Myotube Assembly on Nanofibrous and Micropatterned Polymers

Huang, Ngan F.; Patel, Shyam; Thakar, Rahul; Wu, Jun; Hsiao, Benjamin S.; Chu, Benjamin; Lee, Randall J.; Li, Song

doi:10.1021/nl060060o

Cited by 300 publications

(281 citation statements)

References 25 publications

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“…After washing away the unpolymerized photoresist, PDMS was prepared according to the manufacturer's protocol (Sylgard 184; Dow Corning, Midland, MI), spin-coated onto the patterned silicon wafers to desired thickness (Ϸ250 m), degassed under vacuum, and cured at 70°C for 1 h. The resulting micropatterned membranes were removed from the template, cut to appropriate dimensions (for assembly into custom-built stretch chambers), and thoroughly washed and sonicated before use. The surface topography of micropatterned PDMS membranes was examined by scanning electron microscopy (27). Mechanical testing showed that PDMS membranes were elastic under cyclic uniaxial strain (Ͻ30% strain, 1 Hz).…”

Section: Methodsmentioning

confidence: 99%

Anisotropic mechanosensing by mesenchymal stem cells

Kurpinski

Chu

Hashi

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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362

316

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Mesenchymal stem cells (MSCs) are a potential source for the construction of tissue-engineered vascular grafts. However, how vascular mechanical forces regulate the genetic reprogramming in MSCs is not well understood. Mechanical strain in the vascular wall is anisotropic and mainly in the circumferential direction. We have shown that cyclic uniaxial strain on elastic substrates causes the cells to align perpendicularly to the strain axis, which is different from that in the vascular wall. To simulate the vascular cell alignment and investigate the anisotropic mechanical sensing by MSCs, we used soft lithography to create elastomeric membranes with parallel microgrooves. This topographic pattern kept MSCs aligned parallel to the strain axis, and the cells were subjected to 5% cyclic uniaxial strain (1 Hz) for 2-4 days. DNA microarray analysis revealed global gene expression changes, including an increase in the smooth muscle marker calponin 1, decreases in cartilage matrix markers, and alterations in cell signaling (e.g., down-regulation of the Jagged1 signaling pathway). In addition, uniaxial strain increased MSC proliferation. However, when micropatterning was used to align cells perpendicularly to the axis of mechanical strain, the changes of some genes were diminished, and MSC proliferation was not affected. This study suggests that mechanical strain plays an important role in MSC differentiation and proliferation, and that the effects of mechanotransduction depend on the orientation of cells with respect to the strain axis. The differential cellular responses to the anisotropic mechanical environment have important implications in cardiovascular development, tissue remodeling, and tissue engineering. stem cell engineering ͉ differentiation ͉ genetic reprogramming ͉ uniaxial mechanical strain ͉ micropatterning

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Section: Methodsmentioning

confidence: 99%

Anisotropic mechanosensing by mesenchymal stem cells

Kurpinski

Chu

Hashi

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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362

316

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“…Samples were fi xed in 4% paraformaldehyde and immunofl uorescently stained, as described previously [ 21 ], for the antibodies sGCα 3 (Sigma, St. Louis, MO), sGCβ 3 (Sigma), eNOS, or CD144 (BD Biosciences, San Diego, CA). Total nuclei were stained by Hoechst 33342 (Invitrogen, Carlsbad, CA) dye before imaging on a fl uorescent microscope (Nikon, Burlingame, CA), and images were captured with a SPOT RT color camera (Diagnostic Instruments, Sterling Heights, MI).…”

Section: Immunofl Uorescence Stainingmentioning

confidence: 99%

Role of Nitric Oxide Signaling in Endothelial Differentiation of Embryonic Stem Cells

