Zebrafish have become the animal model of choice for research in many areas. Now, researchers from Rice University trace roots of cancer development using genetically modified zebrafish. These fish produce fluorescent tags in migratory embryonic nerve precursor cells, enabling them to trace neuroblastoma and other forms of cancer.

Neurodevelopmental biologists from the University of Illinois at Chicago and California Institute of Technology set out to find the origins of neuroblastoma, the third-most common pediatric cancer in the US. Rosa Uribe, assistant professor of biosciences, was part of the team which created the novel transgenic fish and published the new study in the journal Genesis. Their zebrafish line produces fluorescent tags that glow in different colors, depending on the behavior of a gene called SOX10. It belongs to the family of embryonic development genes and is active in neural crest cells, embryonic stem cells from which many other cell types in the body develop.

Uribe traced the movements of green neural crest cells in a time-lapse movie playing on the computer in her office. “You can see they’ve already started to migrate that way, and a lot of them are transitioning,” said Uribe for RICE News.

Neural crest cells are the point of origin for neuroblastoma in human. Uribe and her research group are hoping the new fish can provide detailed clues about the disease.

“You can see that a lot of them are transitioning,” she said of the cells on her screen. “Some of them are dividing there, there, there. And then they turn off the green, which means they’re done the dividing.”

Why the cells stop dividing is one of the questions they hope to find an answer to. SOX proteins have more than 20 varieties. They all regulate rapid cell division in fast-growing embryos. The research group put them under close inspection since they are often activated in cancer cells. Inactivation of SOX10 in neural crest cells could be one way of treating cancers where SOX proteins play a role. Uribe and her lab also study the enteric nervous system which forms from these same cells.

“Neural crest cells are stem cells that form from the earliest portion of our central nervous system, the neural tube,” said Uribe. “They express SOX10 in addition to a bunch of other really important genes.”

The research group keeps a stock of breeder fish in hundreds of tanks. Embryos are removed from the tanks by hand each day and brought to the laboratory to observe neural crest cells during embryonic development.

Zebrafish are excellent model organisms. SOX genes in the fish are virtually identical to genes that serve the same basic roles in humans. Also, it is useful that biologists have a huge knowledge of zebrafish. Furthermore, zebrafish breed and develop quickly. For research, the transparent body of fish is a useful property too. It enables researchers to watch what is happening inside them non-invasively and while they are alive.

The research group immobilizes live embryos and takes photographs to trace their development over a period of a few hours. The neural crest cells were tracked and observed for up to four. They first appear in zebrafish embryos about 12 hours after fertilization.

“Neural crest cells also do something else that’s relevant to cancer,” Uribe said. “They undergo something called the epithelial-to-mesenchymal transition, or EMT, shortly after they form, and this is what allows them to break away and migrate to the various places in the embryo.”

EMT allows cells to revert backward along their developmental path and become more like stem cells. This allows embryonic cells to form new tissues but researchers have found many metastatic cancers that use the same genetic circuitry. Unfortunately, EMT is the switch employed by many cancer cells to break away and become metastatic.

One way to understand neural crest cells is to observe them from the moment they form until they finish migrating. The research group will also use other cell lines that have different colored tags for different reporter genes. The goal is to mix and match genes in new strains of zebrafish to test what happens when cells make either too much of a specific protein or too little.

Uribe’s lab has seven microscopes capable of gathering time-lapse images of the variously colored glowing cells. These include latest robotic instruments with tiny hoses and pumps that can draw a single embryo from a numbered test chamber up through a hose. They then transport it to the microscope’s focal plane, bring it into focus, rotate it for image acquisition from any angle and then return it and repeat the process for up to 95 more embryos.

“For the migration time-lapse images there is software that’s capable of following individual cells for hours,” Uribe said. “We can get angles and trajectories, maps of routes taken by one cell or groups of cells, and we can get quantitative data, like velocities and proliferation rates.”

Uribe hopes the new lab equipment, bought with help of CPRIT funding, will help her trace the origins of neuroblastoma in neural crest cells and make progress in understanding cancers.

Learn about genetically modified zebrafish with rainbow skin in the video below:

Scientists in Portugal have demonstrated that zebrafish larvae can be used as avatars for people, in that they can predict the response of human cancer tumors to various drugs. Learn more about new cancer studies in the video below:

By Andreja Gregoric, MSc