Study On The Correlation Between Pancreatic Stone Protein/regenerating Protein And The Pathogenesis Of Multiple Organ Dysfunction Syndrome In Sepsis

Background :Sepsis is a life-threatening condition characterized by a dysregulated host response to infection,which is often accompanied by organ dysfunction and is a major cause of preventable mortality in critically ill patients.Sepsis is a highly heterogeneous syndrome with complex immune-pathophysiological changes,and if left untreated,can progress to multiple organ dysfunction syndrome(MODS).MODS is defined as progressive physiological dysfunction of two or more organ systems,requiring intensive care unit(ICU)intervention to maintain internal homeostasis.Once diagnosed with MODS,the mortality rate of septic patients can reach 28-56%,making it the final stage of a dysregulated systemic inflammatory response.Neutrophils are innate immune cells that play a crucial role in the host defense against infectious pathogens,being the first line of defense against pathogens.Excessive activation of neutrophils during sepsis and their infiltration into vital organs have been found to be key mechanisms of sepsis-induced multiple organ injury.Pancreatic stone protein(PSP/Reg)is a C-type lectin protein that is overexpre-ssed during sepsis,and related studies have found its upregulation in various inflammatory diseases and has the ability to bind to neutrophils directly,but its correlation with MODS is less studied.This study aims to explore the role of PSP/Reg in sepsis-induced multiple organ failure and its potential regulatory mechanisms through clinical research,establishment of sepsis animal models,and in vivo and in vitro experiments,in order to provide new treatment strategies and insights for clinical treatment.Please help me to refine and translate this paragraph into academic English.Chapter I The clinical significance of pancreatic stone protein in patients with sepsis.Objective:To investigate the predictive value of peripheral blood pancreatic stone protein(PSP/Reg)concentration for the occurrence and prognosis of multiple organ dysfunction syndrome(MODS)in patients with sepsis.Methods:1.Prospective collection of adult patients(n=141)who met the diagnostic criteria for sepsis and were admitted to the intensive care unit of the First Affiliated Hospital of Nanchang University between September 2021 and October 2022.2.Upon patient admission,clinical data was collected,and within 24 hours,blood samples were also collected for ELISA analysis,in order to determine peripheral blood PSP/Reg concentration.Standard treatment interventions were administered to all patients,with clinical physicians making the necessary treatment decisions.In order to eliminate any potential bias,the clinical physicians were kept blind to the testing results.Additionally,Sequential Organ Failure Assessment(SOFA)was evaluated daily,along with organ support therapy conditions.Finally,organ damage biomarkers,including liver and kidney function,myocardial enzymes,and oxygenation index,were all carefully recorded.3.All enrolled patients were classified into severe sepsis group and septic shock group according to the Sepsis 3.0 criteria.Based on whether MODS progressed within 48-72 hours after admission,patients were divided into MODS group and non-MODS group.Patients were also categorized into survival group and death group based on whether they survived within 28 days after admission.4.Statistical analysis was performed to investigate the relationship between peripheral blood pancreatitis-associated protein(PSP/Reg)concentration at admission and the severity of sepsis,organ support therapy,progression of multiple organ dysfunction syndrome(MODS),and prognosis in sepsis patients.Additionally,we evaluated the predictive value of PSP/Reg for MODS progression and prognosis.Result:1.The circulating PSP/Reg level in septic shock group(347.1 ng/m L [179.1,665.2])was significantly higher than that in severe sepsis group(82 ng/m L [62,297.7])(p < 0.001).2.Based on the grouping according to the SOFA score at admission,a positive correlation was observed between PSP/Reg levels and SOFA scores(p < 0.0001).3.The Spearman rank correlation test revealed correlation between PSP/Reg levels and the degree of dependence on organ support therapy in patients,including a high demand for vasopressors on admission(r=0.496;p<0.0001),prolonged use of vasopressors(r=0.545;p<0.0001),the need for mechanical ventilation(r=0.607;p<0.01),and the need for RRT treatment(r=0.360;p=0.015).4.The circulating levels of PSP/Reg were significantly higher in the MODS group compared to the non-MODS group(427.2ng/m L [268,951.2] vs.213.8ng/m L[136.8,536.4];p < 0.001).5.The circulating PSP/Reg level was significantly higher in the death group compared to the survival group(427.2ng/m L [289,842.1] vs 208.1ng/m L [135.3,516.2];p < 0.001).6.After adjusting for gender and age,the logistic regression analysis revealed that circulating PSP/Reg levels were independent risk factors for the development of MODS and 28-day mortality in patients(OR: 1.012 [1.003,1.020],p = 0.005;OR:1.006 [1.002,1.010],p < 0.001).7.The ROC curve analysis showed that the AUC for predicting MODS progression at 48-72 hours after ICU admission in sepsis patients based on PSP/Reg was 0.714(p =0.001),and the AUC for predicting 28-day mortality in sepsis patients based on PSP/Reg was 0.734(p <0.001).Conclusion:1.In adult sepsis patients,peripheral blood PSP/Reg levels at admission are closely related to disease severity,organ support therapy,MODS progression,and poor prognosis.2.Measurement of PSP/Reg concentration within 24 hours of admission is helpful in predicting the progression and prognosis of MODS in sepsis patients,identifying patients with a higher risk of death due to organ damage,and improving prognosis.Chapter II The role of pancreatic stone protein in multi-organ damage in septic mice Objective:Explore the role of pancreatic stone protein in multi-organ injury in a mouse model of sepsis