Correction to: Cluster identification, selection, and description in Cluster randomized crossover trials: the PREP-IT trials

An amendment to this paper has been published and can be accessed via the original article

Potential importance of glomerular citrate synthase activity in remnant nephropathy

Potential importance of glomerular citrate synthase activity in remnant nephropathy.BackgroundAldosterone fosters progressive renal injury, but the mechanism is unknown. Both Wistar-Furth rats, which are resistant to aldosterone actions, and adrenalectomized Sprague-Dawley rats, which lack aldosterone, are characterized by resistance to remnant nephropathy and by reduced whole kidney citrate synthase activity. Increase in citrate synthase activity is a well-characterized, specific renal response to aldosterone. Therefore, we performed experiments to test the hypothesis that enhanced citrate synthase activity contributes to remnant nephropathy.MethodsRat models included Wistar (control for Wistar-Furth), Wistar-Furth (resistant to aldosterone), Sprague-Dawley (normal), adrenalectomy (lacking aldosterone), and 5/6 nephrectomy (renal injury). Glomeruli were obtained by differential sieving. Citrate synthase activity was determined spectrophotometrically. Binding characteristics of cytosolic mineralocorticoid receptors were determined by equilibrium competition binding between tritiated and unlabeled aldosterone. Gene sequencing was performed with reverse transcription-polymerase chain reaction (RT-PCR) and fluorescent dye terminators.ResultsIn glomeruli isolated from adrenalectomized Wistar rats with intact renal mass, aldosterone stimulated a threefold increase in citrate synthase activity; this stimulation was not observed in glomeruli from Wistar-Furth rats. Similarly, citrate synthase activity in glomeruli isolated from adrenally intact Sprague-Dawley rats was 65% greater than that from adrenalectomized Sprague-Dawley rats. Compared to sham surgery, subtotal nephrectomy resulted in 100% greater glomerular citrate synthase activity in Sprague-Dawley rats. In Wistar-Furth rats, mineralocorticoid receptor binding was not reduced, and mutations in the mineralocorticoid receptor DNA binding segment were not found.ConclusionCitrate synthase activity is elevated in remnant glomeruli, and experimental models characterized by reduced glomerular citrate synthase activity (Wistar-Furth rats, adrenalectomized Sprague-Dawley rats) are protected from remnant nephropathy

Plasma Membrane Nucleolin Is a Receptor for the Anticancer Aptamer AS1411 in MV4-11 Leukemia Cells

AS1411 is a DNA aptamer that is in phase II clinical trials for relapsed or refractory acute myeloid leukemia and for renal cell carcinoma. AS1411 binds to nucleolin, a protein that is overexpressed in the cytoplasm and on the plasma membrane of some tumor cells compared with normal cells. Studies were performed to determine whether cell surface nucleolin is a receptor for AS1411 in the acute myeloid leukemia cell line MV4-11. Biotinylation of MV4-11 cell surface proteins followed by immunoblotting of the biotinylated proteins showed that full-length (106 kDa) and truncated forms of nucleolin were present on the cell surface. In contrast, K-562 cells, which are 4-fold less sensitive than MV4-11 cells to AS1411, showed no full-length nucleolin and lesser amounts of the truncated forms of nucleolin on the cell surface. Incubation of MV4-11 cells with [32P]AS1411 and immunoprecipitation of the plasma membrane fraction with anti-nucleolin antibody demonstrated the presence of [32P]AS1411-nucleolin complexes. Anti-nucleolin antibody inhibited binding of fluorescein isothiocyanate (FITC)-AS1411 to plasma membrane nucleolin 56 ± 10% SE (P < 0.01) compared with cells incubated with FITC-AS1411 only. Cellular uptake of [32P]AS1411 into MV4-11 cells was blocked by a 20-fold excess of unlabeled AS1411 but not by a 20-fold excess of the biologically inactive oligonucleotide CRO-26. Uptake was approximately 3-fold faster into MV4-11 cells than into K-562 cells. Partial knockdown of plasma membrane and cytosolic nucleolin in MCF-7 cells resulted in a 3-fold decrease in AS1411 uptake. These results provide evidence that plasma membrane nucleolin is a functional receptor for AS1411 in MV4-11 cells

