Study On The Mechanism Of Excessive Activation Of M1 Microglia In Mood Disorders Leading To Exacerbation Of Severe Acute Pancreatitis In Mice

Background and Objective: Acute pancreatitis(AP)is an inflammatory reaction of pancreatic tissue self-digestion,edema,hemorrhage and even necrosis after the activation of pancreatin in the pancreas due to various causes.However,the pathological mechanism of AP is not yet clear.When AP is combined with single or multiple organ dysfunction,more than 48 h,it is severe acute pancreatitis(SAP).SAP is a common clinical emergency and critical illness,with sudden onset,dangerous condition,rapid progress,and mortality as high as 30%-45%.Clinically,we found that when patients have SAP when they are in Mood disorder(MD),the condition is more serious and the mortality rate is higher.In the past,our research group found that MD can increase the incidence of SAP and aggravate the severity of the disease in mice,but the mechanism is still unclear.Studies have shown that M1 type microglia(MG)in the hippocampus of the brain will be over-activated during MD.Over-activated M1 type MG can release inflammatory cytokines and cause tissue inflammatory damage.Therefore,we speculate that when MD is complicated by SAP MG may be over-activated by M1 type MG,secreting inflammatory cytokines,leading to high inflammation of the body,triggering systemic inflammatory response syndrome(SIRS),leading to aggravation of SAP.This study intends to prepare an MD-SAP mouse model,immunofluorescence technology to detect the activation status of mouse M1 MG,Western-blot to detect its activated secretion product inducible nitric oxide synthase(i NOS),and ELISA to detect serum hormones and inflammatory factors.Changes,combined with the histopathological changes of pancreas,lung,and colon,explore the mechanism of SAP aggravated by the excessive activation of M1 microglia during MD.Methods: SPF male C57BL/6J mice(n=42)were randomly divided into two groups A and B(group A,n=18;group B,n=24).After 1 week of adaptive feeding,group A was small The mice were fed in a normal environment,and the mice in group B were fed in parallel and given chronic stress stimulation for 3 weeks.Mice in group A were randomly divided into blank control group(CON group,n=6),severe acute pancreatitis group(SAP group,n=12),mice in group B were randomly divided into mood disorder group(MD group,n= 12)Mood disorder-severe acute pancreatitis group(MD-SAP group,n=12).Mice in the SAP and MD-SAP groups were injected with 20% L-arginine salt solution(3.5g/kg)into the abdominal cavity to induce severe acute pancreatitis.Mice in the CON and MD groups were given the same amount of sterile saline at the same time point(0.02ml/g)intraperitoneal injection constitutes a control.After 24 hours of modeling,observe the state of mice in each group and obtain materials.Collect eye blood for the detection of serum amylase(AMS),tumor necrosis factor-α(TNF-α),interleukin 1β(IL-1β),interleukin 6(IL-6),and adrenergic corticotropin(ACTH),cortisol(CORT),serotonin(5-HT)content,pancreas,lung,and colon tissues were stained with hematoxylin-eosin(HE)to observe the pathological changes.The brain tissues were collected and used immunofluorescence technology to detect the Iba1 and CD86 protein of hippocampus microglia,and Western-blot technology was used to detect the expression of i NOS protein in microglia of the hippocampus.Result: 1.The weight of mice in group B was significantly reduced compared with mice in group A(p<0.05);the incidence of mice in the SAP group and MD-SAP group was 66.67% and 100%,respectively,and the mortality rate was 25% and 41.67% respectively.2.The resting time of mice in group B was longer in forced swimming experiment than group A(p<0.05);the expression levels of serum ACTH and CORT in group B were significantly higher than those in group A(p<0.05),while the expression level of 5-HT was significantly lower In group A(p<0.05).3.The expression levels of serum AMS in SAP and MD-SAP groups were higher than those in CON and MD groups(p<0.05),but there was no significant difference in AMS expression levels between SAP and MD-SAP groups;the SAP group was small The expression levels of serum TNF-α,IL-1β and IL-6 in mice were significantly higher than those in the CON group(p<0.05).Compared with the CON group,the expression levels of serum TNF-α,IL-1β and IL-6 in the MD group were significantly higher Increased(p<0.05).Compared with the SAP group,the expression levels of serum TNF-α,IL-1β and IL-6 in the MD-SAP group were significantly increased(p<0.05).4.The pathological scores of pancreas,lung,and colon in the SAP group were significantly higher than those in the CON group(p<0.05).Compared with the SAP group,the pathological scores of the pancreas,lung and colon in the MD-SAP group were increased(p<0.05),there was no significant difference in the structure of pancreatic tissue and ileum in the CON and MD groups.5.Compared with the CON and SAP groups,the number of activated microglia in the hippocampus of the MD and MD-SAP groups increased significantly,among which the M1 type increased(p<0.05).The expression of i NOS protein in the hippocampus of the MD and MD-SAP groups was higher than that in the CON and SAP groups(p<0.05).Conclusion: 1.Three weeks of chronic stress stimulation can cause mood disorders in mice,and intraperitoneal injection of 20% L-arginine salt solution(3.5g/kg)can build a mouse model of severe acute pancreatitis;2.Mood disorder aggravates mouse SAP by over-activating the M1 type microglia in the hippocampus of the mouse,which increases the level of inflammatory factors in the brain,which in turn leads to inflammation such as TNF-α,IL-1β,and IL-6 in the periphery.The secretion of factors increases,leading to the occurrence of SIRS,thereby aggravating the severity of SAP.