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. 2023 Nov 27;12(23):2718. doi: 10.3390/cells12232718

Figure 1.

Figure 1

Figure 1

Small hepatocytes (SHs) in vitro and in vivo. (A) Photos of SHs isolated from a rat liver (Aa–Ac) and a human liver tissue (Ad–Af). A photo of an SH colony consisting of small-sized cells showing a flat surface 13 days after plating (Aa). A colony consisting of piled-up cells is surrounded by NPCs cultured for 27 days (Ab). A colony of rat HPPCs of the third passage cultured on thin-Matrigel 21 days after replating (Ac). Primary human hepatocytes isolated from a healthy part of the surgically dissected liver tissue are cultured for 50 days. Many mononuclear SHs are proliferating (Ad). Primary human hepatocytes proliferate to form a large colony and part of the cells pile up on the colony 31 days after plating (Ae). Primary human hepatocytes are cultured in the serum-free medium on a hyaluronan-coated dish for 21 days. A colony consisting of SHs is formed (Af). (B) Cells of an SH fraction isolated from a healthy rat liver are plated on the dishes coated with rat tail collagen (Ba), collagen type IV (Bb), fibronectin (Bc), thin-Matrigel (Bd), and collagen gel (Be,Bf). Cells were cultured in the modified Dulbecco’s modified Eagle medium (DMEM)/F12 medium with 10% fetal bovine serum (FBS) for 20 days. From 4 days after plating, 1% DMSO was added to the medium (Ba–Bd,Bf). Cells were fixed with absolute ethanol at day 20 and immunocytochemically stained with Krt8. Nuclei were stained with hematoxylin. SH colonies show Krt8-positive (Brown). (C) SHs appear in liver tissue (in vivo). (Ca) A cluster of small hepatocyte-like progenitor cells (SHPCs) is observed in the rat liver treated with retrorsine (Ret) and 2/3 PH (HE-staining). (Cb) A photo of the rat liver treated with d-galactosamine at 3 days after administration. (Cc) A focus of SHs observed in the 99-week-old rat liver. (Cd) Magnified image of the area surrounded by white lines in the photo (Cb). (Ce) A photo of the rat liver treated with d-galactosamine at 4 days after administration. (Cb,Cd,Ce) White arrowheads may indicate OC-SHs emerging near elongating bile ductules and yellow arrows may show MH-SHs near resident MHs. Bars show 50 μm.