Role of high mobility group box-1 and protection of growth hormone and somatostatin in severe acute pancreatitis

Abstract

In this study, we investigated the potential role of high-mobility group box 1 (HMGB1) in severe acute pancreatitis (SAP) and the effects of growth hormone (G) and somatostatin (S) in SAP rats. The rats were randomly divided into 6 groups of 20 each: sham-operated, SAP, SAP+saline, SAP+G, SAP+S and SAP+G+S. Ileum and pancreas tissues of rats in each group were evaluated histologically. HMGB1 mRNA expression was measured by reverse transcription-PCR. Levels of circulating TNF-α, IL-1, IL-6, and endotoxin were also measured. In the SAP group, interstitial congestion and edema, inflammatory cell infiltration, and interstitial hemorrhage occurred in ileum and pancreas tissues. The levels of HMGB1, TNF-α, IL-1, IL-6 and endotoxin were significantly up-regulated in the SAP group compared with those in the sham-operated group, and the 7-day survival rate was 0%. In the SAP+G and SAP+S groups, the inflammatory response of the morphological structures was alleviated, the levels of HMGB1, TNF-α, IL-1, IL-6, and endotoxin were significantly decreased compared with those in the SAP group, and the survival rate was increased. Moreover, in the SAP+G+S group, all histological scores were significantly improved and the survival rate was significantly higher compared with the SAP group. In conclusion, HMGB1 might participate in pancreas and ileum injury in SAP. Growth hormone and somatostatin might play a therapeutic role in the inflammatory response of SAP.

Figure 1. Morphometric analysis of pancreatic tissue (40×). A, SAP group, necrosis and hemorrhage were observed in half of the samples. After 24 h, the interstitial pancreatic tissue was significantly widened and the lobular gap was increased. There was great inflammatory cell infiltration and focal or patchy necrosis of pancreas under a light microscope. B, SAP+NS group, pathological changes were the same as those in the SAP group. C, Sham-operated group, no necrosis or hemorrhage was found, and the pancreas morphological structure appeared normal. D, SAP+G+S group, edema and inflammatory cell infiltration were found occasionally, and there was a significant difference compared to the SAP group. E, SAP+G group, and F, SAP+S group, although inflammatory changes were still visible, inflammatory response was attenuated compared to the SAP group. The images were obtained from at least three experiments. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin.

Figure 2. Morphometric analysis of the ileum. A, SAP group (40×), interstitial ileum exhibited edema and inflammatory cell infiltration; blood vessels and lymph vessels were dilated and congested. B, SAP group (40×), villus was edematous, highly shortened, disarranged and necrotic. Mucosal thickness became thin. C, SAP+G group (40×), villus epithelium proliferated and mucosal thickness increased. Inflammatory response was visible. D, SAP+S group (40×), inflammatory changes were decreased, and epithelial hyperplasia was observed, but active degree was less significant than that in the SAP+G group. E, SAP+G+S group (40×), intestinal inflammatory response was mild, and microvilli were twisted and became longer. F, SAP+G+S group (100×), the cells were highly columnar, goblet cells were increased, and nuclei were overlapped significantly. G, SAP+G+S group (40×), villus was coarse and terminal was expanded. H, SAP+G+S group (40×), villus was twisted and branched. The images were obtained from at least three experiments. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin.

Figure 3. Ileum mucosa at 24 h under transmission electron microscopy. A, Sham-operated group, the cells were dense; microvilli were arranged compactly and regularly, and organelles were integrated. B, SAP group, cell hydrops was obvious; intracellular particles were loose; organelles broke down; microvilli were disarranged; many necrotic cells were seen; and part of membrane damaged. C, SAP+G group, most of the form and structure were normal; microvilli were slightly hypertrophic, and intercellular junction was integrated. D, SAP+S group, intracellular matter was slightly loose, and organelles were still integrated. E, SAP+S+G group, the microvilli were arranged orderly; structure was intact; organelle structure was complete and clear. The images were obtained from at least three experiments. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin.

Figure 4. High-mobility group box 1 (HMGB-1) mRNA expression. A, HMGB-1 mRNA expression in all groups: M: marker; 1: Sham-operated group; 2: SAP+NS group; 3: SAP group; 4: SAP+G group; 5: SAP+S group; 6: SAP+G+S group. B, HMGB-1 mRNA expression at 24 h. Data are reported as means±SD. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin. *P<0.05, SAP, SAP+NS, SAP+G, and SAP+S groups compared to the sham-operated group. After being treated with G+S, the level of HMGB-1 was obviously decreased compared to the SAP group (^#P<0.05, one-way ANOVA).

Figure 5. Serum endotoxin levels. The results are reported as means±SD. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin. The levels of endotoxin in SAP and SAP+NS groups were sharply elevated compared to those in the sham-operated group (*P<0.05). While in the SAP+G, SAP+S, and SAP+G+S groups, the endotoxin was reduced significantly compared to the SAP group (^#P<0.05), there was no significant difference compared to the sham-operated group (one-way ANOVA).

Figure 6. Serum cytokines TNF-α (A), IL-6 (B), and IL-1β (C) expression. The results are reported as means±SD. SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin. The levels of TNF-α, IL-6, and IL-1β were all elevated in the SAP and SAP+NS groups compared to the sham-operated group (*P<0.05). The levels in the SAP+G, SAP+S, and SAP+G+S groups were decreased compared to the SAP group (^#P<0.05). There were no significant differences in SAP+G, SAP+S, and SAP+G+S groups compared to the sham-operated group (one-way ANOVA).

Figure 7. Survival rate. Survival rates of rats are described with different symbols and colors. The results showed no death in the sham-operated group, and the survival rates of rats in the SAP and SAP+NS groups were significantly different from that in the sham-operated group (P<0.01). SAP+G, SAP+S, and SAP+G+S groups had significantly different survival rates compared to the SAP group (P<0.05, P<0.05, and P<0.01, respectively). *P<0.05 and **P<0.01 compared to the sham-operated group; ^#P<0.05 and ^##P<0.01 compared to the SAP group (one-way ANOVA). SAP: severe acute pancreatitis; NS: normal saline; G: growth hormone; S: somatostatin.