Cell cycle effects of iron depletion on T-47D human breast cancer cells

T-47D human breast cancer cells grown in culture medium containing low concentrations of fetal calf serum (FCS) proliferated very slowly, with an accumulation of cells in the G2 phase of the cell cycle, increased polyploid cells, and increased expression of transferrin receptors. Cell proliferation was stimulated by the addition of human transferrin or ammonium ferric citrate to the medium. Growth inhibition and accumulation of G2-phase cells could also be produced in T-47D cells grown in medium containing 10% FCS by the addition of the iron chelator, desferrioxamine. It is concluded that cellular deprivation of iron and/or transferrin is the major cause of reduced proliferation rates and G2-phase arrest which accompany the culture of these cells in medium supplemented with low concentrations of FCS.     

Horizon scan of global conservation issues for 2011

This review describes outcomes of a 2010 horizon-scanning exercise building upon the first exercise conducted in 2009. The aim of both horizon scans was to identify emerging issues that could have substantial impacts on the conservation of biological diversity, and to do so sufficiently early to encourage policy-relevant, practical research on those issues. Our group included professional horizon scanners and researchers affiliated with universities and non- and inter-governmental organizations, including specialists on topics such as invasive species, wildlife diseases and coral reefs. We identified 15 nascent issues, including new greenhouse gases, genetic techniques to eradicate mosquitoes, milk consumption in Asia and societal pessimism.     

Impact of nature reserve establishment on deforestation: a test

Establishing protected areas is one of the most widely used tools for biodiversity conservation, but the effectiveness of this is disputed. We used satellite-based maps of land cover to compare forest cover and rate of forest loss before and after 28 Mexican protected areas were decreed. Forest cover declined over time, both inside and outside reserves, but the loss was lowest in designated core areas. No significant difference was found in the mean rate of change in forest cover between external buffer, decreed areas and areas not yet decreed. There was also no significant difference in forest loss among protected areas before and after being decreed. The centre of protected areas (whether or not it corresponds to core areas) had a lower rate of forest cover loss. Population and road density was significantly lower within than outside protected areas and in regulated core areas than in the rest of the area. Isolation from the boundary appears to reduce deforestation as much as the presence of regulated core areas, illustrating the importance of size and design and the importance of protecting large areas with adequate core areas for a more effective conservation management.     

Suppression of neurotoxic lesion-induced seizure activity: evidence for a permanent role for the hippocampus in contextual memory

Damage to the hippocampus (HPC) using the excitotoxin N-methyl-D-aspartate (NMDA) can cause retrograde amnesia for contextual fear memory. This amnesia is typically attributed to loss of cells in the HPC. However, NMDA is also known to cause intense neuronal discharge (seizure activity) during the hours that follow its injection. These seizures may have detrimental effects on retrieval of memories. Here we evaluate the possibility that retrograde amnesia is due to NMDA-induced seizure activity or cell damage per se. To assess the effects of NMDA induced activity on contextual memory, we developed a lesion technique that utilizes the neurotoxic effects of NMDA while at the same time suppressing possible associated seizure activity. NMDA and tetrodotoxin (TTX), a sodium channel blocker, are simultaneously infused into the rat HPC, resulting in extensive bilateral damage to the HPC. TTX, co-infused with NMDA, suppresses propagation of seizure activity. Rats received pairings of a novel context with foot shock, after which they received NMDA-induced, TTX+NMDA-induced, or no damage to the HPC at a recent (24 hours) or remote (5 weeks) time point. After recovery, the rats were placed into the shock context and freezing was scored as an index of fear memory. Rats with an intact HPC exhibited robust memory for the aversive context at both time points, whereas rats that received NMDA or NMDA+TTX lesions showed a significant reduction in learned fear of equal magnitude at both the recent and remote time points. Therefore, it is unlikely that observed retrograde amnesia in contextual fear conditioning are due to disruption of non-HPC networks by propagated seizure activity. Moreover, the memory deficit observed at both time points offers additional evidence supporting the proposition that the HPC has a continuing role in maintaining contextual memories.     

