CLEVELAND, Ohio — More than being able to bend their knees or wiggle their toes again, paralyzed people long to be able to use the bathroom normally. Regaining bladder control would end embarrassing accidents, awkward catheters, costly medical care and dangerous, sometimes deadly infections.
A Cleveland team’s research has moved that goal closer to reality.
In rats whose spinal cords were completely cut – the worst kind of paralysis injury – scientists from Case Western Reserve University and the Cleveland Clinic were able to prod nerve fibers to gradually grow across gaps the width of a pencil and reconnect. The tendrils’ months-long journey was aided by scaffolding soaked in growth-boosting and scar-busting chemicals.
The rejoined neurons made it possible for the treated rats to urinate almost normally, which they do frequently as they roam. It’s a medical first in the field of nerve regeneration after decades of work and sometimes dashed hopes. The project’s director, CWRU neuroscientist Jerry Silver, used a similar method two years ago to revive breathing muscles in rats with less severe spinal injuries.
The painstaking grafting process, in which Clinic neuroscientist Yu-Shang Lee borrowed nerve tracts from the rats’ chests and spliced them into the severed cords, did not restore the much more complex connections required for walking, nor was it intended to. The Cleveland team is focusing on more achievable, incremental improvements to paralyzed patients’ quality of life, rather than the elusive “holy grail” of walking.
“It’s clear that some primitive functions may be able to come back,” Silver said. “We’ve shown for the very first time that we can promote long-distance regeneration in the adult spinal cord, across a complete spinal cord lesion. It’s a step in the right direction.”
Before the scientists can attempt the bladder repair in humans, they must show their method can re-grow nerves over longer distances in animals larger than rats, and that the fibers can overcome the entrenched obstacles of scarring in old spinal injuries, not just in fresh ones.
Still, the results to be published Wednesday in the Journal of Neuroscience are stirring excitement. The findings “look really, really impressive,” said Susan Howley, executive vice president and research director of the Christopher and Dana Reeve Foundation, a national spinal cord injury research and advocacy group named for the late “Superman” actor and his wife.. “From my lay perspective, this is an important study that pushes the field forward.”
Silver’s and Lee’s research is “exceptionally rigorous,” and the pair’s findings have “potential clinical importance,” said neuroscientist Phillip Popovich, director of Ohio State University’s Center for Brain and Spinal Cord Repair.
Daunting challenge of repairing the cord
Until fairly recently, many scientists had doubted that the fragile, intricate human spinal cord, once badly damaged, could be fixed.
As an extension of the brain, the cord’s densely bundled nerve fibers are a vast, fast two-way information conduit. They relay commands – some voluntary, some automatic – from the brain to the body’s muscles and organs, and transmit sensation and other vital updates back to the brain. The cord also has the built-in ability to trigger certain quick-action protective reflexes on its own, such as jerking your hand away from a hot stove or straightening your knee to keep your balance.
Some animals with simpler nervous systems are able to recover from disastrous spinal cord injuries. Sever a fish’s cord and eventually it will regain the ability to flap its tail and swim away. Even though the re-knitted nerve connections don’t appear numerous or normal, they’re enough to get the job done.
And human peripheral nerves – the far-flung strands that branch out from the cord to control muscles and organs – can slowly regenerate when damaged, albeit imperfectly. With some luck and a good surgeon, people who’ve had a finger re-attached after an accident may get some feeling and function back when peripheral nerves in the digit re-link.
But the nerves in the human brain and spinal cord (and those of other large mammals) appear unable to fix themselves. Cut them with a knife or a bullet, or crush them with a sharp blow, and if left alone, they won’t recover. Researchers aren’t sure why the disparity exists. Perhaps it’s the sheer complexity of large animals’ central nervous systems, or some evolutionary bias against the time and resources it would take to heal.
Whatever the reason, about 273,000 Americans are living with permanent spinal cord injuries; of those whose care is tracked, roughly 18 percent have no feeling or movement below the chest or waist, while 12 percent are completely paralyzed from the neck down.
