目的 探讨高迁移率族蛋白B1（high modility groupbox 1，HMGB1）抑制剂对脓毒症T淋巴细胞和单核细胞的作用机制。 方法 将30只C57BL/6雄性小鼠随机分为3组，每组10只，依次为假手术组、模型组、抑制剂组。模型组、抑制剂组采用盲肠结扎穿孔手术构建脓毒症小鼠模型，假手术组小鼠仅暴露盲肠后即进行伤口缝合，不结扎。抑制剂组小鼠术后腹腔注射HMGB1特异性抑制剂甘草酸（10 mg/kg），每6 h注射1次，连续4次，假手术组和模型组腹腔注射等剂量的生理盐水。处死小鼠后，无菌分离小鼠的胸腺组织，常规提取胸腺中的T淋巴细胞和单核细胞。MTT法与流式细胞术测定T淋巴细胞的增殖活性和凋亡情况，Transwell趋化实验与ELISA法测定单核细胞的趋化活性以及分泌炎性因子TNF-α、IL-6及IL-10的情况，Western blot检测胸腺组织中HMGB1和第10号染色体同源丢失性磷酸酶-张力蛋白（phosphatase and tensin homolog deleted onchromosometen，PTEN）的表达水平。 结果 与假手术组相比，模型组小鼠胸腺组织T淋巴细胞的增殖活性和凋亡率、单核细胞的趋化活性和TNF-α、IL-6、IL-10蛋白的表达、HMGB1和PTEN的表达明显升高（均P<0.05）；与模型组相比，抑制剂组小鼠胸腺组织T淋巴细胞凋亡率、TNF-α、IL-6蛋白的表达、HMGB1和PTEN的表达明显降低，T淋巴细胞的增殖活性、单核细胞的趋化活性、IL-10蛋白的表达明显升高（均P<0.05）。 结论 HMGB1抑制剂可降低脓毒症小鼠胸腺T细胞的凋亡率，增强T细胞的增殖活性和单核细胞的趋化活性，提高单核细胞抗炎因子IL-10的分泌，抑制促炎因子TNF-α、IL-6的分泌。
Abstract
Objective To investigate the mechanism of the high mobility group box protein 1 (HMGB1) inhibitor on T lymphocytes and monocytes in sepsis. Methods Thirty C57BL/6 male mice were randomly divided into 3 groups, 10 in each group, followed by a sham operation group, a model group, and an inhibitor group. In the model group and inhibitor group, a mouse model of sepsis was replicated by cecal ligation and puncture (CLP). In the sham operation group, the wound was sutured only after exposure to the cecum, and no ligation was performed. The mice in the inhibitor group were intraperitoneally injected with glycyrrhizic acid, a specific inhibitor of HMGB1 (10 mg/kg), and injected once every 6 hours for 4 times. The mice in the sham operation group and the model group were intraperitoneally injected with the same amount of normal saline. After the mice were sacrificed, the thymus tissue of the mice was aseptically isolated, and the T lymphocytes and monocytes in the thymus were routinely extracted. MTT assay and flow cytometry were used to detect the proliferation activity and apoptosis of T lymphocyte. Transwell chemotaxis assay and ELISIA method were used to detect the chemotactic activity of monocytes and the expression levels of TNF-α, IL-6 and IL-10. Western blot was used to detect the expression levels of HMGB1 and phosphatase and tensin homolog deleted onchromosometen (PTEN) in each group. Results Compared with the sham operation group, the proliferation activity and the apoptosis rate of T lymphocytes in the thymus tissue of the model group, the chemotactic activity of monocytes and the expression levels of TNF-α, IL-6, IL-10 and the expression of HMGB1 and PTEN significantly increased (all P<0.05). Compared with the model group, the apoptosis rate of T lymphocytes, the expression levels of TNF-α, IL-6, HMGB1 and PTEN significantly reduced in inhibitor group mice, the proliferation activity of T lymphocytes, the chemotactic activity of monocytes, and the expression of IL-10 protein significantly increased (all P<0.05). Conclusions HMGB1 inhibitor can reduce the apoptosis rate of thymic T cells in septic mice, enhance the proliferation activity of T cells and the chemotactic activity of monocytes, increase the secretion of monocyte anti-inflammatory factor IL-10, and inhibit the pro-inflammatory factor TNF -α, IL-6 secretion.
