New Molecular Target is Key to Enhanced Brain Plasticity

As Alzheimer’s disease progresses, it kills brain cells mainly in the hippocampus and cortex, leading to impairments in “neuroplasticity,” the mechanism that affects learning, memory, and thinking. Targeting these areas of the brain, scientists hope to stop or slow the decline in brain plasticity, providing a novel way to treat Alzheimer’s. Groundbreaking new research has discovered a new way to preserve the flexibility and resilience of the brain.

The study, led by Tel Aviv University’s Prof. Illana Gozes and published in Molecular Psychiatry, reveals a nerve cell protective molecular target that is essential for brain plasticity. According to Prof. Gozes, “This discovery offers the world a new target for drug design and an understanding of mechanisms of cognitive enhancement.”

Prof. Gozes is the incumbent of the Lily and Avraham Gildor Chair for the Investigation of Growth Factors and director of the Adams Super Center for Brain Studies at the Sackler Faculty of Medicine and a member of TAU’s Sagol School of Neuroscience. Also contributing to the study were Dr. Saar Oz, Oxana Kapitansky, Yanina Ivashco-Pachima, Anna Malishkevich, Dr. Joel Hirsch, Dr. Rina Rosin-Arbersfeld, and their students, all from TAU. TAU staff scientists Dr. Eliezer Gildai and Dr. Leonid Mittelman provided the state-of-the-art molecular cloning and cellular protein imaging necessary for the study.

Building on past breakthroughs

The new finding is based on Prof. Gozes’ discovery of NAP, a snippet of a protein essential for brain formation (activity-dependent neuroprotective protein [ADNP]). As a result of this discovery, a drug candidate that showed efficacy in mild cognitive impairment patients, a precursor to Alzheimer’s disease, is being developed. NAP protects the brain by stabilizing microtubules — tiny cellular cylinders that provide “railways and scaffolding systems” to move biological material within cells and provide a cellular skeleton. Microtubules are of particular importance to nerve cells, which have long processes and would otherwise collapse. In neurodegenerative diseases like Alzheimer’s, the microtubule network falls apart, hindering cellular communication and cognitive function.

"Clinical studies have shown that Davunetide (NAP) protects memory in patients suffering from mild cognitive impairment preceding Alzheimer’s disease," said Prof. Gozes. "While the mechanism was understood in broad terms, the precise molecular target remained a mystery for years. Now, in light of our new research, we know why and we know how to proceed."

Stabilizing microtubules

The breakthrough was the discovery of the mechanism promoting microtubule growth at the tips of the tubes (“rails”). The researchers found that the NAP structure allows it to bind to the tip of the growing microtubule, the emerging “railway,” through specific microtubule end-binding proteins, which adhere to microtubules a bit like locomotors to provide for growth and forward movement, while the other end of the microtubule may to be disintegrating. These growing tips enlist regulatory proteins that are essential for providing plasticity at the nerve cell connection points, the synapses.

"We have now revealed that ADNP through its NAP motif binds the microtubule end binding proteins and enhances nerve cell plasticity, providing for brain resilience. We then discovered that NAP further enhances ADNP microtubule binding," said Prof. Gozes.

Researchers hope their discovery will help move Davunetide (NAP) and related compounds into further clinical trials, increasing the potential of future clinical use. Prof. Gozes is continuing to investigate microtubule end-binding proteins to better understand their protective properties in the brain.

Novel Protein Fragments May Protect Against Alzheimer’s

The devastating loss of memory and consciousness in Alzheimer’s disease is caused by plaque accumulations and tangles in neurons, which kill brain cells. Alzheimer’s research has centered on trying to understand the pathology as well as the potential protective or regenerative properties of brain cells as an avenue for treating the widespread disease.

Now Prof. Illana Gozes, the incumbent of the Lily and Avraham Gildor Chair for the Investigation of Growth Factors and director of the Adams Super Center for Brain Studies at the Sackler Faculty of Medicine and a member of Tel Aviv University’s Sagol School of Neuroscience, has discovered novel protein fragments that have proven protective properties for cognitive functioning.

