Hematological Changes as Prognostic Indicators of Survival: Similarities Between Gottingen Minipigs, Humans, and Other Large Animal Models

The animal efficacy rule addressing development of drugs for selected disease categories has pointed out the need to develop alternative large animal models. Based on this rule, the pathophysiology of the disease in the animal model must be well characterized and must reflect that in humans. So far, manifestations of the acute radiation syndrome (ARS) have been extensively studied only in two large animal models, the non-human primate (NHP) and the canine. We are evaluating the suitability of the minipig as an additional large animal model for development of radiation countermeasures. We have previously shown that the Gottingen minipig manifests hematopoietic ARS phases and symptoms similar to those observed in canines, NHPs, and humans.We establish here the LD50/30 dose (radiation dose at which 50% of the animals succumb within 30 days), and show that at this dose the time of nadir and the duration of cytopenia resemble those observed for NHP and canines, and mimic closely the kinetics of blood cell depletion and recovery in human patients with reversible hematopoietic damage (H3 category, METREPOL approach). No signs of GI damage in terms of diarrhea or shortening of villi were observed at doses up to 1.9 Gy. Platelet counts at days 10 and 14, number of days to reach critical platelet values, duration of thrombocytopenia, neutrophil stress response at 3 hours and count at 14 days, and CRP-to-platelet ratio were correlated with survival. The ratios between neutrophils, lymphocytes and platelets were significantly correlated with exposure to irradiation at different time intervals.As a non-rodent animal model, the minipig offers a useful alternative to NHP and canines, with attractive features including ARS resembling human ARS, cost, and regulatory acceptability. Use of the minipig may allow accelerated development of radiation countermeasures

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference

Differences in dispersion of influenza virus lipids and proteins during fusion

Digitally enhanced low-light-level fluorescence video microscopy and immunochemical staining were used to examine influenza virus envelope lipid and protein redistribution during pH-induced fusion. Video microscopy was performed using viruses labeled with either the lipid analogue octadecylrhodamine B (R18) or fluorescein isothiocyanate (FITC) covalently linked to envelope proteins. Viruses were bound to human red blood cells, and the pattern and intensity of fluorescence were monitored for 30 min while cell-virus complexes were perfused with pH 7.4 or 4.8 media at temperatures either above or below 20°C. R18 showed complete redistribution and dequenching by 30 min at all incubation temperatures, confirming reports that viral fusion occurs at subphysiological temperatures. FITC-labeled protein showed spatial redistribution at 28°C but no change at low temperature. Electron microscopy observations of immunochemical staining of viral proteins confirmed both that protein redistribution at 37°C was slower than R18 and the failure of movement within 30 min at 16°C. Video microscopy monitoring of RNA staining by acridine orange of virus-cell complexes showed redistribution to the RBCs at all temperatures but only after low pH-induced fusion. The results are consistent with differential dispersion of viral components into the RBC and the existence of relatively long-lived barriers to diffusion subsequent to fusion pore formation

Evaluation of the Gamma-H2AX Assay for Radiation Biodosimetry in a Swine Model

Abstract: There is a paucity of large animal models to study both the extent and the health risk of ionizing radiation exposure in humans. One promising candidate for such a model is the minipig. Here, we evaluate the minipig for its potential in γ-H2AX-based biodosimetry after exposure to ionizing radiation using both Cs137 and Co60 sources. γ-H2AX foci were enumerated in blood lymphocytes and normal fibroblasts of human and porcine origin after ex vivo �-ray irradiation. DNA double-strand break repair kinetics in minipig blood lymphocytes and fibroblasts, based on the γ-H2AX assay, were similar to those observed in their human counterparts. To substantiate the similarity observed between the human and minipig we show that minipig fibroblast radiosensitivity was similar to that observed with human fibroblasts. Finally, a strong γ-H2AX induction was observed in blood lymphocytes following minipig total body irradiation. Significant responses were detected 3 days after 1.8 Gy and 1 week after 3.8 and 5 Gy with residual γ-H2AX foci proportional to the initial radiation doses. These findings show that the Gottingen minipig provides a useful in viv

Histological examination of crypts and villi in the duodenum of irradiated minipigs (1.6 to 2.0 Gy).

<p>Samples were collected and fixed at necropsy. Panel A (1.6 Gy, H&E 100×). Villar length is consistent throughout the section, and is of normal length, lined by a single layer of columnar epithelium with basilar nuclei and abundant eosinophilic apical cytoplasm. Underlying crypts are densely packed. Panel B (1.9 Gy, H&E 100×). Villar length remains consisted in length and in morphology of the overlying epithelium as well as the abundance and morphology of the underlying villar crypts. Panel C (2.0 Gy, H&E 200×). Multifocally throughout the section, villi are blunted and fused (asterix) with multiple villi recovering by denudation by being covered by a single layer of columnar epithelium. Underlying crypts (arrows) have piled epithelium and occasionally are tortuous; however there is no apparent loss of crypts.</p

Spearman rank correlation (ρ) of ANC counts- and CRP-based parameters with survival in irradiated G. minipig (n = 30).

<p>ANC: absolute neutrophil counts; CRP: C-reactive protein; CRP:plt : CRP-to-platelets ratio; p value (Students' T-test).</p

Compiled published data for hematological dynamics in human, minipig, NHP and canine after irradiation.

<p>REFERENCES: Human data from: ref <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Fliedner3" target="_blank">[33]</a>. Swine data from: ref <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Moroni2" target="_blank">[4]</a>. NHP data from: refs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-MacVittie1" target="_blank">[19]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-MacVittie2" target="_blank">[34]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Herodin1" target="_blank">[40]</a>. Canine data from: refs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Deeg1" target="_blank">[17]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Burstein1" target="_blank">[18]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-MacVittie4" target="_blank">[41]</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025210#pone.0025210-Nothdurft1" target="_blank">[42]</a>.</p