induced by cecal ligation and puncture(CLP).Methods:1.A mouse model of sepsis was induced by cecal ligation and puncture(CLP),and recombinant pancreatic stone protein(PSP/Reg)was used to treat the mice.The animals were divided into three groups: control group(CLP followed by intravenous injection of an equal volume of PBS at 30 minutes),low-dose PSP/Reg40 group(CLP followed by intravenous injection of 40 ng/kg of recombinant PSP/Reg protein at 30minutes),and high-dose PSP/Reg400 group(CLP followed by intravenous injection of 400 ng/kg of recombinant PSP/Reg protein at 30 minutes).2.At 24 hours post-CLP surgery,mice in each group were evaluated using a blinded method based on the "Mouse Sepsis Score".The score assessed the health status of mice based on seven aspects: appearance,level of consciousness,activity,responsiveness to stimuli,eye opening,respiratory rate,and quality of respiration.3.A new batch of mice was obtained and assigned to groups in the same manner described previously,with identical treatments administered.The survival rate of each mouse was evaluated every 12 hours during the 7 days following CLP surgery,and survival curves were plotted.4.At 24 hours post-CLP,the mice in each group were euthanized and the levels of peripheral blood inflammatory cytokines(TNF-α,IL-6,IL-1β)and organ damage markers(lactate dehydrogenase,creatinine,cardiac troponin I)were measured using enzyme-linked immunosorbent assay(ELISA).The degree of pulmonary edema in each group was determined by calculating the lung wet/dry(W/D)ratio.5.Collect tissue samples from the lungs,heart,liver,and kidneys of each group of mice and prepare paraffin sections.Use a TUNEL staining kit to detect the level of cell apoptosis in the major organs of the mice.Result:1.Compared to the PBS group,both low and high dose PSP/Reg groups showed a significant increase in sepsis scores,with the high dose PSP/Reg group having a significantly higher score than the low dose group.PSP/Reg exacerbated the severity of the disease in a dose-dependent manner.2.Compared to the PBS group,the median survival time of the low dose PSP/Reg group was significantly reduced,with a further reduction in the high dose PSP/Reg group.3.Compared to the PBS group,the levels of inflammatory cytokines TNF-α,IL-6,and IL-1β were significantly increased in the low dose PSP/Reg group.Compared to the low dose PSP/Reg group,the high dose PSP/Reg group further increased the levels of TNF-α and IL-6,but did not further increase the expression of IL-1β.4.Compared to the PBS group,the levels of organ injury markers lactate dehydrogenase,creatinine,troponin I,and lung wet/dry weight ratio were significantly increased in the low dose PSP/Reg group.The high dose PSP/Reg group further increased the levels of these organ injury markers.5.Compared to the PBS group,the TUNEL-positive levels in lung,heart,liver,and kidney tissues were significantly increased in the low dose PSP/Reg group,with a further increase in TUNEL-positive levels in tissue sections from the high dose PSP/Reg group.Conclusion:Based on the animal experiments,it can be concluded that PSP/Reg exacerbates systemic inflammation and multi-organ damage in septic mice in a dose-dependent mannerChapter Ⅲ: Mechanisms of Pancreatic Stone Protein Aggravating Multiple Organ Failure Objective:To explore the relevant mechanisms of recombinant pancreatic stone protein(PSP/Reg)aggravating sepsis-induced multiple organ failure.Methods:1.CLP animal modeling and grouping were performed as previously described.The infiltration degree of neutrophils in the lungs,heart,liver,and kidneys was detected using MPO(myeloperoxidase)assay kit and immunofluorescence staining of tissue sections.2.Neutrophils were isolated from each group of mice using a mouse neutrophil isolation kit.The expression levels of CD29 and ICAM-1 activation markers on neutrophils were analyzed using flow cytometry.3.Neutrophils were isolated from C57BL/6 mice using a mouse neutrophil isolation kit and divided into control group(LPS+equal volume of PBS stimulation),low-dose PSP/Reg group(LPS+40 ng/ml Lrecombinant PSP/Reg protein stimulation),and high-dose PSP/Reg group(LPS+400 ng/m L recombinant PSP/Reg protein stimulation).4.Analyze the expression levels of CD29 and ICAM-1 activation markers in neutrophils using flow cytometry.5.Detect the m RNA expression levels of inflammatory cytokines(TNF-α,IL-1β,IL-6)in neutrophils from different groups of mice using q-PCR.6.Measure the level of intracellular NETs using SYTOX detection kit in each group of cells.7.Measure the level of intracellular ROS using Cell RIS detection kit in each group of cells.Result:1.Compared to the PBS group,both low and high doses of PSP/Reg treatment significantly increased the levels of oxidative stress and neutrophil tissue infiltration in important organs of septic mice.2.Compared to the PBS group,both low and high doses of PSP/Reg treatment significantly increased the expression levels of surface activation factors CD29 and ICAM-1 on isolated bone marrow neutrophils in mice.3.In cell experiments,compared to the PBS group,both low and high doses of PSP/Reg treatment significantly increased the expression levels of surface activation factors CD29 and ICAM-1 on bone marrow neutrophils in mice.4.Compared to the PBS group,both low and high doses of PSP/Reg treatment significantly increased the m RNA expression levels of inflammatory cytokines(TNF-α,IL-1β,IL-6)in bone marrow neutrophils in mice.5.Compared to the PBS group,high-dose PSP/Reg stimulation upregulated the production of NETs in bone marrow neutrophils of mice.6.Compared to the PBS group,both low and high doses of PSP/Reg treatment significantly increased the ROS levels in bone marrow neutrophils of mice.Conclusion:1.PSP/Reg treatment significantly increased the activation and tissue infiltration levels of neutrophils in important organs of septic mice.2.In cell experiments,PSP/Reg treatment exacerbated the inflammatory response of bone marrow neutrophils in LPS-induced mice.