Project description:Human intestinal organoids (hIOs) derived from human pluripotent stem cells (hPSCs) have immense potential as a source of intestines. However, due to problems obtaining mature adult-like cells from hPSCs, effective strategies for the maturation of hIOs with improved functionalities must be developed. We established conditions that enable the in vitro maturation of hIOs derived from hPSCs, recapitulating the essential features of intestinal tissue. The maturity of the hPSC-derived hIOs was enhanced by the in vitro maturation in our co-culture system, evidenced by transcriptional changes observed in our microarray data. Global transcription of in vitro-matured hIOs is highly similar to that of adult human small intestine (hSI). Compared to control hIOs, in vitro-matured hIOs exhibited significant increases in the expression of intestinal maturation markers, major transporters, and key enzymes for drug absorption and first-pass metabolism.
Project description:In vitro human pluripotent stem cell derived intestinal organoids (HIOs) are immature and lack for diverse differentiated secretory cell types. We would like to test the hypothesis whether addition of a mesenchyme secreting ligand which is depleted in canonical organoid culture media could increase the maturity and secretory cell type diversity in HIOs in vitro. To do this, we adapted the directed differentiation protocol of HIOs by growing HIOs in media with EGF, NOGGIN, R-spondin-1 (ENR) for 30 days, isolated epithelial cells with dispase, recovered them with adult intestinal enteroid media with Wnt-3A (WENR). Then we introduced the mesenchyme secreting ligand NRG1 to the established enteroid culture (WENR+NRG1) and compared them to the enteroids grown in control condition (WENR).
Project description:Human intestinal organoids (hIOs) resemble the human intestine physiologically and structurally. We recently present an in vitro maturation technique for generating mature and functional hIOs from human pluripotent stem cells (hPSCs). Here, we investigated the mechanisms of STAT3 for inducing in vitro maturation of hIOs. Using CRISPR/Cas9-mediated gene editing, STAT3 knockout (KO) human embryonic stem cell (hESC) lines were generated and characterized. By genome-wide microarray analyses, STAT3 KO hIOs showed markedly different profiles from the in vitro matured hIOs and human small intestine and the majority of genes, which are associated with intestinal development and functions, were downregulated by STAT3 KO. This study reveals important signaling pathways for the maturation of hIOs derived from hPSCs.
Project description:Here, we used single cell RNA-sequencing (scRNA-seq) to profile pluripotent stem cell derived human intestinal organoids (HIOs) grown in matrigel or a non-adhesive alginate hydrogel after 28 days of in vitro growth. Additionally, we used scRNA-seq to profile HIOs derived in the presence of Neuregulin 1 (NRG1) and/or EGF after 40 days of in vitro growth.
Project description:Here, we used single-cell RNA-sequencing (scRNA-seq) to profile pluripotent stem cell derived human intestinal organoids (HIOs) grown in suspension culture after 28 days of in vitro growth. Grown in minigut media supplemented with EGF.
Project description:Here, we used single-cell RNA-sequencing (scRNA-seq) to profile pluripotent stem cell-derived human intestinal organoids (HIOs) grown in media comprised of minigut media + varying concentrations of Epiregulin (EREG) 1 ng/ml, 10 ng/ml, 100 ng/ml after 28 days of in vitro growth.
Project description:Here, we used single-cell RNA-sequencing (scRNA-seq) to profile pluripotent stem cell derived human intestinal organoids (HIOs) grown in an alginate matrix after 3, 7, and 14 days of in vitro growth. Samples were grown in minigut media supplemented with either ENR or EGF.
Project description:To compare miRNA expression profiles between freshly isolated intestinal epithelial cells and cultured organoids in mice. Intestinal organoids largely resembled intestinal epithelial cells in their miRNA profiles. Although the expression levels of some miRNAs were different between crypt and villus epithelial cells, such expression patterns were not reproduced during the maturation of intestinal organoids.
Project description:Retinoic acid promotes the in vitro growth, patterning and improves the cellular composition of human pluripotent stem-cell-derived intestinal organoids