Just because human brain and spinal cord nerves can’t self-repair doesn’t mean they don’t try. In the early 1900s, the renowned Spanish neuroscientist Santiago Ramon y Cajal observed that newly cut nerve fibers sprouted tentative shoots, though their progress stalled and “dried up irrevocably” within a few days.
Cajal guessed – correctly, as it turned out – that the budding nerves needed some kind of nurturing chemical to help them grow, though he couldn’t identify the substance. He suspected that peripheral nerves, which can regenerate on their own, might contain the missing ingredient. Cajal’s lab tried using transplanted snippets of rabbits’ sciatic nerves as a sort of trellis through which the animals’ cut brain nerve fibers might spread, but the results were inconclusive. The concept languished for decades.
Then, in 1981, armed with better nerve growth detection methods, Montreal researchers Albert Aguayo and Samuel David tried Cajal’s bridging technique again, this time in rats’ spinal cords. Nerve fibers were able to cross the bridge, sprouting an inch or more and proving there was no inherent reason spinal cord nerves couldn’t regenerate. But their progress stalled as they exited the splice and tried to re-enter the injured spinal cord, a powerful hint that other obstacles were at work.
Scientists, including CWRU’s Silver, began to probe those impediments. They discovered that the stalled nerve fibers are reacting to strong chemical and physical roadblocks that the body throws up as part of its attempts to prevent further damage after a spinal cord injury. Some labs worked to understand growth-spurring chemicals that might aid the nerves’ advancement. Silver focused on the toxic injury site, and on net-like proteins in the scar that physically ensnare re-growing nerve fibers, stopping them cold. He experimented with an enzyme, chondroitinase, nicknamed “chase,” that clipped the protein straitjacket and freed the nerve fibers. But getting them to form working connections remained a problem.
Putting the neural bridge to work
In 1996, an electrifying report from Sweden seemed to show that researchers had finally overcome all the regeneration barriers and actually repaired a spinal cord injury. In paralyzed rats whose cords were completely severed, neuroscientist Lars Olson and neurosurgeon Henrich Cheng paired Cajal’s nerve bridges with a growth-boosting chemical called FGF and were able to restore some movement and weight-bearing ability in the rats’ hind legs.
Though the work initially was hailed as a breakthrough, other scientists weren’t able to replicate the results, possibly because Cheng’s meticulous microsurgical grafting on the rats’ spinal cords was so demanding. The only one who reported a successful outcome was Yu-Shang Lee, a Taiwanese neuroscientist then at the University of California, Irvine, who had trained with Cheng and learned the difficult bridging surgery. His rats regained some hind limb function too, but it was far from walking again.
Silver, meanwhile, figured he might have a better chance with the nerve grafting approach if he combined it with chase, the scar-busting enzyme, and if he aimed for something less complicated than restoring walking. Breathing seemed like a reasonable target. It mainly required re-activation of a single muscle, the diaphragm. In 2011 Silver’s lab reported their partially paralyzed rats regained “robust” breathing ability when treated with the nerve bridges and chase.
Restoring urinary control seemed the next logical target, since the bladder is a relatively simple system, too – a purse string-like sphincter muscle that pinches the organ shut most of the time, and a muscular wall that squeezes to push out urine when the sphincter relaxes. The brain and spinal cord must work together to coordinate the process.
In paralyzed people, the urinary sphincter locks tight (unlike other muscles, which go limp), and the bladder may overfill, because emptying signals can no longer reach the brain. The bladder grows abnormally large, and the overfilling sometimes triggers sudden spikes in breathing, blood pressure and heart rate, which may be life-threatening. Catheters that manually empty the bladder are conduits for bacteria, and any urine that remains also can breed infection or kidney failure.
Using rats whose bladders were paralyzed, Silver’s lab tried the bridging method that he had previously used to successfully revive breathing. “We failed miserably,” Silver said. “The spinal cord gets very narrow [in the region that controls the bladder] and we were basically destroying the cord with our surgery.”
Fortunately for Silver, Yu-Shang Lee, the young neuroscientist who had mastered the daunting operation under Cheng’s tutelage, had taken a job at the Cleveland Clinic.
