作者投稿
专家审稿
编辑办公
邮件订阅
RSS
Immune Response of Peripheral Blood Mononuclear Cells to Lipopolysaccharide in vitro in Patients with Primary Biliary Cirrhosis
-
摘要:目的 通过观察原发性胆汁性肝硬化(primary biliary cirrhosis, PBC)患者外周血单个核细胞(peripheral blood mononuclear cells, PBMCs)在体外自然状况及在脂多糖(lipopolysaccharide, LPS)刺激下分泌细胞因子的特点, 探讨LPS和细胞因子在PBC发病机制中的作用。方法 采集PBC、原发性干燥综合征(primary Sj gren sjösyndrome, pSS)患者及健康对照者的PBMCs, 分别置于加或不加0.01mg/ml LPS的RPMI1640完全培养基中培养, ELISA法检测PB-MCs上清液中的细胞因子——白细胞介素(interleukin, IL)-1β、IL-2、IL-6、IL-10、肿瘤坏死因子α(tumor necrosisfactor-α, TNF-α)和干扰素γ(interferon-γ, IFN-γ)的水平。结果 (1) 新鲜提取的PBMCs与液氮冻存复苏后的PBMCs, 在体外对LPS刺激的反应具有明显差异。(2)在无LPS刺激培养的PBMCs上清液中, PBC患者的IL-2水平高于健康对照者, 差异有统计学意义(P < 0.05)。(3)在LPS刺激培养的PBMCs上清液中, PBC患者的IL-1β、IL-2和TNF-α水平高于健康对照者, 差异有统计学意义(P < 0.05)。(4) LPS刺激后, PBC患者PBMCs上清液的IL-1β升高幅度和IL-2降低幅度与健康对照者比较, 差异有统计学意义(P < 0.05)。(5) PBC与pSS比较, 无LPS刺激时各细胞因子水平差异无统计学意义; LPS刺激后PBC患者IL-6高于pSS患者, 差异有统计学意义(P < 0.05)。(6) IFN-γ在PBC患者PBMCs上清液中几乎检测不到, 将LPS浓度提高至0.1和1.0mg/ml仍未检测到IFN-γ。结论 PBC发病与细菌感染有关, LPS在PBC的发病中起一定作用; 多种细胞因子与PBC发病有关, 其中以Th1型细胞因子为主。Abstract:Objective To explore the roles of lipopolysaccharide (LPS) and cytokines in the pathogenesis of primary biliary cirrhosis (PBC).Methods The peripheral blood mononuclear cells (PBMCs) were collected from PBC patients, patients with primary Sjögren's syndrome (pSS), and healthy controls and then cultured in vitro in RPMI 1640 medium with or without LPS. The immune responses of PBMCs to 0.01mg/ml LPS in vitro were investigated by measuring the levels of cytokines(IL-1β, IL-2, IL-6, IL-10, TNF-α, IFN-γ) in supernates using enzyme-linked immunosorbent assay (ELISA).Results The response of freshly extracted PBMCs to LPS were remarkably different from that of the resuscitated PBMCs. In the supernates of PBMCs cultured without LPS for 24 hours in vitro, the level of IL-2 was significantly higher in PBC patients than in healthy controls (P < 0.05). In the supernates of PBMCs cultured with 0.01mg/ml LPS for 24 hours in vitro, the levels of IL-1β, IL-2, and TNF-α were significantly higher in PBC patients than in healthy controls (P < 0.05). After the stimulation of LPS, the elevation rate of IL-1β and decrease rate of IL-2 in PBC patients were significantly higher than those in healthy controls (P < 0.05). Compared with the pSS patients, the levels of cytokines in PBC patients showed no difference in culture media without LPS, while IL-6 level was significantly increased in the supernates cultured with LPS in PBC patients. IFN-γ were seldomly detected in the the supernates of PBMCs from PBC patients, even after the LPS concentrations were increased to 0.1 and 1.0 mg/ml.Conclusions The occurence of PBC is related to bacterial infections, in which LPS may play an important role. Many cytokines are involved in the pathogenesis of PBC, especially Th1 subtype.
