Investigating the evolutionary conservation of inflammatory cell migration in vivo
Abstract
Inflammation is a complex, dynamic process, which despite extensive research, is still
far from fully understood. The use of model organisms provides unique insights into
the mechanisms underlying inflammatory cell recruitment.
Previous work using the
fruit fly Drosophila melanogaster has demonstrated that following laser-induced
epithelial wounding, hydrogen peroxide is rapidly produced, and it is this early damage signal that leads to an inflammatory response, via direct activation of the redox sensitive tyrosine kinase Src42a within responding macrophages. Src42a
phosphorylates Draper, the key damage receptor in fly macrophages, which in turn
recruits a downstream kinase, Shark, resulting in macrophage recruitment to wounds.
More recently, the phosphatase Pez has been demonstrated to interact with both
Src42a and Draper and is required for efficient inflammatory macrophage recruitment
to sites of damage in vivo. Importantly, macrophages are insensitive to tissue damage
until they become ‘primed’, by engulfment of an apoptotic corpse, leading to an
increase in Draper expression.
Given macrophage insensitivity to tissue damage prior to priming, we utilised the
tractable genetics of Drosophila and undertook RNA sequencing of both primed and
un-primed cells in vivo, identifying many differentially expressed genes. Selected
candidates were used in macrophage specific, RNAi knock down wounding studies,
revealing novel players in the regulation of macrophage recruitment to wounds.
The mechanism underlying Pez signalling within the macrophage is poorly
understood; we utilised live imaging in Drosophila embryos to investigate Pez
intracellular dynamics during inflammatory cell migration. Microtubule mediated
transport and clathrin mediated endocytosis have been excluded, however, we show
that dynamic Pez puncta correlate with retrograde actin flow providing a potential
mechanism for the regulation of Pez trafficking in macrophages in vivo.
Evolutionary conservation of Pez and Draper signalling in modulating wound induced,
inflammatory cell migration has previously been demonstrated using zebrafish larvae
in a tail fin transection model. Here, we show that the Pez homologue in mouse,
PTPN21, and the murine Draper homologues, MEGF11 and PEAR1, play a role in
blood monocyte and tissue resident macrophage population dynamics through the use
of congenic CD45 bone marrow chimeric mice. Specifically, PEAR1 knock out leads
to a decrease in circulating classical monocytes, whilst PTPN21, MEGF11 and PEAR1
knock out leads to a decrease in alveolar macrophage, but not peritoneal macrophage,
numbers.
The scope for inflammation research is almost unlimited and in order to further dissect the cellular processes that underlie the inflammatory response in vivo, model
organisms must be utilised. Here, we have demonstrated that different model
organisms can be exploited in a synergistic manner, facilitating experiments and
answering questions that would be impossible to undertake using a single system.