A new device developed at the University of Notre Dame uses an innovative method to “eavesdrop” on the conversations of cells.
Scientists have long known that RNA (ribonucleic acid) acts as a messenger inside cells, translating information from DNA to help cells make proteins.
But more recently, scientists have discovered that certain types of RNA venture outside the cell wall. Each of these strands of ‘extracellular RNA’, or exRNA, sits inside a tiny ‘carrier bottle’ and flows along body fluids like a microscopic message in a bottle, carrying information to other cells.
The new appreciation of exRNA also raised a tantalising possibility: Could we use exRNA as a way to “listen in” on cells’ conversations?
“These extracellular RNAs are a gold mine of information,” said Hsueh-Chia Chang, the Bayer Professor of Chemical and Biomolecular Engineering at the University of Notre Dame. “They can carry the early warning signs of cancer, heart disease, HIV and other life-threatening diseases.”
Chang, an expert in nanofluidics, explains that diagnosing a disease using exRNA could prove not only more effective, but also faster and cheaper than existing methods, because there is enough exRNA in a small sample of blood or other bodily fluid to signal the presence of many diseases.
But intercepting and interpreting exRNA messages has been a difficult challenge. Many laboratories have tried to filter them out of blood or other body fluids. Many others have used sophisticated centrifuges to isolate exRNA. These methods have had little success for a simple reason: The different types of “bottles” that carry exRNA messages overlap in size and weight. Even the most advanced filters and centrifuges leave many carriers jumbled together. Labs using these methods have to add extra steps, adding chemicals or small magnetic particles to further sort the carriers into discrete groups.
Four years ago, Chang and a team of researchers at Notre Dame decided to try a radically new approach, and their idea was supported by the National Institutes of Health’s Common Fund, which selects promising “high-risk, innovative endeavors with the potential for extraordinary impact”.
Chang was joined by three other Notre Dame faculty members: Crislyn D’Souza-Schorey, the Morris Pollard Professor of Biological Sciences; David Go, vice president and associate provost for academic strategy and the Viola D. Hank Professor of Aerospace and Mechanical Engineering; and Satyajyoti Senapati, research associate professor in the Department of Chemical and Biomolecular Engineering. Postdoctoral researcher Himani Sharma served as project leader, and chemical and biomolecular engineering graduate student Vivek Yadav assisted with the research.
In a study published in ACS Nano, Sharma, Chang and their colleagues describe the breakthrough device that resulted from their research. The new technology uses a combination of pH (acidity/alkalinity) and electrical charge to separate the carriers. The idea is based on the fact that although the carriers overlap in size and weight, each type has a distinct ‘isoelectric point’ – the pH or level of acidity/basicity at which it has no positive or negative charge.
The device integrates several existing technologies developed at Notre Dame and fits neatly in the palm of the hand.
Through the centre of the device flows what looks like a simple stream of water. But there is something special about the stream that is not visible to the naked eye. On the left side, the water is very acidic, with a pH about the same as a glass of grapefruit juice. On the other side of the stream, the water is very basic, with a pH similar to a bottle of ammonia.
What is special about the device is not just the fact that it has a pH gradient in the stream, but how it achieves that gradient. The technology is able to create the gradient without the addition of any chemicals, making it cheaper, greener and more efficient to operate than designs that rely on the addition of acids and bases.
The gradient comes not from a chemical, but from a double-sided membrane driven by a specially designed chip. The membrane splits the water into two ions (H+ and OH-) and adds a different type of ion to each side of the flow. One side of the membrane releases acidic hydronium ions and the other side releases basic hydroxide ions. When the basic and acidic streams flow together, they create a pH gradient, just as hot and cold streams flowing together would create hot and cold sides with a temperature gradient through the middle of the stream. The team used the two devices running in parallel to select the pH range required for carrier separation, and optimised the process using machine learning.
The pH gradient did what filters and centrifuges could not: It caused the exRNA carriers floating in the stream to sort themselves, like colours of light passing through a prism. The different types of carriers formed lines along their isoelectric points, where they could easily flow into separate outlets.
The new method allowed the research team to produce very pure samples (up to 97 per cent pure) from less than a millilitre of blood plasma, saliva or urine. The process was also lightning fast compared to current methods. While the best existing technologies take about a day to achieve separation, the Notre Dame team was able to comprehensively sort their sample in just half an hour.
“We have filed a patent and hope to commercialise the technology soon to help improve the diagnosis of cancer and other diseases,” said Sharma, who has received several awards for her work on the study from the Harper Cancer Research Institute at Notre Dame.
“Non-communicable diseases are responsible for more than 70 percent of deaths worldwide, and cardiovascular disease and cancer account for most of those deaths,” Sharma said. “Our technology shows a way to improve the way clinicians diagnose these diseases, and that could save an enormous number of lives.”
