Chronic Unpredictable Mide Stress Affects Microglia Activation Under Conditions Of Inflammation Induced By Lipopolysaccharides

ObjectiveTo investigate the effect of the CORT induced by CUMS combined with LPS on the activation of NLRP3 inflammasome and glucocorticoid receptor expression and function, observe the injury of dopamine (DA) neurons mediated by microglia activation, and explore its possible mechanism.MethodsThe rats were randomly divided into 6 groups, including sham group, LPS group, CUMS group and CUMS combined LPS injection 7days,14 days,21days(7d,14d,21d) groups. LPS was injected in rat SNc of LPS and CUMS combined LPS groups to prepare PD model On Day 22, the rest groups were injected with normal saline in the same position. Behavior indexes of each group was observed on day 29. Then on day 43, apomorphine was used to observe rotating case in each group by intraperitoneal injection, then all animals were killed, serum and tissue were collected to be detected. Enzyme-linked immunosorbent assay (ELISA) was used to detect test serum Tumor necrosis factor-a(TNF-a),Interleukin--1β(IL-1 β),Interleukin-6 (IL-6)Corticosterone(CORT),Adrenocorticotropichormone (ACTH) in all rats’ serum. Immunohistochemistry (IHC-P) was used to observe the microglia activation and the damage of DA neurons in SNc of the brain. Western blot (WB) was used to detect the expression of Cysteinyl aspartate specific proteinase-1(Caspase-1), Apoptosis-associated speck-like protein containing CARD(ASC), Nucleotide binding oligomerization domain-like receptor family pyrin domain containing 3(NLRP3), Interleukin-18 (IL-18), Glucocorticoid receptor(GR) and Mineralocorticoid receptor (MR) in SNc.Result1) Behavioral experiments observation:Compared with sham group, spontaneous motor activity and exploratory behavior of rats in LPS, CUMS, CUMS(14d)+ LPS and CUMS(21d)+LPS groups had been decreased.2) Rotating experimental observations:Impaired contralateral rotation occured in LPS and CUMS(14d)+LPS groups, and the number of rotations increased in CUMS(14d)+LPS group compared with LPS group.3) ELISA test results:Compared with sham group, the level of CORT and ACTH in all CUMS groups including CUMS(7d,14d,21d)+LPS groups were increased in different degrees. Compared with sham group.the level of TNF-a, IL-1β, IL-6 in rat serum were significantly increased in rats of LPS group.And compared with LPS group, the level of TNF-a, IL-1β, IL-6 in CUMS(14d)+LPS group were still increased.4) Immunohistochemistry result:A larger number of dopaminergic neurons in SNc of sham and CUMS group rats arranged in neat rows with definite shapes and microglia cell basically stayed in a resting state; In LPS group, most of the dopamine neuron degenerated and disappeared, neuronal cell body was shrinkaged and shape was blurred, while activated microglia cell increased significantly; Compared with LPS group, CUMS(14d)+LPS group increased the degree of dopaminergic neurons injury and the number of activated microglia cell.5) Western blot test results:Compared with sham group, the expression levels of Caspase-1, ASC, NLRP-3 protein and proinflammatory cytokine IL-18, IL-1β in SNc of LPS group had increased, while these elevated degrees enhanced in CUMS(14d)+LPS group compared with LPS group; Compared with the sham group, The GR and MR expression levels in SNc of CUMS group, CUMS(14d)+LPS group and CUMS(21d)+LPS group were decreased to different degrees.ConclusionCUMS can promote inflammation in central nervous system, resulting in DA neurons injury.The mechanism may be CUMS can cause increased activation of NLRP3 inflammasome and microglial,while also lead the decreased expression and dysfunctionthe of GR and MR.