MicroRNA與異煙肼所致小鼠抗結(jié)核藥物性肝損傷關(guān)系的研究
發(fā)布時(shí)間：2018-08-29 19:20
【摘要】：目的采用INH誘導(dǎo)的小鼠抗結(jié)核藥物性肝損傷模型，觀察miR-122、miR-155及氧化損傷指標(biāo)的動(dòng)態(tài)變化，初步探討miR-122、miR-155及氧化應(yīng)激在抗結(jié)核藥物性肝損傷發(fā)生發(fā)展中的作用及關(guān)系，為抗結(jié)核藥物性肝損傷的早期診斷和發(fā)病機(jī)制研究提供實(shí)驗(yàn)依據(jù)。 方法1）急性毒性實(shí)驗(yàn)：（1）實(shí)驗(yàn)對(duì)象：昆明小鼠，對(duì)照組（10只）給予生理鹽水，實(shí)驗(yàn)組（70只）予以INH（180mg/kg）灌胃處理，實(shí)驗(yàn)組分別在給藥后的0.25、0.75、1.5、6、12、18、24h和對(duì)照組24h留取10只小鼠的血清和肝組織標(biāo)本。（2）實(shí)驗(yàn)方法：HE染色法，光學(xué)顯微鏡下觀察肝組織病理學(xué)變化；全自動(dòng)生化分析儀檢測(cè)血清ALT和AST水平；實(shí)時(shí)熒光定量PCR檢測(cè)肝組織中miR-122和miR-155表達(dá)。（3）統(tǒng)計(jì)分析：采用SPSS17.0數(shù)據(jù)分析，數(shù)據(jù)以x s表示，計(jì)量資料采用單因素方差法，相關(guān)性分析采用Pearman相關(guān)性分析，P0.05為差異有統(tǒng)計(jì)學(xué)意義。 2）慢性毒性實(shí)驗(yàn)：（1）實(shí)驗(yàn)對(duì)象：除實(shí)驗(yàn)組每天予以INH（90mg/kg）灌胃，其中實(shí)驗(yàn)組分別在連續(xù)給藥后的1D、3D、5D、1W、2W、3W、4W和對(duì)照組4W留取小鼠的血清和肝組織標(biāo)本，其他同急性毒性實(shí)驗(yàn)。（2）實(shí)驗(yàn)方法：組織病理學(xué)、ALT和AST檢測(cè)同急性毒性實(shí)驗(yàn)，，一般生化分析法檢測(cè)CuZnSOD和MDA的水平；酶聯(lián)免疫吸附吸附法檢測(cè)MT和MRPS11蛋白表達(dá)水平；miR-122表達(dá)水平檢測(cè)同急性毒性實(shí)驗(yàn)。（3）統(tǒng)計(jì)分析：同急性毒性實(shí)驗(yàn)。 結(jié)果1）急性毒性實(shí)驗(yàn)：（1）肝組織病理學(xué)：給藥后的0.25h，可見少量肝細(xì)胞腫脹，隨給藥時(shí)間的變化并逐漸加重。（2）ALT和AST：小鼠血清中ALT和AST在0.75h出現(xiàn)顯著性變化，且兩者均上升到最大值。（3）miR-122和miR-155：給藥后miR-122整體呈下降趨勢(shì)，0.25h即開始發(fā)生顯著性下降為（56.50±27.77）％，（P0.05）；miR-155呈上升趨勢(shì)，0.75h開始出現(xiàn)顯著性上升（11.25±1.43）倍。（4）相關(guān)性分析：miR-122/miR-155比值與ALT和AST均呈負(fù)相關(guān)性（r＝-0.356，r＝-0.246；P0.05），且與肝組織病理學(xué)評(píng)分有很好的負(fù)相關(guān)性（r＝-0.779，P0.01）。 2）慢性毒性實(shí)驗(yàn)：（1）肝組織病理學(xué)：連續(xù)給藥后的第3D，顯微鏡下可見少量肝細(xì)胞腫脹，胞漿稀疏。隨著用藥時(shí)間延長(zhǎng)，肝組織損傷程度逐漸加重。（2）ALT和AST：兩者分別在2W和3W出現(xiàn)顯著性變化。（3）CuZnSOD和MDA：與對(duì)照組相比5D發(fā)生了顯著性變化。（4）miR-122：給藥后整體呈下降趨勢(shì)，第3D表達(dá)開始下降（88.72±5.15）％（P0.05）。（5）MT和MRPS11：與對(duì)照組相比兩者均在第3D出現(xiàn)變化，且在2W分別下降到最小值（908.68±152.87）pg/ml和（9.72±1.31）pg/ml（P0.01）。（6）相關(guān)性分析：miR-122與CuZnSOD、MDA和MT存在相關(guān)性（r＝0.240，r＝-0.311，r＝0.415；均P0.05)；MRPS11與CuZnSOD，MDA和MT存在相關(guān)性（r＝0.313，r＝-0.250，r＝0.366；均P0.05）。 結(jié)論急/慢性毒性實(shí)驗(yàn)中，miRNA與ADLI存在相關(guān)性，miR-122低表達(dá)和miR-155高表達(dá)均與ADLI發(fā)生發(fā)展有關(guān)，miR-122/miR-155比值有望成為ADLI早期診斷標(biāo)志物，且miR-122可能通過調(diào)節(jié)MRPS11參與ADLI中的氧化應(yīng)激反應(yīng)，為ADLI發(fā)病機(jī)制研究提供了新的實(shí)驗(yàn)依據(jù)。
[Abstract]:Objective to observe the dynamic changes of miR-122,miR-155 and oxidative damage indexes in mice induced by INH, and to explore the role and relationship of miR-122,miR-155 and oxidative stress in the occurrence and development of liver injury induced by antituberculous drugs. To provide experimental evidence for the early diagnosis and pathogenesis of liver injury induced by antituberculous drugs. Methods 1) Acute toxicity test: (1) Experimental subjects: Kunming mice, control group (10 mice) were given normal saline, experimental group (70 mice) were treated by INH (180mg/kg) intragastric perfusion. The samples of serum and liver tissue of 10 mice were collected in the experimental group at 0.250.75 ~ 1.5U 1.5A ~ (12) ~ (12) ~ (18) h