Hemorrhage enhances cytokine, complement component 3, and caspase-3, and regulates microRNAs associated with intestinal damage after whole-body gamma-irradiation in combined injury

Abstract

Hemorrhage following whole-body γ-irradiation in a combined injury (CI) model increases mortality compared to whole-body γ-irradiation alone (RI). The decreased survival in CI is accompanied by increased bone marrow injury, decreased hematocrit, and alterations of miRNA in the kidney. In this study, our aim was to examine cytokine homeostasis, susceptibility to systemic bacterial infection, and intestinal injury. More specifically, we evaluated the interleukin-6 (IL-6)-induced stress proteins including C-reactive protein (CRP), complement 3 (C3), Flt-3 ligand, and corticosterone. CD2F1 male mice received 8.75 Gy 60Co gamma photons (0.6 Gy/min, bilateral) which was followed by a hemorrhage of 20% of the blood volume. In serum, RI caused an increase of IL-1, IL-2, IL-3, IL-5, IL-6, IL-12, IL-13, IL-15, IL-17A, IL-18, G-CSF, CM-CSF, eotaxin, IFN-γ, MCP-1, MIP, RANTES, and TNF-α, which were all increased by hemorrhage alone, except IL-9, IL-17A, and MCP-1. Nevertheless, CI further elevated RI-induced increases of these cytokines except for G-CSF, IFN- γ and RANTES in serum. In the ileum, hemorrhage in the CI model significantly enhanced RI-induced IL-1β, IL-3, IL-6, IL-10, IL-12p70, IL-13, IL-18, and TNF-α concentrations. In addition, Proteus mirabilis Gram(-) was found in only 1 of 6 surviving RI mice on Day 15, whereas Streptococcus sanguinis Gram(+) and Sphingomonas paucimobilis Gram(-) were detected in 2 of 3 surviving CI mice (with 3 CI mice diseased due to inflammation and infection before day 15) at the same time point. Hemorrhage in the CI model enhanced the RI-induced increases in C3 and decreases in CRP concentrations. However, hemorrhage alone did not alter the basal levels, but hemorrhage in the CI model displayed similar increases in Flt-3 ligand levels as RI did. Hemorrhage alone altered the basal levels of corticosterone early after injury, which then returned to the baseline, but in RI mice and CI mice the increased corticosterone concentration remained elevated throughout the 15 day study. CI increased 8 miRNAs and decreased 10 miRNAs in serum, and increased 16 miRNA and decreased 6 miRNAs in ileum tissue. Among the altered miRNAs, CI increased miR-34 in the serum and ileum which targeted an increased phosphorylation of ERK, p38, and increased NF-κB, thereby leading to increased iNOS expression and activation of caspase-3 in the ileum. Further, let-7g/miR-98 targeted the increased phosphorylation of STAT3 in the ileum, which is known to bind to the iNOS gene. These changes may correlate with cell death in the ileum of CI mice. The histopathology displayed blunted villi and villus edema in RI and CI mice. Based on the in silico analysis, miR-15, miR-99, and miR-100 were predicted to regulate IL-6 and TNF. These results suggest that CI-induced alterations of cytokines/chemokines, CRP, and C3 cause a homeostatic imbalance and may contribute to the pathophysiology of the gastrointestinal injury. Inhibitory intervention in these responses may prove therapeutic for CI and improve recovery of the ileal morp...

Ghrelin therapy mitigates bone marrow injury and splenocytopenia by sustaining circulating G-CSF and KC increases after irradiation combined with wound

“…CI increases caspase-3 more than RI in ileum [ 39 , 40 ]. In this study, we found significant decreases in caspase-3 in the bone marrow of RI and CI animals from day 1 through day 15.…”

“…Caspase-3 is a critical protease in caspase-dependent apoptosis [ 49 , 50 ]. It is increased in ileum after irradiation and combined injury [ 40 ]. In bone marrow, no increased activated caspase-3 levels were found, but RI and CI decreased caspase-3 levels on day 3 and continue to decrease the levels on day 15.…”

A novel therapy, using Ghrelin with pegylated G-CSF, inhibits brain hemorrhage from ionizing radiation or combined radiation injury

“…CI amplifies increases in concentrations of cytokines/chemokines in blood and tissues. 19,23,33,[36][37][38] In cerebellum lysates, we found increases in IL-6, KC, Eotaxin, G-CSF, MIP-2, MCP-1, and MIP-1α, but decreases in IL-18 ( Figure 3a). Ghrelin therapy with pegylated G-CSF did not mitigate these increases except G-CSF and MIP-2 (Figure 3a).…”

“…26 It is evident that CI increases MAPK activation. 27 It was not clear whether CI alters AKT activation. Nevertheless, AKT and MAPK are known to be associated with apoptosis.…”

The effects of N-acetylcysteine on radiotherapy-induced small intestinal damage in rats

“…14,15 Activation of the caspase-3 cascade plays a highly important role in apoptosis and is regarded as a terminal event prior to cell death and increases under the effect of radiation. 16–18…”