猪链球菌2型PPCDC蛋白对血脑屏障通透性的影响

Streptococcus suis type 2 PPCDC protein affects permeability of blood-brain barrier

摘要:

【背景】猪链球菌2型(Streptococcus suis type 2, SS2)是引起全球经济和公共卫生问题的重要人兽共患病病原菌，引发脑膜炎、关节炎和败血症等临床症状。磷酸泛酰半胱氨酸脱羧酶(phosphopantothenoylcyteine decarboxylase, PPCDC)在前期构建的猪体外血脑屏障模型与猪链球菌全基因组噬菌体随机展示文库的相互作用中被筛选出来，但其影响血脑屏障的机制尚不清楚。【目的】探究PPCDC在SS2诱导脑膜炎过程中发挥的作用。【方法】利用大肠杆菌(Escherichia coli)表达系统诱导表达重组蛋白rPPCDC，通过体外构建人脑微血管内皮细胞单层屏障模型和体内小鼠感染模型探究其对血脑屏障通透性的影响，制备rPPCDC疫苗进行小鼠免疫保护试验。【结果】成功纯化出可溶性重组蛋白rPPCDC，发现其在体外可以降低单层屏障模型的细胞跨膜电阻值，增加突破单层屏障的SS2数量。体内可促进SS2对小鼠的致病性。进一步研究发现，接种rPPCDC疫苗的小鼠血清中有更高的rPPCDC抗体水平，存活率升高。【结论】PPCDC能够增强血脑屏障通透性，促进SS2对小鼠血脑屏障的破坏和脑部感染，并具有免疫保护效力，为后续SS2致病机制研究和疫苗研发提供新思路。

Abstract:

[Background] Streptococcus suis type 2 (SS2) is a major zoonotic pathogen leading to global economic and public health problems, causing meningitis, arthritis, and sepsis. Phosphopantothenoylcysteine decarboxylase (PPCDC) was screened out in the previous study about the interaction between the porcine blood-brain barrier model in vitro and S. suis genome-wide phage display random library, while the mechanism of PPCDC in affecting the blood-brain barrier remains unclear. [Objective] To explore the role of PPCDC in the SS2-induced meningitis. [Methods] In this study, the recombinant protein rPPCDC was expressed in Escherichia coli. The monolayer barrier model of human brain microvascular endothelial cells in vitro and the mouse infection model in vivo were established to study the effect of rPPCDC on the permeability of blood-brain barrier. Furthermore, the rPPCDC vaccine was prepared for the immunoprotection test in mice. [Results] The soluble recombinant protein rPPCDC was successfully purified. This protein reduced the transmembrane resistance of cells in the monolayer barrier model, increased the number of SS2 breaking through the monolayer barrier in vitro. In addition, the protein enhanced the pathogenicity of SS2 in mice. The mice vaccinated with rPPCDC had higher serum levels of rPPCDC antibodies and increased survival. [Conclusion] PPCDC can enhance the permeability of blood-brain barrier and promote the destruction of blood-brain barrier and brain infection of SS2 in mice, thus exerting the immunoprotective effect. The findings provide new ideas for the future research on the pathogenic mechanism of SS2 and the development of vaccines.