| Sepsis caused by respiratory tract infection is one of the common emergency clinical diseases with high morbidity and mortality,which brings great burden to people’s health,public health,economy and society.Further research is needed.A large amount of evidence shows that the early immune response of sepsis is impaired,and its severity and duration are closely related to the clinical prognosis of sepsis patients.Therefore,by studying the expression characteristics of immune and inflammatory indicators in patients with sepsis caused by respiratory tract infection and the corresponding expression characteristics after Chinese medicine intervention,this study aims to find the internal relationship between patients with sepsis caused by respiratory tract infection and immune and inflammatory indicators,as well as the influence and effect of Chinese medicine intervention on the expression of immune and inflammatory indicators,so as to provide an objective basis for further clinical intervention of Chinese medicine.At the same time,the animal experimental model of sepsis caused by respiratory tract infection was established through animal experimental studies,and the immune function and inflammatory response of the model were discussed.Jinhua Qinggan formula was selected as the intervention formula,and the two traditional formulas derived from Maxingshigan Decoction and Yinqiao recipe were compared to explore the pharmacodynamic mechanism of Jinhua Qinggan formula on immune disorders of sepsis mice.Objective:1 To explore the expression characteristics of immune,inflammatory and other indicators in patients with sepsis caused by respiratory infection and the expression characteristics after the intervention of traditional Chinese medicine.The aim of this study was to find the internal relationship between patients with sepsis caused by respiratory infection and immune and inflammatory indicators,and the effect of traditional Chinese medicine intervention on the expression of immune and inflammatory indicators in patients with sepsis caused by respiratory infection.To provide an objective basis for further clinical intervention in Chinese medicine.2 By establishing an animal experimental model of sepsis caused by respiratory infection,we investigated the immune function and inflammatory response in a model of sepsis caused by respiratory infection,and we observed and compared the effects of Jinhua Qinggan formula and its two related traditional formulas:Maxingshigan Decoction and Yin Qiao Fang,on the immune function and inflammatory response of this model,and elucidated the mechanisms at the levels of organs,cells,inflammatory factors and TLR4/MyD88 and PD-1/PD-L1 molecular pathways,respectively,to provide a new objective basis for further research and treatment of sepsis.Methods:1 The epidemiological cross-sectional study method was applied to the clinical research.①According to the inclusion and exclusion criteria,patients with sepsis caused by respiratory infection were collected and divided into sepsis group and septic shock group,and according to whether traditional Chinese medicine(TCM)intervention was applied,patients were divided into applied Chinese medicine group and non-applied Chinese medicine group.The analysis of relevant immune indicators such as PD-1 and PD-L1,and inflammatory indicators such as CRP,IL-6,LPS,IL-1β,EGF,TGF-β1,IFN-γ,MIP-1α,TNF-α,and general laboratory indicators such as SOFA score was carried out to explore the expression characteristics of the observed indicators in patients with sepsis caused by respiratory infection and the expression characteristics of related indicators after TCM intervention.②According to the inclusion and exclusion criteria,patients with sepsis caused by respiratory SARS-CoV-2 infection were collected and divided into sepsis group and septic shock group,and were divided into survival group and death group according to clinical outcomes.The analysis of immune function testing,including T lymphocyte subsets and absolute numbers,general laboratory and other indicators was carried out to explore the expression characteristics of the observed indicators in patients with sepsis caused by respiratory SARS-CoV-2 infection and the expression characteristics of the relevant indicators after the intervention of TCM.2 In the experimental research part,mouse model of sepsis caused by respiratory infection was constructed.72 BALB/c mice were evenly divided into 6 groups according to the random number table method:Control group,Model group,Resatorvid group,Maxingshigan(MXSG)group,Yinqiao(YQ)group,and Jinhua-Qinggan(JHQG)group.In the model group and the TCM intervention group,the mouse model of sepsis caused by respiratory infection was established by the LPS nasal drip method.The Resatorvid group was given intraperitoneal injection of Resatorvid solution.Every TCM intervention infection group was administered by gavage with the herbal granule solution of the corresponding formula,the herbal granules were administered continuously for 5 days.The mass of the spleen and thymus of mice was measured separately,and the index of immune organs was calculated and routine blood counts were detected in peripheral whole blood and