| BackgroundPulmonary ischemia-reperfusion injury(IRI)significantly limits the prognosis of lung transplant patients and is a hot and difficult clinical problem that needs to be addressed.Recent studies have been conducted to treat pulmonary IRI from different perspectives,but the overall results are not satisfactory.The mechanism of pulmonary IRI is complex,involving free radical damage,calcium overload,and neutrophil-mediated pathogenesis.Mesenchymal stem cells(MSC)are pluripotent stem cells derived from the mesoderm.After transplantation,MSC are localised in damaged tissues and play a role in immunomodulation and repair of damaged tissues and organs.A number of studies have demonstrated that MSC can reduce IRI after lung transplantation by paracrine secretion of various factors,and reduce the level of inflammatory cell infiltration and protein exudation in prolonged cold ischemia-reperfusion injury.Stanniocalcin-1(STC1)is a glycoprotein hormone first identified in fish that regulates calcium and phosphorus levels and is expressed in a variety of mammalian and human tissues.AMPK is an important regulator of cellular energy metabolism,and its phosphorylation product,phosphorylated AMPK(Phosphorylated Adenosine),is an important regulator of cellular energy metabolism.UCP2 is a transmembrane protein in the inner mitochondrial membrane of lung tissue,whose function is to transport protons from the inner and outer mitochondrial membrane gaps into the inner mitochondrial membrane,a process that does not produce ATP.The process does not produce ATP,but converts the energy produced into heat for release.The mechanism by which MSC ameliorates ischemia-reperfusion injury in lung transplants is not fully understood.ObjectiveIn this study,we aimed to investigate whether MSC injection could reduce oxidative stress injury and thus lung injury in rat lung transplant ischemia-reperfusion injury by constructing a single lung transplantation model,and to investigate whether MSC could promote AMPK phosphorylation and UCP2 expression through paracrine STC1,in order to supplement the understanding of the mechanism of MSC effect in lung acute ischemia-reperfusion injury.MethodsForty-two male SD rats were randomly divided into sham-operated(6),operated(18)and MSC-transplanted(18)groups.The sham-operated group was treated with open-chest treatment only,while the surgical and MSC transplantation groups were established by a modified three-cuff method.All rats were anesthetized 24 hours after operation,and the specimens were collected.The level of pathological damage was assessed by HE staining,the expression of STC1,p AMPK and UCP2 was detected by immunofluorescence staining,immunofluorescence staining tyrosine signal amplification and Elisa method,MDA level was measured by TBA method and SOD level was measured by xanthine oxidase method.Results1.Successful construction of an allogeneic rat left lung in situ transplantation model.The lung transplantation procedure was completed using the triple cuff method.After the operation,the pulmonary artery,bronchus and vein of the rat were checked to be patent,and the model without pulmonary atelectasis could only enter the analysis of the results.A total of 18 pairs of SD rats were used in the establishment of the rat lung transplantation animal model.12 models were successfully established and 6 groups of rats died during the experiment.The main reasons for failure included: tearing of the vessel wall during the recipient’s pulmonary artery;detachment of the anastomosis site or blood leakage;and death due to inability to disengage the ventilator after the lung transplant model was established after chest closure.2.Protective effect of MSC on lung tissue injury in IRI rats after lung transplantation.The results of lung injury scores suggested that the lung injury scores in the sham-operated and MSC transplantation groups were significantly lower than those in the operated group(P <0.01).There was no significant difference in lung injury scores between the sham-operated and MSC transplantation groups(P>0.05).Lung histopathological damage was significantly less in the sham-operated and MSC transplantation groups compared to the surgical group.3.Protective effect of MSC on the level of oxidative stress in IRI rats after lung transplantation.Plasma MDA levels were significantly lower in both the sham-operated and MSC-transplanted groups(P=0.036 and 0.008,respectively),with no statistical difference between the sham-operated and MSC-transplanted groups(P=0.998).Plasma SOD activity levels were lower in the surgical group compared to the sham-operated group(P=0.013)and higher in the MSC transplant group compared to the surgical group(P=0.045),while there was no significant difference between the sham-operated and MSC transplant groups(P=0.323).Lung tissue homogenate MDA was significantly lower in both the sham-operated and MSC-transplanted groups than in the operated group(P=0.001 and0.014,respectively);no significant difference was seen between the sham-operated and MSC-transplanted groups(P=0.280).In contrast,there was no statistically significant difference in SOD activity of lung tissue homogenates between the three groups(P=0.644).4.MSC reduces ischemia-reperfusion injury by promoting STC1 expression.The mean immunofluorescence intensity of STC1 in lung tissue was significantly higher in the MSC transplant group compared to the surgical group(P=0.039),while there was no significant difference in the mean immunofluorescence intensity of STC1 in lung tissue between the sham-operated and MSC transplant groups and between the sham-operated and surgical groups.The plasma STC1 levels in the MSC transplantation group were significantly higher than those in the sham-operated and operated groups(P=0.013 and P=0.017),while the STC1 levels in the sham-operated and operated groups were not significantly different(P=0.990).5.MSC promotes AMPK phosphorylation and UCP2 expression.The comparison of mean immunofluorescence intensity suggested that p AMPK was significantly lower in the surgical group(P=0.015)than in the sham-operated group(P=0.050)and also significantly lower in the MSC transplant group(P=0.050),while the mean immunofluorescence intensity of p AMPK was not significantly different between the sham-operated and MSC transplant groups.There was a significant difference in the mean immunofluorescence intensity of UCP2 between the three groups(P=0.050),with a trend of higher UCP2 expression in the sham-operated group compared to the operated group(P=0.059)and a trend of higher UCP2 expression in the MSC transplanted group compared to the operated group(P=0.112).Conclusion:In this study,a rat left lung transplantation model was established.It was comfirmed that MSC reduce the extent of oxidative stress injury through the production of STC1 in ischemia-reperfusion injury.The mechanism may be related to the promotion of AMPK phosphorylation and UCP2 expression by STC1. |
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