-
世界初级创伤救治(Primary Trauma Care，PTC)理事会于2012年3月28日在阿根廷首都布宜诺斯艾利斯市召开了理事会议，来自卫生部医院管理研究所的阎赢主任和霍添琪，北京协和医院急诊科于学忠教授、王仲教授和麻醉科黄宇光教授出席了此次会议。会议回顾了PTC在全球近50个国家开展工作的情况，包括PTC在中国实施的成功经验。在卫生部医政司和医院管理研究所的领导下，PTC理事会与中国医师协会急诊医师分会、麻醉学医师分会联手合作，自2010年9月5日在中国成功启动PTC，至今已经在全国15个省份为2000多家医院培训了6000多名医学专业人员，取得了良好的培训效果。北京协和医院作为中国医师协会急诊医师分会和麻醉学医师分会的会长单位，为该项目做出了积极贡献。作为PTC理事，黄宇光教授代表中方团队介绍了PTC多学科全方位合作的中国模式。目前中国已经成为PTC项目培训在全球规模最大的国家，受到各位PTC理事和与会代表的好评。会议对如何促进PTC项目持续发展和相关教材的修订进行了讨论，并且决定PTC世界大会将于2013年在中国召开。
(卫生部医院管理研究所 霍添琪)
-
表 1 新鲜采集分离P BMCs与液氮冻存复苏PBMCs上清液的细胞因子比较
培养条件 IL-1β (pg /ml IL-2 (U/ml) IL-6(pg /ml) IL-10(pg /ml) TNF-α(pg /ml) 无LPS刺激 新鲜PBC 1645±1677 2. 36±0. 18 45 763 ±43484 721±701 2962±2944 冻存PBC 1761±1871 1. 97±0. 61 14 982 ±10931 533±349 3063±1499 P值 0. 879 0.452 0.035 0.368 0.723 LPS刺激 新鲜PBC 5196±2641 1. 86±0. 35 73 169 ±46387 1003 ±629 5827±2154 冻存PBCs 2581±2320 2. 30±1. 12 28 229 ±19874 347 ±278 5353±3029 P值 0. 029 0.396 0.014 0.547 0.009 PBMCs :外周血单个核细胞; LPS:脂多糖; I L:白细胞介素; TNF-α:肿瘤坏死因子α 
下载: 导出CSV
表 2 无LPS刺激PBMCs的细胞因子表达
培养条件 IL-1β (pg /ml IL-2 (U/ml) IL-6(pg /ml) IL-10(pg /ml) TNF-α(pg /ml) 健康对照 842± 602 2. 07±0. 30 29 468 ±32446 646±674 1605±1454 pSS 2646±3390 2. 49±1. 39 33 920 ±17500 334±325 3584±4017 PBC 1645±1677 2. 36±0. 18* 45 763 ±43484 721±701 2962±2944 pSS:原发性干燥综合征; PBC:原发性胆汁性肝硬化; 与健康对照组比较, *P﹤0. 05
下载: 导出CSV
表 3 LPS刺激后PBMCs的细胞因子表达
培养条件 IL-1β (pg /ml IL-2 (U/ml) IL-6(pg /ml) IL-10(pg /ml) TNF-α(pg /ml) 健康对照 1525±1022 1. 54±0. 28 57 933 ±54206 1183 ±939 3582±1292 pSS 4054±2742 2. 46±0. 82 13 391 ± 5425 529 ±596 5207±3742 PBC 5196±2641** 1. 86±0. 35* 73169±46 387† 1003±629 5827±2154** 与健康对照组比较, *P < 0. 05, **P < 0. 01;与pSS组比较, †P < 0. 001
下载: 导出CSV
表 4 有无LP S刺激对PBC与健康对照组PBMCs细胞因子的影响
培养条件 IL-1β (pg /ml IL-2 (U/ml) IL-6(pg /ml) IL-10(pg /ml) TNF-α(pg /ml) 健康对照 1525±1022 1. 54±0. 28 57 933 ±54206 1183 ±939 3582±1292 pSS 4054±2742 2. 46±0. 82 13 391 ± 5425 529 ±596 5207±3742 PBC 5196±2641** 1. 86±0. 35* 73169±46 387† 1003±629 5827±2154** 与健康对照组比较, *P < 0. 05, **P < 0. 01;与pSS组比较, †P < 0. 001
下载: 导出CSV
表 5 LPS刺激后 PBMCs上清液细胞因子的变化
(%) 组 别 IL-1β IL-2 IL-6 IL-10 TNF-α 健康对照 90 ± 72 -26 ±13 221 ±297 132 ±100 220 ±139 pSS 276 ±320 16 ±47 -23 ±121 161 ±241 160 ±162 PBC 462 ±580* -21 ±14* 228 ±438 92 ±135 247 ±232 与健康对照组比较 , *P<0.05
下载: 导出CSV
表 6 不同条件培养的 PBMCs上清液中的 IFN-γ水平
(U/ml) 培养条件 健康对照 (n=11) PBC (n=12) 24 h 48 h 24 h 48 h 无 LPS刺 激 -0.71 ±0.42 -0.56 ±0.35 -0.58 ±0.42 -0.57 ±0.45 LPS刺激 (mg/ml) 0.01 -0.76 ±0.49 -0.73 ±0.48 -0.56 ±0.41 -0.51 ±0.41 0.1 -0.81 ±0.50 -0.61 ±0.35 -0.64 ±0.54 -0.59 ±0.45 1.0 -0.86 ±0.59 -0.82 ±0.56 -0.66 ±0.53 -0.66 ±0.54
下载: 导出CSV
-
[1] Wasilenko ST, Mason GE, Mason AL.Primary biliary cirrhosis, bacteria and molecula mimicry:what's the molecule and where's the mimic[J]? Liver Int, 2009, 29:779-782. DOI: 10.1111/j.1478-3231.2009.02051.x
[2] Ballot E, Bandin O, Chazouilleres O, et al.Immune response to lipopolysaccharide in primary biliary cirrhosis and autoimmune diseases[J].J Autoimmun, 2004, 22:153- 158. DOI: 10.1016/j.jaut.2003.11.002
[3] Heathcote EJ.Management of primary biliary cirrhosis[J]. Hepatology, 2000, 31:1005-1013. DOI: 10.1053/he.2000.5984
