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U.S. DoD awards $2M for study to protect neurological function after cardiac surgery

doctors operating

A collaboration between clinical and basic science researchers including Drs. Ishibashi, Hashimoto-Torii, Jonas, and Deutsch, seeks to to understand how caspase enzyme activation plays a role in the development of fine and gross motor skills in children who underwent cardiac surgery for CHD repair.

The U.S. Department of Defense has awarded $2 million to Children’s National Hospital to study how a family of protease enzymes known as caspases may contribute to brain cell degeneration when activated by prolonged anesthesia and cardiopulmonary bypass during cardiac surgery for congenital heart disease.

This U.S. Army Medical Research Acquisition Activity Award, Anesthesia Neurotoxicity in Congenital Heart Disease, is led by principal investigator Nobuyuki Ishibashi, M.D., with both clinical and basic science co-investigators including Kazue Hashimoto-Torii, Ph.D., (Neuroscience), Richard Jonas, M.D., (Cardiovascular Surgery) and Nina Deutsch, M.D., (Anesthesiology).

While the specific cellular and molecular mechanisms of how anesthesia and cardiac surgery impact cortical development are poorly understood, both seem to impact brain growth and development in young children. The most common neurologic deficit seen in children after CHD surgical repair is the impairment of fine and gross motor skills.

Both anesthetic agents and inflammation like that seen as a result of cardiopulmonary bypass have also been shown to contribute to the activation of a specific group of enzymes that play an essential role in the routine (programmed) death of cells: caspases. However, recent pre-clinical research shows that these enzymes may also contribute to other alterations to cells beyond cell death, including making changes to other cell structures. In pre-clinical models, these changes cause impairments to fine and gross motor skills – the same neurological deficits seen in children with CHD who have undergone procedures requiring prolonged anesthesia and cardiopulmonary bypass.

The research team hypothesizes that caspases are extensively activated as a result of cardiac surgery and while that activation is rarely causing reduced numbers of neurons, the changes that caspase enzymes trigger in neurons are contributing to neurological deficits seen in children with CHD after surgery.

While the study focuses specifically on the impacts of cardiac surgery for correction of a heart defect, the findings could have major implications for any pediatric surgical procedure requiring prolonged anesthesia and/or cardiopulmonary bypass.

Kazue Hashimoto-Torii and Masaaki Torii

Center for Neuroscience Research investigators join CIFASD

Kazue Hashimoto-Torii and Masaaki Torii, Collaborative Initiative on Fetal Alcohol Spectrum Disorders

Masaaki Torii, Ph.D., Kazue Hashimoto-Torii, Ph.D., and their research teams are joining Collaborative Initiative on Fetal Alcohol Spectrum Disorders, a consortium supported by the National Institutes of Health.

Kazue Hashimoto-Torii, Ph.D., Masaaki Torii, Ph.D., and the research teams they lead have joined a national research consortium for Fetal Alcohol Spectrum Disorders that is supported by the National Institutes of Health (NIH).

The Collaborative Initiative on Fetal Alcohol Spectrum Disorders (CIFASD) aims to leverage multidisciplinary approaches to develop effective interventions and treatments for Fetal Alcohol Spectrum Disorders.

“Both of our labs have been fortunate in receiving multiple R series research grants from the NIH. I am deeply honored that we now join this prestigious national consortium, which opens additional opportunities to collaborate with other labs with neurobehavioral, genetics and facial dysmorphology expertise as well as other specialized disciplines,” says Hashimoto-Torii, principal investigator in the Center for Neuroscience Research at Children’s National Health System.

Fetal Alcohol Spectrum Disorders are a constellation of conditions that result from exposure to alcohol in the womb that reflect the vastly different ways fetuses respond to that in utero insult. While early intervention is crucial, one challenge that continues to bedevil the field is trying to determine which pregnancies are most at risk.

“It is crucial to develop early and precise biomarkers for predicting children’s risk for cognitive and behavioral problems,” Hashimoto-Torii says. “Our labs will work on developing a novel approach for identifying such biomarkers.”

The Children’s researchers will examine epigenetic changes at the single cell level that may provide the earliest hint of cognitive and learning difficulties – long before children show any symptoms of such problems. Hashimoto-Torii’s lab will perform single-cell droplet digital polymerase chain reaction (PCR) based biomarker analysis of blood samples from experimental models and humans. Meanwhile, the lab run by Torii – also a principal investigator in the Center for Neuroscience Research – will collect blood samples from experimental models, perform comprehensive behavioral analysis, and evaluate potential correlations between behaviors seen in the experimental models and their drop-PCR results.

“Under the auspices of CIFASD, we ultimately hope to link these biomarkers from our lab with results that our colleagues are seeing in children in order to validate their ability to accurately predict outcomes from prenatal alcohol exposure,” she says.

Kazue Hashimoto Torii

A brain’s protector may also be its enemy

Kazue Hashimoto Torii

By looking back to the earliest moments of embryonic brain development, Kazue Hashimoto-Torii, Ph.D. and her collaborators sought to explain the molecular and cellular bases for complex congenital brain disorders that can result from exposure to harmful agents.

When the brain is exposed to an environmental stressor all is not immediately lost. Brain cells have mechanisms that protect them against the ravages of alcohol and other toxic substances. One of these is a protein the cells make, known as Heat Shock Factor 1 (Hsf1), which helps to shield them from damage. The fetal brain also can make Hsf1, which protects its particularly vulnerable cells from environmental stressors that pregnant mothers are exposed to during gestation.

However, a new study suggests that this system is not perfect. Research led by Children’s National Health System scientists suggests that when too much Hsf1 is produced, it actually can impair the brain during development. While this finding was made in a preclinical model, it raises questions about neural risks for human infants if their mothers drink alcohol in the first or second trimester of pregnancy.