after administration for 24 h and in the control group at 24 h. (2) the pathological changes of liver tissue were observed by the method of: (2) the pathological changes of liver tissue were observed under optical microscope, and the levels of ALT and AST in serum were detected by automatic biochemical analyzer. Real-time fluorescence quantitative PCR was used to detect the expression of miR-122 and miR-155 in liver tissue. (3) Statistical analysis: SPSS17.0 data were analyzed, the data were expressed as x s, and the data were measured by single factor variance method. The correlation analysis using Pearman correlation analysis (P0.05) had statistical significance. 2) chronic toxicity experiment: (1) experiment object: except the experimental group was given INH (90mg/kg) daily, The serum and liver tissues of the experimental group were collected after continuous administration of 1DX 3DX 5DX 1W 2WN 3W 4W and the control group 4W respectively. (2) Experimental methods: the detection of alt and AST in histopathology was the same as acute toxicity test. The levels of CuZnSOD and MDA were detected by general biochemical analysis, and the expression levels of MT and MRPS11 protein by Elisa were detected in the same way as acute toxicity test. (3) Statistical analysis: same acute toxicity test. Results 1) Acute toxicity test: (1) liver histopathology: at 0.25 h after administration, a small amount of hepatocyte swelling was observed, which increased gradually with the time of administration. (2) ALT and AST in serum of ALT and AST: mice showed significant changes at 0.75 h. And both of them increased to the maximum. (3) after miR-122 and miR-155: administration, the whole miR-122 showed a downward trend of 0.25 h, which began to significantly decrease to (56.50 鹵27.77), (P0.05); (4) the correlation analysis showed that the ratio of 1: miR-122% miR-155 was negatively correlated with ALT and AST (r-0.356r-0.246). (2) chronic toxicity test: (1) liver histopathology: 3D after continuous administration, a small amount of liver cells were swollen and cytoplasm was sparse under microscope. (2) there were significant changes in ALT and AST: at 2W and 3W, respectively. (3) CuZnSOD and MDA: showed significant changes compared with the control group. (4) the whole level of miR-122: decreased. 3D expression began to decrease (88.72 鹵5.15)% (P0.05). (5) MT and MRPS11: showed changes in 3D compared with the control group, and decreased to the minimum value (908.68 鹵152.87) pg/ml and (9.72 鹵1.31) pg/ml (P0.01). (6) at 2W. There was a correlation between MRPS11 and CuZnSOD,MDA and MT (r = 0.313). Conclusion the low expression of miR-122 and the high expression of miR-155 in acute / chronic toxicity test are related to the occurrence and development of ADLI. The ratio of miR-122 / miR-155 may be the early diagnostic marker of ADLI, and miR-122 may participate in the oxidative stress reaction of ADLI by regulating MRPS11. It provides a new experimental basis for the study of the pathogenesis of ADLI.
