Fig. 5

Tissue cross-linked hydrogels (TxGels) and in vivo CxGels formation. a (left panels) CLSM images of normal mouse tissues treated with FITC-bAlg-DBCO solution. These tissues were carefully excised, then shredded and suspended with 500 µL of FITC-labeled bAlg-DBCO (0.1%) solution at 37 °C for 30 min. After that, tissues were centrifuged and washed with PBS twice, then tissues were re-suspended with 1 mL of Live Cell Imaging Solution and observed by CLSM. Scale bars indicate 100 μm. (middle panels) CLSM images of azide-modified mouse tissues treated with FITC-bAlg-DBCO solution. Mice treated with AC₄ManNAz for continuous 7 days were killed and the tissues were excised carefully. These tissues were shredded and washed with PBS twice. These tissues were carefully excised, then shredded and suspended with 500 µL of FITC-labeled bAlg-DBCO (0.1%) solution at 37 °C for 30 min. After that, tissues were centrifuged and washed with PBS twice, then tissues were re-suspended with 1 mL of Live Cell Imaging Solution and observed by CLSM. Scale bars indicate 100 μm. (right panels) Photographs of tissue cross-linked hydrogels (TxGels) made through click reaction between azide-modified tissues and bAlg-DBCO solution (2%) ex vivo. Scale bars indicate 100 μm. b Photographs of in vivo formed CxGels. LifeAct-GFP-expressing azide-modified C2C12 cells (4.0 × 10⁶) were suspended with 200 μL of bAlg-DBCO solution (2%) and immediately injected subcutaneously into the muscle layer of the back of each mouse. c Force recovery of injured femoral muscle treated with CxGels prepared through click reaction between LifeAct-GFP-expressing azide-modified C2C12 cells (5.0 × 10⁶) and 250 μL of bAlg-DBCO (2%) or C2C12 cells-encapsulating control physical gels, prepared through physical cross-linking reaction between 250 μL of bAlg solution (2%) and CaCl₂ solution (0.5%) in the presence of azide-modified C2C12 cells (5.0 × 10⁶ cells). Error bars: standard deviation (n = 3). d Fluorescence images of LifeAct-GFP-expressing C2C12 cells in the CxGels or the control physical gels acquired 15 days after transplantation into injured muscle. Green: LifeAct-GFP, blue: Hoechst. Scale bars indicate 50 μm