Preparation of Hydrogel Substrates with Tunable Mechanical Properties

Tse, Justin R.; Engler, Adam J.

doi:10.1002/0471143030.cb1016s47

Cited by 856 publications

(963 citation statements)

References 33 publications

Supporting

Mentioning

942

Contrasting

Unclassified

Order By: Relevance

“…1). Mechanically tunable PA gels were made by adjusting the total polymer content (%T) and cross-linker concentration (%C) in solution (33). The elastic moduli of the two gel formulations used in this study were verified by atomic force microscopy (AFM): 10%T, 1%C, 27.5-31.8 kPa (median: 29.07 kPa) and 10%T, 2.5% C, 52.6-67.2 kPa (median: 57.34 kPa) (Table S1).…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Changes in E-cadherin rigidity sensing regulate cell adhesion

Collins

Denisin

Pruitt

et al. 2017

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

View full text Add to dashboard Cite

Mechanical cues are sensed and transduced by cell adhesion complexes to regulate diverse cell behaviors. Extracellular matrix (ECM) rigidity sensing by integrin adhesions has been well studied, but rigidity sensing by cadherins during cell adhesion is largely unexplored. Using mechanically tunable polyacrylamide (PA) gels functionalized with the extracellular domain of E-cadherin (Ecad-Fc), we showed that E-cadherin-dependent epithelial cell adhesion was sensitive to changes in PA gel elastic modulus that produced striking differences in cell morphology, actin organization, and membrane dynamics. Traction force microscopy (TFM) revealed that cells produced the greatest tractions at the cell periphery, where distinct types of actin-based membrane protrusions formed. Cells responded to substrate rigidity by reorganizing the distribution and size of hightraction-stress regions at the cell periphery. Differences in adhesion and protrusion dynamics were mediated by balancing the activities of specific signaling molecules. Cell adhesion to a 30-kPa Ecad-Fc PA gel required Cdc42-and formin-dependent filopodia formation, whereas adhesion to a 60-kPa Ecad-Fc PA gel induced Arp2/3-dependent lamellipodial protrusions. A quantitative 3D cell-cell adhesion assay and live cell imaging of cell-cell contact formation revealed that inhibition of Cdc42, formin, and Arp2/3 activities blocked the initiation, but not the maintenance of established cell-cell adhesions. These results indicate that the same signaling molecules activated by E-cadherin rigidity sensing on PA gels contribute to actin organization and membrane dynamics during cell-cell adhesion. We hypothesize that a transition in the stiffness of E-cadherin homotypic interactions regulates actin and membrane dynamics during initial stages of cellcell adhesion.C ell adhesion is essential for tissue structure and function. Cells use specialized types of adhesions to interact with the surrounding environment, including integrin-based focal adhesions at cell-extracellular matrix (cell-ECM) contacts and cadherin-based adhesions at cell-cell contacts (1). Integrins bind to the ECM and intracellular proteins that link to the actin cytoskeleton and important signaling pathways (2). Similarly, cadherins regulate cellcell recognition and adhesion (3) and, through cytoplasmic adaptor proteins (catenins, vinculin) (4, 5), also link to the actin cytoskeleton and other proteins with signaling and scaffolding functions (6).Initiation of cell-cell adhesion requires significant reorganization of the actin cytoskeleton and is tightly controlled by the activities of actin nucleating proteins and Rho GTPases. Adhesion is initiated when filopodia from opposing cells come into contact with one another (7,8), and this process is regulated by Cdc42 activity (9, 10) and formin-dependent actin polymerization (11)(12)(13)(14). Intermediate stages of cell-cell contact formation involve lateral expansion of the contact by Rac1-induced and Arp2/3-dependent lamellipodial activity (15, 16). Finally, com...

show abstract

Section: Resultsmentioning

confidence: 99%

“…Briefly, PA gels of varying elastic moduli were made by adjusting the amounts of acrylamide and bis-acrylamide in the prepolymer gel solution (33). Gels were functionalized by UV activation of 0.5-2.0 mg/mL SS (72) and subsequent incubation with collagen I or protein A/G and Ecad-Fc.…”

Section: Methodsmentioning

confidence: 99%

Changes in E-cadherin rigidity sensing regulate cell adhesion

Collins

Denisin

Pruitt

et al. 2017

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

View full text Add to dashboard Cite

show abstract

“…Substrate fabrication and surface activation PA gel was made according to the method described by Tse and Engler [22]. Briefly, aminosilanated coverslips (diameter was 22 mm) were obtained by treating with 0.1M NaOH, 3% 3-aminopropyltriethoxysilane (Sigma) in deionized (DI) water, and 0.5% (v/v) glutaraldehyde (Sigma) in phosphate-buffered saline (PBS) as described in Supplementary Fig.…”

