CCK8和PGE2对CVB体外攻击人外周血pDC的调控
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摘要
目的:了解八肽胆囊收缩素(CCK8)是否参与柯萨奇病毒B(CVB)体外攻击人外周血浆细胞样树突状细胞(p DC)的调控。方法:RT-PCR和IF检测CCK受体的表达;FCM分析协同刺激分子的变化,用ELISA和RT-PCR检测IFN-α的分泌。结果:CVB体外攻击p DC,CCK1R和CCK2R的表达增加;CCK8可以抑制CD80、CD86和HLA-DR表达的增高,而PGE2可以进一步促进CD80、CD86和HLA-DR的表达;同时,CCK8抑制CVB攻击后IFN-α的分泌,而PGE2进一步促进IFN-α的分泌。结论:CCK8和PGE2可双向调控CVB体外攻击人外周血p DC的免疫应答。
Objective:To investigate the immunomodulation of CCK8 on the Coxsackievirus B(CVB)-attacked human peripheral blood plasmacytoid dendritic cells(p DC).Methods:Peripheral blood mononuclear cells of healthy volunteers were separated by Ficoll-Hypaque gradient density centrifugation.The p DC was separated and divided into five groups,which were the control group,CVB attacked group,the group of CCK8 treated after CVB attack,the group of PGE2 treated after CVB attack and the group of CCK8+PGE2 treated after CVB attack.100-time TCID50 of CVB was applied for the attack on p DC.Real-time PCR and Immunofluorescence technique were employed to detect the expression of CCK1 R/CCK2 R mRNA and protein.Then,the expression levels of costimulatory molecules such as CD80,CD86,HLA-DR ligand,and the chemokine receptor CCR7 were evaluated by Flow Cytometry Analysis.The supernatants of p DCs were collected,and the content of IFN-α was determined by Enzyme-linked Immunosorbent Assay.Results:CCK1 R and CCK2 R were co-expressed in human peripheral blood p DC,and both were significantly upregulated after CVB attack in vitro.Expression of CD80,CD86,HLA-DR and IFN-α were decreased in the CVB + CCK8 group compared with the CVB group,which suggested that CCK8 may reduce the CVB activation of p DC.Whereas expression of CD80,CD86,HLA-DR,CCR7 and IFN-α were increased in the CVB + PGE2 group compared with the CVB group,which suggested that PGE2 may increase the CVB activation of p DC in vitro.Conclusion:CCK8 repressed the CVB-attacked p DC,while PGE2 activated the CVB-attacked p DC.
Objective:To investigate the immunomodulation of CCK8 on the Coxsackievirus B(CVB)-attacked human peripheral blood plasmacytoid dendritic cells(p DC).Methods:Peripheral blood mononuclear cells of healthy volunteers were separated by Ficoll-Hypaque gradient density centrifugation.The p DC was separated and divided into five groups,which were the control group,CVB attacked group,the group of CCK8 treated after CVB attack,the group of PGE2 treated after CVB attack and the group of CCK8+PGE2 treated after CVB attack.100-time TCID50 of CVB was applied for the attack on p DC.Real-time PCR and Immunofluorescence technique were employed to detect the expression of CCK1 R/CCK2 R mRNA and protein.Then,the expression levels of costimulatory molecules such as CD80,CD86,HLA-DR ligand,and the chemokine receptor CCR7 were evaluated by Flow Cytometry Analysis.The supernatants of p DCs were collected,and the content of IFN-α was determined by Enzyme-linked Immunosorbent Assay.Results:CCK1 R and CCK2 R were co-expressed in human peripheral blood p DC,and both were significantly upregulated after CVB attack in vitro.Expression of CD80,CD86,HLA-DR and IFN-α were decreased in the CVB + CCK8 group compared with the CVB group,which suggested that CCK8 may reduce the CVB activation of p DC.Whereas expression of CD80,CD86,HLA-DR,CCR7 and IFN-α were increased in the CVB + PGE2 group compared with the CVB group,which suggested that PGE2 may increase the CVB activation of p DC in vitro.Conclusion:CCK8 repressed the CVB-attacked p DC,while PGE2 activated the CVB-attacked p DC.
