Latest research led by scientists at Arizona State University has revealed a number of the first detailed molecular clues related to considered one of the leading causes of death and disability, a condition often called traumatic brain injury (TBI).

TBI is a growing public health concern, affecting greater than 1.7 million Americans at an estimated annual cost of $76.5 billion dollars. It’s a number one reason behind death and disability for youngsters and young adults in industrialized countries, and other people who experience TBI usually tend to develop severe, long-term cognitive and behavioral deficits.

“Unfortunately, the molecular and cellular mechanisms of TBI injury progression are multifaceted and have yet to be fully elucidated,” said Sarah Stabenfeldt, an ASU professor and the leader and corresponding writer of the study, which appears within the journal Science Advances. “Consequently, this complexity affects the event of diagnostic and treatment options for TBI; the goal of our research was to deal with these current limitations.”

Their research approach was to perform a “biopanning” search to disclose several key molecular signatures, called biomarkers, identified directly after immediately after the injury event (the acute phase), and in addition the long-term consequences (the chronic phase) of TBI.

For TBI, the pathology evolves and changes over time, meaning that a single protein or receptor could also be upregulated at one phase of the injury, but not two weeks later. This dynamic environment makes developing a successful targeting strategy complicated.”

Sarah Stabenfeldt, ASU professor

To beat these limitations, The ASU scientists, led by Sarah Stabenfeldt utilize a mouse model for his or her study to start to check the foundation causes of TBI by identifying biomarkers—unique molecular fingerprints found with a given injury or disease.

“The neurotrauma research community is a well-established field that has developed and characterised preclinical animal models to higher understand TBI pathology and assess the efficacy of therapeutic interventions,” said Stabenfeldt. “Using the established mouse model enabled us to conduct biomarker discover where the complexity and evolution of the injury pathology was progressing.”

Scientists can often begin to design therapeutic agents or diagnostic devices based on biomarker discovery. Stabenfeldt’s team used a “bottom up” approach to biomarker discovery.

“Top-down” discovery methods are focused on assessing candidate biomarkers based on their known involvement within the condition of interest,” said study first writer Briana, a recent PhD graduate in Stabenfeldt’s lab. “In contrast, a “bottom-up” method analyzes changes in tissue composition and finds a strategy to connect those changes to the condition. It is a more unbiased approach but may be dangerous because you’ll be able to possibly discover markers that aren’t specific to the condition or pathology of interest.”

Next, they utilized several state-of-the-art ‘biopanning’ tools and techniques to discover and capture molecules, including such a “bait” technique for fishing out potential goal molecules called a phage-display system, along with high-speed DNA sequencing to discover protein targets inside the genome, and mass-spectrometers to sequence the peptide fragments from the phase display experiments.

One further roadblock to discovery is the unique physiology of a mesh-like network designed to guard the brain from injury or harmful chemicals, called the blood-brain barrier (BBB).

“The blood-brain barrier (BBB) barrier is a barrier between the vascular and brain tissue,” explains Stabenfeldt. “In a healthy individual, the BBB tightly regulates nutrient and waste exchange from the blood to the brain and vice versa, essentially compartmentalizing the brain/central nervous system.”

‘Nonetheless, this barrier also complicates drug delivery to the brain so that almost all molecules/drugs don’t passively cross this barrier; due to this fact, the drug delivery field has sought out ways to modulate each entry and delivery mechanisms. Similarly, for blood-based biomarkers for TBI or other neurodegenerative diseases, specificity to the pathology and transfer of the molecule (if it originates within the brain) from the brain to blood is a challenge.”

When a TBI occurs, the initial injury can disrupt the BBB, which triggers a cascade of cell death, torn, disrupted tissues and debris.

The long-term injury causes inflammation and swelling, and ends in the immune response to spring into motion, but in addition can result in an impairment of the brain’s energy sources, or can choke off the brain’s blood supply, resulting in more neuronal cell death and everlasting disability.

A key advantage of their suite of experimental tools and techniques of the phage display system is that the molecules and potential biomarkers identified are sufficiently small to slide through the tiny holes inside the meshwork of the BBB—thus, opening the strategy to therapeutics based on these molecules.

So, despite all these obstacles, the team found a way.

“Our study leverages the sensitivity and specificity of phage to find novel targeting motifs,” said Stabenfeldt. “The mix of phage and NGS [next-generation sequencing] has been used previously, thereby leveraging bioinformatic evaluation. The unique contribution of our study is putting all of those tools together specifically for an in vivo model of TBI.”

They found a collection of unique biomarkers related to only the acute or chronic phases of TBI. Within the acute phase, TBI targeting motif recognized targets related to mainly metabolic and mitochondrial (the powerhouse of the cell) dysfunction, whereas, the chronic TBI motif was largely related to neurodegenerative processes.

“Our method for biomarker discovery was sensitive enough to detect injury in brains that were collected at different points within the experiments,” said study first writer Briana Martinez, a recent PhD graduate in Stabenfeldt’s lab. “It was really interesting to see that proteins involved in neurodegenerative diseases were detected at 7 days post-injury, but not at the sooner, 1-day post-injury timepoint. The proven fact that we were in a position to observe these differences really showcases how useful this method may very well be in exploring various features of brain injury.”

It can also begin to clarify why individuals who have had a TBI are more prone to developing neurodegenerative diseases like Parkinson’s and Alzheimer’s later in life.

This successful discovery pipeline will now function the muse for the next-generation targeted TBI therapeutics and diagnostics.

Next, the group plans to further its collaborations with ASU’s clinical partners and expand their studies to start to search for these same molecules in human samples.

Source:

Journal reference:

Martinez, B.I., et al. (2022) Uncovering temporospatial sensitive TBI targeting strategies via in vivo phage display. Science. doi.org/10.1126/sciadv.abo5047.