A global assessment of the species composition and effectiveness of watermelon pollinators and the management strategies to inform effective pollination service delivery

For most food crops the identity and efficiency of pollinators across key growing regions remains a significant knowledge gap that needs to be addressed before we can develop crop-specific approaches for pollination service delivery. Here, we conducted a systematic literature review and meta-analysis on watermelon (Citrullus lanatus (Thunb. Matsum. & Nakai)), a globally important fruit crop, to identify the floral visitors and their efficiency across different growing regions. We found that 265 insect species visit watermelon flowers (including 5 orders, 18 families and 75 genera) across 17 countries and 6 continents. Bees and flies were the most abundant flower visitors overall, but show distinct regional differences. Honey bees were the majority visitor in 53% of growing regions (range: 0 – 94%), whilst wild bee species were more abundant in 42% of regions (range: 3.4 – 100%). Honey bees and other bees were equally effective at depositing pollen on stigmas, but varied in effectiveness for fruit set and seed set. Pollination data from global studies appear to be limited for the largest-scale watermelon producers, namely: China, Turkey, and India, with the majority (56%) of data available from North America. This synthesis identified four key themes for improving pollination in watermelon: increasing honey bee densities on crops where local polices and environmental conditions are suitable; introducing other managed pollinators; identifying key wild pollinator taxa to encourage within crops; and improving local and landscape management practices to support pollinators

Identification of Ebp1 as a component of cytoplasmic bcl-2 mRNP (messenger ribonucleoprotein particle) complexes

The 3′-UTR (untranslated region) of bcl-2 mRNA contains an ARE (AU-rich element) that potentially regulates the stability of bcl-2 mRNA in a cell specific fashion. Previous studies have demonstrated that multiple proteins interact with bcl-2 mRNA in HL-60 (human leukaemia-60) cells, potentially contributing to the overexpression of Bcl-2 protein. Treatment of HL-60 cells with taxol or okadaic acid has been shown to induce destabilization of bcl-2 mRNA, which was associated with decreased binding of trans-acting factors to bcl-2 mRNA. Nucleolin has been identified as one of the bcl-2 mRNA-binding proteins [Sengupta, Bandyopadhyay, Fernandes and Spicer (2004) J. Biol. Chem. 279, 10855–10863]. In an effort to identify additional bcl-2 mRNA-binding proteins, two polypeptides of approx. 45 kDa and 60 kDa were isolated from HL-60 cells by ARE(bcl-2) (transcripts that contain bcl-2 AREs) RNA affinity chromatography. These proteins were identified as the human proliferation associated protein, Ebp1, and human DRBP76 (double stranded RNA-binding protein 76) respectively, by MALDI (matrix-assisted laser-desorption ionization)-MS. RNA electrophoretic mobility shift assays indicated that recombinant Ebp1 binds to ARE(bcl-2) RNA but not to the group 1 ARE present in GM-CSF (granulocyte macrophage-colony stimulating factor) mRNA in vitro. Antibody supershift assays demonstrated that Ebp1 is present in protein–ARE(bcl-2) RNA complexes formed with cytosolic HL-60 extracts. The interaction of Ebp1 with bcl-2 mRNA in HL-60 cells was also demonstrated by RNA co-immunoprecipitation assays. This interaction was not detected in extracts of taxol-treated HL-60 cells. Immunoprecipitation assays further revealed that Ebp1 co-precipitates with nucleolin from HL-60 cytoplasmic extracts. The observation that co-precipitation was decreased when extracts were treated with RNase suggests that Ebp1 and nucleolin are present in the same bcl-2 mRNP (messenger ribonucleoprotein particle) complexes. RNA-decay assays further demonstrated that Ebp1 decreased the rate of decay of β-globin–ARE(bcl-2) transcripts in HL-60 cell extracts. Collectively, these results indicate a novel function for Ebp1 in contributing to the regulation of bcl-2 expression in HL-60 cells