The Wnt3a-treated smooth muscle cells displayed increased extracellular matrix synthesis, as measured by collagen I and III mRNA expression, along with increased expression of MMP2 and MMP9, but decreased TIMP2 levels. The Wnt3a-induced change in cell phenotype was associated with increased expression of the gap junction protein connexin 43. Consistent with this, Wnt3a-treated smooth muscle cells displayed enhanced intercellular communication, as measured by the scrape-loading dye transfer technique. The canonical Wnt antagonist, dickkopf-related protein 1, completely reversed the contractile protein and connexin 43 expression
seen in the Wnt3a-treated cells, suggesting these changes were dependent on canonical Wnt signaling. Collectively, this data suggest Wnt3a promotes a contractile and secretory phenotype in vascular smooth muscle GW4869 manufacturer cells that is associated with increased gap junction communication. Laboratory Investigation (2012) 92, 246-255; doi:10.1038/labinvest.2011.164; published online 21 November 2011″
“Introduction: TNFR2-Fc and IL-1ra-Fc are recombinant cytokine ligands that target TNF and IL-1. TNFR2-Fc-IL-1ra, a dual-domain agent that incorporates both ligands, allows bifunctional binding of
IL-1 receptors and TNF. This study was designed to characterize Tc-99m-labeled selleck compound forms of these ligands, Tc-99m-IL-1ra-Fc (IF), Tc-99m-TNFR2-Fc (TF), and Tc-99m-TNFR2-Fc-IL-1ra (TFI), for inflammation imaging.
Methods: The cytokine ligands were labeled with Tc-99m by a direct approach via 2-iminothiolane (2-IT) reduction at various 2-IT/protein molar ratios. In vivo inflammation targeting studies were carried out in a mouse ear edema model created by topical application of 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on the right ear of ICR mice.
Results: Radiolabeling yields increased
with increasing amounts of 2-IT. When the 2-IT/protein ratio reached 1000, the radiolabeling yield was greater than 90% without significant colloid production. TPA-treated ears showed high radioligand uptake, which was clearly detected by SPECT and autoradiographic imaging. The activities (%ID/g) in the inflamed and control ears at 3 h after injection were 2.76 +/- 0.20 vs. 0.69 +/- 0.12 for IF, 5.86 +/- 0.40 vs. 2.86 +/- 0.61 for TF, and 7.61 +/- 0.86 vs. 1.99 +/- 0.31 for TFI (P<0.05 vs. controls). TFI showed significantly higher uptake others in the inflamed ears compared to TF and IF (P<0.05). Blocking study results indicated specificity of radioligand binding with decreased radioactive uptake in the inflamed ears. Western blotting and ELISA analysis further confirmed a high expression of IL-1 beta and TNF-alpha in the inflamed ears.
Conclusions: Tc-99m-labeled cytokine ligands are a promising approach for detecting and understanding the inflammatory process. TFI may be more useful than the single-domain ligands for noninvasive detection of inflammatory sites. (c) 2012 Elsevier Inc. All rights reserved.