Every three to seven days, the intestinal epithelium undergoes a complete cellular turnover. This process is driven by intestinal stem cells, which have the capacity to generate all intestinal epithelial lineages. However, several intestinal diseases cause or are exacerbated by insufficient tissue regeneration or repair. By deciphering the true nature of the intestinal stem cells, researchers can potentially regenerate diseased cells and use cell therapy for some of these conditions.
NewYork-Presbyterian and Columbia gastroenterologists Timothy C. Wang, MD, Gastroenterology Division chief, and Kelley Yan, MD, recently published two studies in Cell that identify a new intestinal stem cell population, and both challenge the prevailing model of the intestinal stem cell population that is responsible for the renewal of the intestinal epithelium. Below, Drs. Wang and Yan share more about their recent discovery and how these studies will inform future research.
Challenges With the Prevailing Model
Since 2007, the prevailing model identified crypt-base columnar cells marked by expression of Lgr5 as the sole intestinal stem cells. However, over the years data emerged that raised questions about this model, with demonstration that there was intestinal regeneration even when Lgr5+ cells were completely absent. One widely held theory to explain these results was plasticity, where other differentiated cells could become stem cells in the absence of the Lgr5+ cells.
“I've been sort of skeptical of the standard Lgr5+ model for a number of years, as I kept finding inconsistencies with the claims that a few cells at the very bottom of the crypts were the primary stem cells,” says Dr. Wang. “In fact, I was seeing that the main source of proliferation and cell generation was higher up, and we had other results that seemed to suggest that the stem cells were located higher up in the crypts in a different position.”
A New Understanding of Intestinal Crypt Epithelial Cells
The goal of Dr. Wang’s study was to identify intestinal stem cells in an unbiased manner. To do this, his team took a highly purified collection of intestinal crypt epithelial cells and performed single-cell RNA sequencing. They used a computational program known as VIPER, developed by their co-senior author, Andrea Califano, PhD, the Clyde and Helen Wu Professor of Chemical and Systems Biology at Columbia. They used VIPER to fill in any unidentified genes to avoid critical gene dropout issues common with single-cell RNA sequencing. Then, in an unbiased way, the team clustered the cells into different groups and used the CytoTRACE algorithm to identify clusters with the most stem-like properties. Finally, they used lineage tracing with different markers to identify the locations of these cells.
“What we found, surprisingly, was that the clusters with the most stem-like properties did not overlap with what was considered the classical stem cells, the Lgr5+ cells,” says Dr. Wang. “And in fact, when we did a correlation, we found that Lgr5+ did not correlate.”
Further, Dr. Wang and his colleagues found that these cells were located higher up in the crypts than previously thought and had many properties consistent with stem cells. The key conclusion from the study was that the cells that people previously claimed were responsible for plasticity all overlapped with the stem cells higher up in the upper crypts.
Fgrbp1+ Marks Proliferative Upper Crypt Zone Cells
Dr. Yan’s study aimed to understand what cells, other than Lgr5+ cells, reside in the crypt compartment and have regenerative capacity. They also used single-cell RNA sequencing to determine what cell types reside in the tissue when they pharmacologically depleted Lgr5+ cells. From this, they identified a population of cells that were proliferative and completely different from the Lgr5+ cells, and the researchers developed mice lines to examine this population of cells and track what happens to them over time.
“What we found was that these new cells, which were completely different in their location as well as all the genes that they expressed, were actually the mother cells that gave rise to all the Lgr5+ cells that the rest of the world believed were the actual stem cells,” says Dr. Yan.
Further, they found that Fgfbp1+cells that reside in the upper portion of the crypt above the Lgr5+ cells are responsible for daily turnover and persist for the mouse’s lifetime.
Next Steps
Within the next several years, Dr. Wang and Dr. Yan hope to look more closely at how these new hypotheses hold up in human tissue and want to start growing human intestinal stem cells. Says Dr. Wang, “Being at an academic medical center like NewYork-Presbyterian with such great clinicians really gives us the opportunity to begin to bring these discoveries closer to the clinic, which is our long-term goal.”
