| Objective:(1)Objective to evaluate the effect of "Jiawei Yishen Qiqiao Decoction" on brain function and immune function in patients with sepsis-associated encephalopathy(SAE),and to explore its possible mechanism.(2)Objective to establish an animal model of SAE and explore its possible molecular mechanism by observing the effects of Jiawei Yishen Qiqiao Recipe on inflammatory mediators and cholinergic system.Methods:(1)A total of 40 patients with SAE diagnosed in EICU of Jiangsu Provincial Hospital of traditional Chinese medicine from January 2020 to February 2021 were included.They were divided into traditional Chinese medicine group and control group,20 cases in each group.On the basis of conventional treatment,the traditional Chinese medicine group was given "Jiawei Yishen Qiqiao Decoction":(Gypsum 30g,Huangqi 30g,Huangqin 10g,Danggui 10g,Shicangpu 10g,Taoren 10g,Baizhi 10g,Dahuang 5g).The control group was given the same amount of warm water by nasal feeding.Both groups were treated for 5 days.After enrollment,the basic information of the subjects was recorded,including name,initials,gender,age,hospitalization number,start date of the experiment,The systemic inflammatory indexes(white blood cell count,neutrophil count,hypersensitive C-reactive protein,procalcitonin),as well as biochemical markers(NSE,S100?),Serum cytokines(IL-6,IL-1B,IL-2,IL-4,IL-5,IL-8,IL-10,IL-12,IL-17,TNF-?,IFN-?,IFNf-?),cholinergic related indicators(serum ache,chat),T cell subsets count,GCS score,APACHE ? score,and SOFA score;safety evaluation was conducted during the treatment,liver and kidney function,blood routine,coagulation function;follow up to 28 days,record the mechanical ventilation time,vasoactive drug use time and mortality of the two groups.(2)Thirty six female SD rats were randomly divided into normal group,model group and traditional Chinese medicine group,with 12 rats in each group.Except for the normal group,the remaining animals were injected intraperitoneally with CLP to establish sepsis model.After 24 hours of observation,neurobehavioral abnormalities were found in septic rats.Neurobehavioral Evaluation:neuroreflex score ?6 was used to judge the success of the model.? After successful modeling,rats in the three groups were given normal saline by gavage,30 ml/kg/24 hours,once every 2 hours.The traditional Chinese medicine group was given 0.5ml/100g three times by gavage(1h,3h,5h).The blank group and model group were given the same amount of normal saline at each time point.? Six hours after administration:the levels of NSE,S100?,TNF-?,IL-6,ache,chat and nerve reflex scores in abdominal venous blood of the three groups were detected,and 6 rats in each group were randomly killed.Half of the brain tissue homogenate supernatant was used for NSE,S100 ?,TNF-?,IL-6,ache and chat detection;the other half of brain tissue was sectioned and stained to observe the proportion of dense neurons and brain cell apoptosis.? After 12 hours of administration:the serum NSE,S100 ?,TNF-?,IL-6,ache,chat and nerve reflex scores of the remaining rats in the three groups were detected.After all rats were killed,half of the brain homogenate supernatant was taken to detect NSE,S100 ?,TNF-?,IL-6,ache and chat;the other half of the brain tissue was sliced and stained to observe the proportion of dense neurons and brain cell apoptosis.Results:(1)Clinical study showed that:? compared with day1,the WBC,neut and PCT of Day5 in TCM group were significantly decreased(P<0.05),and the PCT of Day5 in TCM group was significantly decreased(P<0.05).Compared with day1,IL-5,IL-6 and IL-17 were significantly decreased(P<0.05),TNF-? was significantly decreased(P<0.01),IL-4 and IL-10 were significantly increased(P<0.05);IL-6,IL-17 and TNF-? in the control group were significantly decreased(P<0.05);and the levels of IL-6 and TNF-? after intervention of traditional Chinese medicine were significantly lower than those in the control group(P<0.05).? Compared with day 1,the levels of NSE and S100 ? in the two groups decreased significantly(P<0.05),and the level of NSE in the traditional Chinese medicine group decreased significantly(P<0.01);after the intervention of traditional Chinese medicine,the level of NSE in the two groups decreased significantly(P<0.05),but there was no significant difference in the level of S100 ? between the two groups(P>0.05).? Compared with dayl,the content of ache in TCM group and control group decreased significantly after treatment(P<0.05),and after TCM Intervention,the content of ache in TCM group decreased significantly compared with control group(P<0.05).There was no significant difference in chat level between the two groups before and after treatment(P>0.05).? After treatment,compared with day1,the GCS scores of the two groups were significantly improved(P<0.01,P<0.05),and the GCS scores of the traditional Chinese medicine intervention group were significantly improved(P<0.01).?The area under the ROC curve(AUC)of GCS score on day 1,Day5 and prognosis of SAE patients in the two groups were 0.624 and 0.784 respectively.According to Youden index,the best cut-off point of GCS score was Day5?8,with sensitivity of 77.78%,specificity of 68.18%and 95%confidence interval of 0.626 and 0.898.The