Rapid determination of 210Pb and 210Po by sequential cloud point extraction for environmental monitoring.

A new sequential cloud point extraction (CPE) procedure was developed and validated for the determination of 210Pb and 210Po by ICP-MS/MS and alpha spectrometry, respectively. Two distinct CPE systems using 4',4''(5'')-di-tert-butyldicyclohexano-18-crown-6 and N,N,N',N'-tetraoctyldiglycolamide as chelating agents were performed sequentially on the same sample. Method detection limits of 13 and 3.5 mBq L-1, were achieved for 210Pb and 210Po, respectively. The sequential approach shows its outstanding versatility and proficiency to rapidly extract, detect, and quantify both radionuclides in various matrices for a wide range of activities.

Determination of polonium-210 in environmental samples using diglycolamide-based cloud point extraction coupled to alpha spectrometry analysis.

This study presents a novel cloud point extraction (CPE) methodology for the separation and enrichment of polonium-210 prior to alpha-spectrometric quantification in water, urine and digested samples. The extractive behaviour of diglycolamide-based ligands towards Po by CPE was determined and optimised in various acidic conditions. The extraction efficiency and selectivity of the CPE systems depend greatly on the choice of the extracting agent and acidic conditions. The thorough optimisation of those specific parameters has led to the development of a suitable cloud point extraction system for the determination of polonium-210 at ultra-trace levels compatible with alpha-spectrometry. To facilitate this coupling, a back-extraction procedure was optimised and performed on the surfactant-rich phase to enable the spontaneous deposition of polonium-210 onto a silver disk; this also avoids making the matrix transfer step mandatory due to the presence of a nitric medium. Method detection and quantification limits of 3.5 and 12 mBq L-1, respectively, were determined by alpha spectrometry. The robustness of the proposed methodology was demonstrated by the determination of polonium ions concentration in various environmental and biological samples and solid certified reference materials.

Determination of Pb in environmental samples after cloud point extraction using crown ether.

In the present study, a new cloud point extraction methodology based on the selective preconcentration and the extraction of stable lead in acidic conditions with 4',4''(5'')-di-tert-butyldicyclohexano-18-crown-6 as a chelating agent was developed, optimized and validated. A mixture of Triton X-114 as non-ionic surfactant and CTAB as cationic surfactant was used to produce micellar structures that incorporate the chelating agent. Phase separation, induced by coacervation, was achieved by increasing the temperature of the system above the cloud point temperature. Pb extraction efficiency was maximized through an optimisation process where the effect of each parameter (i.e. non-ionic and ionic surfactant concentrations, pH, chelating agent concentration and cloud point temperature) on the chemical recoveries of Pb was assessed. Under optimum experimental conditions, the method reaches recoveries greater than 67% for Pb in a variety of complex matrices. In order to facilitate the quantification of Pb by plasma based instrumentations, a back-extraction procedure using aqueous solution of ammonium citrate were performed on the surfactant rich phase in order to reduce the effects on sample introduction and non-spectral interferences. LOD and LOQ of 0.8µgL-1 and 2.6µgL-1, respectively, were determined by ICP-OES for the complete procedure. Using the back-extraction approach, a preconcentration factor of 39 was achieved for an initial sample volume of 195mL. The ruggedness of the methodology was validated by determining Pb concentration in various environmental and biological samples.

CD73 Expression Is an Independent Prognostic Factor in Prostate Cancer.

