Expression Of Toll-like Receptor In Newborn Mice Brain Damage After HCMV Intrauterine Infection

Objectives: To investigate the mechanism of TLR2 in brain damage in neonatal mice by establishing the model of brain damage in neonatal mice and detecting the expression of TLR2,My D88,IL-8 and IFN-? in brain of model which damage after HCMV intrauterine infection.Methods:1.Establishment and evaluation of animal model: Randomly selected 8 10 weeks,clean level?SPF?BALB/c mice were randomly divided into three groups A,B and C whose serum HCMV Ig M and Ig G antibody test were negative.The A group were treated with intraperitoneal injection of HCMV AD169 suspension 0.5ml/?5.0log strain TCID50?,the B group of mice received intraperitoneal injection of cell suspension 0.5ml/,the C group not given virus solution and cell suspension injection.At the end of the 7 day after the treatment were detected the serum HCMV-Ig M and Ig G antibodies of three groups of mice?ELISA method?.After testing,randomly selected from each group of mice treated with the above treatment: A group of 54?male and female 2: 1,serum HCMV-Ig M were positive?,B and C each 15?male and female 2: 1,serum HCMV-Ig M were negative?.Then paired feeding,to be pregnant rats by natural birthspontaneous delivery and get pregnant rats.Observation of three groups of female mice with vaginal plug appeared and newborn mice stillbirth rate.The mice were decapitated whose brain and blood were taken in each group survived?24hours after birth,P1?under hypothermia anesthesia.Half of the brain tissue was used for HCMV-Ig M test and pathological examination?HE staining?,the other half preserve the reserve in-80 refrigerator.ELISA method was used to detect serum HCMV-Ig M and statistics pups intrauterine infection and the incidence of brain damage.2.The expression of TLR2 in brain tissue of brain damage in neonatal mice and impact on downstream cytokines: The brain tissues of the rats in the three groups of A,B and C were selected respectively.Randomly selected brain tissues of group A which serum and brain HCMV-Ig M were all positive specimens,according to the pathology of brain tissue were divided into A1 group and A2 group.B and C group were randomly selected16 cases of spare brain tissue samples were divided into B1?blank control?and C1group?normal control?.Two groups of serum HCMV-Ig M and brain tissue HCMV-Ig M test results were negative and pathological examination confirmed that no brain tissue damage.Three groups of newborn mice were treated and grouped at 7th days after birth as 1st days.The expression of TLR2 m RNA?RT-q PCR?,My D88,IL-8 and IFN-?ELISA?in neonatal mice?first days and seventh days?were detected respectively.Results:?1?Three group weight of mice were no difference?F=0.61,P=0.51?after injection 7 days.While the positive rate of serum HCMVIg M and Ig G antibody in virus injection group was significantly higher than that of the other groups,the difference was statistically significant?P=0.00?.After paired feeding the time?day?of female mice vaginal plug of the 3 group were respectively?8.7±1.50 ? 6.5±1.1 ? 6.4±0.80?,the average newborn mice of delivery each time were?6.5±1.30?8.0±1.20?7.7±1.00?,neonatal mice serum HCMV Ig M positive rate?%?respectively?40.1%?4.0%?3.0%?,neonatal mouse brain tissues of HCMV Ig M positive rate?%?respectively?22.7%?0%?0%?.The experimental group of new brain tissue?HCMV-Ig M positive?showed obvious pathological changes,more than two sets of newborn mouse brain tissue sections showed no pathological changes.?2?The experimental group expression of TLR2 m RNA ? My D88 ? IL-8 ? IFN-?in neonatal mouse brain tissue confirmed damage(1^ST DAY)significantly higher than the control group and blank control group.While expression(7^EDDAY)of TLR2 mRNA ? MyD88 ? IL-8 ? IFN-?,were detected respectively?0.742±0.11,3.77±0.49ng/ml,30.05±1.95pg/ml,21.30±2.20ng/l?,which tlr2 mrna,My D88,IL-8 group difference no statistical significance,the IFN beta difference among the three groups have statistical significance in the experimental group and control group and blank control group differences between no statistical significance.Conclusions:?1?Injection of HCMV virus by intraperitoneal injection,the virus can infected mice,and lead to central nervous system damage through the placenta vertical infection the offspring.?2?TLR2 receptor induce of inflammatory cytokine release,initiates an inflammatory reaction,and participate in the pathological process of brain damage mainly through the My D88 dependent pathway,in neonatal mice induced by intrauterine infection of HCMV.