area under ROC curve(AUC)of day1,Day5 and S100 ? levels for prognosis evaluation of SAE patients in two groups were 0.600.681,respectively.According to Youden index,the best cutoff point of S100? level was>0.37ug/l in Day5,the sensitivity was 94.44%,the specificity was 36.36%,and the 95%confidence interval was 0.514,0.819.? GCS score was negatively correlated with NSE level(P<0.001);GCS score was negatively correlated with S100 ? level(P<0.001).?Compared with day1,the CD4+level of the two groups were significantly increased(P<0.05);after the intervention of traditional Chinese medicine,the CD8+level of the patients was significantly decreased(P<0.05),and the decrease was more obvious than that of the control group(P<0.05).After treatment,the CD4+/CD8+of Day5 was significantly higher than that of Dayl in both groups(P<0.05),and there was a very significant difference between the traditional Chinese medicine group and the control group(P<0.01).? After treatment,Apache ? score and SOFA score of TCM group and control group were significantly decreased(P<0.01),but there was no significant difference between the two groups(P>0.05).Compared with the control group,the use time of vasoactive drugs in the traditional Chinese medicine group was significantly reduced(P<0.05).(2)Animal experiment showed that:? pathological observation,the number of ganglion cells in the model group at 6h and 12h was significantly higher than that in the normal group(P<0.01);the number of nuclear pyknosis in the traditional Chinese medicine group was significantly lower than that in the model group at 6h and 12h after the intervention of traditional Chinese medicine(P<0.01);at the same time,there was a significant difference between 6h and 12h after the intervention of traditional Chinese medicine(P<0.05).?TUNEL results showed that the apoptotic rate of brain cells in the model group at 6h and 12h was significantly higher than that in the normal group(P<0.01);the apoptotic rate of brain cells in the traditional Chinese medicine group was significantly lower than that in the model group at 6h and 12h after treatment with traditional Chinese Medicine(P<0.05);the apoptotic rate of brain cells in the traditional Chinese medicine group at 6h and 12h also had significant difference(P<0.05).? ELISA results showed that compared with the normal group,the levels of IL-6,TNF-?,NSE and S100 ? in serum and brain homogenate of the model group were significantly higher at 6h and 12h(P<0.01);the levels of IL-6,TNF-?,NSE and S100 p in serum and brain homogenate of the traditional Chinese medicine group were significantly lower than those of the model group at 6h and 12h after the intervention of traditional Chinese medicine(P<0.05),There was also a significant difference in the measurement level between 6h and 12h of TCM Intervention(P<0.05).?IHC staining results showed that the protein expression of IL-6 and TNF-? in brain tissue of model group at 6h and 12h was significantly higher than that of normal group(P<0.01);compared with model group,the protein expression of IL-6 and TNF-? in brain tissue of traditional Chinese medicine group at 6h and 12h after treatment was significantly inhibited(P<0.05).The protein expression of ache and chat in model brain tissue increased significantly at 6 hours and 12 hours(P<0.01);after the intervention of traditional Chinese medicine,the protein expression levels of ache and chat in brain tissue of traditional Chinese medicine group at 6 hours and 12 hours were significantly lower than those of model group(P<0.05);at the same time,the protein expression levels of ache and chat were also significantly different at 6 hours and 12 hours after the intervention of traditional Chinese medicine(P<0.05).Conclusions:(1)Sepsis encephalopathy is one of the most common complications of sepsis.Neuro immune inflammatory reaction is the main cause of SAE injury.(2)"Jiawei Yishen Qiqiao Decoction" can improve the systemic inflammatory response,reduce the release of pro-inflammatory factors,increase the level of anti-inflammatory factors,and reduce the brain injury mediated by uncontrolled inflammatory response in patients with SAE.(3)"Jiawei Yishen Qiqiao Decoction" can exert anti-inflammatory effect by regulating cholinergic system.(4)There is a certain degree of immunosuppression in patients with SAE."Jiawei Yishen Qiqiao Decoction" can improve the immunosuppressive state of patients,and play a role by reducing inhibitory T lymphocytes and increasing helper T lymphocytes.(5)"Jiawei Yishen Qiqiao Decoction" can improve the brain function of SAE patients and shorten the time of using vasoactive drugs,but it has no effect on the mortality and 28 day survival rate.(6)There was a negative correlation between S100? and the severity of brain function,by detecting S100? Level diagnosis and evaluation of SAE is feasible.(7)Sepsis can induce neuroinflammation in rats,and "Jiawei Yishen Qiqiao Decoction" can protect brain by improving neuroinflammation in sepsis.The damage of cholinergic system can lead to brain damage in rats with sepsis."Jiawei Yishen Qiqiao Decoction" can improve the damage of cholinergic system in rats with sepsis. |
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