ObjectiveTo study the expression of zonula occluden-1 (ZO-1) in ileum tissues and the possible mechanism of intestinal mucosal barrier injury in rats with severe acute pancreatitis (SAP). MethodsFifty SD male rats were randomly divided into sham operation group and SAP group, then SAP group was divided into four subgroups with 10 rats in each subgroup according to the sampling time of 3, 6, 12, and 24 h. The SAP model was made by injecting 5% bovine sodium deoxycholate into biliarypancreatic duct with Aho’s method. The rats were killed at 3, 6, 12, and 24 h after making model. The rats in the sham operation group were killed directly. Tumor necrosis factor-α (TNF-α), diamine oxidase (DAO), and histological changes in pancreatic and intestinal pathologies were observed. At the same time, the ZO-1 protein and mRNA expressions of ileum tissues were detected by immunohistochemistry and RT-PCR, respectively. ResultsCompared with the sham operation group 〔TNF-α: (10.83±0.96) ng/L; DAO: (354.79±3.67) U/L; ZO-1 protein: (10.40±0.45) score; ZO-1 mRNA: 0.878±0.014 8〕, the levels of TNF-α at different time 〔3 h: (125.30±0.94) ng/L; 6 h: (181.89±4.93) ng/L; 12 h: (230.58±1.28) ng/L; 24 h: (198.89±4.83) ng/L〕 were significantly higher (Plt;0.05), the activities of DAO 〔3 h: (235.77±0.67) U/L; 6 h: (117.22±5.58) U/L; 12 h: (106.69±1.39) U/L; 24 h: (91.18±1.09) U/L〕 were significantly lower (Plt;0.05), ZO-1 protein 〔3 h: (8.70±0.22) score; 6 h: (3.73±0.19) score; 12 h: (3.92±0.22) score; 24 h: (4.29±0.30) score〕 and mRNA (3 h: 0.806±0.020 7; 6 h: 0.370±0.015 8; 12 h: 0.502±0.019 2; 24 h: 0.562±0.030 3) expressions of the ileum tissues were significantly lower (Plt;0.05) in the SAP group; Meanwhile, the necrosis of ileum mucous membrane chorioepithelium, angiorrhexis and hemorrhage, and inflammatory cell infiltration in the pancreatic and ileum tissues were also observed. ConclusionThe decrease of expression of ZO-1 in ileum tissues is one of the vital causes for mucosal barrier injury in SAP, probably through acts the excessive release of inflammatory cytokines TNF-α and the decrease of DAO activity.
Objective To observe the protective effects of unfractionated heparin (UFH) on high-mobility group box-1 protein (HMGB1) induced increased permeability of endothelial cells, and investigate the protective mechanism of UFH on HMGB1 induced defective expression of zonula occludens-1 (ZO-1). Methods Human umbilical vascular endothelial cells (HUVECs) were culturedin vitro and divided into 4 groups (n=5), namely a control group, a HMGB1 group (100 ng/ml), a heparin group (UFH 10 U/ml), a HMGB1/heparin group (100 ng/ml HMGB1 + UFH 10 U/ml). Endothelial cell viability was measured by methyl thiazolyl tetrazolium (MTT) colorimetric method. Endothelial permeability was determination by Transwell chamber method. Immunofluorescence and laser confocal microscopy were used to assess the distribution of ZO-1. The protein expressions of tight junction protein ZO-1 and nuclear factor kappa B (NF-κB) were detected by Western blot. Results HMGB1 (100 ng/ml) had no inhibitory effect on endothelial cell viability (P＞0.05). UFH pretreatment could reduce the permeability increment of endothelial cells induced by HMGB1. UFH pretreatment could reduce the close loop reduction and damage of ZO-1 induced by HMGB1, enhance the fluorescence intensity and expression of ZO-1, and decrease the NF-κB translocation. Conclusions UFH can protect HMGB1-mediated defect of ZO-1 expression and increased permeability of the endothelial cells. The mechanism may be related to the decreased nuclear translocation of NF-κB.