Organoid-Based Models for Traumatic Brain Injury: Challenges, Innovations, and Future Perspectives

Abstract

Traumatic brain injury (TBI) is a major health concern impacting millions of individuals across the globe. Understanding how TBI damages the brain and finding better treatments are urgent needs. Traditional laboratory models, such as cell cultures and animal studies, often cannot fully represent the complex nature of human brain injuries. Recently, organoid-based models have become exciting new tools in brain research. These models are three-dimensional clusters of brain cells grown from human stem cells, which closely mimic the structure and function of the human brain. Because they come from human cells and grow in three dimensions, brain organoids provide a more accurate and detailed way to study how the brain develops and responds to injury compared with older methods. Brain organoids reproduce several essential characteristics of the developing human brain, making them highly valuable for scientific research. They allow scientists to explore the detailed processes involved in TBI and test new treatment approaches in a controlled laboratory setting. This flexibility and closeness to human biology make organoids a promising platform for advancing TBI research. In this review, we discuss the latest progress in using brain organoids to model TBI. We look at how well these models mimic the injury processes seen in real brain trauma and highlight both the challenges and future possibilities for improving and applying organoid-based models. Overall, organoid technology represents a significant step forward in understanding TBI and developing effective therapies.

 

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Keywords

3D cell cultures - brain organoids - injury models - mechanical injury - traumatic brain injury (TBI)

Introduction

Traumatic brain injury (TBI) remains a global health challenge, affecting approximately 10 million people annually. While many individuals survive, they often endure lifelong disabilities.[1] The pathology of TBI is highly complex and multifactorial, typically categorized into primary and secondary injuries.[2] Primary injury occurs at the moment of impact and, depending on severity, can result in structural damage, inflammation, axonal shear, and cell death. This may lead to complications such as headaches, contusions, hemorrhage, loss of consciousness, skull fractures, cerebral atrophy, or even death.[3] Secondary injury unfolds over time, involving a cascade of metabolic, inflammatory, and degenerative processes,[4] which can contribute to various neurodegenerative diseases. The neurological and cognitive deficits caused by TBI affect the quality of life and place a substantial burden on society.[5]

Despite extensive research, there are currently no definitive therapies for TBI-associated deficits. Although significant progress has been made in understanding the molecular and cellular pathways underlying TBI, the connection between these mechanisms and biomechanical forces remains insufficiently explored.[6] The processes driving posttraumatic neuroinflammation and neurodegeneration, as well as the impact of TBI on neural activity and large-scale brain networks, remain poorly understood. Additionally, strategies for repairing neural circuit damage following TBI continue to be elusive.

TBI has long posed a serious risk to human health, with its long-term consequences drastically impairing patients' quality of life. Despite growing awareness, current clinical interventions primarily focus on neuroprotection and mitigating secondary injuries, offering only limited benefits in terms of prognosis and overall recovery.[7] Regenerative approaches, particularly cell-based therapies, have gained considerable attention as potential avenues for enhancing TBI treatment outcomes.

Although substantial advancements have been made in neuroscience over the past decades, effective treatments for certain neurological disorders remain challenging. To improve our understanding of disease mechanisms, refine disease models, and enhance therapeutic efficacy, the development of more advanced technologies is imperative.

Rodent models have been widely used to investigate the intricate cascade of biological events involved in TBI.[8] However, the mouse brain differs significantly from the human brain in terms of structural complexity, regional organization, and gene expression profiles. While rodent TBI models effectively recapitulate certain pathological features, the spatial and temporal interplay of diverse cell types and signaling networks necessitates human-specific models for improved translational relevance.[9] [10]

To address these limitations, injury-based in vitro culture systems have been developed to model TBI in neurons derived from human-induced pluripotent stem cells (iPSCs).[11] Recent technological advances have enabled the development of cerebral organoids, three-dimensional (3D) brain-like structures generated in vitro. These models closely replicate the gene expression patterns and epigenetic signatures of the human brain, offering a promising platform for studying TBI pathophysiology and potential therapeutic interventions.[12]

 

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TBI Causes and Diagnosis

TBI results from an external force impacting the head, leading to temporary or permanent neurological dysfunction.[2] The primary causes of TBI include falls, motor vehicle accidents, sports-related injuries, and violent assaults, with falls being the most common among older adults and young children. High-impact trauma can cause focal injuries, such as contusions and hemorrhages, or diffuse damage, including axonal shearing and neuroinflammation. The diagnosis of TBI relies on a combination of clinical assessment, neurological examinations, and imaging modalities. Initial evaluation includes the Glasgow Coma Scale (GCS), which assesses consciousness level, along with cognitive and motor function tests. Advanced imaging techniques, such as computed tomography (CT) and magnetic resonance imaging (MRI), are crucial for detecting structural injuries, hemorrhages, and cerebral edema.[13] Emerging diagnostic tools, including blood-based biomarkers like S100B, GFAP, and neurofilament light chain (NfL), offer promising noninvasive approaches for early detection and prognosis.[14] Despite these advancements, TBI diagnosis remains challenging due to the heterogeneity of injury patterns, highlighting the need for improved diagnostic strategies and biomarkers for better patient management.