The "magic numbers" of metallothionein

Metallothioneins (MT) are a family of small cysteine rich proteins, which since their discovery in 1957, have been implicated in a range of roles including toxic metal detoxification, protection against oxidative stress, and as a metallochaperone involved in the homeostasis of both zinc and copper. The most well studied member of the family is the mammalian metallothionein, which consists of two domains: a β-domain with 9 cysteine residues, which sequesters 3 Cd(2+) or Zn(2+) or 6 Cu(+) ions, and an α-domain with 11 cysteine residues and, which sequesters 4 Cd(2+) or Zn(2+) or 6 Cu(+) ions. Despite over half a century of research, the exact functions of MT are still unknown. Much of current research aims to elucidate the mechanism of metal binding, as well as to isolate intermediates in metal exchange reactions; reactions necessary to maintain homeostatic equilibrium. These studies further our understanding of the role(s) of this remarkable and ubiquitous protein. Recently, supermetallated forms of the protein, where supermetallation describes metallation in excess of traditional levels, have been reported. These species may potentially be the metal exchange intermediates necessary to maintain homeostatic equilibrium. This review focuses on recent advances in the understanding of the mechanistic properties of metal binding, the implications for the metal induced protein folding reactions proposed for metallothionein metallation, the value of "magic numbers", which we informally define as the commonly determined metal-to-protein stoichiometric ratios and the significance of the new supermetallated states of the protein and the possible interpretation of the structural properties of this new metallation status. Together we provide a commentary on current experimental and theoretical advances and frame our consideration in terms of the possible functions of MT.     

Recruitment and activation of pancreatic stellate cells from the bone marrow in pancreatic cancer: a model of tumor-host interaction

Background and Aims

Chronic pancreatitis and pancreatic cancer are characterised by extensive stellate cell mediated fibrosis, and current therapeutic development includes targeting pancreatic cancer stroma and tumor-host interactions. Recent evidence has suggested that circulating bone marrow derived stem cells (BMDC) contribute to solid organs. We aimed to define the role of circulating haematopoietic cells in the normal and diseased pancreas.

Methods

Whole bone marrow was harvested from male β-actin-EGFP donor mice and transplanted into irradiated female recipient C57/BL6 mice. Chronic pancreatitis was induced with repeat injections of caerulein, while carcinogenesis was induced with an intrapancreatic injection of dimethylbenzanthracene (DMBA). Phenotype of engrafted donor-derived cells within the pancreas was assessed by immunohistochemistry, immunofluorescence and in situ hybridisation.

Results

GFP positive cells were visible in the exocrine pancreatic epithelia from 3 months post transplantation. These exhibited acinar morphology and were positive for amylase and peanut agglutinin. Mice administered caerulein developed chronic pancreatitis while DMBA mice exhibited precursor lesions and pancreatic cancer. No acinar cells were identified to be donor-derived upon cessation of cerulein treatment, however rare occurrences of bone marrow-derived acinar cells were observed during pancreatic regeneration. Increased recruitment of BMDC was observed within the desmoplastic stroma, contributing to the activated pancreatic stellate cell (PaSC) population in both diseases. Expression of stellate cell markers CELSR3, PBX1 and GFAP was observed in BMD cancer-associated PaSCs, however cancer-associated, but not pancreatitis-associated BMD PaSCs, expressed the cancer PaSC specific marker CELSR3.

Conclusions

This study demonstrates that BMDC can incorporate into the pancreas and adopt the differentiated state of the exocrine compartment. BMDC that contribute to the activated PaSC population in chronic pancreatitis and pancreatic cancer have different phenotypes, and may play important roles in these diseases. Further, bone marrow transplantation may provide a useful model for the study of tumor-host interactions in cancer and pancreatitis.     

Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients

Naturally acquired blood-stage infections of the malaria parasite Plasmodium falciparum typically harbour multiple haploid clones. The apparent number of clones observed in any single infection depends on the diversity of the polymorphic markers used for the analysis, and the relative abundance of rare clones, which frequently fail to be detected among PCR products derived from numerically dominant clones. However, minority clones are of clinical interest as they may harbour genes conferring drug resistance, leading to enhanced survival after treatment and the possibility of subsequent therapeutic failure. We deployed new generation sequencing to derive genome data for five non-propagated parasite isolates taken directly from 4 different patients treated for clinical malaria in a UK hospital. Analysis of depth of coverage and length of sequence intervals between paired reads identified both previously described and novel gene deletions and amplifications. Full-length sequence data was extracted for 6 loci considered to be under selection by antimalarial drugs, and both known and previously unknown amino acid substitutions were identified. Full mitochondrial genomes were extracted from the sequencing data for each isolate, and these are compared against a panel of polymorphic sites derived from published or unpublished but publicly available data. Finally, genome-wide analysis of clone multiplicity was performed, and the number of infecting parasite clones estimated for each isolate. Each patient harboured at least 3 clones of P. falciparum by this analysis, consistent with results obtained with conventional PCR analysis of polymorphic merozoite antigen loci. We conclude that genome sequencing of peripheral blood P. falciparum taken directly from malaria patients provides high quality data useful for drug resistance studies, genomic structural analyses and population genetics, and also robustly represents clonal multiplicity.