A strategy to track and examine a single blood stem cell in its microenvironment using multiple imaging techniques has been developed. By integrating confocal microscopy, X-ray microscopy, and serial block-face scanning electron microscopy, the approach allows scientists to examine once-elusive cell-cell interactions occurring in this space.

“Using our new techniques, we can now see not only the stem cell, but also all the surrounding niche cells that are in contact,” explained Owen Tamplin, PhD, an assistant professor at the University of Wisconsin–Madison who co-led the work with Mark Ellisman, PhD, a professor of neuroscience at the University of California, San Diego. They published their work in eLife in an article entitled, “Defining the ultrastructure of the hematopoietic stem cell niche by correlative light and electron microscopy.”

The niche is a microenvironment found within tissues like the bone marrow that contain the blood stem cells that support the blood system. It is where specialized interactions between blood stem cells and their neighboring cells occur every second, but these interactions are hard to track and not clearly understood.

“Currently, we look at stem cells in tissues with a limited number of markers and at low resolution, but we are missing so much information,” Tamplin said. “This [new strategy] has allowed us to identify cell types in the microenvironment that we didn’t even know interacted with stem cells, which is opening new research directions.”

Although blood stem cells are difficult to locate in most living organisms, the zebrafish larva, which is transparent, offers researchers a unique opportunity to view the inner workings of the blood stem cell niche more easily. In this case, Tamplin, Ellisman, and their team studied early migration events of hematopoietic stem and progenitor cells (HSPCs) into the presumptive adult kidney marrow (KM) niche in zebrafish larvae.

“That’s the really nice thing about the zebrafish and being able to image the cells,” Tamplin said of the animal’s transparent quality. “In mammals, blood stem cells develop in utero in the bone marrow, which makes it basically impossible to see those events happening in real time. But, with zebrafish, you can actually watch the stem cell arrive through circulation, find the niche, attach to it, and then go in and lodge there.”

Using their imaging strategy, they found it possible to track a single putative HSPC deep in live tissue and observe the larval KM niche during the earliest stages of HSPC colonization.

“First, we identified single fluorescently labeled stem cells by light sheet or confocal microscopy,” explained Tamplin. “Next, we processed the same sample for serial block-face scanning electron microscopy. We then aligned the 3D light and electron microscopy datasets. By intersecting these different imaging techniques, we could see the ultrastructure of single rare cells deep inside a tissue. This also allowed us to find all the surrounding niche cells that contact a blood stem cell. We believe our approach will be broadly applicable for correlative light and electron microscopy in many systems.”

Tamplin hopes that this approach can be used for many other types of stem cells, such as those in the gut, lung, and the tumor microenvironment, where rare cells need to be characterized at nanometer resolution.

“This research will really help us to understand how stem cells behave and function,” Tamplin said. “A better understanding of stem cell behavior, and regulation by surrounding niche cells, could lead to improved stem cell-based therapies.”

“Transplanted blood stem cells are used as a curative therapy for many blood diseases and cancers, but blood stem cells are very rare and difficult to locate in a living organism,” he added. “That makes it very challenging to characterize them and understand how they interact and connect with neighboring cells.”

The results of their work indicated that there is heterogeneity in the niches where the HSPCs lodged. “We found a variety of different configurations of HSPCs and surrounding niche cells, suggesting there could be functional heterogeneity in sites of HSPC lodgement,” they wrote. “Our approach also allowed us to identify dopamine beta-hydroxylase (dbh) positive ganglion cells as a previously uncharacterized functional cell type in the HSPC niche.”

Future work will provide more information to determine the degree and functional significance of heterogeneity.