Bacterial Lipoproteins Can Disseminate from the Periphery to Inflame the Brain

Abstract

The current view is that bacteria need to enter the brain to cause inflammation. However, in mice infected with the spirochete Borrelia turicatae, we observed widespread cerebral inflammation despite a paucity of spirochetes in the brain parenchyma at times of high bacteremia. Here we studied the possibility that bacterial lipoproteins may be capable of disseminating from the periphery across the bloodbrain barrier to inflame the brain. For this we injected normal and infected mice intraperitoneally with lanthanide-labeled variable outer membrane lipoproteins of B. turicatae and measured their localization in blood, various peripheral organs, and whole and capillary-depleted brain protein extracts at various times. Lanthanide-labeled nonlipidated lipoproteins of B. turicatae and mouse albumin were used as controls. Brain inflammation was measured by TaqMan RT-PCR amplification of genes known to be upregulated in response to borrelial infection. The results showed that the two lipoproteins we studied , LVsp1 and LVsp2 , were capable of inflaming the brain after intraperitoneal injection to different degrees: LVsp1 was better than LVsp2 and Bt1 spirochetes at moving from blood to brain. The dissemination of LVsp1 from the periphery to the brain occurred under normal conditions and significantly increased with infection. In contrast, LVsp2 disseminated better to peripheral organs. We conclude that some bacterial lipoproteins can disseminate from the periphery to inflame the brain. The traditional belief is that bacteria need to cross the blood brain barrier (BBB) to cause brain inflammation. However, during studies in mice persistently infected with the relapsing fever spirochete Borrelia turicatae, we noticed widespread cerebral microgliosis that could not be explained by the presence of spirochetes in the brain parenchyma.

IL-10 Prevents Apoptosis of Brain Endothelium during Bacteremia

“…In this research, we studied whether the major outer membrane lipoprotein of Bt2, LVsp2, was sufficient to cause apoptosis of HBMEC (14,27,32). HBMEC were incubated with increasing amounts of LVsp2 for various times and examined for the presence of apoptosis by Annexin V and PI staining using FACS.…”

“…After incubation at 55˚C for 1 h, the lysis mixture was centrifuged at 20,000 3 g for 60 min at 35˚C, and the supernatant containing outer membrane proteins was dialyzed using Slide-A-Lyzer Dialysis Cassettes (catalog number 66380; Pierce) and 2 l distilled H 2 O at 4˚C for 48 h. Outer membrane proteins were concentrated by centrifugation using Amicon ultracentrifugal filter devices (catalog number UFC903008; Millipore), and the protein concentration was measured using the BCA Protein Assay Kit (catalog number 23227; Pierce). SDS-PAGE and Western blot analysis were used to assess the identity and purity of the Vsp preparations (14,27,28). The outer membrane protein extract from Bt2 was found by Coomassie brilliant blue-stained SDS-PAGE to contain single bands of the expected m.w.…”

“…Variable surface lipoprotein 2 (LVsp2), the major outer membrane lipoproteins of Bt2, was prepared by detergent extraction, followed by purification as described previously (9,14,25,28). Briefly, suspensions containing ∼5 3 10 9 spirochetes were centrifuged at 8000 3 g for 20 min, washed in 30 ml Tris-EDTA buffer (50 mM Tris [pH 7.6] and 5 mM EDTA), and centrifuged a second time, and the pellet was resuspended in 20 ml lysis and solubilization solution consisting of 2% octylglucopyranoside (O8001; Sigma-Aldrich) and 50 mM DTT in Tris-NaCl-EDTA buffer (50 mM Tris [pH 7.6], 150 mM NaCl, and 5 mM EDTA) (28).…”

“…Our laboratory has been studying the neurologic complications of bacterial infection by using a neurotropic strain of the RF spirochete Borrelia turicatae (9)(10)(11)(12)(13)(14). Several previous studies of relapsing-remitting or persistent high-level infection in mice failed to find any evidence of cerebrovascular injury (9)(10)(11)(12)(15)(16)(17).…”

Peptidoglycan-associated lipoprotein ofAggregatibacter actinomycetemcomitansinduces apoptosis and production of proinflammatory cytokines via TLR2 in murine macrophages RAW 264.7in vitro

“…We previously demonstrated through an ex- vivo study design that living A. actinomycetemcomitans cells release free-soluble AaPAL [14,27]. Similar to other bacterial lipoproteins [46,47], it can be expected that this small molecule can cross blood vessel barriers while the bacteria remain growing locally. As most data of the systemic effects of bacterial components in periodontitis are derived from studies on LPS, our results convey knowledge of the pro-inflammatory capability of another conserved outer membrane component in periodontal species.…”