Colloidal Templating of Highly Ordered Gelatin Methacryloyl- Based Hydrogel Platforms for Three-Dimensional Tissue Analogues

Transcription

1 Electronic Supplementary Information for Colloidal Templating of Highly Ordered Gelatin Methacryloyl- Based Hydrogel Platforms for Three-Dimensional Tissue Analogues Bae Hoon Lee 1,+, Hitomi Shirahama 1,+, Myung Hee Kim 1,+, Jae Ho Lee 1, Nam-Joon Cho 1,2*, Lay Poh Tan 1* 1. School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore School of Chemical and Biomedical Engineering Nanyang Technological University 62 Nanyang Avenue, Singapore Present Address: Dr. Bae Hoon Lee: Institute of Biomaterials and Engineering, Wenzhou Medical University, Wenzhou, Zhejiang , China + These authors contributed equally to the study. *Corresponding authors Prof. Lay Poh Tan School of Materials Science and Engineering Nanyang Technological University 50 Nanyang Avenue , Singapore LPTan@ntu.edu.sg (LPT) Prof. Nam-Joon Cho School of Materials Science and Engineering Nanyang Technological University 50 Nanyang Avenue , Singapore njcho@e.ntu.edu.sg (NJC) Prof. Nam-Joon Cho School of Chemical and Biomedical Engineering Nanyang Technological University 62 Nanyang Avenue , Singapore njcho@e.ntu.edu.sg (NJC) Submitted to NPG Asia Materials

2 Table S1. List of the primer sequences used in amplification. Target genes Forward (5-3 ) Reverse (5-3 ) AAT GATGCTGCCCAGAAGACAGA GGAGTTCCTGGAAGCCTTCA Albumin CTGCACAGAATCCTTGGTGA CTCCTTATCGTCAGCCTTGC CYP3A4 ACCGTGACCCAAAGTACTGG TTCAGGGGGATCTGTGTTTC CYP3A7 AAGTGGACCCAGAAACTGCA GGCTCCACTTACGGTCTCAT G6Pase TTCCTGTTCAGCTTCGCCAT TCAAAGACGTGCAGGAGGAC HNF4 TCGTTGAGTGGGCCAAGTAC TGTCATCGATCTGCAGCTCC HNF6 TTGAGCCATTGAGCGGACAT GGCAGGTTCAAACGTTAGGC E-cadherin AGGCCAAGCAGCAGTACATT AAATGTGTCTGGCTCCTGGG N-cadherin CCTTTCAAACACAGCCACGG TGTTTGGGTCGGTCTGGATG Claudin-1 TGGAAGACGATGAGGTGCAG GCTGGAAGGTGCAGGTTTTG ZO-1 AAACAGCCAGCCGTTAGTCA CTTCATACATGGGGACGCGA

3 Figure S1. The viscosity of GelMA solutions with different DS versus temperature. The sample solutions (30 w/v% in distilled water) contain 1% I2959 and were measured at 50 s -1. GelMA solutions showed the lowest viscosity above 37 o C. GelMA solutions can be easily infiltrated into interstices between polystyrene lattices at 40 o C at rpm.

4 Figure S2. Optical images of Huh-7.5 cell-laden GelMA (DS 98%) ICC scaffolds at day 1, 3, 6, and 9. Cell-laden GelMA ICC scaffolds showed adequate structural integrity over the culture to be easily handled with a tweezer.

5 Figure S3. Storage moduli of GelMA ICC scaffold with different DS at 30% w/v. They were measured at 0.1 Hz at 0.1 % strain. GelMA ICC scaffolds exhibited a range of 1-4 kpa storage moduli, depending on the DS of GelMA. They are relatively soft because of their porous structure but possess sufficient structural integrity.

6 Figure S4. Accelerated enzymatic degradation study of GelMA ICC scaffolds. Micro-scale surface morphology observation of GelMA ICC scaffolds with different DS at 4x (scale bars = 200 m) during enzymatic degradation at 1 mg/ml collagenase Type 1A solution (125 U/mL). The morphology of the microarchitecture of GelMA ICC scaffolds with a lower DS eroded more rapidly than that of GelMA ICC scaffolds with a higher DS.

7 Figure S5. Cell distribution in different layers (depths) of cross-sectional GelMA ICC from the top (the first layer) to bottom. The nucleus of Huh7.5 cells was stained with DAPI for imaging via fluorescence microscopy. The images are displayed in black and white for visual clarification. At day 1, cells were infiltrated into different layers and then the cells were accumulated dominantly in the first and second layers over time.

8 Figure S6. Cell proliferation in 3D GelMA ICC scaffolds and on 2D GelMA substrates, quantified by CCK-8 assay for 30 min incubation on day 1, 3, 6, and 9. Data were normalized to day one absorbance value of each group. The absorbance value of Huh7.5 cells in 2D GelMA substrates gradually increased up to day 6 and then dropped abruptly at day 9. On the other hand, the CCK values of cells in 3D GelMA ICC scaffolds changed little over the period of culture. Cells in 3D GelMA ICC scaffolds seemed to respond to CCK more slowly than cells in 2D GelMA substrates.