“Yu-Shang is a master surgeon. Here he is in my backyard!” Silver exclaimed. “We thought it would be a no-brainer to combine our resources” – Silver’s knowledge of the growth-blocking scar and how to defuse it with chase, and Lee’s surgical talent and experience with growth-boosting FGF.
So they launched the bladder study, with Lee performing two or three of the grueling five-hour cord-splicing surgeries per day. First, under anesthesia, the test rats’ spinal cords were completely severed at the chest level. The full cut, called a transection, was necessary rather than a partial injury so that, if the rats regained any bladder control, the researchers could be certain it was due to their repair and not some residual connections.
Lee carefully installed the graft in the tiny gap. The splice consisted of 18 to 21 peripheral nerve tracts taken from each rat’s chest wall. The peripheral nerves die, but the nurturing tubes that surrounded them serve as a tunnel for the re-growing spinal nerves to burrow through on the way to hoped-for reconnection.
Lee connected each individual fiber to the cord’s cut ends, using a dab of natural fibrin glue, the sticky stuff in clotted blood. The fibrin was seeded with chase and FGF, to encourage nerve re-growth. Lee also injected chase in the spinal cord stumps, where scarring would occur. He wired the rats’ vertebrae together so the delicate grafts wouldn’t be pulled apart when the animals awoke. Then the researchers waited to see what happened.
The ‘full monty’ therapy shows promise
Rats don’t pee like people do. They urinate not just to get rid of waste, but to mark their territory, something they do dozens of times a night, in small, carefully controlled spurts, guided by stuttering nerve firing patterns known as bursting. The complexity of rat urination actually is a good test; if scientists can restore it, that boosts confidence in an eventual human repair.
Though it took three to six months, the 15 rats that Silver and Lee gave what they called the “full monty” treatment – the spinal cord splice, scar-busting chase and growth-spurring FGF – showed “markedly improved” bladder function compared with rats whose cords were cut but got lesser or no repairs.
The full monty rats urinated more often, and in smaller volumes (indicating their bladders weren’t overfilling) than their counterparts. Their sphincter muscles regained the rapid-fire, contract-and-relax bursting pattern typical of healthy rats, and their bloated bladders shrank in size and weight. All told, the results weren’t perfectly normal peeing, but close.
When the scientists examined the full monty rats’ spinal cords, they found that only a few hundred nerve fibers had crossed the bridge and rewired themselves. Those that made it were nerve types originating in the brain stem and involved in bladder control and other crude, reflexive movements, not the ones responsible for precise, complex activities like walking or piano-playing.
It’s not clear why there wasn’t wholesale re-growth of all sorts of nerves. “Those primitive functions may be quite special,” Silver said, powered by nerves that have an amped-up potential to re-grow, with a little help.
Re-cutting the full monty rats’ repaired spinal cords wiped out their regained bladder control, as did giving them drugs that blocked nerves involved in urination. That convinced the scientists it was the grafts and drug treatment, and the reconnected nerve fibers, that had caused the improvements.
Next up, Silver and Lee want to know if their splicing method works in older spinal cord injuries, where scarring and other obstacles to nerve regeneration have been in place for months and presumably are harder to overcome.
They’ll test various ways of “waking up” severed nerve fibers in the cord prior to installing the splice, using drugs, genetic tinkering and electric stimulation. “You can be as creative as you want,” Silver said. “And then, once the [nerve fibers] are in full growth mode, like a dragster spinning its wheels, you put in the bridge and hopefully away they go.”
“We still don’t know why they have this capacity to re-grow,” Lee said. “This will be worthwhile to continue to find out, in terms of implications for future human patients.”
Trying the repair strategy in people must wait for evidence that it works in larger animals, such as cats or pigs, both researchers said. Human neurons grow fast, but they would have to grow much longer distances than the regenerating rat nerves, because of the difference in body size. “That’s what we have to work on,” Silver said, “accelerating the growth.”
By John Mangels, The Plain Dealer
