Our cells are constantly communicating, and scientists have developed an efficient way to understand what messages they send in protein-filled biological suitcases called exosomes.
These spherical exosomes, which reside in the inner membrane of a cell but will eventually travel outside to enter another cell, transport large molecules such as proteins, a major building block in the body and drivers of biological activity, and RNA that produces protein.
“It’s an ongoing process,” says Dr. Sang-Ho Kwon, a cell biologist in the Department of Cell Biology and Anatomy at the Medical College of Georgia at Augusta University, and there’s growing evidence that this happens both in healthy, as well as disease.
“We’re trying to figure out this puzzle of what exosomes do in different scenarios,” Kwon says. He is the corresponding author of a study in Journal of Extracellular Vesicles detailed a labeling technique he and his research team developed to analyze the content of exosomes from each specific cell type to better understand their role in well-being and disease.
“Their contents can help tell us what our cells are saying to each other,” Kwon says, and will likely give us early clues that we’re getting sick and help us better understand how we get sick.
Cargo is thought to be loaded early in the formation of exosomes from their precursor endosomes, near the cell membrane, which work much like stuffing the mail truck at the post office before it goes on its route. The exosomes will remain there until they are released from the cell to travel to other cells.
Kwon and his team wanted to catch the cargo early in the process.
Currently, the main way to study the contents of exosomes is to first take the exosomes out of context, isolate them, a rather laborious process that can lead to conflicting results. In fact, it can isolate a different type of vesicles, basically biological compartments in our body, of which exosomes are just one type.
The MCG team has developed a more efficient method that allows examining only the contents of exosomes and investigating where they are located.
Their labeling system includes a variant of APEX, or ascorbate peroxidase, that is fused to another protein known to seek out exosomes. “APEX is kind of the rocket that gets me in,” Kwon says. APEX has a high affinity for biotin, a B vitamin that attaches to nearby proteins, such as those carried by the developing exosome, labeling them and thus helping to identify them. Biotin can also pass through the cell membrane behind which exosomes reside. Another protein, streptavidin, which binds naturally to biotin, allowed them to purify and clearly identify the protein cargo, as well as the RNA that would produce future proteins, using analysis provided by mass spectrometry.
Kwon’s focus is on kidney damage, and they used their system to show that oxidative stress, a byproduct of oxygen use that is excessive and destructive in disease states, changes the cargo content of exosomes made by kidney cells and found in urine. For example, the expression levels of some proteins changed, and some proteins even disappeared.
Their technique should facilitate the development of databases of the usual contents of different cell types, which will enable comparative studies of what happens to their contents in different disease states such as Kwon’s kidney damage or cancer.
“It turns out that by looking at the exosomes in urine or blood and looking at what’s inside, we can tell if the cell is damaged or a healthy cell,” he says.
Their first use of the labeling system was in living kidney cells in culture. Now they want to use it in an animal model of kidney disease.
The research team says the labeling system could further help track how exosome content changes over time and potentially how cells respond to treatment in the event of disease.
Exosomes are known to play a key role in cellular communication, both between cells of the same type and with other types. Again, there is growing evidence for the role that exosomes play in disease, including sharing with other cells the news that they are sick and potentially even helping the disease spread. “It’s not just delivering good news. Delivering bad news, too,” Kwon says.
He notes that their cargo no doubt varies in these different scenarios, an important reason why we can detect what exosomes carry. The changes could eventually serve as a good way to monitor response to treatment, another aspect of exosome research that is “exploding,” Kwon says. Scientists are also exploring the potential of using exosomes to actually deliver treatment by filling these biological packages with drugs that can be delivered directly to the desired site.
In fact, immune cells, which are key to health and disease, also release exosomes. These biological compartments also appear to play an important role in the removal of cellular debris and other debris from the cells.
“It’s an emerging field right now,” Kwon says. Proteins are the main resident because they can send signals, but they can also bind to other proteins and change their function, he says. RNA can do the same, and small microRNA can alter gene expression and therefore cell function.
Kwon’s interest in exosomes was sealed when, as a postdoctoral fellow at the University of California, San Francisco, he grew kidney tubules, which return vital nutrients to the blood and eliminate unwanted substances in the urine, into a dish and found evidence that exosomes play a key role in changing se gene dynamics there.
He calls the focus on exosomes “reverse science,” with most people looking at how the cell changes, while he and a growing number of colleagues look at the packets the cell sends out to understand what the cell is doing. Although it might not seem like it to most people, he says it’s actually a less complicated way to approach cellular activity because you’re looking at a smaller package with far fewer proteins.
The research was supported by the National Institutes of Health.