DefiniGEN’s intestinal monolayer product INTReady constitutes a ground-breaking new system to model the permeability of the human intestine and investigate drug efflux.
DefiniGEN’s intestinal monolayer product INTReady constitutes a ground-breaking new system to model the permeability of the human intestine and investigate drug efflux. The cells are grown as a monolayer in a similar manner to standard Caco-2 immortalized cell lines with the key benefit of greatly increased biological relevance, as the cells have primary human intestinal cell function, metabolism and genetics.
Figure 2. Def-INT Ready permeability assessment. Monolayer evaluation is consistent when using both TEER measurements and checking permeability of low permeability compounds such as Lucifer Yellow.
Biologically relevant system
Figure 3. Def-INTESTINAL monolayer cells express similar gene expression profiles to primary human intestinal cells.
Efflux transporter activities
Figure 4. Functional ABCB1 (MDR1) and BCRP transporter activity. INTReady monolayer cells exhibit functional efflux transporter activities when Digoxin and Hoechst 33324 were used as ABCB1 (A) and BCRP substrates respectively (B). These results indicate the presence of a polarized monolayer with functional apical transporter activities being observed.
CYP3A4 activity assay
Figure 5. CYP3A4 activity was measured in the Def-INT monolayer via their ability to hydrolyze testosterone into 6B-hydroxytestosterone. This activity was reduced to less than 40% derivative in the presence of a recognized inhibitor Ketoconazole.
Def-INT monolayer secretes mucus-forming proteins
Figure 6. Def-INT monolayer secretes mucus-forming proteins making it a more physiological model. Immunofluorescence shows presence of MUC2, a specific marker for goblet cells. Alcian blue staining demonstrates the presence of acid mucins while PAS staining shows presence of both acid (dark purple) and neutral mucins (bright magenta).
For validation, a set of compounds were screened with Caco-2 and Def-INT cells grown in monolayer. Permeability assay over a range of permeabilities show a good correlation with Caco-2. Def-INT grown monolayer shows better predictability for Furosemide than Caco-2.
Transepithelial electrical resistance analysis
Interaction of Salmonella enterica Serovar Typhimurium with Intestinal Organoids Derived from Human Induced Pluripotent Stem Cells
Generation of Multipotent Foregut Stem Cells from Human Pluripotent Stem Cells