Effects Of B7-H3on The Expression Of MMP-9in Mice With Streptococcus Pneumonia Meningitis

Objective:Bacterial meningitis (BM) is one kind of the most common infectious diseases of the central nervous system in childhood; its definite immunopathogenesis is needed to be identified further. Matrix metalloproteinases-9(MMP-9) played an important role in the disruption of blood-brain barriers (BBB). B7homology3(B7-H3) is a new member of the B7costimulatory molecules family.B7-H3had a critical contribution to the regulation of immune response. This study discussed the effection of B7-H3on the expression of MMP-9and the progress of Streptococcus pneumonia (S. pneumonia) meningitis, to provide a new idea and some experimental data for the immune intervention of BM therapy in vivo and in vitro.Methods:Part1(in vivo), to study the effects of B7-H3on the change of behavior score、the body weight and the expression of MMP-9in the brain of mice with S. Pneumonia meningitis.36male healthy BALB/c (1.5-2-month) mice were randomly divided into three groups:the control group (CON, n=12), the S. pneumonia group (SP, n=12), the B7-H3-treated S. pneumonia group (SP+B7-H3, n=12).The animals with anesthesia underwent a lateral ventricle puncture receiving10μL saline and5μL S. pneumonia suspension as the SP group. The SP+B7-H3group were injected with5μL S. pneumonia suspension and10[μL B7-H3protein suspension. The CON group was injected with15μL saline instead. At6h、24h after the induction of S. Pneumococcal meningitis, the behavior changes of the mice were observed with Loffler standard and their body of weight was recorded. Then the mice were killed and the brain was removed. The brain was used for the observation of brain biopsy structure changes by hematoxylin-eosin staining and the determination of the expression of MMP-9by Immunohistochemistry.Part2(in vitro), to detect the effects of B7-H3on the expression of MMP-9in the microglia supernatant treated with S. pneumonia. Fetal (postnatal1-3days) BALB/c mice cerebrum were used to acquire the primary microglia with high purity, then microglia cells were inoculated into24-well culture plates at the concentration1×105/well.The microglia cells were randomly divided into three groups:the control group (CON, n=20), the S. pneumonia group (SP, n=32), the B7-H3-treated S. pneumonia group (SP+B7-H3, n=32), all of the cells were treated with the same amount of ceftriaxone (100μgmL"1) to prevents the overgrowth of S. pneumonia. The culture supernatant was collected at51、24h、48h and detected the expression of MMP-9by ELISA. Each experiment at each time point set3multiple holes, and repeated two independent experiments.Results:Part1(in vivo),1、Neuroethology evaluation:Both score of SP group were significantly lower than that of CON group at6h、24h after induction of the meningitis(P<0.01);Both score of SP+B7-H3group were significantly lower than that of SP group at6h、24h after induction of the meningitis(P<0.05).2、The change of body weight:Both of SP group had an decrease compared to CON group in the body weight at6h、24h(P<0.01、P<0.05separately); Both of SP+B7-H3group had a decrease compared to SP group in the body weight at6h (P<0.05).3、Hematoxylin and Eosin(H&E) staining:The SP+B7-H3group compared with SP group revealed that leptomeninges blood vessels were dilated, subarachnoid space was extremely expanded, a few of inflammatory cell infiltrationed into leptomeninges and subarachnoid space at6h; The SP+B7-H3group compared with SP group revealed that leptomeninges blood vessels were highly dilated, subarachnoid space was extremely expanded or opened, full off inflammatory cells infiltrationed into leptomeninges and subarachnoid space at24h.4、Immunohistochemistry analysis:The expression of MMP-9were at a low level in CON group at6h and24h.In SP group, the expression of MMP-9compared with CON group were significantly increased at6h(P<0.01),the expression of MMP-9compared with CON group were significantly increased at24h(P<0.01);In SP+B7-H3group, the expression of MMP-9compared with SP group were significantly increased at6h(P<0.05),the expression of MMP-9compared with SP group were significantly increased at24h(P<0.05).Part2(in vivo), All of the expression of MMP-9in SP group were significantly higher than that of CON group at5h、24h、48h (P<0.05);All of the expression of MMP-9in SP+B7-H3group were significantly higher than that of SP group at5h(P<0.01、P<0.05、 P<0.05separately).Conclusions:1、The ideal model of S. Pneumonia meningitis was established by inoculate S. pneumonia suspension through lateral ventricle.2、Both of results in vitro and in vivo were demonstrated that MMP-9may be participate in the damage of the pathology and inflammation in mice with S.pneumonia meningitis.3、B7-H3aggravated the inflammation pathological in brain tissue of S. Pneumonia meningitis, advanced the inflammatory cell infiltration and up-regulated the expression of MMP-9.This outcome indicated that B7-H3enhanced the damage of the pathology and inflammation in S. pneumonia meningitis.