Screening of infection conditions for brain microvascular endothelial cells infected by Streptococcus suis

Abstract

Streptococcus suis is a pathogen that causes swine meningitis, sepsis, and other diseases. There are 34 serotypes, of which type 2 is the most pathogenic. During the infection process of Streptococcus suis, several major virulence factors are involved and play a different roles. Streptococcal meningitis is caused by the bacteria’s ability to cross the blood-brain barrier and enter the central nervous system. Therefore, studying the interaction between Streptococcus suis and cerebral microvascular endothelial cells will help reveal meningitis's pathogenic mechanism. When studying the interaction between bacteria and cells, the number of infected bacteria and the time of infection are very important. In this study, Streptococcus suis serotype two was made into bacteria liquid and counted. Then bacteria were used to infect mouse brain microvascular endothelial cells with different multiplicity of infection (1,10,100 and 200). Cells were harvested at six h, nine h,12h,18h, and 24h after infection. The total RNAs of harvested cells were extracted, and the concentration of RNA was detected. The OD260/OD280 was between 1.8~2.4, OD260/OD230 was 1.5~2.4, and the concentration was greater than 100ng/µL. Total RNAs were reverse transcribed to cDNAs used to perform quantitative PCR to detect the mRNA expression of IL-18, IL-1beta, IL-6, and IL-10. The results showed that each MOI group's mRNA expression is higher than the control group with different infection times. When the multiplicity of infection is at 1, each group's relative expression of cytokines reaches a peak at 18hrs after infection. When the multiplicity of infection is at 10, each group's relative expression of cytokines reaches a peak at 12hrs after infection. When the multiplicity of infection is at 100, the relative expression of cytokines reaches a peak at 12hrs after infection. When the multiplicity of infection is at 200, the relative expressions of each cytokine reach a peak at 6hrs after infection. Based on the mRNA relative expression of each cytokine under different conditions, the optimal multiplicity of infection was 100, and the optimal infection time was 12h. The result provides a basis for the study of the pathogenic mechanism of meningitis.

Keywords

• screen, streptococcus suis 2, infection, mouse brain microvascular endothelial cells