Depleted uranium-catalyzed oxidative DNA damage: absence of significant alpha particle decay

Depleted uranium (DU) is a dense heavy metal used primarily in military applications. Published data from our laboratory have demonstrated that DU exposure in vitro to immortalized human osteoblast cells (HOS) is both neoplastically transforming and genotoxic. DU possesses both a radiological (alpha particle) and a chemical (metal) component. Since DU has a low-specific activity in comparison to natural uranium, it is not considered to be a significant radiological hazard. In the current study we demonstrate that DU can generate oxidative DNA damage and can also catalyze reactions that induce hydroxyl radicals in the absence of significant alpha particle decay. Experiments were conducted under conditions in which chemical generation of hydroxyl radicals was calculated to exceed the radiolytic generation by one million-fold. The data showed that markers of oxidative DNA base damage, thymine glycol and 8-deoxyguanosine could be induced from DU-catalyzed reactions of hydrogen peroxide and ascorbate similarly to those occurring in the presence of iron catalysts. DU was 6-fold more efficient than iron at catalyzing the oxidation of ascorbate at pH 7. These data not only demonstrate that DU at pH 7 can induced oxidative DNA damage in the absence of significant alpha particle decay, but also suggest that DU can induce carcinogenic lesions, e.g. oxidative DNA lesions, through interaction with a cellular oxygen species.     

A Hematogenous Route for Medulloblastoma Leptomeningeal Metastases

1. Download high-res image (170KB)
2. Download full-size image

     

Dynamics of G6PD activity in patients receiving weekly primaquine for therapy of Plasmodium vivax malaria

BackgroundAcute Plasmodium vivax malaria is associated with haemolysis, bone marrow suppression, reticulocytopenia, and post-treatment reticulocytosis leading to haemoglobin recovery. Little is known how malaria affects glucose-6-phosphate dehydrogenase (G6PD) activity and whether changes in activity when patients present may lead qualitative tests, like the fluorescent spot test (FST), to misdiagnose G6PD deficient (G6PDd) patients as G6PD normal (G6PDn). Giving primaquine or tafenoquine to such patients could result in severe haemolysis.MethodsWe investigated the G6PD genotype, G6PD enzyme activity over time and the baseline FST phenotype in Cambodians with acute P. vivax malaria treated with 3-day dihydroartemisinin piperaquine and weekly primaquine, 0·75 mg/kg x8 doses.ResultsOf 75 recruited patients (males 63), aged 5–63 years (median 24), 15 were G6PDd males (14 Viangchan, 1 Canton), 3 were G6PD Viangchan heterozygous females, and 57 were G6PDn; 6 patients had α/β-thalassaemia and 26 had HbE.Median (range) Day0 G6PD activities were 0·85 U/g Hb (0·10–1·36) and 11·4 U/g Hb (6·67–16·78) in G6PDd and G6PDn patients, respectively, rising significantly to 1·45 (0·36–5·54, p<0.01) and 12·0 (8·1–17·4, p = 0.04) U/g Hb on Day7, then falling to ~Day0 values by Day56. Day0 G6PD activity did not correlate (p = 0.28) with the Day0 reticulocyte counts but both correlated over time. The FST diagnosed correctly 17/18 G6PDd patients, misclassifying one heterozygous female as G6PDn.ConclusionsIn Cambodia, acute P. vivax malaria did not elevate G6PD activities in our small sample of G6PDd patients to levels that would result in a false normal qualitative test. Low G6PDd enzyme activity at disease presentation increases upon parasite clearance, parallel to reticulocytosis. More work is needed in G6PDd heterozygous females to ascertain the effect of P. vivax on their G6PD activities.Trial registrationThe trial was registered (ACTRN12613000003774) with the Australia New Zealand Clinical trials (https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=363399&isReview=true).     

