LPS干預(yù)對軍團菌肺炎作用的實驗研究
發(fā)布時間：2018-11-15 17:44
【摘要】：目的 1.通過動物實驗探討LPS干預(yù)對TLR4表達的影響。 2.通過比較LPS干預(yù)后TLR4蛋白及其信號轉(zhuǎn)導(dǎo)通路中相關(guān)細胞因子的表達,結(jié)合病理切片的結(jié)果,探討LPS干預(yù)對軍團菌感染的影響。 方法 1.以C3H/HeN小鼠(6-8周齡)作為實驗動物,每組8只,雌雄各半。設(shè)立LPS干預(yù)染菌組、LPS未干預(yù)染菌組、.對照組三個實驗組。首先對LPS干預(yù)組小鼠利用大腸桿菌脂多糖,以1mg/kg的劑量,進行腹腔注射干預(yù),其他組腹腔注射生理鹽水。24h后與LPS未干預(yù)組一起氣管注射軍團菌懸液染菌,對照組給予等量的生理鹽水,分別于染菌后12h、24h、48h處死小鼠。 2.摘除眼球取血約1ml,采用密度梯度離心分離外周血單核細胞(peripheral blood mononuclear cel, PBMC),FCMS檢測各組中PBMC TLR4水平,比較三組各時間點TLR4蛋白表達水平的變化；并取左肺上葉約0.02g組織固定做病理切片。 3.取左肺中葉約0.02g組織勻漿,ELISA法檢測各組肺組織中TNF-a和IL-1β的含量。 結(jié)果 ①肺組織病理切片結(jié)果顯示：LPS干預(yù)染菌組、LPS未干預(yù)染菌組均出現(xiàn)中性粒細胞、嗜酸粒細胞、單核巨噬細胞等炎性細胞,且肺泡血管有出血現(xiàn)象。同時兩染菌組在24h可見肺組織炎性滲出液,對照組未出現(xiàn)炎癥反應(yīng),表明對照組未受到染菌組的污染。LPS干預(yù)染菌組肺泡血管出血較少,且炎性浸潤略輕于LPS未干預(yù)染菌組。經(jīng)析因分析LPS干預(yù)染菌組、LPS未干預(yù)染菌組和對照組之間肺臟臟器系數(shù)差異有統(tǒng)計學意義(F=5.699,P=0.006),各個時間點之間差異有統(tǒng)計學意義(F=10.648,P0.001),分組和時間點之間交互作用差異無統(tǒng)計學意義(F=0.668,P0.05)。經(jīng)LSD法兩兩比較,LPS干預(yù)染菌組和LPS未干預(yù)染菌組的臟器系數(shù)均高于對照組(P0.05),但LPS干預(yù)染菌組與LPS未干預(yù)染菌組間無統(tǒng)計學差異。 ②三組的TLR4蛋白水平表達有統(tǒng)計學差異(F=47.144,P0.001),各個時間點差異有統(tǒng)計學意義(F=141.543,P0.001),分組和時間點之間交互作用差異也有統(tǒng)計學意義(F=29.902,P0.001)。經(jīng)LSD法兩兩比較,LPS干預(yù)染菌組TLR4‘表達水平在12h、24h與對照組相比,差異有統(tǒng)計學意義,且均高于對照組(P0.05)；LPS未干預(yù)組TLR4表達水平在12h高于對照組(P0.05)。LPS干預(yù)染菌組與LPS未干預(yù)染菌組TLR4蛋白均在12h達到高峰,且LPS干預(yù)組TLR4水平高于LPS未干預(yù)組(P0.05),同時兩染菌組TLR4水平隨著染菌時間的延長而下降,在24h、48h兩染菌組TLR4表達水平差別無統(tǒng)計學意義。 ③三組間TNF-α濃度差異具有統(tǒng)計學意義(F=25.817,P0.001),時間的主效應(yīng)也具有統(tǒng)計學意義(F=8.3933,P0.001),分組和時間點之間未發(fā)現(xiàn)交互作用(F=I.460,P0.05)。經(jīng)兩兩比較,LPS干預(yù)染菌組與對照組未發(fā)現(xiàn)統(tǒng)計學差異,但LPS未干預(yù)染菌組與對照組、LPS干預(yù)染菌組之間的TNF-α濃度差異具有統(tǒng)計學意義,LPS未干預(yù)染菌組各個時間點TNF-α的分泌水平均高于LPS干預(yù)染菌組,兩染菌組TNF-α濃度均在感染24h時達到高峰后開始下降。三組間IL-1β濃度差異具有統(tǒng)計學意義(F=18.990,P0.001),時間的主效應(yīng)無統(tǒng)計學意義(F=I.323,P0.05),分組和時間點之間也未發(fā)現(xiàn)交互作用(F=0.382,P0.05)。LSD法兩兩比較后發(fā)現(xiàn),LPS干預(yù)染菌組與對照組的IL-1β濃度未發(fā)現(xiàn)統(tǒng)計學差異,但LPS未干預(yù)染菌組與對照組、LPS干預(yù)染菌組之間的IL-1β濃度差異具有統(tǒng)計學意義,LPS未干預(yù)染菌組各個時間點IL-1β的分泌水平均高于LPS干預(yù)染菌組。 結(jié)論 ①LPS干預(yù)可顯著調(diào)高小鼠PBMC TLR4表達水平,且具有短時效應(yīng)。 ②LPS未干預(yù)組染菌組TNF-α、IL-1β濃度均高于對照組,提示軍團菌感染后機體可能通過上調(diào)了TNF-α和IL-1β的表達水平以抵抗軍團菌感染。但LPS干預(yù)組染菌組TNF-α、IL-1β濃度均與對照組無明顯差別,說明LPS耐受可能會減輕軍團菌感染的炎性癥狀。
[Abstract]:Purpose 1. The expression of TLR4 by LPS intervention was discussed by animal experiment. Influence. 2. By comparing the expression of the related cytokines in the TLR4 protein and its signal transduction pathway after LPS intervention, the results of the pathological sections were combined to study the effect of LPS on Legionella. infection Effect of method 1. C3H/ HeN mice (6-8 weeks old) as an experimental activity In each group, 8 animals and half of each group were male and female. The LPS intervention group was established. and LPS did not interfere with the stain. In the control group, three experimental groups were divided into three groups: first, the mice of the LPS intervention group were treated with E. coli lipopolysaccharide, and the injection of the abdominal cavity was carried out at a dose of 1 mg/ kg, and the other groups were injected with normal saline in the abdominal cavity. After 24 h, the suspension of the Legionella pneumophila was injected with the LPS without intervention group, and the control group was given the same amount of physiological saline, respectively. after the staining of the bacteria for 12h, The expression of TLR4 in the peripheral blood mononuclear cells (PBMC) and peripheral blood mononuclear cells (PBMC) were measured by density gradient centrifugation, and the expression levels of TLR4 protein in each group were compared in three groups. 0. 02g of tissue was fixed to pathological section. 3. The lung of each group was detected by ELISA in the middle of the left lung. organization The contents of TNF-a and IL-1 were found in the rats. Results The results of the pathological sections of the lung tissue of the rats showed that the LPS intervention group and LPS did not interfere with the bacterial group, and the neutrophils and eosinophils were found. The inflammatory cells, such as single-core macrophages, and the blood vessels of the alveoli were bleeding. At the same time, the two groups of bacteria were found in the pulmonary tissue inflammatory exudation liquid at the same time, and the control group was used as the control group. No inflammatory reaction was observed in the group, indicating that the control group was not contaminated by the stain group. The LPS interfered with the pulmonary alveolar blood of the bacterial group. There was a significant difference in lung organ coefficient between LPS and control group (F = 5.699, P = 0. 