Foregut Stem Cells


Scientists from Cambridge University have designed a new protocol that will convert pluripotent stem cells into primitive gut stem cells that have the capacity to differentiate into liver, pancreas, or some other gastrointestinal structure.

Nicholas Hannan and his colleagues at the University of Cambridge Welcome Trust MRC Stem Cell Institute have developed a technique that allows researchers to grow a pure, self-renewing population of stem cells that are specific to the human foregut, which is the upper section of the human digestive system. These types of stem cells are known as “foregut stem cells” and they can be used to make liver, pancreas, stomach, esophagus, or even parts of the small intestine. Making these types of gastrointestinal tissues can provide material for research into gastrointestinal abnormalities, but might also serve as a source of material to treat type 1 diabetes, liver disease, esophageal and stomach cancer, and other types of severe gastrointestinal diseases.

“We have developed a cell culture system which allows us to specifically isolate foregut stem cells in the lab,” said Hannan. “These cells have huge implications for regenerative medicine, because they are the precursors to the thyroid upper airways, lungs, liver, pancreas, stomach, and biliary systems.”

Hannan did this work in the laboratory of Ludovic Vallier, and they think that their technique will provide the means to analyze the precise embryonic development of the foregut in greater detail. “We now have a platform from which we can study the early patterning events that occur during human development to produce intestines, liver, lungs, and pancreas,” said Hannan.

To make foregut stem cells, Hannan begins with a pluripotent stem cell line; either an embryonic stem cell line or an induced pluripotent stem cell line. Then he differentiated them into definitive endoderm by treating them with CDM-PVA and activin-A (100 ng/ml), BMP4 (10 ng/ml), bFGF (20 ng/ml), and LY294002 (10 mM) for 3 days. Once they differentiated into endoderm, the endodermal cells were grown in RPMI+B27 medium with activin-A (50 ng/ml) for 3-4 days in order to generate foregut stem cells.

(A) GFP-expressing hPSCs were differentiated into hFSCs. (B) Single GFP-positive hFSCs were seeded onto a layer of non-GFP hFSCs and then expanded for five passages. The resulting population was then split into culture conditions inductive for liver or pancreatic differentiation. (C and D) GFP-hFSCs differentiated for 25 days were found to respectively generate cells expressing liver markers (ALB, LDL-uptake) and pancreatic markers (PDX1, C-peptide) from both hESC-derived (C) and hIPSC-derived (D) hFSCs.
(A) GFP-expressing hPSCs were differentiated into hFSCs.  (B) Single GFP-positive hFSCs were seeded onto a layer of non-GFP hFSCs and then expanded for five passages. The resulting population was then split into culture conditions inductive for liver or pancreatic differentiation.  (C and D) GFP-hFSCs differentiated for 25 days were found to respectively generate cells expressing liver markers (ALB, LDL-uptake) and pancreatic markers (PDX1, C-peptide) from both hESC-derived (C) and hIPSC-derived (D) hFSCs.

These foregut stem cells (FSCs) can self-renew, and can also differentiate into any part of the foregut. Thus, FSCs can grow robustly in culture, and they can also differentiate into foregut derivatives. However, these cells also do not form tumors. When injected into mice, they failed to form tumors.

(A) Large cystic hFSC outgrowth under the kidney capsule of a NOD-SCID mouse. (B) Cryosection of a hFSC outgrowth showing large cystic structures lined with epithelial cells. (C) Immunocytochemistry showing foregut outgrowths expressing EpCAM, PDX1, AFP, and NKX2.1. Scale bars, 100 μm or 50 μm as shown. See also Figure S4.
(A) Large cystic hFSC outgrowth under the kidney capsule of a NOD-SCID mouse.  (B) Cryosection of a hFSC outgrowth showing large cystic structures lined with epithelial cells.  (C) Immunocytochemistry showing foregut outgrowths expressing EpCAM, PDX1, AFP, and NKX2.1.  Scale bars, 100 μm or 50 μm as shown. See also Figure S4.

