| Chronic pancreatitis(CP) is due to a variety of reasons which induces pancreatic progressive local or diffuse inflammation, eventually leading to pancreatic tissue and(or) of pancreatic exocrine and endocrine function of irreversible damage. The basic pathological changes is the diffuse fibrosis or calcification.Extracellular matrix(ECM) is mainly produced by activated pancreatic stellate cells(PSCs).TGF-?1 is a strong stimulus for PSCs activation in vitro and TGF-?1 signaling pathway plays an important role in fibrogenesis. Smurf2 and Smad7 are the main inhibitory genes in TGF-?1 signaling pathway. The abnormal expression of Smurf2 and Smad7 have been proved in a variety of human and animal fibrosis diseases which involved in the pathogenesis of fibrosis. But at present, the report on its expression and the role in CP patients or animal models have not been found. The pathogenesis of pancreatic fibrosis has not been fully elucidated and it is lack of effective treatment means to delay or reverse the pancreatic fibrosis. The present study shows that oxidative stress and(or) sustained proinflammatory cytokines(such as IL-1?, IL-6, TNF- ?) release and extensive activation of PSCs is one of the main pathogenesis of CP fibrosis. The antioxidant and plant polyphenols which have an effect of anti-inflammatory and anti-oxidation can improve the symptoms of patients with CP, reduce the pancreatic inflammation and fibrosis degree of CP animal models. Omega-3 is a kind of essential fatty acids, its metabolites EPA and DHA is important for human cell membrane phospholipid composition, which plays an important role in the regulation of various functions of the body. Many studies have confirmed that Omega-3 can inhibit the body's various acute and chronic inflammation, alleviate liver, kidney, lung and myocardial fibrosis of animal models through the anti-inflammatory, antioxidant effect. But the treatment effect of Omega-3 on CP has not been reported.Part? Preparation and validation of CP model Objective Preparation and evaluation of rat CP model for the subsequent in vivo study. Methods 20 male Wistar rats were randomly and equally divided into CP group and control group. CP group were treated with DBTC, 100% ethanol and sterile glycerol mixed solvent via rat tail vein injection(8mg.ml-1.kg-1), and the control rats were intravenously treated with equal volume of 100% ethanol and sterile glycerol mixture solvent.The body weight of all rats were measured at 0 days, 7 days, 14 days, 21 days, 28 days and killed at 4 weeks after injection. Histological scores for HE staining, Masson staining and the content of hydroxyproline for collagen, ?-SMA immunohistochemical staining and Real-time RT-PCR assay for PSCs activation were compared between the two groups of rat pancreatic tissue. Result After 4 weeks, the rats in control group were all survival with linear growth, the general morphology of pancreas, histological scores, Masson staining was normal. CP model success rate is 60%, the body weight increased slowly, HE staining and Masson staining presented with typical CP changes. The ?-SMA expression levels and hydroxyproline content in pancreatic tissues in CP group were significant higher than control group(P < 0.05). Conclusion DBTC induced rats CP is a simple and economic method with acceptable success rate, which is one of the ideal model for the study of CP pathogenesis and intervention at present.Part? The expressions of Smurf2 and Smad7 in CP rats Objective To determinate the expressions of Smurf2 and Smad7 in pancreas in CP rats and investigate the possible mechanism. Methods 12 male Wistar rats were randomly and equally divided into CP group and control group. CP group were treated with DBTC, 100% ethanol and sterile glycerol mixed solvent via rat tail vein injection(8mg.ml-1.kg-1), and the control rats were intravenously treated with equal volume of 100% ethanol and sterile glycerol mixture solvent. After 4 weeks, all rats were killed, and histological scores for HE staining, Masson staining and the content of hydroxyproline for collagen, ?-SMA immunohistochemical staining and Real-time RT-PCR assay for PSCs activation were determinate in pancreatic tissues. The expression levels of Smurf2 and Smad7 protein and m RNA were measured and compared by immunohistochemistry and Real-time RT-PCR. Result The expression levels of Smurf2 protein and m RNA, Smad7 m RNA in pancreatic tissues in CP rats were significantly higher but Smad7 protein were significantly lower than that in control group(P < 0.05). Conclusion Dysregulation of Smad7 and Smurf2 may involve in the pathogenesis of pancreatic fibrosis via TGF-? signaling pathway.Part? The mechanism of Omega-3 alleviates CP in a rat model Objective To observe the treatment effect of Omega-3 on the CP and clarify the possible mechanism. Methods 18 male Wistar rats were randomly and equally divided into CP group,CP+Omega-3group and control group. CP group and CP+Omega-3 group were treated with DBTC, 100% ethanol and sterile glycerol mixed solvent via rat tail vein injection(8mg.ml-1.kg-1), and the control rats were intravenously treated with equal volume of 100% ethanol and sterile glycerol mixture solvent. At three days after injection, CP+Omega-3 group were given with Omega-3(1000mg. Kg-1. day-1) daily by gavage, CP group and control group were filled with equal dose of soybean oil. All rats were killed at 4weeks after gavage daily, and histological scores, Masson staining, malondialdehyde, total superoxide dismutase(SOD) activity, the content of hydroxyproline in pancreatic tissues were determinate. Expression levels of ?-SMA and TGF-?1 were tested by immunohistochemistry and Real-time RT-PCR. Result Compared with CP group, Omega-3 significantly reduced histological scores and Collagen fibers deposition, decreased the content of malondialdehyde and enhanced the total activity of SOD in rat pancreatic tissue. The expression levels of ?-SMA and TGF-?1 protein and m RNA in pancreas were significantly lower than that in CP group. Conclusion Omega-3 can reduce chronic inflammation and collagen deposition in rat pancreas induced by DBTC via anti-inflammatory, anti-oxidative stress and via inhibiting the high expressions of ?-SMA and TGF-?1.Above results suggest that the rat CP model induced by DBTC has obvious advantages, and it is one of ideal models currently for the study of CP pathogenesis and treatment; Smurf2 and Smad7 may be involved in the CP and fibrosis process through TGF-? signaling pathway; Omega-3 can reduce chronic inflammation and collagen deposition in rat pancreas induced by DBTC via anti-inflammatory, anti-oxidative stress and via inhibiting the high expressions of ?-SMA and TGF-?1. |
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