Effects Of Inflammation Induced By LPS On BBB And Its Possible Mechanism

Objective: 1.Establish a mode of intracerebral inflammatory response induced by lipopolysaccharide(LPS), to observe the variation characteristics of blood-brain barrier(BBB) in inflammatory area.2.Research the possible activation mode of astrocytes--VEGF-A pathway in the leakge of BBB induced by inflammation related to LPS.Methods: 1.In vivo study: in this experiment, adult male C57B6 mice for research were randomly divided into four groups : Sham group, LPS group, LRS(LPS inhibitor) + LPS group and Vandetanib(VEGFR2 inhibitor) + LPS group. Subsequently, after giving LPS, LRS and Vandetanib through the lateral ventricle, we measure EB permeability,tight junction protein expression levels,albumin leakage and brain edema for evaluation of blood-brain barrier destruction.Meanwhile,detect astrocyte activation status, the expression of vascular endothelial growth factor(VEGF-A) and IL-1?. 2.In vitro study: primary astrocytes cells were divided into four groups: control group(CON),LPS group,Sham group(normal BV2 cell supernatant), treatment group(BV2 cell supernatant after stimulation by lps). Respectively, at the gene level and protein level detecting changes in the expression level of VEGF-A.Results: 1.In vivo study: after intracerebroventricular injection of LPS, as compared with the control group, the mice brain tissue water content increased, EB and albumin leakage increased, Claudin-5 expression levels dropped, astrocyte marker(GFAP), inflammatory cytokines IL-1? expression levels elevated,above differences were statistically significant(P <0.05).After giving LRS pretreatment,there is no significant difference in the EB permeability and expression levels of GFAP and tight junctions protein Claudin-5 compared with sham. Meanwhile, after administration of LPS stimulation we can observe increased VEGF-A protein levels, the difference was statistically significant(P <0.05). After giving Vandetanib pretreated, compared with LPS group mice brain tissue water content, EB permeability, Claudin Claudin-5 expression level of astrocyte marker(GFAP),expression levels of inflammatory cytokines IL-1? have significant changes, this differences were statistically significant(P <0.05). 2.In vitro study: Primary astrocytes had successful cultured, LPS group compared with the control group, VEGF-A were not significantly changed at gene level or protein expression levels(P> 0.05). But primary astrocytes were cocultured with supernatant of microglia activated by LPS, VEGF-A expression levels were significantly increased, the difference was statistically significant(P <0.05). If administered the cell culture supernatant of normal microglia, VEGF-A expression levels did not change significantly(P> 0.05).Conclusion: 1.LPS can lead to the destruction of BBB, astrocyte activation and increased secretion of VEGF-A, VEGF-A receptor 2 blocker Vandetanib can significantly weaken the above changes, suggesting that LPS lead to abnormal BBB open via increased action of astrocytes--VEGF-A path. 2.LPS can directly cause astrocyte activation, but this time VEGF-A expression of activated astrocytes had no significant change; activated microglia activated by LPS can promote the expression of VEGF-A in astrocytea by secreting certain cytokines(IL-1??TNF-?).