Engineering 'lego-like' proteins to create color ultrasounds

Ultrasound technology can be a pretty versatile tool. Just this week, we learned that doctors from the Salk Institute have successfully used it to 'jump-start' a coma patient's brain, and in the past it's been utilized for improving drug delivery, healing chronic wounds, and it might even help treat Alzheimer's. Caltech researchers have now come up with a method for improving ultrasound imaging, providing a means of peering deeper into the body to visualize individual cells and molecules in distinct colors.

The secret of the Caltech team's method lies in tiny protein-shelled, gas-filled structures called gas vesicles. The structures occur naturally in certain water-dwelling single-celled organisms, but in 2014, the researchers realized that the vesicles reflect sound waves during ultrasound imaging – a discovery that they subsequently verified using laboratory mice.

To make the vesicles truly useful, the team set about altering a protein found on the surface of the structures, called gas vesicle protein C, or GvpC. The protein gives the vesicle its mechanical strength, and can be engineered to change the properties of the nanostructures, with longer versions creating stronger vesicles.

"It's somewhat like engineering with molecular Legos," said senior paper author Mikhail Shapiro. "We can swap different protein 'pieces' on the surface of gas vesicles to alter their targeting properties and to visualize multiple molecules in different colors."

First, the team tried removing the protein entirely, and found that when administered to lab mice, the weakened vesicles actually vibrated significantly more when exposed to ultrasound waves. Resonating with harmonic frequencies, the altered structures were easy to pick out in ultrasound images.

Another experiment saw the researchers altering the nanostructures to include certain features that would bind them to certain kinds of cells, thus allowing them to be aimed at particular types of tissue inside the body. For example, they added an amino acid sequence, which recognizes a type of protein called integrins, which are abundant in tumor cells.

Lastly, the team considered the possibility of using altered gas vesicles to create colored ultrasound images. They created three different types of gas vesicles, each with varying strengths of GvpC, giving them differing abilities to resist collapse under pressure. The different nanostructures can be picked out during imaging by increasing the ultrasound pressure, causing them to collapse in groups.

While all the cells appear the same in the actual imaging – in black and white – it's possible to artificially color them by linking pigments to the time of their collapse.

The researchers believe that by combining alternations made to the gas vesicles over the course of the study – both the addition of features like amino acid sequences, and adjustments made to GvpC – it should be possible to create color ultrasound images that visualize cells in different colors, and describe important processes inside the body.

"You might be able to see tumor cells versus the immune cells attacking the tumor, and thus monitor the progress of a medical treatment." said Shapiro.

The team published a paper on the work in the journal ACS Nano.

Source: Caltech