Huang

Fleißner

Sun

et al. 2010

Stem Cells and Development

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Signaling pathways that govern embryonic stem cell (ESCs) differentiation are not well characterized. Nitric oxide (NO) is a potent vasodilator that modulates other signaling pathways in part by activating soluble guanylyl cyclase (sGC) to produce cyclic guanosine monophosphate (cGMP). Because of its importance in endothelial cell (EC) growth in the adult, we hypothesized that NO may play a critical role in EC development. Accordingly, we assessed the role of NO in ESC differentiation into ECs. Murine ESCs differentiated in the presence of NO synthase (NOS) inhibitor NG-nitroarginine methyl ester ( l -NAME) for up to 11 days were not signifi cantly different from vehicle-treated cells in EC markers. However, by 14 days, l -NAME-treated cells manifested modest reduction in EC markers CD144, FLK1, and endothelial NOS. ESC-derived ECs generated in the presence of l -NAME exhibited reduced tube-like formation in Matrigel. To understand the discrepancy between early and late effects of l -NAME, we assessed the NOS machinery and observed low mRNA expression of NOS and sGC subunits in ESCs, compared to differentiating cells after 14 days. In response to NO donors or activation of NOS or sGC, cellular cGMP levels were undetectable in undifferentiated ESCs, at low levels on day 7, and robustly increased in day 14 cells. Production of cGMP upon NOS activation at day 14 was inhibited by l -NAME, confi rming endogenous NO dependence. Our data suggest that NOS elements are present in ESCs but inactive until later stages of differentiation, during which period NOS inhibition reduces expression of EC markers and impairs angiogenic function.

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“…11 Consequently, changes in muscle tissue architecture in physiological (e.g., fiber rotation during shortening of pennate muscle 12 ) or pathological (e.g., muscle disarray and fibrosis in dystrophic muscle 13 ) conditions can significantly affect generation of muscle contractile force. To recreate native muscle architecture in vitro, researchers have previously applied different topographical, 14 chemical, [15][16][17] or physical 18 cues to align cells in 2D culture, or utilized cylindrically shaped cell-laden hydrogels or self-assembled organoids formed under static uniaxial tension to align muscle cells in 3D culture. 4,8,9,19 However, no current tissue engineering methodologies allow for precise and reproducible control of local 3D myofiber alignment that would enable systematic studies of structure-function relationship encountered in healthy, diseased, or developing skeletal muscle.…”

Section: Introductionmentioning

confidence: 99%

Local Tissue Geometry Determines Contractile Force Generation of Engineered Muscle Networks

Bian

Juhas

Pfeiler

et al. 2012

Tissue Engineering Part A

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The field of skeletal muscle tissue engineering is currently hampered by the lack of methods to form large muscle constructs composed of dense, aligned, and mature myofibers and limited understanding of structure-function relationships in developing muscle tissues. In our previous studies, engineered muscle sheets with elliptical pores (''muscle networks'') were fabricated by casting cells and fibrin gel inside elastomeric tissue molds with staggered hexagonal posts. In these networks, alignment of cells around the elliptical pores followed the local distribution of tissue strains that were generated by cell-mediated compaction of fibrin gel against the hexagonal posts. The goal of this study was to assess how systematic variations in pore elongation affect the morphology and contractile function of muscle networks. We found that in muscle networks with more elongated pores the force production of individual myofibers was not altered, but the myofiber alignment and efficiency of myofiber formation were significantly increased yielding an increase in the total contractile force despite a decrease in the total tissue volume. Beyond a certain pore length, increase in generated contractile force was mainly contributed by more efficient myofiber formation rather than enhanced myofiber alignment. Collectively, these studies show that changes in local tissue geometry can exert both direct structural and indirect myogenic effects on the functional output of engineered muscle. Different hydrogel formulations and pore geometries will be explored in the future to further augment contractile function of engineered muscle networks and promote their use for basic structure-function studies in vitro and, eventually, for efficient muscle repair in vivo.

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Myotube Assembly on Nanofibrous and Micropatterned Polymers

Cited by 300 publications

References 25 publications

Anisotropic mechanosensing by mesenchymal stem cells

Anisotropic mechanosensing by mesenchymal stem cells

Role of Nitric Oxide Signaling in Endothelial Differentiation of Embryonic Stem Cells

Local Tissue Geometry Determines Contractile Force Generation of Engineered Muscle Networks

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