Neutrophils on the hunt : Migratory strategies employed by neutrophils to fulfill their effector function

Thesis

Abstract

Acknowledgments

About the Author

List of Publications

Table of Contents

List of Figures

List of Tables

List of Symbols/Abbreviations

1 General Introduction

2 Run and Fumble - What Eukaryotes Learnt from Bacteria

3 Make Your Body Count - How Local Environment can Help Neutrophils Shape the Local Gradient

4 Additional Work

5 Materials and Methods

6 References

7 Appendix

During my Ph.D. research, I managed a series of projects, each focused on the
mechanisms underlying cell migration. My work involved an in-depth examination of
the complex strategies employed by neutrophils, with a specific focus on their ability to
synchronize spatial-temporal cues and optimize their gradient perception. However, it
is essential to acknowledge that not all projects yielded successful results, as some
ideas were discontinued and are archived for future reference within this thesis.
My main project investigated how neutrophils decode spatial cues for precise navigation. Human neutrophils showcased distinct movement patterns based on source
type – linear or point-like. By combining single-cell tracking in 3D environments with
proxy dyes, this project linked cell behaviors to gradient changes, revealing a stronger
response to semi-exponential gradients from point sources. In addition, neutrophils
exhibited oscillating migration speeds, using speed minima to adjust trajectories toward sources. Experiencing continuous concentration changes, they accelerated over
time and employed a "Run and Fumble" strategy, alternating between consistent runs
and strategic "tumbles" for efficient navigation.
The project extended to the possibility of cells amplifying perceived gradients by
enclosing their immediate surroundings, pushing attractants forward for enrichment
while depleting it at the cell rear. Microfluidic devices were employed, and various experimental parameters configurations were optimized. Although significant differences
in migratory efficacy were detected across pore sizes and device heights, quantifying
gradient manipulation effects proved challenging.
The "Laser-Assisted Protein Adsorption by Photobleaching" (LAPAP) project was
promising, as it allowed the printing of gradients. Initially successful with dendritic cells,
we aimed to adapt it for neutrophils. Through extensive experimentation with multiple
parameters, we attempted to trigger responses from neutrophils. Despite these efforts
and collaboration, the project failed due to practical challenges and limitations.
Facing a lack of neutrophil-like cells at IST, we initially established the SCF-HoxB8
primary murine cell line. Despite their existence, their migratory behavior was largely
unexplored due to potential limitations. Through differentiation protocol refinements we
enhanced their migratory capabilities, though their capacity still lagged behind human
neutrophils. Despite this, the improved migration potential of these cells pointed toward
their utility for in vitro murine neutrophil migration studies.