Circulating mediators of inflammation and immune activation in AIDS-related non-Hodgkin lymphoma

Background: Non-Hodgkin lymphoma (NHL) is the most common AIDS-related malignancy in developed countries. An elevated risk of developing NHL persists among HIV-infected individuals in comparison to the general population despite the advent of effective antiretroviral therapy. The mechanisms underlying the development of AIDS-related NHL (A-NHL) are not fully understood, but likely involve persistent B-cell activation and inflammation. Methods: This was a nested case-control study within the ongoing prospective Multicenter AIDS Cohort Study (MACS). Cases included 47 HIV-positive male subjects diagnosed with high-grade B-cell NHL. Controls were matched to each case from among participating HIV-positive males who did not develop any malignancy. Matching criteria included time HIV+ or since AIDS diagnosis, age, race and CD4+ cell count. Sera were tested for 161 serum biomarkers using multiplexed beadbased immunoassays. Results: A subset of 17 biomarkers, including cytokines, chemokines, acute phase proteins, tissue remodeling agents and bone metabolic mediators was identified to be significantly altered in A-NHL cases in comparison to controls. Many of the biomarkers included in this subset were positively correlated with HIV viral load. A pathway analysis of our results revealed an extensive network of interactions between current and previously identified biomarkers. Conclusions: These findings support the current hypothesis that A-NHL develops in the context of persistent immune stimulation and inflammation. Further analysis of the biomarkers identified in this report should enhance our ability to diagnose, monitor and treat this disease. © 2014 Nolen et al

Inaccurate DNA Synthesis in Cell Extracts of Yeast Producing Active Human DNA Polymerase Iota

Mammalian Pol ι has an unusual combination of properties: it is stimulated by Mn2+ ions, can bypass some DNA lesions and misincorporates “G” opposite template “T” more frequently than incorporates the correct “A.” We recently proposed a method of detection of Pol ι activity in animal cell extracts, based on primer extension opposite the template T with a high concentration of only two nucleotides, dGTP and dATP (incorporation of “G” versus “A” method of Gening, abbreviated as “misGvA”). We provide unambiguous proof of the “misGvA” approach concept and extend the applicability of the method for the studies of variants of Pol ι in the yeast model system with different cation cofactors. We produced human Pol ι in baker's yeast, which do not have a POLI ortholog. The “misGvA” activity is absent in cell extracts containing an empty vector, or producing catalytically dead Pol ι, or Pol ι lacking exon 2, but is robust in the strain producing wild-type Pol ι or its catalytic core, or protein with the active center L62I mutant. The signature pattern of primer extension products resulting from inaccurate DNA synthesis by extracts of cells producing either Pol ι or human Pol η is different. The DNA sequence of the template is critical for the detection of the infidelity of DNA synthesis attributed to DNA Pol ι. The primer/template and composition of the exogenous DNA precursor pool can be adapted to monitor replication fidelity in cell extracts expressing various error-prone Pols or mutator variants of accurate Pols. Finally, we demonstrate that the mutation rates in yeast strains producing human DNA Pols ι and η are not elevated over the control strain, despite highly inaccurate DNA synthesis by their extracts

Nanoscopic Molecular Cluster V15: High-Field EPR and Magnetization at Ultra-Low Temperatures

Tsukerblat B, Tarantul A, Müller A. Nanoscopic Molecular Cluster V15: High-Field EPR and Magnetization at Ultra-Low Temperatures. Chem. J. Moldova. 2007;2(1 (in memoriam Prof. Y. E. Perlin):17-35

Aptamers: Problems, Solutions and Prospects

Aptamers are short single-stranded oligonucleotides that are capable of binding various molecules with high affinity and specificity. When the technology of aptamer selection was developed almost a quarter of a century ago, a suggestion was immediately put forward that it might be a revolutionary start into solving many problems associated with diagnostics and the therapy of diseases. However, multiple attempts to use aptamers in practice, although sometimes successful, have been generally much less efficient than had been expected initially. This review is mostly devoted not to the successful use of aptamers but to the problems impeding the widespread application of aptamers in diagnostics and therapy, as well as to approaches that could considerably expand the range of aptamer application.</jats:p

A Hybrid Detection Method Based on Peroxidase-mediated Signal Amplification and Click Chemistry for Highly Sensitive Background-free Immunofluorescent Staining

The copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction is increasingly used for detection of various macromolecules and metabolites in biological samples. Here, we present a detailed analysis of the CuAAC reaction conditions in cells and tissue sections. Using the optimized CuAAC conditions, we have devised a highly sensitive immunostaining technique, based on the tyramide signal amplification/catalyzed reporter deposition (TSA/CARD) method with a novel alkyne tyramide substrate. The described method offers improved detection threshold compared to conventional immunofluorescent staining and produces significantly lower non-specific background than TSA/CARD with fluorescent tyramides. </jats:p