Résumé
Objective:To explore the effect of probiotics Lactiplantibacillus plantarum(LP) WCFS1 by gavage on acute necrotizing pancreatitis (ANP) and associated ileum injury in mice. Methods:Twenty-four healthy male mice were gavaged with broad-spectrum antibiotics for 3 weeks to establish microbiota-depleted mice, and then randomly divided into control group (CON), ANP model group (ANP), LP gavage group (LP) and LP gavage and ANP induced group (LP+ ANP) , with 6 mice in each group. Mice in LP and LP+ ANP group were treated by gavage of LP (1×10 9 CFU/ml, 0.2 ml/day per mouse) for 1 week, while CON and ANP were gavaged with sterile phosphate buffered saline for 1 week instead. The ANP model was induced by intraperitoneal injection with caerulein (100 μg/kg) for 10 times with 1-hour interval between two injections and the 10th injection with lipopolysaccharide(LPS) 5 mg/kg intraperitoneally, and the mice were sacrificed 2 h later. Levels of LP in stool and ileal mucosa were detected by real-time PCR; the pancreas and ileum were collected for pathological examination to observe the extent of tissue inflammation and to score the pathology. Serum amylase activities were determined by enzymatic kinetic chemistry; serum inflammators levels and intestinal permeability were detected by ELISA; levels of inflammators in pancreatic and ileal tissues were detected by real-time PCR; ileal tight-junction proteins (occludin, claudin-1 and ZO-1) were measured by immunofluorescence staining. Results:LP levels in the stool and ileal mucosa of mice were significantly increased after LP gavage, and the differences were statistically significant (913.30±39.12 vs 2.39±1.39, 23.11±0.50 vs 1.38±0.28, all P value <0.05). The pathological scores of pancreatic tissue of CON, LP, ANP and LP+ ANP group were (0.26±0.41), (0.17±0.26), (8.55±0.46) and (6.30±0.45); the serum amylase activities were (219.70±19.73), (217.60±11.30), (2896.24±98.32) and (1837.13±131.60)U/L, IL-1β were (0.87±0.28), (1.4±0.85), (67.41±6.45) and (36.33±5.65)pg/ml, IL-6 were (0.74±0.27), (0.16±0.16), (280586.12±39163.92) and (107912.75±31283.47)pg/ml, IL-10 were (35.52±5.27), (50.99±15.34), (2008.45±184.83) and (3070.35±403.71)pg/ml; the expression level of pancreatic IL-1β mRNA was 1.42±0.39, 0.95±25, 20.53±0.50 and 10.69±1.01, IL-6 mRNA was 1.31±0.44, 0.93±0.023, 21.97±1.71 and 11.54±1.75, IL-10 mRNA was 0.93±0.14, 0.75±0.15, 0.99±0.21 and 1.76±0.19; there was no significant difference between LP and CON group, and pancreatic pathological scores, serum amylase、IL-1β and IL-6 levels, and the expression level of pancreatic IL-1β and IL-6 mRNA were significantly decreased in LP+ ANP group compared with those in ANP group, while serum IL-10 levels and the expression level of pancreatic IL-10 mRNA were significantly increased compared with ANP group, and all the differences were statistically significant (all P values <0.05). The pathological scores of ileal tissue of CON, LP, ANP and LP+ ANP group were 0, 0, (3.17±0.41) and (1.67±0.52); the levels of serum DAO of CON, LP, ANP and LP+ ANP group were (0.03±0.03), (0.02±0.02), (0.50±0.05) and (0.49±0.06)ng/ml; LPS levels were (2.75±0.35), (3.74±0.28), (7.19±0.92) and (5.88±0.38)ng/ml; the expression level of ileal IL-1β mRNA was 1.21±0.20, 1.17±0.09, 1.81±0.25 and 1.63±0.21; IL-6 mRNA was 1.01±0.29, 2.83±0.42, 54.45±8.50 and 16.87±4.42; IL-10 mRNA was 1.12±0.41, 6.09±2.51, 11.65±1.47 and 29.86±2.93. There was no significant difference between LP and CON group, except that the ileal IL-10 mRNA expression was significantly higher than that of CON group. Ileal pathological scores, serum LPS levels and the expression level of ileal IL-6 mRNA were significantly lower in LP+ ANP group than those in ANP group, while the expression level of ileal IL-10 mRNA was significantly higher than that of ANP group; the expression of ileal tight junction proteins (ocludin, claudin-1, ZO-1) was significantly higher than those in ANP group, and all the differences were statistically significant (all P values <0.05). Conclusions:LP WCFS1 gavage could ameliorate the injury of pancreatic and ileal barrier in caerulein-induced ANP mice.
Résumé
Objectives To observe the changes of autophagy of acinar cells in hypertriglyceridemia (HTG) associated acute necrotic pancreatitis(ANP) rats,and investigate the effects of HTG on autophagy and AP. Methods Forty SD rats were randomly divided into 4 groups using random number method, including control group,ANP group,hyperlipemia(HL) group,hyperlipidemia associated acute necrotizing pancreatitis (HANP) group. Control group and ANP group were fed with normal diet for 2 weeks,while the rats in HL and HANP group were fed with a high-fat diet for 2 weeks,and after the blood collection ANP and HANP rats were induced by 3.5% sodium taurocholate retrograde injection into the pancreaticobiliary duct to establish the ANP model. Rats in control group and HL group were intraperitoneally injected with equal volume of normal saline solution. ALL rats were sacrificed at 48 h after the model establishment. Serum triglyceride (TG), total cholesterol (TC) and serum amylase levels were measured; pancreatic tissue was histologically examined and scored; Transmission electron microscope were used to observe the intracellular ultrastructure changes and calculate the number of autophagosome and autophagic lysosome;the expressions of autophagy related proteins such as LC3,LC3Ⅱ, LC3Ⅰ, p62 and Lamp-2 were determined by Western-blot. Results After 2-week high fat diet, serum levels of TG and TC in HANP and HL groups were obviously higher than those in the control and ANP group;Serum amylase levels in HANP group and ANP group were higher than the control and HL group,and the differences were statistically significant (all P<0.05). Transmission electron microscope showed that the number of autophagosomes in ANP group was significantly higher than those in other three groups and the difference was statistically different(all P<0.05);and the autophagy activity in HANP group was impaired compared with ANP group. Compared with the control group, in ANP group the protein expressions of LC3 Ⅱ/Ⅰ ratio and p62 in pancreatic tissue were up-regulated (2.11 ± 0.03 vs 1.70 ± 0.08, 9 420 ± 624 vs 4 973 ± 577),but LAMP-2 was down-regulated(20 533 ± 578 vs 23 231 ± 1 155). Compared with ANP group,the proteins expression levels of LC3Ⅱ/Ⅰand p62 in HANP group was obviously decreased (20 533 ± 578 vs 23 231 ± 1 155), and the difference was statistically significant (all P<0.05). Conclusions Hypertriglyceridemia can worsen the pathology injury of AP,and the mechanism may be related to the decreased autophagy related proteins LC3Ⅱ/Ⅰ and p62 and decreased number of autophagosome.