| Part 1 The effect of TNF-α/RIP3 mediated cell necrosis in acute pancreatitis and its mechanism researchObjective To explore the effect of TNF- α /RIP3 mediated cell necrosis in acute pancreatitis and its mechanism research.Methods 84 cases of patients with AP were collected. 52 cases of mild acute pancreatitis(MAP) and 32 cases of severe acute pancreatitis(SAP) were included. Gather their clinical data and 5ml peripheral blood to extract mononuclear cells after 1, 3, 5d of AP attack and semi-quantitative RT-PCR to detect the expression of TNF-αand RIP3. We used Real-time PCR to detect mi R155 and ELISA to test VIP. Meanwhile collect plasma of SAP to detect endotoxin. And the correlation analysis was performed among TNF- α, RIP3, mi R155,VIP and endotoxin. 20 cases of healthy people were gathered as control.Results(1) The expression of TNF-αand RIP3 after the onset of AP were significantly increased, and reached to the highest at 3d after AP onset, the MAP group 0.564±0.088 and 0.422±0.086, SAP group 0.968±0.158 and 0.882±0.122 respectively. And then their expression decreased at 5d. The expression of TNF- α and RIP3 in SAP group were significantly higher than MAP group at the same day of AP onset(P < 0.05). The endotoxin level was greatly higher in SAP patients, and reached to the highest of 74.026±18.146pg/ml at 3d after SAP onset,and then dropped at 5d after onset, and endotoxin level was associated with RIP3 expression in SAP.(2) The expression of mi R155 was obviously increased after AP onset compared to control group and reached to the peak at 3d after onset(P < 0.05). The expression of mi R155 in MAP and SAP at 3d after AP onset was3.494±1.189 and 6.369±1.807. The expression in SAP was more significantly increased compared to MAP(P < 0.05).And the expression of mi R155 had relationship with RIP3.(3)The expression of VIP was obviously elevated after AP onset compared to control group. VIP was obviously elevated at 1d of AP onset and then decreased at 3d.And it elevated again at 5d. The expression of VIP in SAP was more significantly elevated at the same day of AP onset compared to MAP(P < 0.05).The expression of VIP had correlation with mi R155 and RIP3.Conclusion TNF-α/RIP3 may participate in the onset of AP and may be associated with the severity of AP. Gut barrier dysfunction maybe further raise RIP3 expression in SAP.Mir155 and VIP maybe participate in the regulation of TNF- α /RIP3 mediated cell necrosis in AP.Part 2 The effects of exogenous VIP on TNF-α/RIP3 mediated cell necrosis in severe acute pancreatitisObjective To explore the effects of exogenous VIP on TNF-α/RIP3 mediated cell necrosis in severe acute pancreatitis.Methods 45 SD rats were selected and divided into SAP, VIP and SO group. Establish SAP rat model and intraperitoneal injection of 5×10-9mol VIP as VIP group. Kill the rats after 1 h, 6 h, 12 h and detect the plasma endotoxin level, TNF-α/RIP3 and mi R155 expression with semi-quantitative RT-PCR and real-time fluorescence quantitative RTPCR in peripheral blood, pancreas tissue and intestinal tissue respectively.Results(1) In SAP group, the TNF-α/RIP3 expression in peripheral blood, pancreas and intestinal tissue was obviously elevated compared to SO group and reached to the peak at1 h, the expression was 0.984±0.179/0.785±0.139, 0.992±0.182/0.804±0.143 and0.948±0.195/0.812±0.152, and it declined later. And the expression of mi R155 was increased obviously compared to SO group, and reached to the peak at 1h, the expression was 3.042 ± 1.015,4.205 ± 1.132 and 3.453 ± 0.937, and decreased later. The plasma endotoxin level was significantly higher in SAP group compared to SO group, and showed a trend of progressive increase, and it reached to 52.058±6.356 at 12 h.(2) In VIP group,TNF-α/RIP3 expression in peripheral blood, pancreas and intestinal tissue was obviously elevated compared to SO group, but it was decreased compared to SAP group at the same time point, especially at 6h, p < 0.05. And mi R155 expression increased obviously compared to SO group, and was higher than SAP group at the same time point, especially at 6h, p < 0.05. Compared to SAP group, the plasma endotoxin level in VIP group was decreased at the same time point, especially at 6h, p < 0.05.Conclusion VIP may down-regulate the expression of TNF-α/RIP3 to relieve SAP inflammation, and mi R155 may assist to negatively down-regulate the expression of TNF-α/RIP3 to inhibit SAP inflammation. |
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