本文選題：EGCG + 肝缺血再灌注　； 參考：《廣西醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的：觀察表沒食子兒茶素沒食子酸酯[(-)-Epigallocatechin-3-gallate, EGCG]對Balb/c小鼠肝缺血再灌注損傷的保護(hù)性作用,研究其作用與血紅素氧合酶-1(Hemeoxygenase-1,HO-1)、Toll樣受體-4(Toll-like receptor4, TLR4)/核因子kappa B P65 (Nuclear fator-κB P65, NF-κB P65)及相關(guān)信號分子的關(guān)系,探討其作用機(jī)制。方法：健康雄性Balb/c小鼠18只,隨機(jī)分3組(n=6),分別為：假手術(shù)組(Sham組)、缺血再灌注組(I/R組)和I/R+EGCG處理組(EGCG組)。各組分別于再灌注末摘除眼球取血樣,處死小鼠、收集肝組織。測定血清中谷丙轉(zhuǎn)氨酶(ALT)、谷草轉(zhuǎn)氨酶(AST)水平；缺血再灌注肝組織蘇木素-伊紅(Hematoxylin-Eosine, HE)染色后光鏡下觀察其病理學(xué)改變；免疫組織化學(xué)測定TLR4、HO-1和NF-κB P65蛋白的表達(dá)量；Western Blot進(jìn)一步驗(yàn)證TLR4、HO-1和NF-κB P65蛋白水平表達(dá)的變化; Real time-PCR檢測EGCG作用后小鼠左外葉肝組織中TLR4、HO-1、IL-1β、IL-6及TNF-α等信號分子mRNA表達(dá)水平的變化。結(jié)果：經(jīng)EGCG預(yù)處理后,EGCG組血清ALT水平較I/R組均明顯降低[(823.00-±200.25)vs(1143.33±268.66),(715.33±159.15)vs(1295.00±214.39),P0.05],與Sham組比較,EGCG組血清AST、 ALT水平均顯著升高[(823.00±200.25)vs(349.00±39.24),(715.33±159.15)vs(205.00±28.35),P0.01]。從病理學(xué)上觀察,根據(jù)Suzuki評分標(biāo)準(zhǔn),EGCG組(6.50±1.05)顯著高于Sham組(0.43±0.38)(P0.01),I/R組(8.17±1.47)明顯高于EGCG組(6.50±1.05)(P0.05),EGCG組血再灌注肝組織損傷程度明顯小于IR組,但較Sham組顯著增大。依據(jù)Soslow評分標(biāo)準(zhǔn)對免疫組化結(jié)果進(jìn)行評分：Sham組、EGCG組、I/R組TLR4指標(biāo)得分分別為2.83±0.98分、5.67±1.51分、8.33±2.25分,Sham組、I/R組、EGCG組指標(biāo)得分分別為3.00±0.98分、6.67±1.97分、9.00±1.55分,Sham組、I/R組、EGCG組HO-1指標(biāo)得分分別為2.92±0.85分、6.56±1.44分、9.43±2.31分,與I/R組比較,EGCG組TLR4、NF-kB P65蛋白表達(dá)量明顯降低(P0.05),EGCG組HO-1蛋白表達(dá)量明顯升高(P0.05)；與Sham組比較,EGCG組TLR4、NF-kB P65蛋白表達(dá)量顯著增高(P0.01),EGCG組HO-1蛋白表達(dá)量明顯增高(P0.01)。以β-actin為內(nèi)參照,Western Blot結(jié)果示,EGCG組HO-1蛋白相對表達(dá)量明顯高于I/R組[(0.90±0.06)vs(0.71±0.06),P0.05],同時(shí)I/R組HO-1蛋白的相對表達(dá)量明顯高于Sham組[(0.71±0.06)vs(0.52±0.05),P0.05]；I/R組TLR4、NF-κB P65蛋白的相對表達(dá)量明顯高于EGCG組[(1.24±0.09)vs(0.98±0.07),(1.56±0.06)vs(1.27±0.06),P0.05],且EGCG組TLR4、NF-κB P65蛋白相對表達(dá)量明顯高于Sham組[(0.98±0.07)vs(0.60±0.07),(1.27±0.06)vs(0.59±0.05),P0.05]。Real time-PCR結(jié)果示,HO-1 mRNA的相對表達(dá)水平EGCG組(5.86±0.94)的較I/R組(3.32±0.823)明顯增高(P0.05),I/R組較Sham組(1.00±0.00)顯著增高(P0.01); TLR4、TNF-α、IL-6及IL-lb mRNA的相對表達(dá)水平I/R組[(5.86±0.78)、(4.43±1.39)、(5.06±1.43)、(6.53±1.62)]較EGCG組[(3.82±1.42)、(2.74±0.73)、(2.93±0.80)、(4.36±1.19)]明顯增高(P0.05),EGCG組較Sham組(1.00±0.00)顯著增高(P0.01)。結(jié)論：EGCG可通過抑制炎癥反應(yīng),減少肝組織缺血再灌注損傷,其機(jī)制：一方面可能與EGCG下調(diào)TLR4/NF-κB P65信號通路及通路下游TNF-α、IL-1β、IL-6等炎癥信號分子表達(dá)有關(guān),減少炎癥介質(zhì)介導(dǎo)的炎癥反應(yīng)對肝缺血再灌的損傷；另一方面其機(jī)制可能是EGCG抑制TLR4的表達(dá),從而上調(diào)HO-1的表達(dá),發(fā)揮對肝缺血再灌注損傷的保護(hù)作用。
[Abstract]:Objective: To observe the protective effect of epigallocatechin gallate (-) (-) -Epigallocatechin-3-gallate, EGCG] on hepatic ischemia reperfusion injury in Balb/c mice, and to study the role of epigallocatechin gallate (EGCG]) and the role of heme oxygenase (Hemeoxygenase-1, HO-1), Toll like receptor -4 (Toll-like receptor4, TLR4) / nuclear factor kappa. The relationship between kappa B P65) and the related signal molecules. Methods: 18 healthy male Balb/c mice were randomly divided into 3 groups (n=6), respectively: sham operation group (group Sham), ischemia reperfusion group (group I/R) and I/R+EGCG treatment group (EGCG group). Each group was removed from the end of reperfusion and removed the blood sample of the eyeball, killed the mice and collected the liver tissue. The serum was collected and serum was measured. The level of middle glutamic pyretransaminase (ALT) and glutamic oxaline aminotransferase (AST); the pathological changes were observed under the