006), and there was a significant difference between the time points (F = 10.648, P0.001), group and time. point-to-point interaction There was no significant difference (F = 0. 668, P0.05). Compared with the control group (P <0.05), the organ coefficients of the LPS-treated group and the LPS-free group were higher than those in the control group (P0.05). There was no significant difference in the level of TLR4 protein in the three groups (F = 47. 144, P 0.001), and the difference of time point was statistically significant (F = 141.543, P0.001), and the interaction between the group and time point was statistically significant (F = 141.543, P 0.001). The difference was also of statistical significance (F = 29. 902, P0. The expression level of TLR4 in LPS-treated group was significantly higher than that of control group (P <0.05). The expression of TLR4 in LPS-free group was higher than that of control group (P <0.05), and the level of TLR4 expression in LPS-free group was higher than that of control group (P0.05). The TLR4 protein in both group and LPS group reached the peak at 12h, and the level of TLR4 in LPS group was higher than that of LPS group (P0.05). There was no significant difference in the expression of TLR4 in the two groups. The difference between the three groups was statistically significant (F = 25. 817, P 0.001). The primary effect of time was also of statistical significance (F = 8.3933, P 0.001). There was no interaction between the group and the time point (F = I. 460, P0.05). There was a significant difference in the concentration of TNF-1 between the group and the LPS-treated group, and the levels of TNF-1 in the LPS-free group were higher than that of the LPS-treated group. The concentration of TNF-1 in the two groups was decreased after 24 h of infection. There was no significant difference in the concentration of IL-1 in the three groups (F = 18. 990, P. 001). The primary effect of time was not significant (F = I. 323, P0.05). There was no interaction between the group and the time point (F = 0.382, P0.05). After the LSD method, the levels of IL-1 in the LPS-treated group and the control group were not found to be statistically different, but the LPS did not interfere with the control group and the control group. There was no significant difference in the concentration of IL-1 in the group and LPS intervention group, and LPS did not interfere. bacteria The levels of IL-1 at all time points in the group were higher than that of the LPS-treated group. The expression level of TLR4 in the high-dose mice was significantly increased by LPS intervention and short-time effect. The concentration of TNF-1 and IL-1 in the group-infected cell group was higher than that of the control group. After the infection of Legionella, the expression level of TNF-1 and IL-1 was up-regulated in order to resist the infection of Legionella.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2013
【分類號】：R563.1
本文編號：2333996
[Abstract]:Purpose 1. The expression of TLR4 by LPS intervention was discussed by animal experiment. Influence. 2. By comparing the expression of the related cytokines in the TLR4 protein and its signal transduction pathway after LPS intervention, the results of the pathological sections were combined to study the effect of LPS on Legionella. infection Effect of method 1. C3H/ HeN mice (6-8 weeks old) as an experimental activity In each group, 8 animals and half of each group were male and female. The LPS intervention group was established. and LPS did not interfere with the stain. In the control group, three experimental groups were divided into three groups: first, the mice of the LPS intervention group were treated with E. coli lipopolysaccharide, and the injection of the abdominal cavity was carried out at a dose of 1 mg/ kg, and the other groups were injected with normal saline in the abdominal cavity. After 24 h, the suspension of the Legionella pneumophila was injected with the LPS without intervention group, and the control group was given the same amount of physiological saline, respectively. after the staining of the bacteria for 12h, The expression of TLR4 in the peripheral blood mononuclear cells (PBMC) and peripheral blood mononuclear cells (PBMC) were measured by density gradient centrifugation, and the expression levels of TLR4 protein in each group were compared in three groups. 