Signaling through TLR5 mitigates lethal radiation damage by neutrophil-dependent release of MMP-9

Craig M Brackett,Scott I Abrams,Ivan Molodtsov,Bojidar M Kandar,Lyudmila G Burdelya,Nicholas H Trageser,Srabani Pal,Kellee F Greene,Andrei V Gudkov,Alyssa R Aldrich

doi:10.1038/s41420-021-00642-6

Craig M Brackett, Scott I Abrams + Show 8 more

Open Access

https://doi.org/10.1038/s41420-021-00642-6

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Abstract

Acute radiation syndrome (ARS) is a major cause of lethality following radiation disasters. A TLR5 agonist, entolimod, is among the most powerful experimental radiation countermeasures and shows efficacy in rodents and non-human primates as a prophylactic (radioprotection) and treatment (radiomitigation) modality. While the prophylactic activity of entolimod has been connected to the suppression of radiation-induced apoptosis, the mechanism by which entolimod functions as a radiomitigator remains poorly understood. Uncovering this mechanism has significant and broad-reaching implications for the clinical development and improvement of TLR5 agonists for use as an effective radiation countermeasure in scenarios of mass casualty resulting from accidental exposure to ionizing radiation. Here, we demonstrate that in contrast to radioprotection, neutrophils are essential for the radiomitigative activity of entolimod in a mouse model of lethal ARS. Neutrophils express functional TLR5 and rapidly exit the bone marrow (BM), accumulate in solid tissues, and release MMP-9 following TLR5 stimulation which is accompanied by an increase in the number of active hematopoietic pluripotent precursors (HPPs) in the BM. Importantly, recombinant MMP-9 by itself has radiomitigative activity and, in the absence of neutrophils, accelerates the recovery of the hematopoietic system. Unveiling this novel TLR5-neutrophil-MMP-9 axis of radiomitigation opens new opportunities for the development of efficacious radiation countermeasures to treat ARS following accidental radiation disasters.

Highlights

Exposure to ionizing radiation causes significant DNA damage in many cells of the organism and is the underlying cause of multiple pathologies including acute radiation sickness (ARS)
The remarkable efficacy of TLR5 agonists as a radiomitigator has been recognized for many years, but a lack of mechanistic knowledge regarding which components of this multifunctional pathway rescue lethally irradiated organisms has significantly impeded the clinical development of TLR5 agonists as a radiation countermeasure following accidental exposure to ionizing radiation
We provide evidence supporting the key role for the release of MMP-9 by TLR5-stimulated Nφ as an essential mediator of radiomitigation

Summary

INTRODUCTION

Exposure to ionizing radiation causes significant DNA damage in many cells of the organism and is the underlying cause of multiple pathologies including acute radiation sickness (ARS). Transferred Nφ were improving HPS recovery and the release of matrix released from the BM and recruited to the liver and lungs, two of metalloproteinase-9 (MMP-9) by Nφ has been linked to stimulating the most prominent sites of recruitment post-entolimod treathematopoiesis [21], we hypothesized that entolimod mitigates ment (Fig. 2B) These results indicate that entolimod stimulates Nφ radiation damage to the HPS through Nφ-dependent release of release from the BM and recruitment to tissues through a blood-. Since Nφ differentiation occurs across a linear path from pre-Nφ radiomitigative but not radioprotective abilities of entolimod, (ii) and to immature and mature Nφ [23], we sought to express functional TLR5 but undergo minimal transcriptional understand whether entolimod stimulates the release of certain changes post-entolimod suggesting that Nφ mitigate ARS through Nφ populations from the BM. “sandwich” platform that utilizes biotinylated-entolimod (b-entolimod) and a streptavidin-conjugated fluorochrome in combination with antibodies to identify which immune cell subsets express

RESULTS

DISCUSSION

MATERIALS AND METHODS