Recovery from hematopoietic injury by modulating prostaglandin E2 signaling post-irradiation

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Abstract

While high dose total body irradiation (TBI) is used therapeutically, the proliferation of nuclear weapons, increasing use of nuclear power, and worldwide radical terrorism underscore the need to develop countermeasures to a radiological mass casualty event. The hematopoietic syndrome of the acute radiation syndrome (HS-ARS) results from severe compromise to the hematopoietic system, including lymphocytopenia, neutropenia, thrombocytopenia, and possible death from infection and/or hemorrhage. Given adequate time to recover, expand, and appropriately differentiate, bone marrow hematopoietic stem cells (HSC) and progenitor cells (HPC) may overcome HS-ARS and restore homeostasis of the hematopoietic system. Prostaglandin E2 (PGE2) has been shown to have pleiotropic effects on hematopoiesis, acting to inhibit apoptosis and promote self-renewal of HSC, while inhibiting HPC proliferation. We assessed the radio-mitigating potential of modulating PGE2 signaling in a mouse model of HS-ARS. Treatment with the PGE2 analog 16,16 dimethyl PGE2 (dmPGE2) 6 h post-irradiation or inhibition of PGE2 synthesis via delayed administration of the non-steroidal anti-inflammatory drug (NSAID) Meloxicam resulted in increased survival of lethally irradiated mice. Both early dmPGE2 and delayed Meloxicam treatment were associated with increased HPC activity 35 days following irradiation, demonstrating enhanced recovery of hematopoiesis. Our results define two different treatment modalities that are highly effective and safe to administer, and can be readily available.

Introduction

With the proliferation of nuclear weapons, increasing use of nuclear power and the advent of worldwide radical terrorism, there is an increasing need and research emphasis on developing countermeasures in the event of a radiological mass casualty event [33], [36], [47]. Radiation accidents at Chernobyl and Fukushima serve as examples of the complexities of containment and triage even in the absence of a nuclear detonation. The highly proliferative nature (production of upwards of a trillion cells per day) of the hematopoietic system [35] required to maintain homeostasis and respond to stress demands make them highly radiosensitive [4], [9], [10], [17], [53]. Substantive damage to bone marrow causes the hematopoietic syndrome of the acute radiation syndrome (HS-ARS), with subsequent hematologic compromise affecting systemic oxygenation, nutrient delivery and detoxification, vascular integrity and anti-infective capacity. Survival, self-renewal, proliferation, differentiation, and migration of HSC and HPC are regulated by interacting networks of cytokines, chemokines, other regulatory molecules and the bone marrow microenvironment [6], [50] and successful radiation countermeasures will need to account for the hematopoietic system. Life threatening effects of radiation exposure result from DNA and other cellular damage that triggers processes leading to modification of cell cycle checkpoints, arrest/repair and apoptosis, influencing genomic stability or epigenetic processes in descendent cells, and from indirect (bystander) toxic effects mediated via abnormal microenvironmental nurturing and effector functions [4], [11], [12], [31], [53].

HS-ARS is characterized by life-threatening lymphocytopenia, neutropenia, and thrombocytopenia, and possible death due to infection and/or bleeding. Doses < 2 Gy do not cause significant bone marrow damage [2]. However, at 2–8 Gy the acute radiation syndrome develops proportional to radiation dose, resulting in cytopenias and marrow failure in ensuing weeks [9], [17], [54], and without treatment, results in the sequelae of infection, bleeding, deficient wound healing, and even death [11], [12]. While bone marrow HSC and HPC are susceptible to radiation exposure, surviving populations of these cells can recover hematopoiesis, given time to repair DNA damage, self-renew, expand and differentiate. Allogeneic hematopoietic transplant is not considered a viable or practical treatment for HS-ARS [3], necessitating the need for development of alternative approaches to treat affected individuals. The unpredictability of a mass casualty radiation event requires development and utilization of post exposure mitigators of radiation injury with appropriate ease of administration, stability for purposes of stockpiling, ability for rapid distribution and a window of efficacy. In addition, faced with the complexities of a mass casualty event and difficulty of individual dosimetry and triage, interventions that can mitigate or reduce the severity of exposure, but that are benign to those individuals with limited or no exposure, are required.

We have previously reported on the positive effects on HSC by prostaglandin E2 (PGE2) treatment, both decreasing apoptosis through up-regulation of the endogenous anti-apoptotic protein Survivin and increasing self-renewal division and homing/engraftment in the bone marrow [23]. In addition, previous work from our laboratory demonstrated that PGE2 dose-dependently inhibits mouse and human myeloid progenitor proliferation in semisolid culture assays [38], [39] and that in vivo administration of PGE inhibits HPC proliferation [14], [15], while inhibition of PGE synthesis in vivo enhances HPC number [37]. Based on our previous findings of the pleiotropic effects of PGE2 signaling, we hypothesized that administration of PGE2 early post-irradiation exposure would enhance survival and self-renewal of HSC and HPC, while inhibition of cyclooxygenase (COX) with non-steroidal anti-inflammatory drugs (NSAIDs), which inhibit PGE2 biosynthesis, later post-irradiation would abrogate PGE2 inhibition of myelopoiesis, leading to marrow myeloid HPC expansion. Here, we tested this hypothesis and show that treatment with 16,16 dimethyl-PGE2 (dmPGE2), a long acting derivative of PGE2, shortly following irradiation or delayed post-irradiation administration of the NSAID Meloxicam significantly enhance animal survival and hematopoietic recovery.

Section snippets

Mice

Male and female C57Bl/6 mice were purchased from Jackson Laboratories at 10–12 weeks of age. Mice were housed in microisolator cages (5 mice per cage) with sterilized direct contact bedding (Alpha Dri). Animal holding rooms were maintained at 21 ± 3 °C with 30–80% relative humidity, with at least 10 air changes per hour of 100% fresh air, and a 12 h light/dark cycle. Mice were fed ad libitum with commercial rodent chow (Harlan 2018SXC) in cage hoppers and acidified (pH 2.0–3.0) water in sipper tube

PGE2 treatment increases survival post-irradiation

PGE2 biosynthesis is increased following γ-radiation and can result from up-regulation of cytoplasmic phospholipase A2 (cPLA2) [8] or COX activity [24]. In rats, spinal cord irradiation elevates PGE2 levels within 3–24 h that persists for 3 days [52]. In mice, brain irradiation induces COX-dependent PGE2 production and elevated levels of PGE synthases [32]. In breast cancer patients, radiation therapy triggers monocyte PGE2 production [7] and in leukemia and lymphoma patients undergoing

Discussion

These studies outline two different pharmaceutical strategies for radio-mitigation. In the context of a radiation incident, particularly one in a densely populated area where there are numerous affected (and non-affected) individuals, strategies that can be employed quickly and safely to the masses are ideal. While hematopoietic transplantation may be able to treat patients after exposure to radiation, the logistics and timing involved in a mass casualty situation make transplantation

Acknowledgments

These studies were supported by NIH grant HL096305 (LMP) and contract HHSN272201000046C (CMO). Flow cytometry was performed in the Flow Cytometry Resource Facility of the Indiana University Simon Cancer Center (NCI P30 CA082709). Additional core support was provided by the Center of Excellence in Hematology grant P01 DK090948. JH and KNS were supported by Training Grant HL007910.

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