- 魏 B inhibitor inhibited the increase of DCs migration and CCR7 expression induced by LPS (p0.01). Epinephrine inhibited the expression of phosphorylated p38-MAPK- 魏 B stimulated by LPS. Conclusion: epinephrine may down-regulate the expression of CCR7 on the surface of mature DCs through p38 MAPK-NF- 魏 B pathway, thereby inhibiting the migration of mature DCs. The increased secretion of catecholamine neurotransmitters during chronic stress may be one of the factors regulating the migration of DCs in atherosclerotic (Atherosclerosis, AS). This may partly explain the role of chronic stress in the pathogenesis of AS.
【学位授予单位】：第二军医大学
【学位级别】：博士
【学位授予年份】：2010
【分类号】：R392

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