Scientists Are Creating Mutant Fish With Curvy Spines to Study Scoliosis

Scientists Are Creating Mutant Fish With Curvy Spines to Study Scoliosis

A revolution in how we understand a debilitating disease that affects the spine is happening in a laboratory in downtown Toronto, bathed in an eerie pink light.

The colour of the light is necessary to properly see the twitching figures in a microscope of tiny fish embryos, genetically modified to glow lime-green under fluorescent light. That green light is a sign that Curtis Boswell’s experiments are working.

Curtis Boswell in his Toronto lab. All images by Paul Tadich

At this ultra-modern lab, the 25-year-old molecular biologist is breeding genetically modified zebrafish with super curvy spines in order to better understand a debilitating condition: Scoliosis, a spine-twisting ailment that affects some three percent of humanity. It can cause all sorts of problems, from reduced heart and lung function in extreme cases, to chronic pain.

Until recently, scientists had little idea about what causes scoliosis, which usually begins to afflict its sufferers in their teens. Researchers have identified some genes linked to the disease, but there was no consensus on the mechanism behind it. Boswell and his collaborators, who include other researchers in Toronto and at Princeton University, are changing all that, by looking at fish.

Boswell is a doctoral student in the department of molecular genetics at the University of Toronto and a researcher at the Hospital for Sick Children, working withBrian Ciruna. Well put-together in a crisp white lab coat when I met with him recently in his lab, he told me how the humble zebrafish became his preoccupation in the hunt for a cause of scoliosis in humans.

“Scoliosis a disease we really don’t know much about,” he explained. “It’s characterized by a three-dimensional, torsional spinal curvature. Patients can have a lot of social issues with scoliosis. People with the condition often have to wear braces up to 22 hours a day.”

If it gets bad enough, the patient will have to undergo seriously invasive surgery in which steel rods are used to straighten the spine.

Zebrafish in the Toronto lab

Boswell’s lab was already working with zebrafish—tiny creatures only a couple of millimetres long, which are transparent save a streak of colour—because they’re a popular model organism to study developmental biology.

These fish develop from a fertilized egg to a complex embryo with a beating heart, in a matter of days. They also have spinal columns that, thanks to evolution’s parsimonious nature, develop in a very similar way to us humans.

The lab stumbled onto scoliosis because of a serendipitous discovery. They were interested in finding out what happens when key genes are absent during the earliest days of life. So they used genetic engineering techniques like CRISPR to disrupt several genes that are important for normal embryonic development in zebrafish.

One is called ptk7, and it’s important in determining how cells migrate during the formation of the embryo. To Boswell’s surprise, zebrafish in which ptk7 had been eliminated had normal embryonic lives. But when they reached their “teenage” years, they developed severe curves in their spines, just like humans with scoliosis. The similarities even extended to the fact that more females—fish and humans—are biased toward the disease, and they typically suffer from a more severe form of it.

“We thought this fish could provide us with the first genetically-defined animal model of scoliosis,” said Boswell.

Boswell examining zebrafish embryos

So he and his collaborators started digging. They already knew that a proper ptk7 gene is important for the correct functioning of motile cilia. These are whip-like appendages on the surfaces of some cells that beat back and forth in a rhythmic fashion. They line the inside surface of the windpipe, whipping in concert to drive foreign matter from the lungs.

Another place many cilia are found is on the inside of the spinal column. Here, their purpose during development is to evenly bathe nervous system cells in the cerebrospinal fluid, or CSF. It’s a nutrient-rich concoction that delivers essential nutrients to the nervous system.

Boswell thought a lack of movement in the CSF might be related to the fish developing scoliosis. So he devised an experiment to find out. He injected tiny fluorescent beads into the spinal columns of normal zebrafish embryos and compared how they moved in the CSF—like bottles bobbing in a river—relative to embryos that were missing the ptk7 gene.

What he found was striking. In normal fish, the tiny beads, and thus the CSF, flowed smoothly through the spinal column in an orderly flow. But in zebrafish where ptk7 was missing, the beads swirled around in random spirals, producing truncated trails on fluorescent microscope footage.

Looking through the literature, Boswell found that humans with scoliosis often suffer from an ailment called primary ciliary dyskinesia, meaning their cilia beat poorly or are unsynchronized. This suggested a genetic mechanism underlying scoliosis in humans.

“One of the challenges in studying this disease is that there hasn’t be an single gene associated with it,” said he. The next step is to screen people with scoliosis to see which genetic mutations they harbour. If ptk7 is involved, Boswell’s knowledge of how the gene fits into the matrix of development will help to design drugs that can repair its function.

All because of a chance discovery of fish with curvy spines.

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