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Analysis of Lymphocyte Extravasation Using an In Vitro Model of the Human Blood-brain Barrier
Published on: April 5, 2017
TLR4 induces CCR7-dependent monocytes transmigration through the blood-brain barrier
Alexandre Paradis1, Stéphane Bernier1, Nancy Dumais1
1Département de Biologie, Faculté des Sciences, Université de Sherbrooke, Sherbrooke, QC J1K 2R1, Canada.
Abstract:
In this study, we examined whether bacterial pathogen-associated molecular patterns recognized by toll-like receptors (TLRs) can modify the CCR7-dependent migration of human monocytes. MonoMac-1 (MM-1) cells and freshly isolated human monocytes were cultivated in the presence of agonists for TLR4 (which senses lipopolysaccharides from gram-negative bacteria), TLR1/2 (which senses peptidoglycan from gram-positive bacteria), and TLR9 (which recognizes bacterial DNA rich in unmethylated CpG DNA). CCR7 mRNA transcription was measured using quantitative reverse transcription polymerase chain reaction and protein expression was examined using flow cytometry. CCR7 function was monitored using migration and transmigration assays in response to CCL19/CCL21, which are natural ligands for CCR7. Our results show that TLR4 strongly increases monocyte migratory capacity in response to CCL19 in chemotaxis and transmigration assays in a model that mimics the human blood-brain barrier, whereas TLR1/2 and 9 have no effect. Examination of monocyte migration in response to TLRs that are activated by bacterial components would contribute to understanding the excessive monocyte migration that characterizes the pathogenesis of bacterial infections and/or neuroinflammatory diseases.
Insights
Toll-like receptor 4 (TLR4) activation significantly enhances human monocyte migration toward CCL19, crucial for neuroinflammation. Other TLRs studied (TLR1/2, TLR9) did not affect this monocyte migration.
Area of Science:
- Immunology
- Neuroscience
- Microbiology
Background:
- Toll-like receptors (TLRs) recognize pathogen-associated molecular patterns, initiating immune responses.
- Monocyte migration, guided by chemokine receptor CCR7, is critical in inflammatory diseases.
- Understanding how TLRs influence monocyte migration is key to addressing bacterial infections and neuroinflammation.
Purpose of the Study:
- To investigate the impact of bacterial pathogen-associated molecular patterns, sensed by TLRs, on CCR7-dependent human monocyte migration.
- To determine if TLR4, TLR1/2, and TLR9 activation affects monocyte migration in response to CCL19/CCL21.
Main Methods:
- Human monocytes (MonoMac-1 cells and primary cells) were stimulated with agonists for TLR4, TLR1/2, and TLR9.
- CCR7 messenger RNA (mRNA) and protein expression were quantified using RT-qPCR and flow cytometry.
- Monocyte migration and transmigration assays were performed in response to CCL19/CCL21, including a blood-brain barrier model.
Main Results:
- TLR4 activation significantly enhanced monocyte migratory capacity towards CCL19 in chemotaxis and transmigration assays.
- TLR4-mediated enhancement of migration was observed in a human blood-brain barrier model.
- TLR1/2 and TLR9 activation did not significantly affect CCR7-dependent monocyte migration.
Conclusions:
- TLR4 activation plays a critical role in augmenting monocyte migration, particularly in contexts relevant to neuroinflammation.
- These findings contribute to understanding excessive monocyte infiltration in bacterial infections and neuroinflammatory conditions.
- Targeting TLR4 signaling could offer therapeutic strategies for managing inflammatory diseases involving monocyte recruitment.
