摘要
在明确雷公藤红素(CEL)干预肥胖-抑郁共病小鼠杏仁核(AMY)及中缝背核(DRN)mRNA表达异同基础上,进一步揭示CEL干预肥胖-抑郁共病中枢炎症的药效靶标群.C57BL/6J小鼠随机分为正常(Chow)组,肥胖-抑郁共病(COM)组,CEL低、中、高剂量(CEL-L、CEL-M、CEL-H,0.5、1.0、2.0 mg-kg-1)组.Chow组给予普通饲料,其余组给予高脂饲料结合潮湿垫料慢性应激.饲养10周后灌胃给药3周,然后取Chow组、COM组和CEL-H组小鼠的AMY及DRN进行转录组分析,并将2个核团的目标差异基因以Venn图取交集.将交集基因导入STRING,进行蛋白-蛋白相互作用(PPI)分析,以DAVID进行基因本体论(GO)功能富集分析,明确CEL调控AMY、DRN的核心靶点.在独立样本中,通过qPCR对上述交集基因进行验证.结果显示,CEL调控AMY及DRN的共有基因为趋化因子家族Ccl2、Ccl5、Ccl7、Cxcl10、Cxcr6和Hsp70家族Hspa1a、Hspa1b及Myd88、Il2ra、Irf7、Slc17a8、Drd2、Parp9、Nampt.GO 分析显示,前五位节点 Ccl2、Cxcl1O、Myd88、Ccl5、Irf7 均参与免疫-炎症调控(P＜0.01).独立样品qPCR结果显示,在AMY中,与Chow组相比,COM组趋化因子家族、Hsp70、Myd88、Il2ra、Irf7、Slc17a8、Parp9、Nampt表达显著上调,Drd2有降低的趋势;与COM组相比,CEL-H组上述病理变化显著改善.在DRN中,与Chow组相比,COM组趋化因子家族、Hsp70、Myd88、Il2ra、Irf7、Parp9、Nampt表达显著下调,Slc17a8表达显著上调;与COM组相比,CEL-H组Cxcr6、Irf7、Drd2表达显著上调,Slc17a8表达显著下调.在AMY及DRN中,CEL对Irf7的表达抑制及激活均呈现剂量依赖性(R2分别为0.709 8、0.917 2).上述研究结果显示,CEL可通过调控相同靶标蛋白的双向表达,干预肥胖-抑郁共病小鼠AMY的免疫激活及DRN的免疫抑制状态,从而有效改善神经炎症.
Abstract
This study aims to further elucidate the efficacy targets of celastrol(CEL)intervention in central inflammation in mice with obesity-depression comorbiditiy,based on the differential mRNA expression in the amygdala(AMY)and dorsal raphe nucleus(DRN)after CEL intervention.C57BL/6J mice were randomly divided into a normal diet group(Chow),a obesity-depression comor-bidity(COM)group,and low-,medium-,and high-dose CEL groups(CEL-L,CEL-M,CEL-H,0.5,1.0,2.0 mg-kg-1).The Chow group received a normal diet,while the COM group and CEL-L,CEL-M,CEL-H groups received a high-fat diet combined with chronic stress from wet bedding.After 10 weeks of feeding,the mice were orally administered CEL for three weeks.Subsequently,the AMY and DRN of mice in the Chow,COM,and CEL-H groups were subjected to transcriptome analysis,and the intersection of target differentially expressed genes in both nuclei was visualized using a Venn diagram.The intersected genes were then imported into STRING for protein-protein interaction(PPI)analysis,and Gene Ontology(GO)analysis was performed using DAVID to identify the core targets regulated by CEL in the AMY and DRN.Independent samples were subjected to quantitative real-time PCR(qPCR)to validate the intersection genes.The results revealed that the common genes regulated by CEL in the AMY and DRN included chemo-kine family genes Ccl2,Ccl5,Ccl7,Cxcl1O,Cxcr6,and Hsp70 family genes Hspa1a,Hspa1b,as well as Myd88,Il2ra,Irf7,Slc17a8,Drd2,Parp9,and Nampt.GO analysis showed that the top 5 nodes Ccl2,Cxcl10,Myd88,Ccl5,and Irf7 were all involved in immune-inflammation regulation(P＜0.01).The qPCR results from independent samples showed that in the AMY,compared with the results in the Chow group,chemokine family genes,Hsp70,Myd88,Il2ra,Irf7,Slc17a8,Parp9,and Nampt were significantly up-regulated in the COM group,with Drd2 showing a decreasing trend;these pathological changes were significantly improved in the CEL-H group compared to the COM group.In the DRN,compared with the results in the Chow group,chemokine family genes,Hsp70,Myd88,I12ra,Irf7,Parp9,and Nampt were significantly down-regulated,while Slc17a8 was significantly up-regulated in the COM group;compared with those in the COM group,Cxcr6,Irf7,and Drd2 were significantly up-regulated,while Slc17a8 was signifi-cantly down-regulated in the CEL-H group.In both the AMY and DRN,the expression of Irf7 by CEL showed both inhibition and acti-vation in a dose-dependent manner(R2were 0.709 8 and 0.917 2,respectively).These findings suggest that CEL can effectively im-prove neuroinflammation by regulating bidirectional expression of the same target proteins,thereby intervening in the immune activation of the AMY and immune suppression of the DRN in COM mice.
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