T-lymphocyte subsets in peripheral blood and splenic were detected by flow cytometry.The levels of EPI,IFN-α,IFN-β,IL-1β,IL-10 and TNF-α in peripheral blood and IFN-α,IL-1βand TNF-α in alveolar lavage fluid were measured by ELISA.The expression of TLR4,Myeloid differential protein-88(MyD88)and PD-1 and PD-L1 in lung tissues was observed by immunohistochemistry(IHC).The expression of TLR4,MyD88,soluble TNF receptor-associated factor 6(TRAF6),TGF-β-activated kinase 1(TAK1)and nuclear factorκB(NF-κB)in lung tissues was detected by Western-blot.Results:1 The expression characteristics of general laboratory indicators in patients with sepsis caused by respiratory infection:the expression of PLT,HGB,HCT,pH,HCO3,ALB,TT3 and TSH in sepsis patients was higher than that septic shock patients(P<0.05).The expression of PCT,NLR,PLR,LAC,TNI,BNP,CR,UA,ALT,AST,GGT,DD,FDP,PT,PT%,APTT,TT,CRP/ALB ratio,PCT/ALB ratio and SOFA score in sepsis patients was lower than that in septic shock patients(P<0.05).2 The expression characteristics of inflammatory indicators in patients with sepsis caused by respiratory infection:The expression of LPS,IL-1β,TGF-β1,IFN-γ,IL-6 and CRP in sepsis patients was lower than that in septic shock patients(P>0.05).3 The expression characteristics of immune indicators in patients with sepsis caused by respiratory infection:the expression of PD-1 in sepsis patients was lower than that in septic shock patients(P<0.05),and the expression of PD-L1 was higher than that in septic shock patients(P<0.05).The expression of PD-1 in patients with sepsis and septic shock was higher than that in normal volunteers(P<0.05),and the expression of PD-L1 in patients with sepsis and septic shock was lower than that in normal volunteers(P<0.05).The expression of C4 in sepsis patients was higher than that in septic shock patients(P<0.05),and the expression of IgE in sepsis patients was lower than that in septic shock patients(P<0.05).4 The expression characteristics of general laboratory,immune and inflammatory indicators in patients with sepsis caused by respiratory SARS-CoV-2 infection:IgE,IgM and C3 in sepsis patients were lower than those in septic shock patients(P<0.05),the levels of CD3+,CD3+CD4+and CD4+/CD8+ in sepsis patients were higher than those in septic shock patients(P<0.05).LY#,CD3+,CD3+CD8+,CD3+CD4+,CD4+/CD8+ in the death group were lower than those in the survival group(P<0.05).CRP,D-D,IgE and IgM in the death group were higher than those in the survival group(P<0.05).5 The expression characteristics of observation indicators after Chinese herbal medicine intervention in sepsis patients:①The expression of LPS,IL-1β,IL-6,PD-1,IgE and CRP in sepsis caused by respiratory infection in the application of traditional Chinese medicine group was lower after intervention than before(P<0.05),and the expression of EGF,PD-L1,CD3+,CD3+CD4+,CD3+CD8+,and CD4+/CD8+in sepsis group with traditional Chinese medicine after intervention was higher than that before intervention(P<0.05),There was no significant difference between the above indicators after intervention and before intervention in sepsis group without traditional Chinese medicine(P>0.05).② After intervention,IgG and IgE in TCM injection group of sepsis patients caused by respiratory SARS-CoV-2 infection were lower than before intervention(P<0.05),and the CD3+,CD3+CD8+and CD3+CD4+in TCM injection group were higher than before unused TCM injection group(P<0.05).6 Experimental research:①Index of immune organs:All experimental groups were higher than those in Control group(P<0.001);The increase of immune organ index in Resatorvid group,MXSG group and JHQG group was lower than that in Model group(P<0.05).② Peripheral blood cell count:There was no statistical difference in total white blood cell count,red blood cell count and platelet count among all groups(P>0.05).③T lymphocyte subsets in the peripheral blood:The ratio of CD3+CD4+in YQ group and JHQG group was higher than that in Model group and Resatorvid group(P<0.05).The ratio of CD3+CD8+in YQ group and JHQG group was lower than that in Control group and Resatorvid group(P<0.05).The rate of CD4+CD8+in Model group was lower than that in Control group(P<0.05),while that in Resatorvid group was higher than that in Model group(P<0.05).The CD4+/CD8+ratio of YQ group and JHQG group was higher than that of Control group(P<0.05),and JHQG group was higher than that of Model group and Resatorvid group(P<0.05).④Results of ELISA detection in peripheral blood:The level of EPI in Model group was higher than that in Control group(P<0.001),while that in Resatorvid group,YQ group and JHQG group was lower than that in Model group(P<0.001).The levels of IFN-α and IFN-βin Model group were lower than those in Control group(P<0.001),while the level of IFN-αin Resatorvid group and JHQG group was higher than that in Model group(P<0.001),and the level of IFN-β in Resatorvid group and MXSG group was higher than that in Model group.The level of IL-1β in Model group was significantly higher than that in Control group(P<0.001).The level of IL-1β in each drug intervention group was lower than that in Model group(P<0.001),and the level in MXSG group was lower than that in Resatorvid group,YQ group and JHQG group(P<0.001).The level of IL-10 in Model group was lower than that in Control group(P<0.05),while that in MXSG group and YQ group was higher than that in Model group(P<0.05).In addition,the levels of IL-10 in MXSG group and YQ group were higher than those in Resatorvid group and JHQG group(P<0.05).The level of TNF-α in Model group was higher than that in Control group(P<0.001),while that in MXSG group was lower than that in Model group(P<0.001).