Human Intestinal iPSC Organoids - Definigen

Intestinal Organoids

DefiniGEN’s iPSC-derived intestinal organoids provide a unique in vitro system to model the human intestine.
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Product Overview

DefiniGEN’s iPSC-derived intestinal organoids provide a unique in vitro system to model the human intestine. The organoids display a polarized epithelium and harbor a mixture of cell types normally present in the primary intestinal epithelial barrier in vivo, including goblet cells, Paneth cells, enterocytes, and enteroendocrine cells. The cells can be used for drug absorption, metabolism, induction of transporters, and the modelling of infectious disease.

Benefits
  • Highly standardized cell product containing human intestinal organoids with consistent performance and biologically relevant data
  • Wild-type donor genetics and karyotype verified
  • Multiple cell types identified in the intestinal organoids including epithelial cells, enterocytes, goblet cells, enteroendocrine cells, and Paneth cells
  • Cells display multiple key gut markers OLFM4, CHGA, MUC2, Villin and KRT19
  • Cells display CYP450 induced activities
  • Organoids can be maintained long-term in culture through passaging
Technical Data
Intestinal cell morphology

Typical intestinal organoid morphology is observed in Def-INTESTINAL cells. The organoids initially form spheroid structures which over successive passages develop the crypt architecture characteristic of primary human intestinal organoids.

DefiniGEN intestinal morphology
Figure 1. Typical intestinal morphology is observed in Def-INTESTINAL cells. Organoids grown encapsulated in matrigel in a 24 well plate.
Immunocytochemistry analysis

Immunocytochemistry analysis has demonstrated that the Def-INTESTINAL organoids display a polarized epithelium and are composed of differentiated cell types with distinct morphologies. The organoids contain absorptive enterocytes as well as the major secretory lineage cell types including Paneth cells, goblet cells, and enteroendocrine cells.

DefiniGEN intestinal immunocytochemistry
Figure 2. Organoids display specific marker gene expression profiles. Organoids display positive staining of key intestinal cell markers including: epithelial cells (E-cadherin), enterocytes (villin), goblet cells (mucin), enteroendocrine (somatostatin), and Paneth cells (lysozyme).

Key intestinal cell marker analysis

DefiniGEN intestinal marker analysis
Figure 3. Def-INTESTINAL organoids have been demonstrated to display multiple key gut markers. Gene expression analysis shows the intestinal markers KRT19, Villin, and CHGA have similar expression profiles in Def-INTESTINAL and the primary control relative to GAPDH.
Metabolic enzyme activity
DefiniGEN CYP activity in intestinal organoids
Figure 4. A range of cytochrome P450 enzymes have been identified in Def-INTESTINAL organoids. Gene expression analysis shows cytochromes CYP3A4, CYP2D6, and CYP2J2 expression against primary human control.


Phase I and Phase II metabolism

Phase I and II Metabolism DefiniGEN Intestinal Organoids
Figure 5. CYP3A4 gene expression and activity in Def-INT cells. Organoids show upregulation of both gene expression (A) and activity (B) of CYP3A4 when induced Rifampicin and Vitamin D3 for 48 and 72h in culture respectively. Activity measured with CYP3A4 P450-Glo assay.
Phase I and Phase II metabolism

CES2 activity GST activity UGT activity in DefiniGEN intestinal organoids
Figure 6. A) Intestinal-specific isoform Carboxylesterase 2 is active in Def-INT organoids as their ability to hydrolyse CDCFDA into CDCF can be specifically inhibited by Loperamide, a well known CES2 inhibitor. B) GST activity in Def-INT organoids lysates is comparable to that of human small intestine S9 fraction. Activity measured by conjugation of CDNBC into CDNB-SG. C) UGT activity measured by glucuronidation of 4-MU in Def-INT organoids and Monolayer cells are higher to values of Caco-2 found in the literature.

Drug transporter analysis

Down-regulation or inhibition of ABC efflux transporters in the intestine can be used as a strategy to improve oral drug bioavailability of known substrates as these transporters prevent drug molecules from being absorbed. The SLC (solute carrier) family have an important role in physiological processes ranging from the cellular uptake of nutrients to the absorption of drugs and other xenobiotics. SLCs are primarily involved in the uptake of small molecules into cells.

Drug transporter analysis DefiniGEN Intestinal Organoids
Figure 7. Def-INTESTINAL ABCB1 and SLCO2B1 gene expression analysis. (A) Gene expression analysis shows transporter ABCB1 expression profiles against primary control. (B) Gene expression analysis shows transporter SLCO2B1 expression profiles against primary control.

A)                                                                                                                      B)

MDR1 transporter localization DefiniGEN intestinal organoids
Figure 8. Def-INTESTINAL MDR1 transporter analysis. (A) Def-INTESTINAL organoids can transport Rhodamine 123, a specific substrate of MDR1. MDR1 activity is inhibited by Verapamil a specific inhibitor. (B) Immunostaining of intestinal organoids showing localization of the MDR1 transporter protein within highly folded crypt structures.