| Objective:By using cecal ligation and puncture (CLP), to build an acute lung injury (ALI)model, test the transcribing of Nrf-2, the protein expression levels and the transcribinglevels of superoxide dismutase(SOD), heme oxygenase1(HO-1) and catalase(CAT) byintervening with nuclear factor-erythroid2-related factor (Nrf-2)’agonist sulforaphane(SFN) and Nrf-2’ inhibitor all-trans retinoic acid (ATRA), and research on the protectiveeffect of Nrf-2signal pathway on the rats’ ALI.Methods:A total of128SPF healthy adult male Sprage-Dawley rats were used and randomlydivided them into four groups:SHAM group (32rats), CLP group (32rats), CLP+SFNgroup (32rats) and CLP+ATRA group (32rats). Each group was also randomly dividedinto four subgroups (8rats) by observing hours of8h,16h and24h and the survivalconditions. The SHAM group received the intraperitoneal injection of0.5ml/100g corn oil,was opened on the abdomen and then was closed after the caecum was found. The CLPgroup received the same amount of corn oil of intraperitoneal injection. Then the abdomenwas opened, the caecum was found, the ligation at50%of total length in caecum wasmade with the remote vertex of the caecum as the starting point, pierced two holes at themidpoint and the1/4part from the vertex point of the side of the caecum ligation in themesenterium with a20G needle, squeezed a small amount of intestinal contents and thenthe abdomen was closed. The CS and CA received the intraperitoneal injection of SFN（5mg/kg） andATRA（1mg/kg）30minutes before modeling. Then the rats were killedat corresponding time points by hemorrhagic shock method (bloodletting from theabdominal aorta), and their bilateral lung tissues were removed. The left two lobipulmonis was tested of the wet-dry ratio value(W/D), the right lobi inferior was testedusing HE staining and immunohistochemistry, the lobi medius pulmonis was tested ofNrf-2protein level using the Western-blotting method, and the total RNA of the lobisuperior was extracted for testing the level of Nrf-2,SOD1,HO-1and CAT mRNA by theRT-PCR method. Results:(1) The Appearance of lung tissues and the pathology by HE staining: in the SHAMgroup, the appearance of lung tissues was normal, the edge structures of the bronchus, thealveolus and the alveolus structure were clear and no clear pathology was found. In theCLP, CS and CA groups, the appearance and the pathology of lung tissues had varyingdegrees of damage compared with the SHAM group. At each experiment time point, thelung damage in the CS group was smaller than the CLP group, and the lung damage in theCA group was bigger than that of the CLP group.(2) The wet-dry ratio value of the left lung tissues: The W/D value of the CLP groupwas higher than that of the SHAM group. There was a significant difference（P<0.05）atthe experimental time points of8h,16h and24h. There was a significant difference inW/D value between the CS groupand SHAM group（P<0.05）at the experimental timepoints of16h. At each experimental time point, the W/D value of the CS group wassmaller than that of the CLP group （P<0.01，P<0.01，P<0.05）. There was a significantdifference in W/D value beween the CA group and the SHAM group（P<0.01）at theexperimental time points of8h and16h. There was a significant difference in W/D valuebetween the CA group, CLP group and CS group（P＜0.01，P＜0.01）at the experimentaltime points of8h and16h.(3) The semi-quantitative analysis of Nrf-2by IHC-P: The expression of Nrf-2of thelung tissues in the SHAM group was less. The expressions of a small part of the bronchialepithelial cells and alveolar epithelial cells were not obvious and showed as faint yellowgranular. They were weakly positive expression, most of which were cytoplasmic stainingand no nuclear staining. The expression of Nrf-2of the CLP group was stronger than thatof the SHAM group. There were expressions in bronchial epithelial cells, inflammationinfiltrated cells and alveolar epithelial cells, among which the alveolar epithelial cellsexpressed strongest, most as nucleus expression, showed as brown, were strong positiveexpression and declined with the time points compared within the group. The condition ofthe Nrf-2expression in the lung tissues of the group CA was as same as that of the SHAMgroup.The positive coefficient of Nrf-2in the lung tissues: The positive coefficient of theCLP group was higher than that of the SHAM group. There was a significant difference（P<0.05，P＜0.01，P＜0.01）at the experimental time points of8h,16h and24h. Thepositive coefficient of the CS group was higher than that of the SHAM group. There was a significant difference（P<0.01，P<0.01，P<0.01）at the experimental time points of8h,16hand24h. The positive coefficient of the CS group was higher than that of the CLP group.There was a significant difference （P<0.01，P<0.05，P<0.01）at the experimental timepoints of8h,16h and24h. The positive coefficient of the CA group was higher than that ofthe SHAM group. There was a significant difference （P<0.01，P<0.01，P<0.01）at theexperimental time points of8h,16h and24h. The positive coefficient of the CA group washigher than that of the CLP group and the CS group. There was a significant difference（P<0.01）at the experimental time points of8h,16h and24h.