NOD2 Expression in Streptococcus pneumoniae Meningitis and Its Influence on the Blood-Brain Barrier

Hindawi Canadian Journal of Infectious Diseases and Medical Microbiology Volume 2018, Article ID 7292084, 8 pages https://doi.org/10.1155/2018/7292084 Research Article NOD2 Expression in Streptococcus pneumoniae Meningitis and Its Influence on the Blood-Brain Barrier Ying Wang,1,2 Xinjie Liu ,1 and Qi Liu2 1 2 Department of Pediatrics, Qilu Hospital, Shandong University, 107# Wen Hua Xi Road, Jinan, Shandong 250012, China The People’s Hospital in Zoucheng, 59# Qian Quan Road, Zoucheng, Shandong 273500, China Correspondence should be addressed to Xinjie Liu; Received 10 May 2018; Revised 26 June 2018; Accepted 3 July 2018; Published 13 August 2018 Academic Editor: Pietro Mastroeni Copyright © 2018 Ying Wang et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Streptococcus pneumoniae meningitis is one of the most common disorders seen in clinical practice. It is believed that the brain tissue immune injury is caused by the expression of pattern-recognition receptors (PRR) which can further induce the release of other cytokines and inflammatory cascades. The aim of this study is to investigate the expression of nucleotide-binding oligomerization domain 2 (NOD2) and inflammatory factors in rat brain tissues infected with Streptococcus pneumoniae and its influence on the blood-brain barrier (BBB) permeability. Rats were given an intracranial injection of Streptococcus pneumoniae to construct the Streptococcus pneumoniae meningitis rat models. The expression change curves of NOD2 and inflammatory factors at different time points (0 h, 12 h, 24 h, 48 h, and 7 d) after Streptococcus pneumoniae were evaluated by enzyme-linked immunosorbent assay (ELISA). Western blotting analysis and quantitative real-time polymerase chain reaction (qRT-PCR) were engaged to examine the expression of NOD2. Furthermore, the changing processes of pathological characteristics, nervous system score, cerebral oedema, and BBB permeability were observed. Our results showed that NOD2 expression began to increase in the 12 h after Streptococcus pneumoniae infection group, while the remaining inflammatory factors were not obviously increased. Meanwhile, the levels of NOD2, as well as inflammatory factors IL-1β, TNF-α, and IL-6 were markedly elevated in 24 h and 48 h infection groups, which were consistent with the increases in BBB permeability and BWC, and the positive expression of NOD2 in the infected rat brain tissues was observed using immunohistochemistry (IHC). This study suggests that NOD2 might be related to the activation of inflammation pathways and the damage to the blood-brain barrier. NOD2 and inflammatory factors have played vital roles in the pathogenesis of Streptococcus pneumoniae meningitis. 1. Introduction Streptococcus pneumoniae is the common central nervous system infectious pathogen in clinical practice [1, 2]. The overall mortality and prognosis for survivors remain unsatisfactory regardless of the application of effective antibiotics and the promotion of effective vaccination [3, 4]. It is currently believed that pattern-recognition receptors (PRR) are expressed and maybe produce inflammatory factors initiating the brain tissue immune injury [5]. Inflammatory factors can lead to secondary immune responses, brain oedema, and BBB dysfunction. Among all the pathomechanisms of cerebral injury, the effects of the NLRs protein family have attracted high attention in recent years [6]. So far 20 members of the NOD family have been reported, such as apoptotic protease-activating factor-1 (APAF-1), nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and disease-resistance proteins (R-protein), most representative of which is NOD1 and NOD2 [7–9]. The difference between NOD1 and NOD2 is that the caspaseassociated recruitment domain (CARD) of the aminoterminal domain [10, 11]. NOD1 and NOD2 proteins can identify peptidoglycan, but their recognition dose not overlap: the NOD2 protein can recognise GlcNAc-MurNAc tripeptide muropeptide (GM-Trilys) only in Gram-positive bacteria, except for identifying GlcNAc-MurNAc dipeptide (GM-di) [6]. In this section, we focused on NOD2 because Streptococcus pneumoniae belongs to Gram-positive bacteria. NLRs can induce the activation of the NF-κB or MAPK pathway to induce inflammatory reaction [13, 14]. Typically, 2 Canadian Journal of Infectious Diseases and Medical Microbiology NOD2 protein is involved in both of these two canonical pathways [15]. Research in vitro suggests that NOD2 protein can induce microglial cell hyperplasia and astrocyte demyelination and elevate the inflammatory cytokine levels such as the activating tumor necrosis factor (TNF) [16]. Other NLRs family members also participate in the proinflammatory effect of NOD2 in signal transduction. For instance, the intracellular pattern recognition receptor can activate the caspase ligand and apoptosis-associated specklike protein, which will result in caspase-1 activation stimulation and IL-1β secretion [17]. These immune cells release proinflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), which readily cross the blood-brain barrier into the brain parenchyma and promote the immune inflammatory damage of brain tissue [16]. IL-6 is one kind of the interleukin, which is a cytokine that has been verified to be highly expressed in numerous infection models [18]. Therefore, TNF-α, IL-1β, and IL-6 were selected as the related inflammatory factors in this experiment, to further explore the expression of NOD2 as well as the related inflammatory factors in Streptococcus pneumoniae meningitis, as well as their influence on the BBB. A Streptococcus pneumoniae-induced meningitis model was established in this study. The changing processes of pathological characteristics, the nervous system score, cerebral oedema, and BBB permeability were observed. In the meanwhile, expression change curves of NOD2 and inflammatory factors were recorded at different time points after Streptococcus pneumoniae. This research result provides a theoretical basis for further exploring the mechanism of NOD2 in Streptococcus pneumoniae meningitis. 2. Materials and Methods 2.1. Preparation of Streptococcus pneumoniae Suspension. Streptococcus pneumoniae serum type III standard bacteria were provided by the National Institute for the Control of Pharmaceutical and Biological Products (Beijing). The bacteria were inoculated on the sheep blood agarose medium under 37°C and 5% CO2 environments overnight. Subsequently, single colonies were selected and inoculated on the VITAL AER broth overnight. The culture solution was centrifuged and precipitated, and Streptococcus pneumoniae suspension at the concentration of 1.5 × 108 cfu·ML−1 was diluted with sterile normal saline for intracisternal injection [19, 20]. 2.2. Construction of Streptococcus (...truncated)