| Severe acute pancreatitis(SAP) is a inflammatory disease caused by trypsin activation.The disease will cause pancreatic edema, necrosis and hemorrhage and result in systemic organ failure. Although endoscopic sphincterotomy in treatment of the patients with gallstones, using selective intestinal decontamination and antibiotic prophylaxis may be beneficial in the treatment of acute pancreatitis, but there is no specific drug. Intestinal barrier function plays an important role in SAP. Research on the influence factors of the intestinal barrier and reducing the gut injury in SAP is helpful for the treatment of SAP. In recent years,more and more attention is paid to the effect of curcumin on SAP.Curcumin is a polyphenol compound with antioxidant, anti-inflammatory, antiviral, antitumor effect. Curcumin can cause changes of growth factors, protein kinase and transcription factors expression. EF24 is the analogues of curcumin.The recent studies about EF24 focuses on promoting tumor cell apoptosis and the molecular target of anti-tumor. Vivek R. Yadav et al. report that EF24 can inhibit inflammatory reaction during hemorrhagic shock, our study is to analyze the effect of EF24 on intestinal mucosal barrier in acute pancreatitis from the perspective of inflammatory response.Objective: the experiment aimed to explore the effect and mechanism of biphenyl fluorine ketone EF24 on pancreatitis intestinal injury by observing the changes of SAP pancreas and intestinal histological, detecting the expression of intestinal tissue p65, Claudin-1 protein and mRNA and the concentration of serum IL-6, TNF-α, which can provide new ideas for treatment of intestinal injury in SAP.Methods:1 All the 60 Sprague-Dawley(SD) rats were randomly divided into control group(CON), severe acute pancreatitis group(SAP) and EF24 group(EF24).Then each group was further divided into two subgroups(6h group and 12 h group)according to time points, 10 rats in each group.2 SAP group and EF24 group rats were injected intraperitoneally with 20% L- arginine 3.0g/kg. After an hour, they were injected with the same dose of L- arginine. The CON group rats were injected intraperitoneally with the same dose of 0.9% NaCl, after an hour, once again injected with the same dose of NaCl. In SAP group, the rats were injected intraperitoneally with EF24(0.4mg/kg) 2 h after induction of acute pancreatitis. Con group and EF24 group rats were intraperitoneally injected with the same dose of polyethylene glycol(PEG)400(PEG: 0.9%NaCl=1:1).3 The pancreas, intestinal tissues and blood were taken 6 or 12 hours after induction of acute pancreatitis. The level of serum IL-6 and TNF-α were evaluated by ELISA. The histopathology changes of intestinal and pancreatic tissues were measured by Hematoxylin and Eosin(HE) staining. The expression of intestine tissue’s p65,Claudin-1 protein and mRNA were detected by Western-blot and RT-PCR.4 All data were presented as mean±S(standard deviation) and analyzed by a commercially available statistical software package(SPSS13.0). The significance was assumed at P<0.05.Results:1 Histopathological changes of intestinal and pancreatic tissues: in SAP group, the damages of intestinal and pancreatic tissue aggravated gradually with time passing. The intestinal and pancreatic lesions in EF24 group were alleviated than in SAP group at each time point and were heavier than in CON group at each time point.2 The levels of serum TNF-α: in SAP group, the concentration of serum TNF-α increased significantly at 6h and 12h(P<0.01),and the levels of serum TNF-α were up-regulated at 12 h compared with that at 6 h(P<0.05). Compared with CON group, the expression of serum TNF-α in EF24 group were up-regulated(P<0.05). The levels of serum TNF-α in EF24 group decreased compared with that in SAP group(P<0.05). The concentration of serum TNF-α in EF24 group and in SAP group was the same at each time point(P>0.05).3 The levels of serum IL-6: the concentration of serum IL-6 in SAP group significantly increased compared with that in the other two groups at each time point(P<0.01). Compared with CON group, the expression of serum IL-6 in EF24 group were up-regulated(P<0.05). The levels of serum IL-6 in EF24 group decreased compared with that in SAP group(P<0.05). In all groups, no significant difference was found between the subgroups(P>0.05).4 The expression of intestinal Claudin-1 mRNA: compared with each subgroup of CON group, Claudin-1 mRNA in SAP group decreased significantly(P<0.01).Claudin-1 mRNA in EF24 group was lower than that in CON group in each subgroup(P<0.05). The expression of Claudin-1 mRNA in EF24 group elevated compared with that in SAP group(P<0.05). In SAP group, Claudin-1 mRNA in 6h group was significantly higher than 12 h group(P<0.01).5 The expression of intestinal p65 mRNA:p65 mRNA in SAP group and EF24 group up-regulated significantly compared with each subgroup of CON group(respectively P<0.01, P<0.05). P65 mRNA in EF24 group was lower than that in SAP group in each subgroup(P<0.05). Between the two subgroups in each group, p65 mRNA amount in 12 h group was significantly higher than that of the 6h group(P<0.05) in SAP-S group; there was no significant difference between EF24 group and CON group(P>0.05).6 The expression of intestinal Claudin-1 protein: The level of Claudin-1 protein in SAP declined significantly compared with CON group in the two subgroups(P<0.01).The expression of Claudin-1 protein in EF24 group was iower than that in each subgroup of CON group(P<0.05).But the level of Claudin-1 protein in EF24 group elevated compared with SAP group(P<0.05). There was significant difference between the subgroups in SAP group; The expression of Claudin-1 protein in 6h group was higher than that of the12 h group(P<0.05).7 The expression of intestinal p65 protein: SAP group was obviously higher than that of CON group in each subgroup(P<0.01).EF24 group were reduced compared with SAP group(P<0.05).P65 protein level significantly increased in EF24 group compared with CON group(P<0.05). No significant difference was found between the subgroups in CON group(P>0.05).In EF24 group,6h group was higher than 12h(P<0.05); on the contrary, 12 h group was higher than 6h in SAP group(P<0.01).Conclusion:1 Expression of intestinal Claudin-1 protein increased significantly in EF24 group, suggesting that EF24 could improve the intestinal integrity in SAP and alleviate the damage of intestinal mucosa.2 EF24 could inhibit the expression of proinflammatory cytokine TNF-α, IL-6 and the inflammatory reaction of SAP, which promoted the recovery of intestinal injury in SAP.3 The mechanism of EF24 involving in SAP intestinal injury might be inhibiting NF-κB activation and down-regulating the concentration of proinflammatory factor TNF-α, IL-6. |
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