0. 02g of tissue was fixed to pathological section. 3. The lung of each group was detected by ELISA in the middle of the left lung. organization The contents of TNF-a and IL-1 were found in the rats. Results The results of the pathological sections of the lung tissue of the rats showed that the LPS intervention group and LPS did not interfere with the bacterial group, and the neutrophils and eosinophils were found. The inflammatory cells, such as single-core macrophages, and the blood vessels of the alveoli were bleeding. At the same time, the two groups of bacteria were found in the pulmonary tissue inflammatory exudation liquid at the same time, and the control group was used as the control group. No inflammatory reaction was observed in the group, indicating that the control group was not contaminated by the stain group. The LPS interfered with the pulmonary alveolar blood of the bacterial group. There was a significant difference in lung organ coefficient between LPS and control group (F = 5.699, P = 0. 006), and there was a significant difference between the time points (F = 10.648, P0.001), group and time. point-to-point interaction There was no significant difference (F = 0. 668, P0.05). Compared with the control group (P <0.05), the organ coefficients of the LPS-treated group and the LPS-free group were higher than those in the control group (P0.05). There was no significant difference in the level of TLR4 protein in the three groups (F = 47. 144, P 0.001), and the difference of time point was statistically significant (F = 141.543, P0.001), and the interaction between the group and time point was statistically significant (F = 141.543, P 0.001). The difference was also of statistical significance (F = 29. 902, P0. The expression level of TLR4 in LPS-treated group was significantly higher than that of control group (P <0.05). The expression of TLR4 in LPS-free group was higher than that of control group (P <0.05), and the level of TLR4 expression in LPS-free group was higher than that of control group (P0.05). The TLR4 protein in both group and LPS group reached the peak at 12h, and the level of TLR4 in LPS group was higher than that of LPS group (P0.05). There was no significant difference in the expression of TLR4 in the two groups. The difference between the three groups was statistically significant (F = 25. 817, P 0.001). The primary effect of time was also of statistical significance (F = 8.3933, P 0.001). There was no interaction between the group and the time point (F = I. 460, P0.05). There was a significant difference in the concentration of TNF-1 between the group and the LPS-treated group, and the levels of TNF-1 in the LPS-free group were higher than that of the LPS-treated group. The concentration of TNF-1 in the two groups was decreased after 24 h of infection. There was no significant difference in the concentration of IL-1 in the three groups (F = 18. 990, P. 001). The primary effect of time was not significant (F = I. 323, P0.05). There was no interaction between the group and the time point (F = 0.382, P0.05). After the LSD method, the levels of IL-1 in the LPS-treated group and the control group were not found to be statistically different, but the LPS did not interfere with the control group and the control group. There was no significant difference in the concentration of IL-1 in the group and LPS intervention group, and LPS did not interfere. bacteria The levels of IL-1 at all time points in the group were higher than that of the LPS-treated group. The expression level of TLR4 in the high-dose mice was significantly increased by LPS intervention and short-time effect. The concentration of TNF-1 and IL-1 in the group-infected cell group was higher than that of the control group. After the infection of Legionella, the expression level of TNF-1 and IL-1 was up-regulated in order to resist the infection of Legionella.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2013
【分類號】：R563.1
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