Enhancement of inosine-mediated A2AR signaling through positive allosteric modulation.

Inosine is an endogenous nucleoside that is produced by metabolic deamination of adenosine. Inosine is metabolically more stable (half-life 15h) than adenosine (half-life <10s). Inosine exerts anti-inflammatory and immunomodulatory effects similar to those observed with adenosine. These effects are mediated in part through the adenosine A2A receptor (A2AR). Relative to adenosine inosine exhibits a lower affinity towards the A2AR. Therefore, it is generally believed that inosine is incapable of activating the A2AR through direct engagement, but indirectly activates the A2AR upon metabolic conversion to higher affinity adenosine. A handful of studies, however, have provided evidence for direct inosine engagement at the A2AR leading to activation of downstream signaling events and inhibition of cytokine production. Here, we demonstrate that under conditions devoid of adenosine, inosine as well as an analog of inosine 6-S-[(4-Nitrophenyl)methyl]-6-thioinosine selectively and dose-dependently activated A2AR-mediated cAMP production and ERK1/2 phosphorylation in CHO cells stably expressing the human A2AR. Inosine also inhibited LPS-stimulated TNF-α, CCL3 and CCL4 production by splenic monocytes in an A2AR-dependent manner. In addition, we demonstrate that a positive allosteric modulator (PAM) of the A2AR enhanced inosine-mediated cAMP production, ERK1/2 phosphorylation and inhibition of pro-inflammatory cytokine and chemokine production. The cumulative effects of allosteric enhancement of adenosine-mediated and inosine-mediated A2AR activation may be the basis for the sustained anti-inflammatory and immunomodulatory effects observed in vivo and thereby provide insights into potential therapeutic interventions for inflammation- and immune-mediated diseases.

Culture phases, cytotoxicity and protein expressions of agarose hydrogel induced Sp2/0, A549, MCF-7 cell line 3D cultures.

Advancements in cell cultures are occurring at a rapid pace, an important direction is culturing cells in 3D conditions. We demonstrate the usefulness of agarose hydrogels in obtaining 3 dimensional aggregates of three cell lines, A549, MCF-7 and Sp2/0. The differences in culture phases, susceptibility to cisplatin-induced cytotoxicity are studied. Also, the 3D aggregates of the three cell lines were reverted into 2D cultures and the protein profile differences among the 2D, 3D and revert cultures were studied. The analysis of protein profile differences using UniProt data base further augment the usefulness of agarose hydrogels for obtaining 3D cell cultures.

3D cell culture systems: advantages and applications.

Cell cultures are important material of study for the variety of advantages that they offer. Both established continuous cell lines and primary cell cultures continue to be invaluable for basic research and for direct applications. Technological advancements are necessary to address emerging complex challenges and the way cells are cultured in vitro is an area of intense activity. One important advancement in cell culture techniques has been the introduction of three dimensional culture systems. This area is one of the fastest growing experimental approaches in life sciences. Augmented with advancements in cell imaging and analytical systems, as well as the applications of new scaffolds and matrices, cells have been increasingly grown as three dimensional models. Such cultures have proven to be closer to in vivo natural systems, thus proving to be useful material for many applications. Here, we review the three dimensional way of culturing cells, their advantages, the scaffolds and matrices currently available, and the applications of such cultures in major areas of life sciences.

Biosynthesis of silver nanoparticles using citrus sinensis peel extract and its antibacterial activity.

Biosynthesis of silver nanoparticles (AgNPs) was achieved by a novel, simple green chemistry procedure using citrus sinensis peel extract as a reducing and a capping agent. The effect of temperature on the synthesis of silver nanoparticles was carried out at room temperature (25°C) and 60°C. The successful formation of silver nanoparticles has been confirmed by UV-vis, FTIR, XRD, EDAX, FESEM and TEM analysis and their antibacterial activity against Escherichia coli, Pseudomonas aeruginosa (gram-negative), and Staphylococcus aureus (gram-positive) has been studied. The results suggest that the synthesized AgNPs act as an effective antibacterial agent.