PURPOSE: CD73 is an adenosine-generating ecto-enzyme that suppresses antitumor immunity in mouse models of cancer, including prostate cancer. Although high levels of CD73 are associated with poor prognosis in various types of cancer, the clinical impact of CD73 in prostate cancer remains unclear. EXPERIMENTAL DESIGN: We evaluated the prognostic value of CD73 protein expression and CD8(+) cell density in 285 cases of prostate cancer on tissue microarray (TMA). Normal adjacent and tumor tissues were evaluated in duplicates. RESULTS: Univariate and multivariate analyses revealed that high levels of CD73 in normal adjacent prostate epithelium were significantly associated with shorter biochemical recurrence (BCR)-free survival. Notably, CD73 expression in normal epithelium conferred a negative prognostic value to prostate-infiltrating CD8(+) cells. Surprisingly, high levels of CD73 in the tumor stroma were associated with longer BCR-free survival in univariate analysis. In vitro studies revealed that adenosine signaling inhibited NF-κB activity in human prostate cancer cells via A2B adenosine receptors. Consistent with these results, CD73 expression in the prostate tumor stroma negatively correlated with p65 expression in the nuclei of prostate tumor cells. CONCLUSIONS: Our study revealed that CD73 is an independent prognostic factor in prostate cancer. Our data support a model in which CD73 expression in the prostate epithelium suppresses immunosurveillance by CD8(+) T cells, whereas CD73 expression in the tumor stroma reduces NF-κB signaling in tumor cells via A2B adenosine receptor signaling. CD73 expression, including in normal adjacent prostate epithelium, can thus effectively discriminate between aggressive and indolent forms of prostate cancer.

CD73 is associated with poor prognosis in high-grade serous ovarian cancer.

The cell surface nucleotidase CD73 is an immunosuppressive enzyme involved in tumor progression and metastasis. Although preclinical studies suggest that CD73 can be targeted for cancer treatment, the clinical impact of CD73 in ovarian cancer remains unclear. In this study, we investigated the prognostic value of CD73 in high-grade serous (HGS) ovarian cancer using gene and protein expression analyses. Our results demonstrate that high levels of CD73 are significantly associated with shorter disease-free survival and overall survival in patients with HGS ovarian cancer. Furthermore, high levels of CD73 expression in ovarian tumor cells abolished the good prognosis associated with intraepithelial CD8(+) cells. Notably, CD73 gene expression was highest in the C1/stromal molecular subtype of HGS ovarian cancer and positively correlated with an epithelial-to-mesenchymal transition gene signature. Moreover, in vitro studies revealed that CD73 and extracellular adenosine enhance ovarian tumor cell growth as well as expression of antiapoptotic BCL-2 family members. Finally, in vivo coinjection of ID8 mouse ovarian tumor cells with mouse embryonic fibroblasts showed that CD73 expression in fibroblasts promotes tumor immune escape and thereby tumor growth. In conclusion, our study highlights a role for CD73 as a prognostic marker of patient survival and also as a candidate therapeutic target in HGS ovarian cancers.

Cell cycle-dependent localization of CHK2 at centrosomes during mitosis.

BACKGROUND: Centrosomes function primarily as microtubule-organizing centres and play a crucial role during mitosis by organizing the bipolar spindle. In addition to this function, centrosomes act as reaction centers where numerous key regulators meet to control cell cycle progression. One of these factors involved in genome stability, the checkpoint kinase CHK2, was shown to localize at centrosomes throughout the cell cycle. RESULTS: Here, we show that CHK2 only localizes to centrosomes during mitosis. Using wild-type and CHK2-/- HCT116 human colon cancer cells and human osteosarcoma U2OS cells depleted for CHK2 with small hairpin RNAs we show that several CHK2 antibodies are non-specific and cross-react with an unknown centrosomal protein(s) by immunofluorescence. To characterize the localization of CHK2, we generated cells expressing inducible GFP-CHK2 and Flag-CHK2 fusion proteins. We show that CHK2 localizes to the nucleus in interphase cells but that a fraction of CHK2 associates with the centrosomes in a Polo-like kinase 1-dependent manner during mitosis, from early mitotic stages until cytokinesis. CONCLUSION: Our findings demonstrate that a subpopulation of CHK2 localizes at the centrosomes in mitotic cells but not in interphase. These results are consistent with previous reports supporting a role for CHK2 in the bipolar spindle formation and the timely progression of mitosis.