Fig. 1: Programmable presentation of integrin and N-cadherin adhesive domains to study hMSCs mechanosensing. | Nature Communications

Fig. 1: Programmable presentation of integrin and N-cadherin adhesive domains to study hMSCs mechanosensing.

From: Programmable integrin and N-cadherin adhesive interactions modulate mechanosensing of mesenchymal stem cells by cofilin phosphorylation

Fig. 1

a Schematic of RGD conjugation to PEG hydrogels by DNA hybridization. DNA strands were drawn as lines with arrows at the 3′ end. “*” indicates complementary sequences. b “ON” substrate allows for integrin-RGD adhesive interactions. c Characterization of RGD conjugation and hMSCs adhesion on “OFF” and “ON” substrates, as shown by (i) fluorescence images of FAM-labeled RGD (this experiment was repeated independently 3 times with similar results), (ii) bright-field, (iii) live-dead staining (green for live cells and red for dead cells) and (iv) immunostaining for paxillin (green), F-actin (red), and nuclei (blue). d Cell density (from left to right n = 15, 17 samples) and e spreading area of hMSCs (from left to right n = 163, 194 cells) on “ON” and “OFF” substrates. f Young’s moduli of “ON” and “OFF” substrates (n = 5 samples per group). g Quantification of hMSCs viability on “ON” and “OFF” substrates (from left to right n = 11, 13 samples). h Schematic of RGD removal from “ON” hydrogels using a fully complementary displacement strand through a toehold-mediated strand displacement reaction. Toehold domain was labelled with lowercase “c”. i Fluorescence images of removal of FAM-labeled RGD from the “ON” substrates (this experiment was repeated independently 3 times with similar results). j Schematic of programmable presentation of RGD and HAVDI. A hydrogel was modified with two different primary strands to bind to RGD and HAVDI, respectively. RGD-DNA1 and HAVDI-DNA2 bearing orthogonal toeholds allowed selective removal of either signal via its complementary displacement strand. Toehold domains were labelled with lowercase letters “c” and “f”. k “Dual ON” substrates allow for RGD/integrin and HAVDI/N-cadherin ligations. l Fluorescent tagging allowed for visualization of peptide conjugation and removal on the PEG substrates (this experiment was repeated independently 3 times with similar results). Data are presented as mean ± s.e.m., and p values were obtained using one-way ANOVA followed by Tukey’s post hoc test (d, e, f, g). Scale bars: 100 µm (c, i, l). Source data are provided as a Source Data file.

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