Effects Of Transplantation Of The Bone Mesenchymal Stem Cells On Autophagy In Rats With Severe Acute Pancreatitis And Relevant Mechanism

Objective: To validate whether allogenic transplantation of bone marrow mesenchymal stem cells(MSCs) depress autophagy of acinal cells in rat with severe pancreatitis, and explore in further potential signal pathway underlying this effect.Methods: Five-to-six-week old female Sprague-Dawley(SD) rats weighing 250±50 g were randomly and equally divided into normal, SAP model and SAP+MSCs groups. All animals were injected intravenously with double-labeled adenovirus particlemRFP-GFP-LC3(2×109/rat), and then, were housed in room(22-28 ℃; relative humidity: 40-70%). One week later, each rat in SAP+MSCs group underwent intraperitoneal injection with 20% L-arginine and intravenous transplantation of 5×106MSCs derived from male rats one hour later, and the MSCs were instead with physiological saline in SAP model group. As to control animals, L-arginine and MSCs were instead with same volume of physiological saline. At 6 h, 12 h, 24 h, 48 h and 72 h after operation, the amylase, trypsinogen activation peptide(TAP), TNF-α and IL-6 in serum were quantified, and pathological changes of pancreas, lung and kidney were scored based on HE slides. The autophagy was traced by GFP-LC3 under fluorescence microscope. And,expressions of LCII, PI3 K, p-PI3 K, mTOR and p-mTOR in rat pancreases were detected at operative 12 h by Western Blotting and then quantified using Image J software. The data exhibited as mean±SD were statistically analyzed with t-test using SPSS 18.0, and p value lesser than 0.05 was considered as statistical significance.Results: Compared to those in the control, the hyperemia, edema and infiltration of inflammatory cells were found in pancreas from SAP animals. Furthermore, the physiological structure of pancreas in a few SAP rats was destructed or lost. And, the pathological scores, and serum levels of amylase, TAP, TNF-α and IL-6 from SAP ratswere significantly higher than those in control(p<0.05). In addition, SAP animals produced much more ultrastructure related autophagy, and statistical up-regulation in LC3-II expression at postoperative 6 h and 12 h(p<0.05). Compared to those from SAP rats, transplantation of MSCs caused significant decreases in pathological scores and in serum levels of amylase, TAP, TNF-α and IL-6(p<0.05). And, expression LC3-II was down-regulated(p<0.05). However, statistical differences were undetectable on postoperative 24 h, 48 h and 72h(p>0.05). The quantization of PI3 K, p-PI3 K, mTOR and p-mTOR on postoperative 12 h showed that no obvious differences were found in both PI3 K and mTOR proteins between three groups. But, p-PI3 K and p-mTOR were upregulated in SAP animals compared to control rats(p<0.05), whereas those two proteins in SAP+MSCs rats were down-regulated statistically compared those in SAP(p<0.05).Conclusion: Acute severe pancreatitis causes autophagy, which activate the trypsinogen and thereafter participate in the injury of pancreas. Implantation of bone marrow MSCs could suppress autophagy via PI3K-AKT-mTOR pathway, and attenuate the pancreatitis and dysfunction of multiple organs.