What are the advantages to FSCs as opposed to making pancreatic cells or liver cells from pluripotent stem cells? These types of experiments always create cultures that are impure. Such cultures are difficult to use because not all the cells have the same growth requirements and they would be dangerous for therapeutic purposes because they might contain undifferentiated cells that might grow uncontrollably and cause a tumor. Therefore, FSCs provide a better starting point to make pure cultures of pancreatic tissues, liver tissues, stomach tissues and so on.

Ludovic Vallier, the senior author of this paper said this of his FSCs, “What we have now is a better starting point – a sustainable platform for producing liver and pancreatic cells. It will improve the quality of the cells that we produce and it will allow us to produce the large number of uncontaminated cells we need for the clinical applications of stem cell therapy.”

Vallier’s groups is presently examining the mechanisms that govern the differentiation of FSCs into specific gastrointestinal cell types in order to improve the production of these cells for regenerative medicine.

Mesenchymal Stem Cells Found Around Blood Vessels in the Liver


Mesenchymal stem cells (MSCs) are found throughout the body and it is possible that every organ in our body has a MSC population. MSCs have the ability to differentiate into three main tissues: bone, fat and cartilage. However, the efficiency of this differentiation differs from one MSC population to another. Also, some MSCs can form smooth muscle for blood vessels and there is even evidence that MSCs can form blood vessels under certain conditions (for example, see Wingate K, Bonani W, Tan Y, Bryant SJ, Tan W. Acta Biomater. 2012 8(4):1440-9. doi: 10.1016/j.actbio.2011.12.032).

One of the places MSCs are usually found is around blood vessels. MSCs like to hang out on the outside of blood vessels in some tissues, and for this reason, MSCs are sometimes called “perivascular” stem cells.

One organ that has a stem cell population is the liver, but there is disagreement as to where they reside. Now a new publication has established that cells that hang out near blood vessels in liver are the MSC population in liver.

Eva Schmelzer from the McGowan Institute for Regenerative Medicine at the University of Pittsburgh has published a fine paper in the journal Stem Cells and Development detailing, with the help of her trusty laboratory colleagues, the characterization of liver MSCs.

Briefly, Schmelzer and her colleagues obtained fetal and adult lover tissue from tissue suppliers and minced them up, digested them with the appropriate enzymes, pushed them through cell strainers and then destroyed all the contaminating red blood cells. The remaining cells were grown in a cell culture medium. The stem cells would outgrow all the other cells, which would make their isolation and purification easy.

To purify the cells, Schmelzer’s co-workers used a technique called “flow cytometry.” When they had purified the liver MSCs, they set about characterizing them.

The liver MSCs grew quite well in culture and also grew quickly. They also expressed lots of surface proteins normally found on MSCs, confirming that they are MSCs. When gene expression experiments examined what genes these MSCs expressed, they expressed some smooth muscle genes and a several other genes enriched in cells near blood vessels. When Schmelzer examined cross sections of liver to determine where these cells are located, she found them curled up next to blood vessels.

In culture, the liver MSCs did not make very good cartilage or fat. However, they did make very good smooth muscle and bone. The efficiency of MSC differentiation tends to depend on where they were isolated. The rule of thumb is that MSCs most easily differentiate into those tissues that are closest to their own tissue of origin. Therefore, we would expect bone marrow MSCs to make better bone and cartilage than fat-based MSCs, and we would expect fat-based MSCs to make better fat than bone or liver-based MSCs. The ability of liver MSCs to be so good and making bone might be a little surprising, but when we consider that bone marrow stem cells begin their lives in the liver before they migrate to the bone marrow, perhaps this finding makes more sense.

In short, the adult and fetal liver contain a MSC population that is found on the outside of the blood vessels and these cells have an excellent capacity to make bone and smooth muscle for blood vessels. Thus liver biopsies might provide do more than provide material for diagnostic purposes – they might secure cells for regenerative purposes.