【學(xué)位授予單位】：河北聯(lián)合大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R575
[Abstract]:Objective to observe the dynamic changes of miR-122,miR-155 and oxidative damage indexes in mice induced by INH, and to explore the role and relationship of miR-122,miR-155 and oxidative stress in the occurrence and development of liver injury induced by antituberculous drugs. To provide experimental evidence for the early diagnosis and pathogenesis of liver injury induced by antituberculous drugs. Methods 1) Acute toxicity test: (1) Experimental subjects: Kunming mice, control group (10 mice) were given normal saline, experimental group (70 mice) were treated by INH (180mg/kg) intragastric perfusion. The samples of serum and liver tissue of 10 mice were collected in the experimental group at 0.250.75 ~ 1.5U 1.5A ~ (12) ~ (12) ~ (18) h after administration for 24 h and in the control group at 24 h. (2) the pathological changes of liver tissue were observed by the method of: (2) the pathological changes of liver tissue were observed under optical microscope, and the levels of ALT and AST in serum were detected by automatic biochemical analyzer. Real-time fluorescence quantitative PCR was used to detect the expression of miR-122 and miR-155 in liver tissue. (3) Statistical analysis: SPSS17.0 data were analyzed, the data were expressed as x s, and the data were measured by single factor variance method. The correlation analysis using Pearman correlation analysis (P0.05) had statistical significance. 2) chronic toxicity experiment: (1) experiment object: except the experimental group was given INH (90mg/kg) daily, The serum and liver tissues of the experimental group were collected after continuous administration of 1DX 3DX 5DX 1W 2WN 3W 4W and the control group 4W respectively. (2) Experimental methods: the detection of alt and AST in histopathology was the same as acute toxicity test. The levels of CuZnSOD and MDA were detected by general biochemical analysis, and the expression levels of MT and MRPS11 protein by Elisa were detected in the same way as acute toxicity test. (3) Statistical analysis: same acute toxicity test. Results 1) Acute toxicity test: (1) liver histopathology: at 0.25 h after administration, a small amount of hepatocyte swelling was observed, which increased gradually with the time of administration. (2) ALT and AST in serum of ALT and AST: mice showed significant changes at 0.75 h. And both of them increased to the maximum. (3) after miR-122 and miR-155: administration, the whole miR-122 showed a downward trend of 0.25 h, which began to significantly decrease to (56.50 鹵27.77), (P0.05); (4) the correlation analysis showed that the ratio of 1: miR-122% miR-155 was negatively correlated with ALT and AST (r-0.356r-0.246). (2) chronic toxicity test: (1) liver histopathology: 3D after continuous administration, a small amount of liver cells were swollen and cytoplasm was sparse under microscope. (2) there were significant changes in ALT and AST: at 2W and 3W, respectively. (3) CuZnSOD and MDA: showed significant changes compared with the control group. (4) the whole level of miR-122: decreased. 3D expression began to decrease (88.72 鹵5.15)% (P0.05). (5) MT and MRPS11: showed changes in 3D compared with the control group, and decreased to the minimum value (908.68 鹵152.87) pg/ml and (9.72 鹵1.31) pg/ml (P0.01). (6) at 2W. There was a correlation between MRPS11 and CuZnSOD,MDA and MT (r = 0.313). Conclusion the low expression of miR-122 and the high expression of miR-155 in acute / chronic toxicity test are related to the occurrence and development of ADLI. The ratio of miR-122 / miR-155 may be the early diagnostic marker of ADLI, and miR-122 may participate in the oxidative stress reaction of ADLI by regulating MRPS11. It provides a new experimental basis for the study of the pathogenesis of ADLI.
【學(xué)位授予單位】：河北聯(lián)合大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R575
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