Genes are not the only factors guiding cells in creating complex structures like tissues and organs. Researchers have highlighted the significance of another key element in development: cell density, which refers to how closely or loosely cells are arranged in a space. Through both computer models and practical lab studies, a team of scientists effectively manipulated cell density to control how mouse cells organized themselves into intricate formations. This research is a step forward in the larger ambition of creating synthetic tissues, which could be used in various medical contexts, including drug testing and providing grafts or transplants.
Genes are not the only forces directing cells to form multicellular structures, tissues, and organs. A recent publication in Nature Communications by USC Stem Cell researcher Leonardo Morsut and Caltech computational biologist Matt Thomson sheds light on another crucial influencing factor: cell density—referring to how tightly or loosely cells are packed together. The scientists employed both computational models and laboratory tests, utilizing cell density as a practical method to regulate how mouse cells arrange themselves into complex patterns.
“This research marks a significant advancement towards our broader goal of engineering synthetic tissues,” stated Morsut, who is an assistant professor in stem cell biology and regenerative medicine, along with biomedical engineering, at the Keck School of Medicine of USC. “The potential applications for synthetic tissues in medicine are vast, including drug testing, therapies, grafts, or transplants for patients.”
The study involved two types of mouse cells—connective tissue cells and stem cells—modified to include a synthetic communication system or “genetic circuit.” This circuit utilizes a component Morsut developed known as “synNotch,” a genetically engineered protein that functions as a “sensor” on the cell surface. This sensor identifies external signals that prompt the cell to respond, typically by activating a specified gene.
In these experiments, researchers leveraged synNotch to trigger a circuit that involved green fluorescence and a mechanism to extend the signal—though it can activate any gene. The fluorescence allowed them to easily monitor the patterns formed by the cells. For instance, the scientists could create green fluorescent rings originating from a central point within a cluster of cells.
Unexpected findings
During the experiments, co-first author Marco Santorelli, a postdoctoral researcher in the Morsut Lab, observed that genetically identical cells did not always yield identical patterns.
“We noticed that the patterning outcomes varied when starting with genetically identical cells at different densities,” Morsut recounted. “It puzzled us at first. Marco once told me that the experiment worked, but only in part of the plate. Upon closer inspection, we discovered a density gradient that corresponded with variations in the patterns formed.”
When cell density increased beyond a certain level, the effectiveness of synNotch diminished, producing inconsistent patterns. Additionally, cell density continually fluctuated as cells multiplied at varying rates, leading to complex interactions with the synNotch genetic circuit.
Does it compute?
Co-first author Pranav S. Bhamidipati, a participant in the USC-Caltech MD-PhD program, sought to develop a computational model to predict and clarify this multifaceted and dynamic cell behavior.
“This marked one of my earliest experiences where computational modeling accurately predicted behaviors mirroring real cell activity,” noted Thomson, who is an assistant professor of computational biology at Caltech and an investigator with the Heritage Medical Research Institute. “It guided us in considering how multiple factors like cell density, proliferation rates, and signaling interrelate.”
Morsut remarked, “We were pleased to have the computational model to explore and understand the range of potential patterns, and how to transition between them.”
With the help of the computational model, the researchers successfully utilized cell density to create a variety of predictable fluorescent patterns that emerged over designated timeframes.
It’s okay to be a little dense
To delve deeper into how cell density influenced these outcomes, co-first author Josquin Courte, a postdoctoral researcher in the Morsut Lab, carried out multiple experiments that led to a surprising conclusion. Higher cell density creates stress, which accelerates the breakdown of not only synNotch but also cell surface sensors in general.
This indicates that cell density serves as a versatile tool for directing both engineered and naturally occurring cells to construct a wide variety of structures, tissues, and organs.
“Nature has utilized cell density together with genetic circuits to produce the remarkable diversity of multicellular structures, tissues, and organs,” said Morsut. “Now we can harness this same approach to enhance our work in creating synthetic multicellular formations—and eventually tissues and organs—for regenerative medicine.”
