Journal: bioRxiv
Article Title: Self-organized vascularized human liver spheroids: Serum-free culture conditions and use as tissue building blocks
doi: 10.1101/2025.10.25.684548
Figure Lengend Snippet: A. Merged bright field and GFP fluorescence microscopy images of spheroids composed of HepaRG, GFP-HUVEC, and MSC cells at a ratio of 5:2:2 that were cultured in ECGM2 2%FCS, EH 2%FCS, or EH 4%SR for 4, 7, or 14 days. B. Projected area of spheroids cultured in ECGM2 2%FCS, EH 2%FCS, or EH 4%SR on days 4, 7, and 14 after cell seeding. A graphical representation with individual data points is shown in Supplemental Figure S1. (C, D). Numbers of Ki-67-positive (C) and E-cadherin-positive (D) cells within spheroids cultured in ECGM2 2%FCS, EH 2%FCS, or EH 4%SR on day 7 after seeding. E. Representative Ki-67 and E-cadherin immunofluorescence images of paraffin sections of spheroids cultured in ECGM2 2%FCS, EH 2%FCS, or EH 4%SR on day 7 after cell seeding. 4′,6-diamidino-2-phenylindole (DAPI) staining marks the cell nuclei. F. Representative fluorescence microscopy images showing GFP-positive vessel-like structures within spheroids cultured in EH 2%FCS or EH 4%SR for 7 days. Means ± SEM are shown. Scale bars: 100 µm. * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, n.s. – non-significant.
Article Snippet: 2.5 mg/ml collagen type I (C3867-1VL, Sigma-Aldrich) was neutralized with ∼9% v/v ice-cold 0.2 N NaOH (1091371000, Merck, Darmstadt, Germany) and 1x Medium 199 (M0650-100ML, Sigma-Aldrich), and mixed at a 1:1 ratio with spheroids in 1.2% methylcellulose (M0512-100G, Sigma-Aldrich, viscosity: 4,000 cP), prepared as described in Tetzlaff et al, 2018 [ ] in ECGM2 medium (C-22211, PromoCell).
Techniques: Fluorescence, Microscopy, Cell Culture, Immunofluorescence, Staining
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