【摘要】： 目的：本研究觀察慢性應(yīng)激的主要神經(jīng)遞質(zhì)兒茶酚胺(Catecholamines, CAs)對人外周血單核細胞來源的樹突狀細胞(Monocyte-derived dendritic cells, MoDCs)遷移功能以及趨化因子受體(Chemokine Receptor-7, CCR7)表達的影響,并進一步研究腎上腺素受體(adrenergic receptors, AR)-絲裂原活化蛋白激酶(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：磁珠分選法純化人外周血單核細胞,經(jīng)rhGM-CSF和rh-IL4誘導(dǎo)分化為不成熟DCs。LPS誘導(dǎo)成熟的基礎(chǔ)上給予不同濃度腎上腺素或去甲腎上腺素刺激,采用transwell小室檢測DCs向MIP-3遷移能力,流式細胞儀及熒光定量逆轉(zhuǎn)錄多聚酶鏈反應(yīng)檢測CCR7基因和蛋白的表達,ELISA檢測細胞因子IL10分泌的變化,Western-blot觀察p38 MAPK及NF-κB通路的蛋白表達變化情況。進一步通過給予α、β1、β2、β等不同腎上腺素受體阻滯劑,以及MAPK、NF-κB等通路抑制劑,研究腎上腺素受體和信號通路在其中的作用。 結(jié)果：腎上腺素和去甲腎上腺素不影響DCs的凋亡和成熟(CD86,HLA-DR),但均可抑制LPS誘導(dǎo)成熟的DCs向MIP-3β遷移(p0.05)。腎上腺素下調(diào)成熟DCs表達CCR7,而去甲腎上腺素上調(diào)成熟DCs表達CCR7并促進DC分泌IL10。D2受體阻滯劑對腎上腺素作用的抑制最明顯,β1受體阻滯劑能抑制甲腎上腺素對IL10分泌的影響。p38 MAPK抑制劑、NF-κB抑制劑可以抑制LPS誘導(dǎo)的DCs遷移能力增強和CCR7表達增加(p0.01)；腎上腺素能抑制LPS刺激的磷酸化p38-MAPK、NF-κB表達。 結(jié)論：腎上腺素可能通過p受體-p38 MAPK-NF-κB通路下調(diào)成熟DCs表面CCR7的表達,從而抑制成熟DCs的遷移功能。慢性應(yīng)激時分泌增多的兒茶酚胺類神經(jīng)遞質(zhì),可能是動脈粥樣硬化(Atherosclerosis, AS)條件下調(diào)節(jié)DCs遷移功能的因素之一,這可部分解釋慢性應(yīng)激在AS發(fā)病中的作用機制。
[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 偽, 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.
【學(xué)位授予單位】：第二軍醫(yī)大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2010
【分類號】：R392

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本文編號：2329909