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Phagocytosis Assays for Borrelia burgdorferi
Juan Anguita1,2, Ana Carreras-González3, Nicolás Navasa3
1CIC bioGUNE, Macrophage and Tick Vaccine Laboratory, Parque Tenolágico de Bizkaia, 48160, Derio, Bizkaia, Spain. janguita@cicbiogune.es.
Abstract:
Phagocytosis of Borrelia burgdorferi, the causative agent of Lyme disease, is a poorly understood process, despite its importance during the host immune response to infection. Thus, macrophages infiltrate the infected tissues, including the base of the heart and phagocytose the spirochete, therefore contributing to their elimination from infected tissues and leading to inflammation. An impaired bacterial clearance will result in bacterial persistence that may interfere with normal physiology of the heart, such as electrical signals from the heart, resulting in an impaired coordination of the beating of the heart or "heart block." This chapter presents a protocol for establishing primary mouse macrophage cultures, a method for lentivirus silencing of primary cells, and a method for the in vitro study of macrophage phagocytosis of fluorescently labeled Borrelia burgdorferi.
Insights
Understanding how macrophages engulf Borrelia burgdorferi (Lyme disease bacteria) is crucial. This study details methods to investigate this process, aiding in understanding Lyme disease pathogenesis and potential treatments.
Area of Science:
- Immunology
- Microbiology
- Cell Biology
Background:
- Phagocytosis of Borrelia burgdorferi, the Lyme disease agent, is critical for immune response but poorly understood.
- Macrophages clear Borrelia burgdorferi from infected tissues, but impaired clearance leads to persistent infection and potential heart complications like heart block.
Purpose of the Study:
- To present a comprehensive protocol for studying macrophage phagocytosis of Borrelia burgdorferi in vitro.
- To establish primary mouse macrophage cultures and methods for lentivirus-mediated gene silencing.
Main Methods:
- Establishment of primary mouse macrophage cultures.
- Lentivirus-mediated gene silencing in primary macrophages.
- In vitro assessment of macrophage phagocytosis using fluorescently labeled Borrelia burgdorferi.
Main Results:
- The study provides a detailed protocol for investigating macrophage-Borrelia burgdorferi interactions.
- Methods for cell culture and genetic manipulation are established for further research.
Conclusions:
- The presented methods facilitate in-depth study of the phagocytic process involving Borrelia burgdorferi and macrophages.
- This research contributes to understanding Lyme disease pathogenesis and host-pathogen interactions.