New Hope for Blast-Damaged Brains

When media outlets report on wartime injuries, footage of soldiers with missing limbs often makes the nightly news and stories on post-traumatic stress disorder grab headlines. However, the most common wounds suffered by military personnel serving in Iraq and Afghanistan have been brain injuries. Traumatic brain injuries (TBI) occur when soldiers are exposed to blasts from improvised explosive devices and roadside bombs. Some sources estimate that almost 16% of all soldiers in both war zones have experienced a mild brain injury from an explosive blast. These injuries are often different from brain injuries suffered by civilians, and the medical community has few effective treatments for them.

After serving a tour of duty in Afghanistan, Kevin Kit Parker, a major in the U.S. Army and a bioengineer at the Harvard School of Engineering and Applied Science in Cambridge, Massachusetts, was inspired to conduct research into brain injuries caused by bomb blasts. In two recently published papers, his lab identified the biological mechanisms that cause neurons to swell and break, and blood vessels to shrink, when brain tissue is damaged in an explosion. The damage is caused by a cell signaling pathway, called the ROCK pathway, which runs amok after the blast. The researchers hope that post-blast treatment with a drug that blocks the pathway from forming may prevent some of the damage. The findings were published in the Proceedings of the National Academy of Sciences, and in PLoS One.

“So many young men and women are returning from military service with brain injuries, and we just don’t know how to help them,” said Parker in a press release. “We have established a toe-hold as we try to climb up on top of this problem. In many ways, this work is just the beginning.”
Blast Research Comes to a Head

A traumatic brain injury (TBI) may occur off the battlefield — for example, in sporting events and car accidents, and in shaken baby syndrome. The injury occurs any time there is an impact or force that shakes the brain or penetrates the skull and disrupts the brain’s normal function. TBIs may range from mild concussions to loss of consciousness followed by amnesia.

TBIs may have a variety of immediate and long-term effects: they may impair the ability to think clearly, as well as the senses of taste, touch and smell. Victims’ communication and language skills, and their emotional state, may also be affected. TBI is also known to cause epilepsy in some cases, and has been linked to Alzheimer’s, Parkinson’s and other degenerative brain diseases.

The type of TBI that has been occurring in soldiers in the wars in Iraq and Afghanistan is called blast-induced TBI; it is caused by the detonation of improvised explosive devices and roadside bombs. These blast forces may cause only mild TBI, but they can still have long-term effects on military personnel. The symptoms often differ from those commonly experienced by civilians, and currently there are few medical treatments specifically designed for blast-induced TBI.

Suffocating Cells

To improve medical treatment for TBI in military personnel, Parker and his lab developed a system that simulates the effects of TBI on nerve cells and blood vessels in the brain. They cultured layers of neurons or blood vessel tissue and attached them to a stretchy polymer. Then, using a “high velocity tissue stretcher” that they had built, they quickly pulled on both sides of the stretchy material to simulate a blast wave going through the cells.

In an article published in the Proceedings of the National Academy of Sciences, the researchers reported that vascular smooth muscle cells — the type of cells that line blood vessels — could account for one common side effect of blast-induced TBI, called cerebral vasospasm. This is a condition where the blood vessels surrounding the nerve cells contract, cutting off the blood supply to the cells. Vasospasm exacerbates brain damage by starving neurons of oxygen. It had long been assumed that vasospasm occurred only when there was hemorrhaging inside the brain, but, since the start of the war in Afghanistan, vasospasm has often been observed in blast-induced TBI with no bleeding.

After giving the smooth muscle cells a carefully measured tug, the researchers studied them for 24 hours to look for changes. The cells appeared intact under the microscope, but an hour after the blast they began taking up high concentrations of calcium ions from the surrounding environment. The calcium ions interacted with molecules called actin, which make up the cytoskeleton inside the cell. In all eukaryotic cells, actin molecules assemble into filaments and form a scaffolding, which determines the cell’s shape and allows it to move. When actin was stimulated by the increased calcium concentration in the muscle cells, it caused the entire cell to contract, resulting in vasospasm.

