MST1-dependent vesicle trafficking regulates neutrophil transmigration through the vascular basement membrane
Angela R.M. Kurz, … , Sergio D. Catz, Markus Sperandio
Angela R.M. Kurz, … , Sergio D. Catz, Markus Sperandio
Published October 4, 2016
Citation Information: J Clin Invest. 2016;126(11):4125-4139. https://doi.org/10.1172/JCI87043.
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Research Article Inflammation

MST1-dependent vesicle trafficking regulates neutrophil transmigration through the vascular basement membrane

Abstract

Neutrophils need to penetrate the perivascular basement membrane for successful extravasation into inflamed tissue, but this process is incompletely understood. Recent findings have associated mammalian sterile 20–like kinase 1 (MST1) loss of function with a human primary immunodeficiency disorder, suggesting that MST1 may be involved in immune cell migration. Here, we have shown that MST1 is a critical regulator of neutrophil extravasation during inflammation. Mst1-deficient (Mst1–/–) neutrophils were unable to migrate into inflamed murine cremaster muscle venules, instead persisting between the endothelium and the basement membrane. Mst1–/– neutrophils also failed to extravasate from gastric submucosal vessels in a murine model of Helicobacter pylori infection. Mechanistically, we observed defective translocation of VLA-3, VLA-6, and neutrophil elastase from intracellular vesicles to the surface of Mst1–/– neutrophils, indicating that MST1 is required for this crucial step in neutrophil transmigration. Furthermore, we found that MST1 associates with the Rab27 effector protein synaptotagmin-like protein 1 (JFC1, encoded by Sytl1 in mice), but not Munc13-4, thereby regulating the trafficking of Rab27-positive vesicles to the cellular membrane. Together, these findings highlight a role for MST1 in vesicle trafficking and extravasation in neutrophils, providing an additional mechanistic explanation for the severe immune defect observed in patients with MST1 deficiency.

Authors

Angela R.M. Kurz, Monika Pruenster, Ina Rohwedder, Mahalakshmi Ramadass, Kerstin Schäfer, Ute Harrison, Gabriel Gouveia, Claudia Nussbaum, Roland Immler, Johannes R. Wiessner, Andreas Margraf, Dae-Sik Lim, Barbara Walzog, Steffen Dietzel, Markus Moser, Christoph Klein, Dietmar Vestweber, Rainer Haas, Sergio D. Catz, Markus Sperandio

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Figure 4

Mst1–/– neutrophils fail to penetrate the BM in vivo and in vitro.

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Mst1–/– neutrophils fail to penetrate the BM in vivo and in vitro. (A)...
(A) Immunostaining of TNF-α–stimulated cremaster muscle whole mounts from WT and Mst1–/– mice for laminin α5 (white, BM), ESAM1 (red, endothelial junctions), and MRP14 (green, neutrophils) (n = 3). Scale bar: 20 μm. (B) 3D reconstruction and illustration of a vessel cross section demonstrating the classification of transmigrating neutrophils respective to their position. Position I, neutrophils in intimate contact with the endothelium; position II, neutrophils between the endothelium and the BM; position III, fully transmigrated neutrophils. Scale bar: 10 μm. (C) Distribution pattern of transmigrating neutrophils respective to their positions (positions I–III) in WT and Mst1–/– mice (n = 45 analyzed vessels from 5 mice per group, scatter plot with mean of >25 analyzed vessels per group, ***P < 0.001, **P < 0.01, 2-way ANOVA, Sidak’s multiple comparisons test). (D–F) Neutrophil transmigration in a Transwell assay with or without CXCL1 through filters coated with BSA (control), laminin 1 (LN-1), or LN-1, PECAM-1, and ICAM-1 (LN-1/P/I) (n = 3 mice, triplicate measurements were performed, mean ± SEM, ***P < 0.001, 2-way ANOVA, Sidak’s multiple comparisons test) (D); with or without CXCL1 through an endothelial monolayer of 1G11 cells (n = 3 mice, triplicate measurements were performed, mean ± SEM, ***P < 0.001, 2-way ANOVA, Sidak’s multiple comparisons test) (E); and to HBSS, 10 ng/ml and 100 ng/ml CXCL1 (n = 3 mice, duplicate measurements were performed, mean ± SEM, 2-way ANOVA, Sidak’s multiple comparisons test) (F).