In a study published in the Journal of Alzheimer’s Disease, Prof. Gozes examined the protective effects of two newly discovered protein fragments in mice afflicted with Alzheimer’s disease-like symptoms. Her findings have the potential to serve as a pipeline for new drug candidates to treat the disease.

NAP time for Alzheimer’s

"Several years ago we discovered that NAP, a snippet of a protein essential for brain formation, which later showed efficacy in Phase 2 clinical trials in mild cognitive impairment patients, a precursor to Alzheimer’s," said Prof. Gozes. "Now, we’re investigating whether there are other novel NAP-like sequences in other proteins. This is the question that led us to our discovery."

Prof. Gozes’ research focused on the microtubule network, a crucial part of cells in our bodies. Microtubules act as a transportation system within nerve cells, carrying essential proteins and enabling cell-to-cell communications. But in neurodegenerative diseases like Alzheimer’s, ALS, and Parkinson’s, this network breaks down, hindering motor abilities and cognitive function.

"NAP operates through the stabilization of microtubules — tubes within the cell which maintain cellular shape. They serve as ‘train tracks’ for movement of biological material," said Prof. Gozes. "This is very important to nerve cells, because they have long processes and would otherwise collapse. In Alzheimer’s disease, these microtubules break down. The newly discovered protein fragments, just like NAP before them, work to protect microtubules, thereby protecting the cell."

Down the tubes

In her new study, Prof. Gozes and her team looked at the subunit of the microtubule — the tubulin — and the protein TAU (tubulin-associated unit), important for assembly and maintenance of the microtubule. Abnormal TAU proteins form the tangles that contribute to Alzheimer’s; increased tangle accumulation is indicative of cognitive deterioration. Prof. Gozes decided to test both the tubulin and the TAU proteins for NAP-like sequences. After confirming NAP-like sequences in both tubulin subunits and in TAU, she tested the fragments in tissue cultures for nerve-cell protecting properties against amyloid peptides, the cause of plaque build up in Alzheimer patients’ brains.

"From the tissue culture, we moved to a 10-month-old transgenic mouse model with frontotemporal dementia-like characteristics, which exhibits TAU pathology and cognitive decline," said Prof. Gozes. "We tested one compound — a tubulin fragment — and saw that it protected against cognitive deficits. When we looked at the ‘dementia’-afflicted brain, there was a reduction in the NAP parent protein, but upon treatment with the tubulin fragment, the protein was restored to normal levels."

Prof. Gozes and her team also measured the brain-to-body mass ratio, an indicator of brain degeneration, and saw a significant decrease in the mouse model compared to normal mice. Following the introduction of the tubulin fragments, however, the mouse’s brain to body ratio returned to normal. “We clearly see here the protective effect of the treatment,” said Prof. Gozes. “We witnessed the restorative and protective effects of totally new protein fragments, derived from proteins critical to cell function, in tissue cultures and on animal models.”

(Image: Getty Images)

Engineering resilience in the brain

Penn researchers model neural structures on the smallest scales to better understand traumatic brain injury

Compared to the monumental machines of science, things like the International Space Station or the Large Hadron Collider, the human brain doesn’t look like much. However, this three-pound amalgam of squishy cells is one of the most complicated and complex structures in the known universe.

With hundreds of billions of neurons, each with its own inner world of organelles and molecular components, understanding the fundamental wiring of the brain is a major undertaking, one that has received a commitment of at least $100 million worth of federal funding from the National Science Foundation (NSF), the National Institutes of Health and the Defense Advanced Research Projects Agency.

And with all of the brain’s interconnected structures, protecting or repairing this complicated machine means thinking like an engineer.

"The idea is really quite simple," says Vivek Shenoy, an NSF-supported professor of materials science and engineering at the University of Pennsylvania’s School of Engineering and Applied Science. "All of the mechanical properties of cells come from their cytoskeleton and the molecules within it. They’re all reinforcing frames, like the frame in a building. Engineers design buildings and other structural objects to make sure they don’t fail, so it’s the same principle: structural engineering on a very, very small level."