Parker attempted to prevent vasospasm in the cells by treating them with a drug that blocks the signaling pathway that is known to regulate cell contraction. A regulator called ROCK controls the movement of actin filaments. The researchers found that when they treated a layer of blasted cells with a drug that blocks ROCK, called HA-1077, vasospasm could be prevented.

The activation of the ROCK pathway and its effect on cell contraction explains why military doctors often observed vasospasm in mild cases of TBI even when they found no evidence of hemorrhaging. “They’ve duplicated [in cell culture] a finding that has been baffling to clinicians,” is what Jack Tsao, a neuroscientist at the Uniformed Services University of the Health Sciences in Bethesda, Maryland, told ScienceNOW.
When the Network Breaks Down

In a second paper, published in the journal PLoS One, the researchers used rat neurons to simulate the damage seen in the nerve cells of soldiers with blast-induced TBI. After “blasting” the rat neurons with their tissue stretcher, they found that the axons — long projections from the neuron that attach to other cells and transmit signals — were broken in some places and had large swellings in others.

The swellings occurred at places along the axons where the cell attached to other cells, or the extracellular matrix (ECM), which is the connective tissue that supports the neurons. At these focal points, molecules called integrins anchor the axon by connecting the actin cytoskeleton inside the cell to the ECM or to the membrane of another neuron. When the integrins are dislodged by a blast, molecular signaling inside the cell runs amok and causes the actin filaments to contract. The contraction pulls in the axons, causing swelling, and breaks up the neural networks that make up the brain.

Since integrin is also regulated by ROCK, the researchers treated the blasted neurons with the same drug. “Encouragingly, we also found that treating the neural tissue with HA-1077, which is a ROCK inhibitor, within the first 10 minutes of injury, reduced the number of focal swellings. We think that further study of ROCK inhibition could lead to viable treatments within the near future,” said Borna Dabiri, a bioengineering graduate student in Parker’s lab, and the lead author on the PLoS One paper, in a press release.

“This is pathological activation of a totally healthy signaling pathway,” said Parker to Nature News. Fortunately, since the same pathway causes the blood vessels and the neurons to contract, the same drug can be used to treat both problems, says Parker. “That’s a really a rich therapeutic opportunity.”
New Treatments for Battlefield Blasts?

Parker cautions that these findings are very preliminary, and notes that further studies will be needed to see if the cells in a soldier’s brain react like the cells in his culture plates. “It would be inappropriate to extrapolate from a dish to some dude’s head,” Parker commented to ScienceNOW. He also hopes to try smaller forces, such as those that would cause a mild concussion, and wants to explore different cellular pathways before conducting drug testing in animals and humans.

Ken Barbee, a bioengineer who studies neuron injury and repair at Drexel University in Philadelphia, Pennsylvania, is intrigued by the paper, but remains unconvinced that the ROCK pathway is the cause of the cellular changes. In his own work, he found that tears in the cell membrane of neurons can cause TBI, and has successfully used drugs to repair the tears. “I think the reality is that when the whole tissue undergoes deformation, you’re probably going to get a combination” of injuries to the neurons and disrupted cell signaling pathways, Barbee commented to Nature News. When asked about Barbee’s claim, Dabiri agreed that membrane tears are common in cases of severe TBI, but claims that they have not been observed in mild TBI cases, such as those simulated by their experiments.

The group’s findings have been hailed by a number of other scientists studying TBI, and their approach is seen as providing new techniques for TBI research. “It’s an elegant demonstration that biomechanical stretch will produce changes in the cytoskeleton,” is what David Hovda, director of the Brain Injury Research Center at the University of California, Los Angeles, told Nature News. “This is an important fundamental discovery.”
Discussion Questions

Would you expect soldiers who have experienced blast-induced TBI to be likely to also suffer from post-traumatic stress disorder? How does the chaos on the battlefield complicate blast-induced TBI treatment? Do you think HA-1077 would be effective in treating TBI caused by sports mishaps or car accidents?


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