The Role Of S100A9 In Sepsis-associated Encephalopathy In Mice

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Establishing of SAE mouse model by CLP and LPS Objective: To construct a mouse model of sepsis-associated encephalopathy(SAE)by using Cecal ligation and puncture(CLP)and intraperitoneal injection of lipopolysaccharide(LPS).Methods: The SAE mouse model was established by CLP and intraperitoneal injection of LPS.The body weight and survival of the mice were monitored daily.The cognitive level of the mice was evaluated by new item identification and Barnes maze.Results: The mortality of CLP and LPS mouse model showed a higher mortality rate.The survival rate of the CLP model group was 46.429%,the survival rate of the LPS model group was 57.143%,and the body weight decreased significantly.In the new item identification experiment,the discrimination index of new articles in both groups of CLP and LPS was significantly lower than that in the control group(p <0.01).In the Barnes maze experiment,the number of mistakes and latency of finding the target hole in the CLP and LPS groups were significant greater than the control group(p < 0.05).Conclusion: CLP and LPS model mice have obvious cognitive impairment,showing typical SAE symptoms.Part ? Transcriptome sequencing of hippocampus of SAE mice Objective: In order to find key functional molecules and potential molecular mechanisms related to SAE,transcriptome sequencing analysis of hippocampus of SAE mice was perform.Methods: The SAE mouse model was established by CLP and intraperitoneal injection of LPS.The mouse hippocampus were obtained at 3 and 7 days after modeling for transcriptome sequencing analysis.According to the previous literature,the differential genes were selected for q PCR validation.Results: In the two SAE mouse models constructed by CLP and LPS,transcriptomic sequencing analysis revealed a total of 17 differentially expressed genes,of which 11 genes were up-regulated: Cybb(cytochrome b-245 beta chain),Galnt15(Polypeptide N-Acetylgalactosaminyltransferase 15),RUNX1(Runt-related transcription factor 1),Ms4a6d(membrane-spanning 4-domains subfamily A member 6D),Gardner-Rasheed feline sarcoma viral(v-fgr)oncogene homolog(Fgr),S100A9(S100 calcium-binding protein A9),Pir B(paired immunity Paired immunoglobulin-like receptor B),Fcgr4(low affinity immunoglobulin gamma Fc region receptor IV),H2-Q7(histocompatibility antigen q7? chain(H-2)Class I histocompatibility antigen,Q7 alpha chain),lyz2(lysozyme 2),Csf2rb(colony stimulat Ing factor 2 receptor ?);of which down-regulated 6 genes together: Fst(Follistatin),Otx2(Orthodenticle homeobox 2),Krt18(Keratin18),IGFBPL1(Insulin-like growth factor binding protein 1),Inhba(inhibin beta-A),Nptx2(neuronal Pentraxin 2).Combined with differential expression multiples(>2)and p-value significance(p < 0.01),and refer to the relevant literature,select 6 genes(Galnt15,Runx1,S100A9,Otx2,IGFBPL1,Nptx2)to further verify.QPCR results was consistent with transcriptomics sequencing results.The difference of S100A9 in the hippocampus of SAE mice was the most obvious.Conclusion: Total 17 differentially expressed genes were found in the hippocampus of mice SAE model constructed by CLP and LPS;The genes closely related to inflammation or neuronal function,including Galnt15,Runx1,S100A9,Otx2,IGFBPL1,Nptx2,were differentially expressed in the model,of which the difference in S100A9 was the most obvious.Part ? S100A9 inhibitor improves cognitive function in SAE mice Objective: To investigate the role of S100A9 in SAE and its possible mechanism.Methods: S100A9 inhibitor Paquinimod was used to intervene in both CLP and LPS SAE mice.The body weight and survival of the mice were monitored daily.The cognitive level of the mice was evaluated by new item identification and Barnes maze.At the same time,the level of neuronal inflammation in hippocampus of mice was detected by immunohistochemistry and q PCR.Results: Paquinimod intervention significantly reduced the mortality of SAE mice constructed with CLP and LPS(p < 0.05),but did not increase the body weight of mice(p > 0.05);In the new item identification experiment,Paquinimod improved the learning and memory of SAE mice,and the recognition index of new articles was significantly increased in mice(p < 0.05).In Barnes maze experiment,Paquinimod could not reverse the spatial learning and memory impairment of SAE mice constructed by CLP and LPS,but it has a certain degree of improvement(p > 0.05).Further studies revealed that Paquinimod significantly down-regulated the expression of S100A9 in the hippocampus of SAE mice(p < 0.05),and also inhibited the expression of Iba1 in microglia(p < 0.01),and promoted the hippocampal M1 type microglia transformed into M2 type microglia in SAE mice(p< 0.05).Conclusion: S100A9 inhibitor Paquinimod significantly reduced the mortality of SAE mice constructed by CLP and LPS;Paquinimod improved learning and memory in SAE mice;Paquinimod inhibits the expression of Iba1 in hippocampal microglia of SAE mice and promotes the transformation of M1 microglia into M2 microglia.