Generation of hierarchical nano- and microwrinkle structure for smooth muscle cell alignment

On the contrary to the complicated and high-cost photolithography based topographical patterning, the non-lithographical strain responsive wrinkling method can generate a variety of wrinkle structures with ease, high uniformity, and cost-effectiveness. The strain responsive wrinkling approach relied on the modulus difference between a thin and stiff film and a soft substrate, resulting in a periodic out-of-plane buckling deformation upon the release of the compressive stress. While the previous reports were dedicated to the micropattern generation, we, in this study, investigated the effect of the UV/O exposure time (10, 20, 30, 40, 50, and 60 min) and the stretching rate (10, 20, 30, and 40%) of the PDMS substrate on the wavelength and the amplitude of the wrinkle patterns in details. Sole nanowrinkle as well as hierarchical nano/microwrinkle patterns could be fabricated through the fine control of those factors. Furthermore, we examined the human aortic smooth muscle cell (HASMCs) alignment on the topographical patterned surface, and found that more than 80% of the HASMCs were cultured and aligned well along with the hierarchical nano/microwrinkle rather than the nanowrinkle pattern.     

Direct Inhibition of GSK3β by the Phosphorylated Cytoplasmic Domain of LRP6 in Wnt/β-Catenin Signaling

Wnt/β-catenin signaling plays a central role in development and is also involved in a diverse array of diseases. Binding of Wnts to the coreceptors Frizzled and LRP6/5 leads to phosphorylation of PPPSPxS motifs in the LRP6/5 intracellular region and the inhibition of GSK3β bound to the scaffold protein Axin. However, it remains unknown how GSK3β is specifically inhibited upon Wnt stimulation. Here, we show that overexpression of the intracellular region of LRP6 containing a Ser/Thr rich cluster and a PPPSPxS motif impairs the activity of GSK3β in cells. Synthetic peptides containing the PPPSPxS motif strongly inhibit GSK3β in vitro only when they are phosphorylated. Microinjection of these peptides into Xenopus embryos confirms that the phosphorylated PPPSPxS motif potentiates Wnt-induced second body axis formation. In addition, we show that the Ser/Thr rich cluster of LRP6 plays an important role in LRP6 binding to GSK3β. These observations demonstrate that phosphorylated LRP6/5 both recruits and directly inhibits GSK3β using two distinct portions of its cytoplasmic sequence, and suggest a novel mechanism of activation in this signaling pathway.