Shenoy applies this approach to a problem very much in the public eye—traumatic brain injury. Even the mildest forms of TBI, better known as concussions, can do irreversible damage to the brain. More serious forms can be fatal.

With a background in mechanical engineering and materials science, one might think that Shenoy’s contribution to this problem involves designing new helmets or other safety devices. Instead, he and his colleagues are uncovering the fundamental math and physics behind one of the core mechanisms of the injury: swelling in axons caused by damage to internal structures known as microtubules. These neural “train tracks” transport molecular cargo from one end of a neuron to another; when the tracks break, the cargo piles up and produces bulges in the axons that are the hallmark of fatal TBIs.

Armed with a better understanding of the mechanical properties of these critical structures, Shenoy and his colleagues are laying the foundations for drugs that could one day bolster neurons’ reinforcing frames, making them more resilient when faced with a TBI-inducing impact.

Train tracks and crossties

The first step toward this understanding was resolving a paradox: Why were the microtubules, the stiffest elements of the axons, the parts that were breaking when loaded with the stress of a blow to the head?

A recent finding from Shenoy’s team shows that the answer rests with a critical brain protein known as tau, which is implicated in several neurodegenerative diseases, including Alzheimer’s. If microtubules are like train tracks, tau proteins are the crossties that hold them together. The protein’s elastic properties help explain why rapid movement of the brain, whether on a football field or a car crash, leads to TBI.

Shenoy’s colleague Douglas Smith, professor of neurosurgery in Penn’s Perelman School of Medicine and director of the Penn Center for Brain Injury and Repair, had previously studied the mechanical properties of axons, subjecting them to strains of different forces and speeds.

"What we saw is that with slow loading rates, axons can stretch up to at least 100 percent with no signs of damage," Smith said. "But at faster rates, axons start displaying the same swellings you see in the TBI patients. This process occurs even with relatively short stretches at fast rates."

To explain this rate-dependent response, Shenoy and Smith had to delve deeper inside the structure of microtubules. Based on Smith’s work, other biophysical modelers had previously accounted for the geometry and elastic properties of the axon during a stretching injury, but they did not have good data for representing tau’s role.

"You need to know the elastic properties of tau," Shenoy said, "because when you load the microtubules with stress, you load the tau as well. How these two parts distribute the stress between them is going to have major impact on the system as a whole."

Elastic properties

Shenoy and his colleagues had a sense of tau’s elastic properties but did not have hard numbers until a 2011 experiment from a Swiss and German research team physically stretched out lengths of tau by plucking it with the tip of an atomic force microscope.

"This experiment demonstrated that tau is viscoelastic," Shenoy said. "Like Silly Putty, when you add stress to it slowly, it stretches a lot. But if you add stress to it rapidly, like in an impact, it breaks."

This behavior is because the strands of tau protein are coiled up and bonded to themselves in different places. Pulled slowly, those bonds can come undone, lengthening the strand without breaking it.

"The damage in traumatic brain injury occurs when the microtubules stretch but the tau doesn’t, as they can’t stretch as far," Shenoy said. "If you’re in a situation where the tau doesn’t stretch, such as what happens in fast strain rates, then all the strain will transfer to the microtubules and cause them to break."

With a comprehensive model of the tau-microtubule system, the researchers were able to boil down the outcome of rapid stress loading to equations with only a handful of variables. This mathematical understanding allowed the researchers to produce a phase diagram that shows the dividing line between strain rates that leave permanent damage versus ones that are safe and reversible.

Next steps

Having this mathematical understanding of the interplay between tau and microtubules is only the beginning.

"Predicting what kind of impacts will cause these strain rates is still a complicated problem," Shenoy said. "I might be able to measure the force of the impact when it hits someone’s head, but that force then has to make its way down to the axons, which depends on a lot of different things.

"You need a multiscale model, and our work will be an input to those models on the smallest scale."