Genetic targeting of Card19 is linked to disrupted NINJ1 expression,impaired cell lysis,and increased susceptibility to Yersinia infection

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-deficient mice (Card19^lxcn) whose macrophages were protected from cell lysis and showed reduced apoptosis and pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as an independently-generated CRISPR/Cas9 Card19 knockout mouse line (Card19^Null) showed no defect in macrophage cell lysis. Notably, Card19 is located on chromosome 13, immediately adjacent to Ninj1, which was recently found to regulate cell lysis downstream of GSDMD activation. RNA-seq and western blotting revealed that Card19^lxcn BMDMs have significantly reduced NINJ1 expression, and reconstitution of Ninj1 in Card19^lxcn immortalized BMDMs restored their ability to undergo cell lysis in response to caspase-dependent cell death stimuli. Card19^lxcn mice exhibited increased susceptibility to Yersinia infection, whereas independently-generated Card19^Null mice did not, demonstrating that cell lysis itself plays a key role in protection against bacterial infection, and that the increased infection susceptibility of Card19^lxcn mice is attributable to loss of NINJ1. Our findings identify genetic targeting of Card19 being responsible for off-target effects on the adjacent gene Ninj1, disrupting the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage and impacting host survival and bacterial control during Yersinia infection.     

Mafb and c-Maf Have Prenatal Compensatory and Postnatal Antagonistic Roles in Cortical Interneuron Fate and Function

1. Download high-res image (175KB)
2. Download full-size image

     

A truncated RHAMM protein for discovering novel therapeutic peptides

The receptor for hyaluronan mediated motility (RHAMM, gene name HMMR) belongs to a group of proteins that bind to hyaluronan (HA), a high-molecular weight anionic polysaccharide that has pro-angiogenic and inflammatory properties when fragmented. We propose to use a chemically synthesized, truncated version of the protein (706–767), 7?kDa RHAMM, as a target receptor in the screening of novel peptide-based therapeutic agents. Chemical synthesis by Fmoc-based solid-phase peptide synthesis, and optimization using pseudoprolines, results in RHAMM protein of higher purity and yield than synthesis by recombinant protein production. 7?kDa RHAMM was evaluated for its secondary structure, ability to bind the native ligand, HA, and its bioactivity. This 62-amino acid polypeptide replicates the HA binding properties of both native and recombinant RHAMM protein. Furthermore, tubulin-derived HA peptide analogues that bind to recombinant RHAMM and were previously reported to compete with HA for interactions with RHAMM, bind with a similar affinity and specificity to the 7?kDa RHAMM. Therefore, in terms of its key binding properties, the 7?kDa RHAMM mini-protein is a suitable replacement for the full-length recombinant protein.     

Plant diversity does not buffer drought effects on early‐stage litter mass loss rates and microbial properties

Human activities are decreasing biodiversity and changing the climate worldwide. Both global change drivers have been shown to affect ecosystem functioning, but they may also act in concert in a non‐additive way. We studied early‐stage litter mass loss rates and soil microbial properties (basal respiration and microbial biomass) during the summer season in response to plant species richness and summer drought in a large grassland biodiversity experiment, the Jena Experiment, Germany. In line with our expectations, decreasing plant diversity and summer drought decreased litter mass loss rates and soil microbial properties. In contrast to our hypotheses, however, this was only true for mass loss of standard litter (wheat straw) used in all plots, and not for plant community‐specific litter mass loss. We found no interactive effects between global change drivers, that is, drought reduced litter mass loss rates and soil microbial properties irrespective of plant diversity. High mass loss rates of plant community‐specific litter and low responsiveness to drought relative to the standard litter indicate that soil microbial communities were adapted to decomposing community‐specific plant litter material including lower susceptibility to dry conditions during summer months. Moreover, higher microbial enzymatic diversity at high plant diversity may have caused elevated mass loss of standard litter. Our results indicate that plant diversity loss and summer drought independently impede soil processes. However, soil decomposer communities may be highly adapted to decomposing plant community‐specific litter material, even in situations of environmental stress. Results of standard litter mass loss moreover suggest that decomposer communities under diverse plant communities are able to cope with a greater variety of plant inputs possibly making them less responsive to biotic changes.