引文
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[17]Maric J,Ravindran A,Mazzurana L,et al.PGE2 suppresses human group 2 innate lymphoid cell function[J].J Allergy Clin Immunol,2018,141(5):1761-1773.
[2]叶丽萍,胡静涛,王春凤.树突状细胞与轮状病毒相互作用机制研究进展[J].中国免疫学杂志,2017,33(6):947-950.Ye LP,Hu JT,Wang CF.Recent progress in the mechanism of interaction between dendritic cells and rotavirus[J].Chin J Immunol,2017,33(6):947-950.
[3]Mc Govern N,Shin A,Low G,et al.Human fetal dendritic cells promote prenatal T-cell immune suppression through arginase-2[J].Nature,2017,546(7660):662-666.
[4]Anguille S,Van de Velde AL,Smits EL,et al.Dendritic cell vaccination as postremission treatment to prevent or delay relapse in acute myeloid leukemia[J].Blood,2017,130(15):1713-1721.
[5]Macri C,Pang ES,Patton T,et al.Dendritic cell subsets[J].Semin Cell Dev Biol,2017,doi:10.1016/j.semcdb.2017.12.009.
[6]Patel VI,Metcalf JP.Identification and characterization of human dendritic cell subsets in the steady state:a review of our current knowledge[J].J Investig Med,2016,64(4):833-847.
[7]Alculumbre S,Pattarini L.Purification of human dendritic cell subsets from peripheral blood[J].Methods Mol Biol,2016,1423:153-167.
[8]Stead SO,Mc Innes SJP,Kireta S,et al.Manipulating human dendritic cell phenotype and function with targeted porous silicon nanoparticles[J].Biomaterials,2018,155:92-102.
[9]Hamalainen S,Nurminen N,Ahlfors H,et al.Coxsackievirus B1 reveals strain specific differences in plasmacytoid dendritic cell mediated immunogenicity[J].J Med Virol,2014,86(8):1412-1420.
[10]Wu F,Fan X,Yue Y,et al.A vesicular stomatitis virus-based mucosal vaccine promotes dendritic cell maturation and elicits preferable immune response against coxsackievirus B3 induced viral myocarditis[J].Vaccine,2014,32(31):3917-3926.
[11]Jia X,Cong B,Zhang J,et al.CCK8 negatively regulates the TLR9-induced activation of human peripheral blood p DCs by targeting TRAF6 signaling[J].Eur J Immunol,2014,44(2):489-499.
[12]Zhang JG,Liu JX,Jia XX,et al.Cholecystokinin octapeptide regulates the differentiation and effector cytokine production of CD4(+)T cells in vitro[J].Int Immunopharmacol,2014,20(2):307-315.
[13]Zhang D,Li H,Geng J,et al.The therapeutic effects of cholecystokinin octapeptide on rat liver and kidney microcirculation disorder in endotoxic shock[J].Immunopharmacol Immunotoxicol,2017,39(1):2-10.
[14]Liao Y,Chen KH,Dong XM,et al.A role of pre-mir-10a coding region variant in host susceptibility to coxsackie virus-induced myocarditis[J].Eur Rev Med Pharmacol Sci,2015,19(18):3500-3507.
[15]Cen Z,Guo Y,Kong Q,et al.IL-10-producing B cells involved in the pathogenesis of Coxsackie virus B3-induced acute viral myocarditis[J].Int J Clin Exp Pathol,2015,8(1):830-835.
[16]Teijaro JR.Type I interferons in viral control and immune regulation[J].Curr Opin Virol,2016,16:31-40.
[17]Maric J,Ravindran A,Mazzurana L,et al.PGE2 suppresses human group 2 innate lymphoid cell function[J].J Allergy Clin Immunol,2018,141(5):1761-1773.