Acute radiation syndrome (ARS) is caused by acute exposure to ionizing radiation that damages multiple organ systems but especially the bone marrow (BM). Researchers from the University of Wisconsin School of Medicine and Public Health have previously shown that human macrophages educated with exosomes from human BM-derived mesenchymal stromal cells (MSCs) primed with lipopolysaccharide (LPS) prolonged survival in a xenogeneic lethal ARS model. The purpose of this study was to determine if exosomes from LPS-primed MSCs could directly educate human monocytes (LPS-EEMos) for the treatment of ARS.
Human monocytes were educated by exosomes from LPS-primed MSCs and compared to monocytes educated by unprimed MSCs (EEMos) and uneducated monocytes to assess survival and clinical improvement in a xenogeneic mouse model of ARS. Changes in surface molecule expression of exosomes and monocytes after education were determined by flow cytometry, while gene expression was determined by qPCR. Irradiated human CD34+ hematopoietic stem cells (HSCs) were co-cultured with LPS-EEMos, EEMos, or uneducated monocytes to assess effects on HSC survival and proliferation.
Treatment with LPS-EEMos educated for 3 days or 24 h significantly improves survival and clinical scores in mice with lethal ARS when infused 4 h after radiation exposure
On day 0, NSG mice received 4 Gy of lethal radiation followed by an i.v. treatment 4 h later with PBS (vehicle control), 1 × 107 EEMos, or 1 × 107 LPS-EEMos. A Survival curve of treated mice after radiation. B Mean clinical scores (percent weight loss, posture, activity, and fur texture) C Mean percent weight change. D On day 0, NSG mice received 4 Gy of lethal radiation followed by an i.v. treatment 4 h later with PBS (vehicle control), 1 × 107 EEMos, or 1 × 107 LPS-EEMos. Survival curve of treated mice after radiation. E Mean clinical scores. F Mean percent weight change. The final mean percent weight change and clinical score were carried over after death to allow for comparison between groups at a given time point by Kruskal-Wallis with a Dunn post-test. Results pooled from three separate experiments, with 10 to 14 mice/group, for survival analysis. A representative clinical score and weight change from one of 3 experiments are shown, with 2 to 6 mice/group. *p ≤ 0.05, **p ≤ 0.01
LPS priming of MSCs led to the production of exosomes with increased expression of CD9, CD29, CD44, CD146, and MCSP. LPS-EEMos showed increases in gene expression of IL-6, IL-10, IL-15, IDO, and FGF-2 as compared to EEMos generated from unprimed MSCs. Generation of LPS-EEMos induced a lower percentage of CD14+ monocyte subsets that were CD16+, CD73+, CD86+, or CD206+ but a higher percentage of PD-L1+ cells. LPS-EEMos infused 4 h after lethal irradiation significantly prolonged survival, reducing clinical scores and weight loss as compared to controls. Complete blood counts from LPS-EEMo-treated mice showed enhanced hematopoietic recovery post-nadir. IL-6 receptor blockade completely abrogated the radioprotective survival benefit of LPS-EEMos in vivo in female NSG mice, but only loss of hematopoietic recovery was noted in male NSG mice. PD-1 blockade had no effect on survival. Furthermore, LPS-EEMos also showed benefits in vivo when administered 24 h, but not 48 h, after lethal irradiation. Co-culture of unprimed EEMos or LPS-EEMos with irradiated human CD34+ HSCs led to increased CD34+ proliferation and survival, suggesting hematopoietic recovery may be seen clinically.
LPS-EEMos are a potential counter-measure for hematopoietic ARS, with a reduced biomanufacturing time that facilitates hematopoiesis.