When fetuses are chronically exposed to harmful agents such as alcohol, ethanol or methyl mercury in utero, the experience can negatively affect fetal brain development in unpredictable ways. Some fetal brains show little or no damage, while others suffer severe damage. By looking at the earliest moments of embryonic brain development, an international research team that includes five Children’s National authors sought to explain the molecular and cellular bases for complex congenital brain disorders that can result from exposure to such harmful agents.

“From a public health perspective, there is ongoing debate about whether there is any level of drinking by pregnant women that is ‘safe,’ ” says Kazue Hashimoto-Torii, Ph.D., principal investigator in the Center for Neuroscience Research at Children’s National and senior author of the paper published May 2 in Nature Communications. “We gave ethanol to pregnant preclinical models and found their offspring’s neural cells experienced widely differing responses to this environmental stress. It remains unclear which precise threshold of stress exposure represents the tipping point, transforming what should be a neuroprotective response into a damaging response. Even at lower levels of alcohol exposure, however, the risk for fetal neural cells is not zero,” Hashimoto-Torii adds.

The cerebral cortex – the thin outer layer of the cerebrum and cerebellum that enables the brain to process information – is particularly vulnerable to disturbances in the womb, the study authors write. To fend off insult, neural cells employ a number of self-preservation strategies, including launching the protective Hsf1-Heat shock protein (Hsp) signaling pathway that is used by a wide range of organisms, from single-cell microbes to humans. Developing fetuses activate Hsf1-Hsp signaling upon exposure to environmental stressors, some to no avail.

To help unravel the neurological mystery, the researchers used a method that allows a single molecule to fluoresce during stress exposure. They tapped specific environmental stressors, such as ethanol, hydrogen peroxide and methyl mercury – each of which are known to produce oxidative stress at defined concentrations. And, using an experimental model, they examined the Hsf1 activation pattern in the developing cerebral cortex by creating a marker, an encoding gene tagged with a type of fluorescent protein that makes it glow bright red.

“Our results suggest that heterogeneous events of abnormal brain development may occur probabilistically – which explains patterns of cortical malformations that vary with each individual, even when these individuals are exposed to similar levels of environmental stressors,” Hashimoto-Torii adds.

Among the more striking findings, neural cells with excessively high levels of Hsf1-Hsp activation due to ethanol exposure experience disruptions to normal development, with delayed migration by immature cortical neurons. For the fetal brain to develop normally, neurons need to migrate to precise places in the brain at just the right time to enable robust neural connections. When neurons fail to arrive at their destinations or get there too late, there can be gaps in the neural network, compromising efficient and effective communication across the brain’s various regions.

“Even a short period of Hsf1 overactivation during prenatal development causes critical neuronal migration deficiency. The severity of deficiency depends on the duration of Hsf1 overactivation,” Hashimoto-Torii says. “Expression patterns vary, however, across various tissues. Stochastic response within individual cells may be largely responsible for variability seen within tissue and organs.”

The research team found one bright spot: Cortical neurons that stalled due to lack of the microtubule-associated molecule Dcx were able to regain their ability to migrate properly when the gene was replenished after birth. A reduction in Hsf1 activity after birth, however, did not show the same ability to trigger the “reset” button on neural development.

“The finding suggests that genes other than microtubule-associated genes may play pivotal roles in ensuring that migrating neurons reach their assigned destinations in the brain at the right time – despite the added challenge of excessive Hsf1 activation,” according to Hashimoto-Torii.

Fetal Brain Cells

Tracking environmental stress damage in the brain

Fluorescence Reporter

A team led by Children’s National developed a fluorescence reporter system in an experimental model that can single out neurons that have survived prenatal damage but remain vulnerable after birth.

What’s known

When fetuses are exposed to environmental stressors, such as maternal smoking or alcohol consumption, radiation or too little oxygen, some of these cells can die. A portion of those that survive often have lingering damage and remain more susceptible to further environmental insults than healthy cells; however, researchers haven’t had a way to identify these weakened cells. This lack of knowledge has made it difficult to discover the mechanisms behind pathological brain development thought to arise from these very early environmental exposures, as well as ways to prevent or treat it.

What’s new

A team led by Kazue Hashimoto-Torii, Ph.D., a principal investigator in the Center for Neuroscience Research at Children’s National Health System, developed a marker that makes a protein known as Heat Shock Factor 1 glow red. This protein is produced in cells that become stressed through exposure to a variety of environmental insults. Gestation is a particularly vulnerable time for rapidly dividing nerve cells in the fetal brain. Tests showed that this marker worked not just on cells in petri dishes but also in an experimental model to detect brain cells that were damaged and remained vulnerable after exposure to a variety of different stressors. Tweaks to the system allowed the researchers to follow the progeny of cells that were affected by the initial stressor and track them as they divided and spread throughout the brain. By identifying which neurons are vulnerable, the study authors say, researchers eventually might be able to develop interventions that could slow or stop damage before symptoms arise.

Questions for future research

Q: How do different environmental insults damage brain cells during gestation?
Q: How does this damage translate into pathology in organisms as they mature?
Q: Do the progeny of damaged brain cells retain the same degree of damage as they divide and spread?
Q: Can this new detection system be used to find and track damage in other organs, such as the heart, eye and liver?

Source: Torii, M., S. Masanori, Y.W. Chang, S. Ishii, S.G. Waxman, J.D. Kocsis, P. Rakic and K. Hashimoto-Torii. “Detection of vulnerable neurons damaged by environmental insults in utero.” Published Dec. 22, 2016 by Proceedings of the National Academy of Sciences. doi: 10.1073/pnas.1620641114