(4) The expressions of Nrf-2,SOD1,CAT and HO-1mRNA:The Nrf-2mRNA expressions: The Nrf-2mRNA expressions of the CLP group washigher than that of the SHAM group at the experimental time points of8h,16h and lowerthan that of the SHAM group at the experimental time points of24h. The result hadstatistic difference （P<0.05，P＜0.01，P＜0.01）. The Nrf-2mRNA expressions of theCS group was higher than that of the SHAM group at the experimental time points of8h,16h and24h. The result had significant statistic difference （P＜0.01）. The Nrf-2mRNAexpressions of the CS group was higher than that of the CLP group at the experimentaltime points of8h,16h and24h. The result had statistic difference （P<0.05，P<0.01，P<0.01）.The Nrf-2mRNA expressions of the CA group had statistic difference with theSHAM group at the experimental time points of16h and24h（P＜0.05，P＜0.01）. TheNrf-2mRNA expressions of the CA group was lower than that of the CLP group at theexperimental time points of8h. The result had statistic difference （P<0.05）. The Nrf-2mRNA expressions of the CA group was lower than that of the CS group at theexperimental time points of8h,16h and24h. The result had significant statistic difference（P＜0.01）.The SOD1mRNA expressions: The SOD1mRNA expressions of the CLP group washigher than that of the SHAM group at the experimental time points of8h,16h and lowerthan that of the SHAM group at the experimental time points of24h. The result hadstatistic difference （P<0.05，P＜0.01，P＜0.01）. The SOD1mRNA expressions of theCS group was higher than that of the SHAM group at the experimental time points of8h,16h and24h. The result had significant statistic difference （P＜0.01）. The SOD1mRNAexpressions of the CS group was higher than that of the CLP group at the experimentaltime points of8h,16h and24h. The result had statistic difference （P<0.05，P<0.01，P<0.01）.The SOD1mRNA expressions of the CA group had statistic difference with the SHAM group at the experimental time points of16h and24h（P＜0.01，P＜0.01）. TheSOD1mRNA expressions of the CA group was lower than that of the CLP group at theexperimental time points of8h. The result had statistic difference （P<0.01）. The SOD1mRNA expressions of the CA group was lower than that of the CS group at theexperimental time points of8h,16h and24h. The result had significant statistic difference（P＜0.01）.The CAT mRNA expressions: The CAT mRNA expressions of the CLP group washigher than that of the SHAM group at the experimental time points of8h,16h and lowerthan that of the SHAM group at the experimental time points of24h. The result hadstatistic difference （P<0.05，P＜0.01，P＜0.01）. The CAT mRNA expressions of the CSgroup was higher than that of the SHAM group at the experimental time points of8h,16hand24h. The result had significant statistic difference （P＜0.01）. The CAT mRNAexpressions of the CS group was higher than that of the CLP group at the experimentaltime points of8h,16h and24h. The result had statistic difference （P<0.05，P<0.01，P<0.01）.The CAT mRNA expressions of the CA group had no statistic difference with theSHAM group at the experimental time points of8h,16h and24h（P＞0.05）. The CATmRNA expressions of the CA group was lower than that of the CLP group at theexperimental time points of8h. The result had statistic difference （P<0.05）. The CATmRNA expressions of the CA group was lower than that of the CS group at theexperimental time points of8h,16h and24h. The result had significant statistic difference（P＜0.01）.The HO-1mRNA expressions: The HO-1mRNA expressions of the CLP group washigher than that of the SHAM group at the experimental time points of8h,16h and lowerthan that of the SHAM group at the experimental time points of24h. The result hadstatistic difference （P<0.05，P＜0.01，P＜0.01）. The HO-1mRNA expressions of theCS group was higher than that of the SHAM group at the experimental time points of8h,16h and24h. The result had significant statistic difference （P＜0.01）. The HO-1mRNAexpressions of the CS group was higher than that of the CLP group at the experimentaltime points of8h,16h and24h. The result had statistic difference （P<0.05，P<0.01，P<0.01）.The HO-1mRNA expressions of the CA group had statistic difference with theSHAM group at the experimental time points of8h and16h（P＜0.05，P＜0.01）. TheHO-1mRNA expressions of the CA group had no statistic difference with the CLP groupat the experimental time points of8h,16h and24h. The HO-1mRNA expressions of the CA group was lower than that of the CS group at the experimental time points of8h,16hand24h. The result had statistic difference（P<0.05，P<0.05，P<0.01）.(5) The test of the protein content of Nrf-2: At the time points of8h,16h, the Nrf-2protein content in the CLP group was significantly higher than that of the SHAM group（P<0.05）, at the time points of24h, the Nrf-2protein content in the CLP group wassignificantly lower than that of the SHAM group（P<0.05）, and each time the Nrf-2protein content in the CS group was significantly higher than that of the SHAM group andthe CLP group（P<0.01，P<0.01，P<0.05; P<0.05，P<0.01，P<0.01）.The Nrf-2proteincontent in the CA group was lower than that of the SHAM group at the time points of24h(P<0.05). The Nrf-2protein content in the CA group was lower than that of the CLPgroup at the time points of8h and16h(P<0.05). The Nrf-2protein content in the CAgroup was significant lower than that of the CS group at the time points of8h and16h(P<0.01).Conclusion:The Nrf-2ARE signal pathway has a protective effect on the rats’ acute lung injury. |
PDF Full Text Request