In the longer term, however, knowing the parameters that lead to irreversible damage could lead to better understanding of brain injuries and diseases and to new preventive measures. It may even be possible to design drugs that alter microtubule stability and elasticity of axons in traumatic brain injury; Smith’s group has demonstrated that treatment with the microtubule-stabilizing drug taxol reduced the extent of axon swellings and degeneration after injuries in which they are stretched.

Ultimately, insights on the molecular level will be inputs to a more comprehensive view of the brain and its many hierarchies of organizations.

"When you’re talking about something’s mechanical properties, stiffness is what comes to mind," Shenoy said. "Biochemistry is what determines that stiffness in the brain’s structures, but that’s only at the molecular level. Once you build it up and formulate things at the appropriate scale, protecting the brain becomes more of a structural engineering problem."

Researchers Model a Key Breaking Point Involved in Traumatic Brain Injury

Even the mildest form of a traumatic brain injury, better known as a concussion, can deal permanent, irreparable damage. Now, an interdisciplinary team of researchers at the University of Pennsylvania is using mathematical modeling to better understand the mechanisms at play in this kind of injury, with an eye toward protecting the brain from its long-term consequences.

Their recent findings, published in the Biophysical Journal, shed new light on the mechanical properties of a critical brain protein and its role in the elasticity of axons, the long, tendril-like part of brain cells. This protein, known as tau, helps explain the apparent contradiction this elasticity presents. If axons are so stretchy, why do they break under the strain of a traumatic brain injury?    

Tau’s own elastic properties reveal why rapid impacts deal permanent damage to structures within axons, when applying the same force more slowly causes them to safely stretch. This understanding can now be used to make computer models of the brain more realistic and potentially can be applied toward tau-related diseases, such as Alzheimer’s.

The team consists of Vivek Shenoy, professor of materials science and engineering in the School of Engineering and Applied Science, Hossein Ahmadzadeh, a member of Shenoy’s lab, and Douglas Smith, professor of neurosurgery in Penn’s Perelman School of Medicine and director of the Penn Center for Brain Injury and Repair. 

“One of the main things you see in the brains of patients who have died because of a TBI is swellings along the axons,” Shenoy said. “Inside axons are microtubules, which act like tracks for transporting molecular cargo along the axon. When they break, there’s an interruption in the flow of this cargo and it starts to accumulate, which is why you get these swellings.”  

Smith had previously studied the mechanical properties of axons as a whole. By patterning axons in culture in parallel tracts, Smith and his colleagues could apply a stretch to the axons at different forces and speeds and measure how they responded.

“What we saw is that with slow loading rates, axons can stretch up to at least 100 percent with no signs of damage,” Smith said. “But at faster rates, axons start displaying the same swellings you see in the TBI patients. This process occurs even with relatively short stretch at fast rates. So the rate at which stretch is applied is the important component, such as occurs during rapid movement of the brain and stretching of axons due to head impact from a fall, assault or automobile crash.”

This observation still did not explain to researchers why microtubules, the stiffest part of the axon, were the parts that were breaking. To solve that puzzle, the researchers had to delve even deeper into their structure.

Microtubules are closely packed together inside axons, somewhat like a bundle of straws. Binding the individual straws together is the protein tau. Other biophysical modelers had previously accounted for the geometry and elastic properties of the axon during a stretching injury based on Smith’s work but did not have good data for representing tau’s role in the overall behavior of the system when it is loaded with stress over different lengths of time. 

“You need to know the elastic properties of tau,” Shenoy said, “because when you load the microtubules with stress, you load the tau as well. How these two parts distribute the stress between them is going to have major impact on the system as a whole.”

Shenoy and his colleagues had a sense of tau’s elastic properties but did not have hard numbers until a 2011 experiment from a Swiss and German research team physically stretched out lengths of tau by plucking it with the tip of an atomic force microscope.

“This experiment demonstrated that tau is viscoelastic,” Shenoy said. “Like Silly Putty, when you add stress to it slowly, it stretches a lot. But if you add stress to it rapidly, like in an impact, it breaks.”

