Topography-driven alterations in endothelial cell phenotype and contact guidance

Bioengineering; Biophysics; Cell biology; Biomedical engineering; Regenerative medicine; Directional topography; Endothelial cells; Vascular-like networks; Contact guidance; Vascularizatio

Topography-Mediated Myotube and Endothelial Alignment, Differentiation, and Extracellular Matrix Organization for Skeletal Muscle Engineering

Understanding the response of endothelial cells to aligned myotubes is important to create an appropriate environment for tissue-engineered vascularized skeletal muscle. Part of the native tissue environment is the extracellular matrix (ECM). The ECM is a supportive scaffold for cells and allows cellular processes such as proliferation, differentiation, and migration. Interstitial matrix and basal membrane both comprise proteinaceous and polysaccharide components for strength, architecture, and volume retention. Virtually all cells are anchored to their basal lamina. One of the physical factors that affects cell behavior is topography, which plays an important role on cell alignment. We tested the hypothesis that topography-driven aligned human myotubes promote and support vascular network formation as a prelude to in vitro engineered vascularized skeletal muscle. Therefore, we used a PDMS-based topography substrate to investigate the influence of pre-aligned myotubes on the network formation of microvascular endothelial cells. The aligned myotubes produced a network of collagen fibers and laminin. This network supported early stages of endothelial network formation.</p

Full humanization of the glycolytic pathway in Saccharomyces cerevisiae

Although transplantation of single genes in yeast plays a key role in elucidating gene functionality in metazoans, technical challenges hamper humanization of full pathways and processes. Empowered by advances in synthetic biology, this study demonstrates the feasibility and implementation of full humanization of glycolysis in yeast. Single gene and full pathway transplantation revealed the remarkable conservation of glycolytic and moonlighting functions and, combined with evolutionary strategies, brought to light context-dependent responses. Human hexokinase 1 and 2, but not 4, required mutations in their catalytic or allosteric sites for functionality in yeast, whereas hexokinase 3 was unable to complement its yeast ortholog. Comparison with human tissues cultures showed preservation of turnover numbers of human glycolytic enzymes in yeast and human cell cultures. This demonstration of transplantation of an entire essential pathway paves the way for establishment of species-, tissue-, and disease-specific metazoan models

Directional topography gradients drive optimum alignment and differentiation of human myoblasts

Tissue engineering of skeletal muscle aims to replicate the parallel alignment of myotubes on the native tissue. Directional topography gradients allow the study of the influence of topography on cellular orientation, proliferation, and differentiation, resulting in yield cues and clues to develop a proper in vitro environment for muscle tissue engineering. In this study, we used a polydimethylsiloxane-based substrate containing an aligned topography gradient with sinusoidal features ranging from wavelength (lambda) = 1,520 nm and amplitude (A) =176 nm to lambda = 9,934 nm and A = 2,168 nm. With this topography gradient, we evaluated the effect of topography on human myoblasts distribution, dominant orientation, cell area, nuclei coverage, cell area per number of nuclei, and nuclei area of myotubes. We showed that human myoblasts aligned and differentiated irrespective of the topography section. In addition, aligned human myotubes showed functionality and maturity by contracting spontaneously and nuclei peripheral organization resembling natural myotubes