Induced Pluripotent Stem Cells Differentiated into Intestinal Cells

Even the liver is the main organize when it comes to the metabolization of drugs, the small intestine also plays an important role in all aspects of drug metabolism. Unfortunately, no laboratory system exists at present that serves as a standardized system for evaluating the way drugs interact with the small intestine.

A new study by Tamihide Matsubara and his colleagues from Nagoya City University in Japan has sought to alleviate this problem. Matsubara and his coworkers used human induced pluripotent stem (iPS) cells to produce functional human intestinal enterocytes and showed that they faithfully recapitulated the drug metabolism of normal, human intestinal enterocytes.

To make intestinal enterocytes from iPS cells, Matsubara and others treated these cells with chemicals called activin A and fibroblast growth factor 2 to drive the cells to become intestinal-like stem cells. These cultured intestinal-like stem cells them differentiated into enterocytes when grown in a culture medium that contained epidermal growth factor and other small-molecule compounds.

The differentiated cells expressed intestinal marker genes and drug transporters. For example, they expressed sucrase-isomaltase, an intestine-specific marker, and enterocyte drug-metabolizing enzymes such as CYP1/2, CYP2C9, CYP2C19, CYP2D6, CYP3A4/5, UGT, and SULT. Inhibitor studies showed that the intestinal oligopeptide transporter SLC15A1/PEPT1 was inhibited by the pain reliever ibuprofen, just like in naturally-occurring enterocytes. Also, active forms of vitamin D increased the expression of the enzymes CYP3A4 and CYP3A4/5, which is also observed in naturally-occurring human enterocytes.

These results show that Matsubara and his colleagues have successfully generated enterocyte-like cells that have the same drug metabolizing capacities as naturally-occurring enterocytes. These cells would be very useful for developing novel evaluation systems to predict individual human intestinal drug metabolism.

Induced Pluripotent Stem Cells Make Lungs

Since my father died of disseminated lung cancer (squamous cell carcinoma), this report has particular meaning to me.

When a person dies, their lungs can be harvested and stripped of their cells. This leaves a so-called “lung scaffold” that can then be used to build new lungs by means of tissue engineering techniques. Lung scaffolds consist of a protein called collagen, and sugar-rich proteins called “proteoglycans” (say that fast five times) and a rubber band-like protein called elastin. Depending on how the lung scaffolds are made more or less of these components can remain in the lung scaffold (see TH Peterson, and others, Cells Tissues Organs. Feb 2012; 195(3): 222–231). The important thing is that the cells are gone and this greatly reduces the tendency for the lung scaffold to be rejected by someone else’s immune system.

Once a lung scaffold is generated from a whole lung, cells can be used to reconstitute the lung. The key is to use the right cell type or mix of cell types and to induce them to form mature lung tissue.

The laboratory of Harald Ott at Harvard University Medical School used a technique called “perfusion decellularization” to make lung scaffolds from the lungs of cadavers. Then he and his co-workers used lung progenitor cells that were derived from induced pluripotent stem cells (iPSCs). This study was published in The Annals of Thoracic Surgery, and it examined the ability of iPSCs to regenerate a functional pulmonary organ

Whole lungs from rat and human cadavers were stripped of their living material by means of constant-pressure perfusion with a strong detergent called sodium dodecyl sulfate (SDS; 0.1% if anyone is interested). Ott and his crew then sectioned some of the resulting lung scaffolds and left others intact, and then applied human iPSCs that had been differentiated into developing lung tissue.

Lung tissue develops from the front part of the developing gut. This tissue is called “endoderm,” since it is in the very innermost layer of the embryo.

Lung Development

Therefore, the iPSCs were differentiated into endoderm with a cocktail of growth factors (FGF, Wnt, Retinoic acid), and then further differentiated in the anterior endoderm (foregut; treated cells with Activin-A, followed by transforming growth factor-β inhibition), and then even further differentiated into anterior, ventral endoderm, which is the precise tissue from which lungs form. In order to be sure that this tissue is lung tissue, they must express a gene called NK2 homeobox 1 (Nkx2.1). If these cells express this gene, then they are certainly lung cells.

Ott and his group showed that their differentiate iPSCs strongly expressed Nkx2.1, and then seeded them on slices and whole lung scaffolds. Then Otts’s group maintained these tissues in a culture system that was meant to mimic physiological conditions.

Those cells cultured on decellularized lung slices divided robustly and committed to the lung lineage after 5 days. Within whole-lung scaffolds and under the physiological mimicking culture, cells upgraded their expression of Nkx2.1. When the culture-grown rat lungs were transplanted into rats, they were perfused and ventilated by host vasculature and airways.

Thus these decellularized lung scaffolds supports the culture and lineage commitment of human iPSC-derived lung progenitor cells. Furthermore, whole-organ scaffolds and a culture system that mimics physiological conditions, allows scientists to enable seeding a combination of iPSC-derived endothelial and epithelial progenitors and enhance early lung fate. Transplantation of these laboratory-grown lungs seem to further maturation of these grafted lung tissues.