Hematological changes as prognostic indicators of survival: similarities between Gottingen minipigs, humans, and other large animal models

Background

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.

Principal Findings

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.

Significance

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.     

Pathology during acute infections: contributions of intracellular pathogens and the CTL response

Previous work has shown how, in the case of cytotoxic T-lymphocyte (CTL) responses to persistent viral infections, pathology may arise as a consequence of cell destruction directly by the virus or indirectly due to the CTL response, leading to maximum pathology at intermediate efficacy of the immune response. We expand these studies to consider pathology arising during acute infections with intracellular pathogens controlled by the CTL response. We show that, in contrast to persistent infections, pathology during acute infections is minimized with increasing efficacy of the immune response. The implications of these results for vaccination are discussed.     

pSAT vectors: a modular series of plasmids for autofluorescent protein tagging and expression of multiple genes in plants

Autofluorescent protein tags represent one of the major and, perhaps, most powerful tools in modern cell biology for visualization of various cellular processes in vivo. In addition, advances in confocal microscopy and the development of autofluorescent proteins with different excitation and emission spectra allowed their simultaneous use for detection of multiple events in the same cell. Nevertheless, while autofluorescent tags are widely used in plant research, the need for a versatile and comprehensive set of vectors specifically designed for fluorescent tagging and transient and stable expression of multiple proteins in plant cells from a single plasmid has not been met by either the industrial or the academic communities. Here, we describe a new modular satellite (SAT) vector system that supports N- and C-terminal fusions to five different autofluorescent tags, EGFP, EYFP, Citrine-YFP, ECFP, and DsRed2. These vectors carry an expanded multiple cloning site that allows easy exchange of the target genes between different autofluorescence tags, and expression of the tagged proteins is controlled by constitutive promoters, which can be easily replaced with virtually any other promoter of interest. In addition, a series of SAT vectors has been adapted for high throughput Gateway recombination cloning. Furthermore, individual expression cassettes can be assembled into Agrobacterium binary plasmids, allowing efficient transient and stable expression of multiple autofluorescently tagged proteins from a single vector following its biolistic delivery or Agrobacterium-mediated genetic transformation. Electronic supplementary material Electronic supplementary material is available for this article at and accessible for authorised users.     

Unraveling prion structures and biological functions

A report on a Joint Cold Spring Harbor Laboratory/Wellcome Trust Conference on 'Prion Biology', Hinxton, UK, 7-11 September 2005.     

The role of models in understanding CD8+ T-cell memory

Immunological memory - the ability to 'remember' previously encountered pathogens and respond faster on re-exposure - is a central feature of the immune response of vertebrates. We outline how mathematical models have contributed to our understanding of CD8(+) T-cell memory. Together with experimental data, models have helped to quantitatively describe and to further our understanding of both the generation of memory after infection with a pathogen and the maintenance of this memory throughout the life of an individual.     

1,8-Anilinonaphthalene sulfonate binds to central cavity of human hemoglobin

Binding of 1,8-anilinonaphthalene sulfonate (1,8-ANS) to main (HbA(1)) and glycosylated (HbA(1C)) forms of human oxyhemoglobin in the presence/absence of inositolhexaphosphate (IHP) in 50 mM potassium phosphate buffer, pH 7.4, was studied by time-correlated single photon counter with subnanosecond time resolution. The redistribution of contributions of the most long-lived and the most short-lived fluorescent decay components in the presence of IHP provides an evidence of the probe binding within oxyhemoglobin central cavity, namely DPG-binding site. Finally, it was shown that the fluorescent probe is extremely sensitive for hemoglobin central cavity modification, provided by the carbohydrate moiety in case of 1,8-ANS interactions with HbA(1C).     

Karyotype of the rare Johnston’s genet<Emphasis Type="Italic">Genetta johnstoni</Emphasis> (Viverridae) and a reassessment of chromosomal characterization among congeneric species

We present herein new data on karyotypes of members of the genusGenetta. G- and C-banded chromosomes of the Johnston’s genetGenetta johnstoni Pocock, 1908 (2n = 50 / FNa = 92) are described for the first time, and compared with those ofG. genetta (2n = 54 / FNa = 92). In addition, the standard karyotype ofG. maculata (2n = 52 / FNa = 96) was studied. A reassessment of taxonomic attribution of previously published material allowed us to characterize (2n, FNa, and chromosome morphology) the karyotypes of three genets, previously unknown (G. pardina, G. letabae andG. tigrina). Our results show that despite a rather low interspecific variability in 2n and FNa, all the species of genets (exceptG. pardina andG. maculata) appear differentiated when chromosomal morphology is taken into account. Although chromosomal banding data are limited, confrontation of G-band karyotypes with preliminary molecular phylogenetic results reveals that karyotypic evolution within the genusGenetta might involve various rearrangements like Robertsonian and tandem translocations, pericentric inversions, and centromere fissions; thus providing at least for some taxa a solid postzygotic isolation. Finally, our study suggests that cytogenetic analyses might constitute a useful tool for questioning interspecific boundaries, especially within the taxonomically debated complex of large-spotted genets.