Session:

Bacterial Infections

Abstract No.:

45.064

Title:

A possible role of ICAM-1 in OspA mediated borrelial adhesion to BMEC surface

Author(s):

L. Pulzova¹, E. Bencurova², R. Mucha³, P. Mlynarcik², T. Csank¹, M. Bhide¹; ¹University of veterinary medicine and pharmacy in Kosice, Department of microbioligy and immunology, Kosice, SR/SK, ²University of veterinary medicine and pharmacy in Kosice, Department of microbiology and immunology, Kosice, SR/SK, ³Slovak Academy of Sciences, Institute of Neuroimmunology, Bratislava/SK

Abstract:

Background: Neuroborreliosis can arise at any time during the course of Lyme disease. Neurological manifestations are attributed in part to penetration of the blood-brain barrier (BBB) and invasion of the central nervous system. The main prerequisite for successful BBB translocation is stationary adhesion to surface of endothelial cells (BMECs) mediated by ligand:receptor interactions. Our previous studies showed that borrelial OspA is able to interact with host CD40 (1) and upregulate ICAM-1. It can be hypothesized that ICAM-1 can be another potential candidate for borrelial BMEC stationary adhesion.
Methods: To elucidate ICAM-1 contribution in BBB adhesion process, His-tagged ICAM-1 of rat was overexpressed in Saccharomyces cerevisiae andpurified with metal affinity chromatography. Further, His-tagged ICAM-1 was immobilized on magnetic-beads (MB-IMAC, Bruker) and hybridized with whole cell lysate of neuroinvasive (SKT-7.1) Borrelia strain. After washing of unbound proteins, ICAM-1 and its ligand/s were eluted from beads and proteins were analyzed with MALDI-mass spectrometry. Results showed that ICAM-1 interacted with one protein candidate of the Borrelia. This borrelial ligand was further identified as OspA with the help of peptide mass fingerprinting. Binding ability of OspA of SKT-7.1 to ICAM-1 was further confirmed by pull down assay based on His-tagged OspA. To demonstrate importance of ICAM-1 in adhesion of Borrelia on BMEC, in situ, BMEC monolayer was incubated first with antibodies against ICAM-1 and then with SKT-7.1. Inhibition of the adhesion was compared to positive control wherein BMECs were not blocked with anti-ICAM-1 antibodies.
Results: Results approved that borrelial adhesion to BMEC can be mediated via OspA:ICAM-1 dyad. Blocking of ICAM-1 on BMECs reduces borrelil adhesion by 82% compared to control.
Conclusion: Taken together, study demonstrated that OspA provides Borrelia with an essential function in adhesion to endothelium. It can be concluded that adhesion process is mediated not only via OspA:CD40 dyad (1) but also via OspA:ICAM-1, and this interaction may plays an intrinsic role in borrelial neuroinvasion.Acknowledgements: Work was performed in collaboration with Dr. E. Chakurkar, at ICAR Goa India, mainly for transfection and in-vivo protein production. Financial support was from APVV-0036-10 and VEGA - 2/0121/11. Reference:  Lucia Pulzova, et al., Nature Scientific Reports 1, 2011.