本文選題：18β-甘草次酸 + 金黃色葡萄球菌　； 參考：《浙江理工大學(xué)》2017年碩士論文

【摘要】：目的:建立小鼠金黃色葡萄球菌(SA)肺炎動(dòng)物模型,研究18β-甘草次酸(18β-GA)對(duì)SA肺炎的保護(hù)作用,并進(jìn)一步探討其分子機(jī)制。方法:6-8周齡C57/BL6雌鼠隨機(jī)分為3組,對(duì)照組:小鼠氣道滴注磷酸鹽緩沖液(PBS);SA模型組:小鼠氣道滴注SA(1×108 CFU);18β-GA干預(yù)組:小鼠在SA氣道滴注前2小時(shí),予腹腔注射18β-GA(50mg/kg)。觀察3組小鼠生存率變化,檢測(cè)各組小鼠肺組織和肝組織病理改變,肺泡灌洗液(BALF)中性粒細(xì)胞計(jì)數(shù),用ELISA檢測(cè)BALF中炎癥因子表達(dá),RT-q PCR測(cè)定肺組織炎癥因子表達(dá),CFU測(cè)定肺組織細(xì)菌負(fù)荷,Western Blot測(cè)定肺組織高遷移率蛋白1(HMGB1)表達(dá)。另一組實(shí)驗(yàn)觀察HMGB1抗體對(duì)SA肺炎小鼠生存率的影響。用18β-GA(20μM)預(yù)處理巨噬細(xì)胞RAW264.7和肝細(xì)胞Hep G2,再加入SA刺激。ELISA檢測(cè)巨噬細(xì)胞RAW264.7上清液中炎癥因子的表達(dá),Western Blot檢測(cè)P38/p-P38MAPK,JNK/p-JNK MAPK,ERK/p-ERK MAPK和P65/p-P65 NF-κB相關(guān)信號(hào)蛋白的表達(dá)。檢測(cè)肝細(xì)胞Hep G2上清的谷丙轉(zhuǎn)氨酶(ALT)和谷草轉(zhuǎn)氨酶(AST)的活性。結(jié)果:體內(nèi)實(shí)驗(yàn)發(fā)現(xiàn):預(yù)先腹腔注射18β-GA對(duì)SA肺炎及繼發(fā)性肝損傷具有保護(hù)作用。與SA模型組比較,18β-GA干預(yù)可以減輕SA引起的肺組織和肝組織病理?yè)p傷,18β-GA預(yù)處理組小鼠BALF中性粒細(xì)胞計(jì)數(shù)顯著降低(P0.05);炎癥因子(腫瘤壞死因子TNF-α,白細(xì)胞介素IL-1β,IL-6)下調(diào)(P0.05),肺組織細(xì)菌載量降低(P0.05)。18β-GA可顯著抑制HMGB1在感染小鼠肺組織中的表達(dá),腹腔注射HMGB1抗體可以顯著提高SA肺炎小鼠的生存率(P0.05)。體外實(shí)驗(yàn)提示:18β-GA通過(guò)抑制巨噬細(xì)胞P65 NF-κB蛋白磷酸化,顯著降低炎癥因子表達(dá)(P0.05),同時(shí)可以直接抑制SA生長(zhǎng);18β-GA也可降低SA誘導(dǎo)的肝細(xì)胞ALT表達(dá)(P0.05)。結(jié)論:18β-GA對(duì)SA肺炎和繼發(fā)性肝損傷有保護(hù)作用。18β-GA通過(guò)減少炎癥介質(zhì)產(chǎn)生和降低損傷模式分子HMGB1表達(dá),發(fā)揮保護(hù)作用。
[Abstract]:Objective: to establish an animal model of SAA pneumonia in mice, to study the protective effect of 18 尾 -glycyrrhetinic acid (18 尾 -GAA) on SA pneumonia and to explore its molecular mechanism. Methods C57/BL6 female mice aged 6-8 weeks were randomly divided into three groups. The control group was treated with SA(1 脳 108CFU 18 尾 -GA. The mice were injected intraperitoneally with 18 尾 -GAN 50 mg / kg before the infusion of SA. The survival rate of the three groups was observed, the pathological changes of lung and liver tissues and the neutrophil count in alveolar lavage fluid (BALF) of each group were detected. Expression of inflammatory factors in BALF was detected by ELISA RT-Q PCR. CFU was used to detect the expression of inflammatory factors in lung tissue. Western Blot was used to detect the expression of high mobility protein 1 (HMGB1) in lung tissue. In another group, the effect of HMGB1 antibody on survival rate of SA pneumonia mice was observed. Macrophages RAW264.7 and hepatocyte Hep G2 were pretreated with 18 尾 -GAN 20 渭 M, and then SA stimulated. Elisa was added to detect the expression of inflammatory factors in the supernatant of macrophage RAW264.7. Western Blot was used to detect the expression of P38 / p-P38 MAPK / JNKP / p-JNK MAPK / ERKP / ERK MAPK and P65/p-P65 NF- 魏 B related signal proteins. The activities of alanine aminotransferase (alt) and aspartate aminotransferase (AST) in the supernatant of Hep G2 were measured. Results: in vivo, 18 尾-GA had protective effect on SA pneumonia and secondary liver injury. Compared with the SA model group, the intervention of TNF- 18 尾 -GA could attenuate the pathological injury of lung and liver tissue induced by SA. The neutrophil count of BALF in the preconditioning group of 18 尾 -GA significantly decreased P0.05, and the inflammatory factor (TNF- 偽, interleukin IL-1 尾 -IL-6) down-regulated P0.05G. The decrease of bacterial load in lung tissue could significantly inhibit the expression of HMGB1 in lung tissue of infected mice. Intraperitoneal injection of HMGB1 antibody could significantly improve the survival rate of SA pneumonia mice (P 0.05). The in vitro experiments showed that the expression of P0.05A was significantly decreased by inhibiting the phosphorylation of p65 NF- 魏 B protein in macrophages, and that the growth of SA was inhibited directly by 18 尾 -GA, and the expression of ALT in hepatocytes induced by SA was also decreased. Conclusion.18 尾 -GA has a protective effect on SA pneumonia and secondary liver injury. 18 尾 -GA plays a protective role by reducing the production of inflammatory mediators and reducing the expression of HMGB1.
【學(xué)位授予單位】：浙江理工大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R563.1

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本文編號(hào)：1950661