⑤ELISA detection results of alveolar lavage fluid:The level of IFN-α in Model group was significantly lower than that in Control group(P<0.001),while that in Resatorvid group and MXSG group was higher than that in Model group(P<0.05).The level of IL-1β in Model Group was higher than that in Control group(P<0.05),while that in Resatorvid group and MXSG group was lower than that in Model group(P<0.05).The level of TNF-α in Model Group was significantly higher than that in Control group(P<0.001),while that in Resatorvid group,MXSG group,and YQ group was lower than that in Model group(P<0.05).⑥The proliferation rate of spleen lymphocytes in Model group was lower than that in Control group(P<0.001),while that in MXSG group,YQ group,and JHQG group was higher than that in Model group(P<0.05).⑦Spleen T lymphocyte subsets:The rate of CD3+CD4+in Resatorvid group,YQ group and JHQG group was higher than that in Model group(P<0.05),and the rate of CD4+CD8+in Resatorvid group and YQ group was lower than that in Control group(P<0.05).⑧Results of IHC:The expression of TLR4 in Model group was higher than that in Control group,and that in drug intervention groups was lower than that in Model group,but there wasn’t statistics difference in them(P>0.05).The expression of MyD88 in Model group was higher than that in Control group(P<0.05),but that in Resatorvid group,MXSG group and JHQG group was lower than that in Model group(P<0.05).The expression of PD-1 in Model group was higher than that in Control group(P<0.05),but that in in Resatorvid group and MXSG group was lower than that in Model group(P<0.05).The expression of PD-L1 in YQ group and JHQG group was lower than that in Model group(P<0.05).⑨Results of Western blot:The expression of TLR4,TRAF6,TAK1 and NF-κB in Model group was higher than that in Control group(P<0.001),while that in Resatorvid group,MXSG group and JHQG group was lower than that in Model group(P<0.05).The expression of MyD88 in all experimental groups was higher than in Control group(P<0.001),while that in all drug intervention groups was lower than that in Model group(P<0.001).Resatorvid group was lower than MXSG group(P<0.001),that in MXSG group was lower than JHQG group(P<0.05).JHQG group was lower than that in YQ group(P<0.001).Conclusions:1 General laboratory indicators such as PLT,HGB,HCT,PH,HCO3,ALB,TT3,TSH,PCT,NLR,PLR,LAC,TNI,BNP,CR,UA,ALT,AST,GGT,DD,FDP,PT,PT%,APTT,TT,CRP/ALB,PCT/ALB,SOFA,etc,have their corresponding expression characteristics in patients with respiratory system infection-induced sepsis and septic shock.Inflammatory indicators such as LPS,IL-1β,TGF-β1,IFN-γ,IL-6,CRP and immune indicators such as PD-1,PD-L1,IgE,C4 also have their own expression characteristics in patients with sepsis and septic shock.Studies have shown that patients with sepsis caused by respiratory infection do have impaired immune function and inflammatory reactions.The immune and inflammatory reactions of septic shock patients are more obvious than those of sepsis patients,and the immune function impairment is more serious,suggesting that the prognosis of patients is poor.The above related indicators have certain clinical guiding significance,because they can be used as a reference index to evaluate the severity of the disease,the severity of inflammation,the impaired immune function,whether the patients with sepsis caused by respiratory infection are complicated with multiple organ failure,and judge the prognosis of patients.2 TCM can significantly improve the immune impairment in patients with sepsis caused by respiratory system infection,improve the corresponding expression of immune indicators,and play a bidirectional regulation role on the immune system.In addition,so that the body can achieve immune balance.TCM can significantly improve the inflammatory response and inflammatory damage in patients with sepsis,and reduce the expression level of inflammatory factors.The results of this study suggest that the application of Chinese medicine has a bright future,and provide an objective clinical basis for further TCM treatment of sepsis.3 The mouse model of sepsis caused by respiratory tract infection showed immune dysfunction,up-regulated expression of inflammatory factors and inflammatory pathways.Maxingshigan Decoction,Yinqiao recipe and Jinhua Qinggan formula can interfere with immune function and inflammatory response of mouse model of sepsis caused by respiratory tract infection.These Chinese medicines can affect the organ index of immune organs,T lymphocyte proliferation and subsets,and the expression of immune and inflammatory factors to varying degrees,which may be related to the regulation of TLR4/MyD88 and PD-1/PD-L1 pathways. |
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