Abstract
Interactions between the different liver cell types are critical to the maintenance or induction of their function in vitro. In this work, human-induced Pluripotent Stem Cells (hiPSCs)-derived Liver Sinusoidal Endothelial Cells (LSECs) and Hepatocytes-Like Cells (HLCs) were cultured and matured in a microfluidic environment. Both cell populations were differentiated in Petri dishes, detached, and inoculated in microfluidic biochips. In cocultures of both cell types, the tissue has exhibited a higher production of albumin (3.19 vs 5.31 μg/mL/106 cells in monocultures and cocultures) as well as a higher inducibility CYP450 over monocultures of HLCs. Tubular-like structures composed of LSECs and positive for the endothelial marker PECAM1, as well as a tissue more largely expressing Stabilin-2 were detected in cocultures only. In contrast, monocultures exhibited no network and less specific endothelial markers. The transcriptomic analysis did not reveal a marked difference between the profiles of both culture conditions. Nevertheless, the analysis allowed us to highlight different upstream regulators in cocultures (SP1, EBF1, and GATA3) and monocultures (PML, MECP2, and NRF1). In cocultures, the multi-omics dataset after 14 days of maturation in biochips has shown the activation of signaling related to hepatic maturation, angiogenesis, and tissue repair. In this condition, inflammatory signaling was also found to be reduced when compared to monocultures as illustrated by the activation of NFKB and by the detection of several cytokines involved in tissue injury in the latter. Finally, the extracted biological processes were discussed regarding the future development of a new generation of human in vitro hepatic models.
Highlights
The differentiation of hepatic cells and the maintenance of their phenotypes in in vitro cocultures of hepatocytes and non-parenchymal cells has been widely studied (Bale et al, 2015)
The liver microvasculature is mostly composed of Liver Sinusoidal Endothelial Cells (LSECs) supported by Hepatic Stellate Cells (HSCs) in the space of Disse which separate them from hepatocytes (Arias, 2009)
To tackle the remaining issues in monocultures of Hepatocytes-Like Cells (HLCs), we propose in this manuscript an extension of our investigations by performing cocultures of human-induced Pluripotent Stem Cells (hiPSCs)-derived LSECs and HLCs in a fluidic microenvironment
Summary
The differentiation of hepatic cells and the maintenance of their phenotypes in in vitro cocultures of hepatocytes and non-parenchymal cells has been widely studied (Bale et al, 2015). These efforts aim at reproducing the complex structure of the liver, comprised of a multitude of cell types in close interaction. Primary hepatocytes (Bale et al 2015) and LSECs (Fraser et al, 1995) are known to rapidly lose their phenotype and metabolic functions in vitro which consist an important remaining issue in in vitro modeling
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