Alternative functional in vitro models of human intestinal epithelia

Abstract

Physiologically relevant sources of absorptive intestinal epithelial cells are crucial for human drug transport studies. Human adenocarcinoma-derived intestinal cell lines, such as Caco-2, offer conveniences of easy culture maintenance and scalability, but do not fully recapitulate in vivo intestinal phenotypes. Additional sources of renewable physiologically relevant human intestinal cells would provide a much needed tool for drug discovery and intestinal physiology. We compared two alternative sources of human intestinal cells, commercially available primary human intestinal epithelial cells (hInEpCs) and induced pluripotent stem cell (iPSC)-derived intestinal cells to Caco-2, for use in in vitro transwell monolayer intestinal transport assays. To achieve this for iPSC-derived cells, intestinal organogenesis was adapted to transwell differentiation. Intestinal cells were assessed by marker expression through immunocytochemical and mRNA expression analyses, monolayer integrity through Transepithelial Electrical Resistance (TEER) measurements and molecule permeability, and functionality by taking advantage the well-characterized intestinal transport mechanisms. In most cases, marker expression for primary hInEpCs and iPSC-derived cells appeared to be as good as or better than Caco-2. Furthermore, transwell monolayers exhibited high TEER with low permeability. Primary hInEpCs showed molecule efflux indicative of P-glycoprotein (Pgp) transport. Primary hInEpCs and iPSC-derived cells also showed neonatal Fc receptor-dependent binding of immunoglobulin G variants. Primary hInEpCs and iPSC-derived intestinal cells exhibit expected marker expression and demonstrate basic functional monolayer formation, similar to or better than Caco-2. These cells could offer an alternative source of human intestinal cells for understanding normal intestinal epithelial physiology and drug transport.

Characteristic Analysis of Intestinal Transport in Enterocyte-Like Cells Differentiated from Human Induced Pluripotent Stem Cells

“…To our knowledge, there are few reports involving drug membrane permeability using human iPS-derived enterocytes (Kauffman et al, 2013;Ozawa et al, 2015). However, this evaluation used only TEER measurements and the permeability of a nonabsorbable marker (fluorescein isothiocyanate-dextran, molecular weight 4 or 150 kDa) in these reports.…”

“…However, the pharmacokinetic functions of organoids and intestinal cells were only briefly explored in these studies. Kauffman et al (2013) and Ozawa et al (2015) have reported the pharmacokinetic characteristics of human iPS-derived enterocyte-like cells, but the characteristics of intestinal transport have been insufficiently investigated.…”

Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology

“…Generation of more biologically-complex intestinal tissue has been accomplished through directed differentiation of (embryonic or induced) pluripotent stem cells to intestinal tissue (Kauffman et al, 2013;Ogaki et al, 2013). In general, pluripotent stem cells can be differentiated into all cells of the three embryonic germ layers.…”

Non-animal approaches for toxicokinetics in risk evaluations of food chemicals

“…Directed differentiation of human embryonic stem cells and induced pluripotent stem cells (iPSC) to intestine-like organoids with crypt-villus physiology and long-term culturing capacity has been achieved (Sato et al, 2011). However, current schemes for human intestine stem cells frequently rely on 3D culture conditions, whereas monolayer cultures are required for absorption experiments (Kauffman et al, 2013). Protocols are currently being optimized to achieve this goal (Astashkina and Grainger, 2014).…”