Section: Methodsmentioning

confidence: 99%

Insights into the Role of Focal Adhesion Modulation in Myogenic Differentiation of Human Mesenchymal Stem Cells

Lui

Xiong

et al. 2013

Stem Cells and Development

View full text Add to dashboard Cite

We report the establishment of a novel platform to induce myogenic differentiation of human mesenchymal stem cells (hMSCs) via focal adhesion (FA) modulation, giving insights into the role of FA on stem cell differentiation. Micropatterning of collagen type I on a polyacrylamide gel with a stiffness of 10.2 kPa efficiently modulated elongated FA. This elongated FA profile preferentially recruited the b 3 integrin cluster and induced specific myogenic differentiation at both transcription and translation levels with expression of myosin heavy chain and a-sarcomeric actin. This was initiated with elongation of FA complexes that triggered the RhoA downstream signaling toward a myogenic lineage commitment. This study also illustrates how one could partially control myogenic differentiation outcomes of similar-shaped hMSCs by modulating FA morphology and distribution. This technology increases our toolkit choice for controlled differentiation in muscle engineering.

show abstract

“…This was suggested to be due to the refractive stretching of the precursor along Z-direction. Pore sizes were in the ideal range of 200-400 μm, known to facilitate osteo-differentiation and functional bone formation [12,57,58]. The translation of the STL model to the structure (see figure 5) was good and reproducible in terms of pores and throat sizes achieved (see table 3).…”

Section: Manufacturing Of 3d Scaffoldsmentioning

confidence: 88%

“…Traditional scaffold fabrication techniques have included the production of porous polymer constructs by freeze-extraction and -gelation [5], electrospinning [6], melt electrospinning [7,8] or with hydrogels [9][10][11][12] as substrates for cell attachment.…”

Section: Introductionmentioning

confidence: 99%

In vitro degradation and mechanical properties of PLA-PCL copolymer unit cell scaffolds generated by two-photon polymerization

et al. 2016

View full text Add to dashboard Cite

The manufacture of 3D scaffolds with specific controlled porous architecture, defined microstructure and an adjustable degradation profile was achieved using two-photon polymerization (TPP) with a size of 2 × 4 × 2 mm3. Scaffolds made from poly(D,L-lactide-co-ɛ-caprolactone) copolymer with varying lactic acid (LA) and ɛ -caprolactone (CL) ratios (LC16:4, 18:2 and 9:1) were generated via ring-opening-polymerization and photoactivation. The reactivity was quantified using photo-DSC, yielding a double bond conversion ranging from 70% to 90%. The pore sizes for all LC scaffolds were see 300 μm and throat sizes varied from 152 to 177 μm. In vitro degradation was conducted at different temperatures; 37, 50 and 65 °C. Change in compressive properties immersed at 37 °C over time was also measured. Variations in thermal, degradation and mechanical properties of the LC scaffolds were related to the LA/CL ratio. Scaffold LC16:4 showed significantly lower glass transition temperature (Tg) (4.8 °C) in comparison with the LC 18:2 and 9:1 (see 32 °C). Rates of mass loss for the LC16:4 scaffolds at all temperatures were significantly lower than that for LC18:2 and 9:1. The degradation activation energies for scaffold materials ranged from 82.7 to 94.9 kJ mol−1. A prediction for degradation time was applied through a correlation between long-term degradation studies at 37 °C and short-term studies at elevated temperatures (50 and 65 °C) using the half-life of mass loss (Time (M1/2)) parameter. However, the initial compressive moduli for LC18:2 and 9:1 scaffolds were 7 to 14 times higher than LC16:4 (see 0.27) which was suggested to be due to its higher CL content (20%). All scaffolds showed a gradual loss in their compressive strength and modulus over time as a result of progressive mass loss over time. The manufacturing process utilized and the scaffolds produced have potential for use in tissue engineering and regenerative medicine applications.

show abstract

Preparation of Hydrogel Substrates with Tunable Mechanical Properties

Cited by 856 publications

References 33 publications

Changes in E-cadherin rigidity sensing regulate cell adhesion

Changes in E-cadherin rigidity sensing regulate cell adhesion

Insights into the Role of Focal Adhesion Modulation in Myogenic Differentiation of Human Mesenchymal Stem Cells

In vitro degradation and mechanical properties of PLA-PCL copolymer unit cell scaffolds generated by two-photon polymerization

Contact Info

Product

Resources

About