关键词
高迁移率族蛋白B1； /
/
第10号染色体同源丢失性磷酸酶-张力蛋白； /
/
脓毒症； /
/
T淋巴细胞； /
/
单核细胞； /
/
小鼠
Key words
High mobility group protein B1; /
Phosphatase and tensin homolog deleted onchromosometen; /
/
Sepsis; /
/
T lymphocyte; /
/
Monocyte; /
/
Mouse
{{custom_sec.title}}
{{custom_sec.title}}
{{custom_sec.content}}
参考文献
[1] 冯莹, 刘伯毅, 方志成, 等. 内源性雌激素调控脓毒症小鼠心肌L型钙离子通道α1C亚基的表达[J]. 中国临床解剖学杂志, 2019, 37(5): 523-527. DOI: 10.13418/j.issn.1001-165x.2019.05.009.
[2] 胡彩珍, 叶立刚, 徐善祥. T细胞亚群监测在基于Sepsis-3的创伤性脓毒症患者中的应用[J]. 中华急诊医学杂志, 2019, 28(2): 181-184. DOI: 10.3760/cma.j.issn.1671-0282.2019.02.010.
[3] Meyer N, Harhay MO, Small DS, et al. Temporal trends in incidence, sepsis-related Mortality, and hospital-based acute care after sepsis[J]. Crit Care Med, 2018, 46(3): 354-360. DOI: 10.1097/CCM.0000000 000002872.
[4] Watanabe E, Thampy LK, Hotchkiss RS. Immunoadjuvant therapy in sepsis: novel strategies for immunosuppressive sepsis coming down the pike[J]. Acute Med Surg, 2018, 5(4): 309-315. DOI: 10.1002/ams2.363.
[5] 梁伟智, 陈灿, 李理, 等. 免疫功能对脓毒症患者预后的影响[J]. 中华危重病急救医学, 2018, 30(12): 1128-1131.
[6] Yao Y, Deng H, Li PF, et al. α-Lactose improves the survival of septic mice by blockade of TIM-3 signaling to prevent NKT cell apoptosis and attenuate cytokine storm[J]. Shock, 2017, 47(3): 337-345. DOI: 10.1097/SHK.0000000000000717.
[7] 许志平, 马红玲, 陈双峰, 等. MCP-1和mHLA-DR检测对脓毒症患者病情危重程度及预后评估的临床意义[J]. 中华重症医学电子杂志, 2019, 5(3): 225-229. DOI: 10.3877/cma.j.issn.2096-1537.2019.03.004.
[8] Ong SM, Hadadi E, Dang TM, et al. The pro-inflammatory phenotype of the human non-classical monocyte subset is attributed to senescence[J]. Cell Death Dis, 2018, 9(3): 266-278. DOI: 10.1038/s41419-018-0327-1.
[9] Wei SW, Gao YG, Dai XG, et al. SIRT1-mediated HMGB1 deacetylation suppresses sepsis-associated acute kidney injury[J]. Am J Physiol Renal Physiol, 2019, 316(1): F20-F31. DOI: 10.1152/ajprenal.00119.2018.
[10]周仪华, 袁影, 盛志勇, 等. HO-1对脓毒症大鼠血浆中HMGB-1表达的影响及其心肌保护机制[J]. 重庆医学, 2019, 48(5): 748-751. DOI: 10.3969/j.issn.1671-8348.2019.05.006.
[11]Zhou M, Fang HS, Du M, et al. The modulation of regulatory T cells via HMGB1/PTEN/β-Catenin axis in LPS induced acute lung injury[J]. Front Immunol, 2019, 10(6): 1612-1626. DOI: 10.3389/fimmu. 2019. 01612.
