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目的:测定脓毒血症鼠Peyer小结内滤泡辅助性T细胞(T follicular helper,Tfh)的变化,分析地塞米松(dexamethasone,Dex)对脓毒血症Tfh细胞的可能作用。方法:昆明小鼠诱导脓毒血症模型,模型诱导成功后随机分2组,脓毒血症组(SE组,n=12)、脓毒血症+Dex处理组(DE组,n=12)。另设对照组(NC组,n=12)。测定血清白介素-6(interleukin-6,IL-6)、降钙素原(procalcitonin,PCT)、中性粒细胞明胶酶相关载脂蛋白(neutrophil gelatinase-associated lipocalin,NGAL)水平。Peyer小结Ⅰ型1-磷酸鞘氨醇(Sphingosine 1-Phosphate 1,S1P1)、CXC趋化因子配体13(CXC chemokine ligand 13,CXCL13)免疫组化染色。Western blot分别检测Peyer小结IL-21、程序性死亡分子-1(programmed death 1,PD-1)、胞嘧啶脱氨酶(enzyme activation-induced cytidine deaminase,AID)蛋白的表达。流式细胞仪测定3组Peyer小结Tfh细胞占T淋巴细胞的百分率。结果:与NC组相比,SE组血清IL-6(19.7±5.20 vs 10.7±3.60 ng·L~(-1))、PCT(1.56±0.92 vs 0.31±0.09μg·L~(-1))、NGAL(0.44±0.11 vs 0.35±0.09 mg·L~(-1))升高(P<0.05或0.01),Peyer小结S1P1(0.22±0.06 vs 0.14±0.04)、CXCL13(0.25±0.07 vs 0.15±0.04)的表达增加(P<0.01),IL-21(0.60±0.08 vs 0.35±0.08)、PD-1(0.30±0.04 vs 0.20±0.05)、AID(0.23±0.05 vs 0.18±0.03)蛋白的表达亦升高(P<0.05或0.01),Tfh细胞占T淋巴细胞(8.30±2.00 vs 5.69 vs 1.64%)的百分率升高(P<0.01)。Dex治疗可降低SE鼠血清IL-6(19.7±5.20 vs 12.8±3.40 ng·L~(-1))、PCT(1.56±0.92 vs 0.71±0.44μg·L~(-1))水平(P<0.05或0.01),降低Peyer小结S1P1(0.22±0.06 vs 0.17±0.05)、CXCL13(0.25±0.07 vs 0.19±0.06)的表达(P<0.05或0.01),下调Peyer小结IL-21(0.60±0.08 vs 0.48±0.09)、PD-1(0.30±0.04 vs 0.26±0.06)、AID(0.23±0.05 vs0.19±0.04)蛋白的表达(P<0.05),降低Tfh细胞占T淋巴细胞(8.30±2.00 vs 6.56±1.59%)的百分率(P<0.05)。结论:Peyer小结Tfh细胞可能参与脓毒血症的发病机制,Dex抑制Peyer小结Tfh的活化,对Peyer小结微环境起保护作用。
Objective: To determine the changes of T follicular helper (Tfh) in septic mice Peyer nodules, and to analyze the possible effect of dexamethasone (Dex) on sepsis Tfh cells. Methods: The model of sepsis was induced in Kunming mice. The rats were randomly divided into 2 groups: sepsis group (SE group, n = 12), sepsis + Dex group (n = 12 ). Another control group (NC group, n = 12). Serum levels of IL-6, procalcitonin (PCT) and neutrophil gelatinase-associated lipocalin (NGAL) were measured. Peyer’s summary immunohistochemical staining of Sphingosine 1-Phosphate 1 (S1P1) and CXC chemokine ligand 13 (CXCL13). Western blot were used to detect the expression of IL-21, PD-1 and AID in Peyer’s nodules. Flow cytometry Tyer cells in three groups Tfh cells accounted for the percentage of T lymphocytes. Results: Compared with NC group, serum IL-6 (19.7 ± 5.20 vs 10.7 ± 3.60 ng · L -1), PCT (1.56 ± 0.92 vs 0.31 ± 0.09 μg · L -1) (P <0.05 or 0.01), NGAL (0.44 ± 0.11 vs 0.35 ± 0.09 mg · L -1) (P <0.05 or 0.01) (P <0.01), the expression of IL-21 (0.60 ± 0.08 vs 0.35 ± 0.08), PD-1 (0.30 ± 0.04 vs 0.20 ± 0.05) and AID (0.23 ± 0.05 vs 0.18 ± 0.03) (P <0.05 or 0.01). The percentage of Tfh cells in T lymphocytes (8.30 ± 2.00 vs 5.69 vs 1.64%) was also increased (P <0.01). Dex treatment reduced serum IL-6 (19.7 ± 5.20 vs 12.8 ± 3.40 ng · L -1) and PCT (1.56 ± 0.92 vs 0.71 ± 0.44 μg · L -1) in SE rats (P < 0.05 or 0.01), and decreased the expression of Peyer’s tubal S1P1 (0.22 ± 0.06 vs 0.17 ± 0.05) and CXCL13 (0.25 ± 0.07 vs 0.19 ± 0.06) (P <0.05 or 0.01) 0.48 ± 0.09), PD-1 (0.30 ± 0.04 vs 0.26 ± 0.06), AID (0.23 ± 0.05 vs0.19 ± 0.04), and Tfh cells (8.30 ± 2.00 vs 6.56 ± 1.59%) (P <0.05). Conclusion: Peyer’s summary Tfh cells may be involved in the pathogenesis of sepsis. Dex inhibits Peyer’s Tfh activation and protects Peyer’s microenvironment.