[4] Vitali C, Bombardieoi S, Jonssin R, et al.Classification criteria for Sjögren's syndrome:a revised version of European criteria proposed by the American European Consensus Group[J].Ann Rheum Dis, 2002, 61:554-558. DOI: 10.1136/ard.61.6.554
[5] Brown SB, Savill J.Pagocytosis triggers macrophage release of Fas ligand and induces apoptosis of bystander leukocytes [J].J Immunol, 1999, 162:480-485. http://europepmc.org/abstract/med/9886423
[6] Nagano T, Yamamoto K, Matsumoto S, et al.Cytokine profile in the liver of primary biliary cirrhosis[J].J Clin Immunol, 1999, 19:422-427. DOI: 10.1023/A:1020511002025
[7] Solís Herruzo JA, Solís Muñoz P, Muñoz Yagǜe T.The pathogenesis of primary biliary cirrhosis[J].Rev Esp Enferm Dig, 2009, 101:413-423.
[8] Mao TK, Lian ZX, Selmi C, et al.Altered monocytes responses to defined TLR ligands in patients with primary biliary cirrhosis[J].Hepatology, 2005, 42:802-808. DOI: 10.1002/hep.20859
[9] Barak V, Selmi C, Schlesinger M, et al.Serum inflammatory cytokines, complement components, and soluble interleukin 2 receptor in primary biliary cirrhosis[J].J Autoimmun, 2009, 33:178-182. DOI: 10.1016/j.jaut.2009.09.010
[10] Parikh-Patel A, Gold EB, Worman H, et al.Risk factors for primary biliary cirrhosis in a cohort of patients from the United States[J].Hepatology, 2001, 33:16-21. DOI: 10.1053/jhep.2001.21165
[11] Sasatomi K, Noguchi K, Sakisaka S, et al.Abnormal accumulation of endotoxin in biliary epithelial cells inprimary biliary cirrhosis and primary sclerosing cholangitis[J].J Hepatol, 1998, 29:409-416. DOI: 10.1016/S0168-8278(98)80058-5
[12] Honda Y, Yamagiwa S, Matsuda Y, et al.Altered expression of TLR homolog RP105 on monocytes hypersensitive to LPS in patients with primary biliary cirrhosis[J].J Hepatol, 2007, 47:404-411. DOI: 10.1016/j.jhep.2007.03.012
[13] Dumoulin FL, Leifeld L, Honecker U, et al.Intrahepatic expression of interleukin-1 beta and tumor necrosis factor-alpha in chronic hepatitis C [J].J Infect Dis, 1999, 180: 1704-1708. DOI: 10.1086/315070
[14] Yamashiki M, Kosaka Y, Nishimura A, et al.Analysis of serum Cytokine Levels in primary biliary cirrhosis patients and healthy adults[J].J Clin Lab Anal, 1998, 12:77-82. DOI: 10.1002/(SICI)1098-2825(1998)12:2<77::AID-JCLA1>3.0.CO;2-G
[15] Lohr HF, Schlaak JF, Gerken G, et al.Phenotypic alanalysis and cytokine release of liver-infiltrating and peripheral blood T lymphocytes from patients with chronic hepatitis of different etiology[J].Liver, 1994, 14:161-166. http://www.ncbi.nlm.nih.gov/pubmed/8078396/
[16] Matsumoto T, Morizane T, Aoki Y, et al.Autoimmune hepatitis in primary Sjögren's Syndrome:pathological study of the livers and labial salivary glands in 17 patients with primary Sjögren's Syndrome[J].Pathol Int, 2005, 55:70- 76. DOI: 10.1111/j.1440-1827.2005.01790.x
[17] Liu B, Zhang FC, Zhang ZL, et al.Interstitial lung disease and Sjögren's syndrome in primary biliary cirrhosis:a causal or casual association [J]? Clin Rheumatol, 2008, 27: 1299-1306. DOI: 10.1007/s10067-008-0917-x
-
期刊类型引用(1)
1. 屈建，刘高峰，朱珠，孙春华. 我国医院药学学科的建设与发展(下). 中国医院药学杂志. 2014(17): 1423-1433 . 
百度学术
其他类型引用(0)
计量
- 文章访问数: 152
- HTML全文浏览量: 83
- PDF下载量: 13
- 被引次数: 1