[12]Piliponsky AM, Shubin NJ, Lahiri AK, et al. Basophil-derived tumor necrosis factor can enhance survival in a sepsis model in mice[J]. Nat Immunol, 2019, 20(2): 129-140. DOI: 10.1038/s41590-018-0288-7.
[13]樊晓光, 彭锦, 洪树坤, 等. 虫草素联合谷氨酰胺对LPS诱导的脓毒症大鼠炎症失衡及肝肺病理变化的影响[J]. 中国临床解剖学杂志, 2019, 37(6): 638-643. DOI: 10.13418/j.issn.1001-165x.2019.06.007.
[14]Jensen IJ, Sjaastad FV, Griffith TS, et al. Sepsis-induced T cell immunoparalysis: the ins and outs of impaired T cell immunity[J]. J Immunol, 2018, 200(5): 1543-1553. DOI: 10.4049/jimmunol.1701618.
[15]任超, 李秀花, 吴瑶, 等. 迷走神经对脓毒症大鼠多器官功能及T淋巴细胞免疫反应的影响[J]. 中华烧伤杂志, 2018, 34(11): 815-820. DOI: 10.3760/cma.j.issn.1009-2587.2018.11.018.
[16]孔令宇, 席錾, 马文婷, 等. 连续性肾替代治疗对脓毒症患者CD8+T淋巴细胞功能的影响[J]. 中国感染控制杂志, 2018, 17(11): 974-978. DOI: 10.3969/j.issn.1671-9638.2018.11.006.
[17]Moghadasian MH, Zhao RZ, Ghazawwi N, et al. Inhibitory effects of North American wild rice on monocyte adhesion and inflammatory modulators in low-density lipoprotein receptor-knockout mice[J]. J Agric Food Chem, 2017, 65(41): 9054-9060. DOI: 10.1021/acs.jafc.7b03216.
[18]Xia Q, Wei L, Zhang YT, et al. Immune checkpoint receptors Tim-3 and PD-1 regulate monocyte and T lymphocyte function in septic patients[J]. Mediators Inflamm, 2018, 2018(11): 1632902-1632910. DOI: 10.1155/2018/1632902.
[19]Ferreira MNV, Brunialti MKC, Santos SS, et al. Immunophenotyping of monocytes during human sepsis shows impairment in antigen presentation: a shift toward nonclassical differentiation and upregulation of FcγRi-receptor[J]. Shock, 2018, 50(3): 293-300. DOI: 10.1097/SHK.0000000000001078.
[20]Zhou M, Zhang YD, Chen XL, et al. PTEN–Foxo1 signaling triggers HMGB1-mediated innate immune responses in acute lung injury[J]. Immunol Res, 2015, 62(1): 95-105. DOI: 10.1007/s12026-015-8639-z.
[21]Deng M, Tang Y, Li W, et al. The endotoxin delivery protein HMGB1 mediates caspase-11-dependent lethality in sepsis[J]. Immunity, 2018, 49(4): 740-753. DOI: 10.1016/j.immuni.2018.08.016.
[22]Wang ZH, Hou L, Yang H, et al. Electroacupuncture pretreatment attenuates acute lung injury through α7 nicotinic acetylcholine receptor-mediated inhibition of HMGB1release in rats after cardiopulmonary bypass[J]. Shock, 2018, 50(3): 351-359. DOI: 10.1097/SHK.0000000000001050.
[23]Bandala-Sanchez E, Bediaga NG, Goddard-Borger ED, et al. CD52 glycan binds the proinflammatory B box of HMGB1 to engage the Siglec-10 receptor and suppress human T cell function[J]. Proc Natl Acad Sci U S A, 2018, 115(30): 7783-7788. DOI: 10.1073/pnas.1722056115.
[24]Ha SD, Cho W, Kim SO. HDAC8 prevents anthrax lethal toxin-induced cell cycle arrest through silencing PTEN in human monocytic THP-1 Cells[J]. Toxins (Basel), 2017, 9(5): 162-176. DOI: 10.3390/toxins905 0162.
PDF(4553 KB)
京ICP备08002354号-3