This behavior is because the strands of tau protein are coiled up and bonded to themselves in different places. Pulled slowly, those bonds can come undone, lengthening the strand without breaking it. 

“The damage in traumatic brain injury occurs when the microtubules stretch but the tau doesn’t, as they can’t stretch as far,” Shenoy said. “If you’re in a situation where the tau doesn’t stretch, such as what happens in fast strain rates, then all the strain will transfer to the microtubules and cause them to break.”

With a comprehensive model of the tau-microtubule system, the researchers were able to boil down the outcome of rapid stress loading to equations with only a handful of variables. This mathematical understanding allowed the researchers to produce a phase diagram that shows the dividing line between strain rates that leave permanent damage versus safe and reversible loading and unloading of stress.

“Predicting what kind of impacts will cause these strain rates is still a complicated problem,” Shenoy said. “I might be able to measure the force of the impact when it hits someone’s head, but that force then has to make its way down to the axons, which depends on a lot of different things.

“You need a multiscale model, and our work will be an input to those models on the smallest scale.”

In the longer term, knowing the parameters that lead to irreversible damage could lead to better understanding of brain injuries and diseases and to new preventive measures. It may even be possible to design drugs that alter microtubule stability and elasticity of axons in traumatic brain injury; Smith’s group has demonstrated that treatment with the microtubule-stabilizing drug taxol reduced the extent of axon swellings and degeneration after stretch injury.    

“Intriguingly, it may be no coincidence that tau is also the same protein that forms neurofibrillary tangles, one of the hallmark brain pathologies of chronic traumatic encephalopathy, or CTE, which is linked to a history of concussions and higher levels of TBI,” said Smith. “Uncovering the role of tau at the time of trauma may provide insight into how it is involved in long-term degenerative processes.”

Scientists learn what makes nerve cells so strong

How do nerve cells — which can each be up to three feet long in humans — keep from rupturing or falling apart?

Axons, the long, cable-like projections on neurons, are made stronger by a unique modification of the common molecular building block of the cell skeleton. The finding, which may help guide the search for treatments for neurodegenerative diseases, was reported in the April 10 issue of Neuron by researchers at the University of Illinois at Chicago College of Medicine.

Microtubules are long, hollow cylinders that are a component of the cytoskeleton in all cells of the body. They also support transport of molecules within the cell and facilitate growth. They are made up of polymers of a building-block substance called tubulin.

“Except for neurons, cells’ microtubules are in constant dynamic flux — being taking apart and rebuilt,” says Scott Brady, professor and head of anatomy and cell biology at UIC and principal investigator on the study. But only neurons grow so long, he said, and once created they must endure throughout a person’s life, as much as 80 to 100 years. The microtubules of neurons are able to withstand laboratory conditions that cause other cells’ microtubules to break apart.

Brady had been able to show some time ago that the neuron’s stability depended on a modification of tubulin.

“But when we tried to figure out what the modification was, we didn’t have the tools,” he said.

Yuyu Song, a former graduate student in Brady’s lab and the first author of the study, took up the question. “It was like a detective story with many possibilities that had to be ruled out one by one,” she said. Song, who is now a post-doctoral fellow at Howard Hughes Medical Institute at Yale School of Medicine, used a variety of methods to determine the nature of the modification and where it occurs.

She found that tubulin is modified by the chemical bonding of polyamines, positively charged molecules, at sites that might otherwise be chinks where tubulin could be broken down, causing the microtubules to fall apart. She was also able to show that the enzyme transglutaminase was responsible for adding the protective polyamines.

The blocking of a vulnerable site on tubulin would explain the extraordinary stability of neuron microtubules, said Brady. However, convincing others required the “thorough and elegant work” that Song brought to it, he said. “It’s such a radical finding that we needed to show all the key steps along the way.”

The authors also note that increased microtubule stability correlates with decreased neuronal plasticity — and both occur in the process of aging and in some neurodegenerative diseases. Continued research, they say, may help identify novel therapeutic approaches to prevent neurodegeneration or allow regeneration.