light microscopy of hematoxylin eosin (Hematoxylin-Eosine, HE) staining in ischemia reperfusion liver tissue; the expression of TLR4, HO-1 and NF- kappa B P65 protein was measured by immunohistochemistry; Western Blot further demonstrated the expression of TLR4, and the level of the protein level. Change; Real time-PCR detected the change of mRNA expression level of TLR4, HO-1, IL-1 beta, IL-6 and TNF- alpha in the left outer lobe liver of mice after EGCG action. Results: after the EGCG pretreatment, the serum ALT level of EGCG group was significantly lower than that of the Group [(1143.33 + 268.66), (715.33 + 159.15), 1295 + 214.39). In group am, serum AST and ALT levels in EGCG group were significantly higher [(823 + 200.25) vs (349 + 39.24), (715.33 + 159.15) vs (205 + 28.35), P0.01]. was observed from pathology. According to Suzuki scoring standard, EGCG group (6.50 + 1.05) was significantly higher than Sham group (0.43 + 0.38) (P0.01). I/R group was significantly higher than that of the group. The degree of hepatic tissue injury in blood reperfusion was significantly smaller than that of the IR group, but it was significantly higher than that in the Sham group. According to the Soslow score, the scores of the immunohistochemical results were 2.83 + 0.98, 5.67 + 1.51, 8.33 + 2.25, Sham, I/R, and EGCG groups were 3 + 0.98, 6.67 + 1.97, 9., respectively. The scores of HO-1 index in group Sham, group I/R and group EGCG were 2.92 + 0.85, 6.56 + 1.44 and 9.43 + 2.31 respectively. Compared with I/R group, EGCG group TLR4, NF-kB P65 protein expression decreased significantly (P0.05), EGCG group HO-1 protein expression increased significantly. The expression of HO-1 protein was significantly higher (P0.01). The results of Western Blot showed that the relative expression of HO-1 protein in group EGCG was significantly higher than that of I/R Group [(0.90 + 0.06) vs (0.71 + 0.06), P0.05], and the relative expression of HO-1 protein in I/R group was significantly higher than that of the Group [(0.71 + 0.06) (0.52 + 0.05). The relative expression was significantly higher than that in the EGCG Group [(1.24 + 0.09) vs (0.98 + 0.07), (1.56 + 0.06) vs (1.27 + 0.06), P0.05], and TLR4 in EGCG group, and the relative expression of NF- kappa B P65 protein was significantly higher than that of Sham Group [(0.98 + 0.07) vs (0.60 + 0.07). Compared with group I/R (3.32 + 0.823), group I/R was significantly higher than that of group Sham (1 + 0) (P0.01), TLR4, TNF- a, IL-6 and IL-lb mRNA were significantly higher than EGCG group (5.86 + 0.78), (4.43 + 1.39), (5.06 + 1.43), (6.53 + 1.62)]. Group (1 + 0) significantly increased (P0.01). Conclusion: EGCG can reduce the liver tissue ischemia reperfusion injury by inhibiting the inflammatory response, its mechanism may be related to EGCG down the TLR4/NF- kappa B P65 signaling pathway and the downstream TNF- a, IL-1 beta, IL-6 and other inflammatory signals, and reduce inflammatory mediators mediated inflammatory reaction to liver ischemia On the other hand, the mechanism may be that EGCG inhibits the expression of TLR4, thereby upregulates the expression of HO-1, and plays a protective role in hepatic ischemia-reperfusion injury.
【學(xué)位授予單位】：廣西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：R657.3

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相關(guān)期刊論文 前2條

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本文編號：2105652