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[Complement 5a regulates the function of dendritic cells to induce pathogenic polarization of regulatory T cell/helper T cell 17 in sepsis].

Zhonghua Wei Zhong Bing Ji Jiu Yi Xue. 2021 Jan;33(1):17-22. doi:10.3760/cma.j.cn121430-20201102-00696
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摘要


OBJECTIVE:To explore the mechanism of complement 5a (C5a) in the pathogenesis of sepsis. METHODS:SPF male C57BL/6J mice were selected and divided into sham operation group (Sham group), cecal ligation and puncture (CLP) group and CLP+anti-C5A monoclonal antibody intervention group (CLP+anti-C5a group) according to random number table with 20 mice in each group. A CLP model was reproduced to induce sepsis, and those in the Sham group only underwent laparotomy without ligation and perforation. In the CLP+anti-C5a group, 0.15 mg of anti-C5a monoclonal antibody was injected intraperitoneally immediately after CLP, and in the Sham group and CLP group were given equal amount of normal saline. The cumulative survival rate was analyzed by Kaplan-Meier method. Serum levels of tumor necrosis factor-α (TNF-α), interleukins (IL-12, IL-4), and interferon-γ (IFN-γ) were measured 24, 48 and 72 hours after operation by enzyme linked immunosorbent assay (ELISA). Immunohistochemical staining was used to observe the expression of C5a receptor (C5aR) in lung and kidney tissues 48 hours after operation. The proportions of dendritic cell (DC), regulatory T cell (Treg) and helper T cell 17 (Th17) in splenic mononuclear cells 48 hours after operation were analyzed by flow cytometry. RESULTS:The 7-day cumulative survival rate of mice in the CLP group was significantly lower than that in the Sham group (30.00% vs. 100.00%; Log-Rank test: χ2 = 47.470, P < 0.001), and the peripheral blood inflammatory mediators TNF-α, IL-12 and IL-4 were increased 24 hours after operation, followed by a significant decreasing at 48 hours, and then gradually increased at 72 hours. IFN-γ gradually increased 24 hours after operation and lasted for 72 hours. Immunohistochemistry showed that a large number of C5aR was expressed in pulmonary and renal endothelial cells 48 hours after operation in the CLP group. Compared with the Sham group, the proportion of DC [(1.80±0.30)% vs. (6.90±1.20)%, P < 0.05] and Treg [(0.38±0.02)% vs. (4.00±0.50)%, P < 0.05] in splenic mononuclear cells was down-regulated in the CLP group, the proportion of Th17 was up-regulated [(0.83±0.08)% vs. (0.32±0.03)%, P < 0.05], and disorder of immune function was found. After anti-C5A monoclonal antibody intervention, the 7-day cumulative survival rate increased significantly compared with the CLP group (54.54% vs. 30.00%; Log-Rank test: χ2 = 28.090, P < 0.001); TNF-α, IL-12 and IFN-γ were further increased, while IL-4 was significantly decreased; the expression of C5aR in lung and kidney tissues were significantly decreased, and the expression of mature DC cells [(5.10±1.20)% vs. (1.80±0.30)%, P < 0.05] and Treg [(2.58±0.05)% vs. (0.38±0.02)%, P < 0.05] in spleen were significantly increased compared with the CLP group, and Th17 was significantly decreased [(0.54±0.05)% vs. (0.83±0.08)%, P < 0.05]. CONCLUSIONS:It is preliminarily concluded that anti-C5A monoclonal antibody may improve the prognosis of sepsis by improving the polarization of mature DC and T cells in the spleen, and C5a plays an important role in the immune regulation of sepsis cells.

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