 

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Management for TBI Patients

TBIs, both primary and secondary injuries, affect how well a person can think, move, and recover. The primary injury happens at the moment of impact and is caused by a direct blow to the head. This damage cannot be undone and starts a chain reaction in the body. The secondary injury happens later and includes problems like low oxygen to the brain, brain swelling, high pressure inside the skull, fluid buildup, infections, and other issues that can make the brain injury worse.[15]

The effects of brain injury can also impact organs outside the brain, leading to multiorgan dysfunction syndrome (MODS), which occurs in more than 68% of patients with moderate to severe TBI within the first 72 hours of hospital admission. When these other organs stop working properly, it can create a harmful cycle that worsens brain damage. A key factor in the development of MODS after TBI is autonomic dysfunction, particularly in cases of severe brain injury.[16] This dysfunction triggers a chain of physiological problems that affect several organs outside the brain, including the lungs, heart and blood vessels, kidneys, and blood clotting systems. As a result, TBI patients who develop MODS often face a poor long-term outlook and have a higher risk of death.[17]

 

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Types of TBI

TBI encompasses a diverse spectrum of injuries categorized based on the mechanism, severity, and pathophysiological impact on brain structures. TBI is generally divided into two main types: closed (blunt) TBI, where the skull remains intact, and penetrating (open) TBI, where an object breaks through the skull and enters the brain. Closed TBI occurs when an external force impacts the head without breaking the skull, often leading to contusions, concussions, diffuse axonal injury (DAI), or intracranial hemorrhages.[18] This type is common in falls, motor vehicle accidents, and sports-related injuries. In contrast, penetrating TBI results from an object, such as a bullet, knife, or skull fragment, breaching the brain tissue, often causing severe focal damage, hemorrhage, and increased risk of infection.[19]

TBI severity is classified into mild, moderate, and severe based on clinical assessments such as the GCS. Mild TBI (concussion), the most frequent form, can result in transient symptoms like headache, dizziness, confusion, and brief loss of consciousness, with potential long-term consequences such as post-concussion syndrome and chronic traumatic encephalopathy after repetitive injuries.[20] Moderate TBI involves more significant neurological impairment, with longer unconsciousness, memory loss, and potential cognitive deficits. Severe TBI often leads to prolonged unconsciousness or coma, extensive brain damage, and long-term disabilities, including motor, cognitive, and behavioral impairments.[21]

TBI can also be classified based on the type and extent of injury. Focal injuries, which involve localized damage to specific brain regions, include cerebral contusions, epidural hematomas (bleeding between the dura mater and skull), subdural hematomas (bleeding between the dura and arachnoid layers), and intracerebral hemorrhages (bleeding within brain tissue). These injuries can cause elevated intracranial pressure, herniation, and significant neurological deficits if untreated.[22]

Additionally, TBI may be categorized into primary and secondary injuries. Primary injury refers to the immediate mechanical damage at the moment of impact, including skull fractures, hemorrhages, and axonal disruption. Secondary injury develops over hours to days due to biochemical and cellular responses, including neuroinflammation, oxidative stress, excitotoxicity, and disruption of the blood–brain barrier (BBB), contributing to progressive neuronal loss and worsening outcomes.

Given the complexity and heterogeneity of TBI, accurate classification is essential for guiding treatment strategies, prognosis assessment, and rehabilitation planning. Advances in neuroimaging, biomarker discovery, and experimental models continue to refine our understanding of TBI subtypes, ultimately improving patient management and therapeutic interventions.

 

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Current Problem and Challenges with Severe TBI

Severe TBI remains a critical global health issue, posing significant challenges in diagnosis, treatment, and long-term patient management.[1] Despite advancements in neurocritical care, severe TBI is associated with high mortality and morbidity, with survivors often experiencing profound neurological deficits, cognitive impairment, and permanent disability.[5] One of the primary challenges is the lack of effective neuroprotective therapies, as current treatments primarily focus on stabilizing the patient and managing secondary injuries rather than directly promoting neural repair. The heterogeneity of severe TBI, including variations in injury mechanisms, anatomical damage, and patient-specific responses, complicates treatment strategies and clinical trials, making it difficult to develop standardized therapeutic protocols.

Another major challenge is the limited ability of conventional neuroimaging techniques such as CT and MRI to fully capture the complex pathophysiological changes occurring in the injured brain, including DAI, microvascular damage, and metabolic dysfunction. The absence of biomarkers makes it more difficult to predict how the condition will progress, and to develop personalized treatment. Additionally, secondary injury mechanisms, including neuroinflammation, excitotoxicity, oxidative stress, and disruption of the BBB, continue to progress long after the initial trauma, exacerbating neuronal loss and worsening patient outcomes. Current therapeutic interventions, such as intracranial pressure management, hypothermia, and decompressive craniectomy, have variable efficacy, and no pharmacological agent has demonstrated consistent neuroprotective benefits in clinical trials.[23]

Long-term rehabilitation and recovery present additional challenges, as many patients require prolonged intensive care, cognitive rehabilitation, and support for motor and sensory deficits. The economic and social burden of severe TBI is immense, affecting not only patients but also caregivers and health care systems, particularly in low-resource settings where access to specialized care and rehabilitation is limited.

 

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The Need for Advanced TBI Models

Despite significant progress in TBI research, the lack of accurate and translational models remains a major barrier to understanding its complex pathophysiology and developing effective treatments. Traditional animal models, while valuable, fail to fully replicate the structural, genetic, and physiological characteristics of the human brain, limiting their ability to predict clinical outcomes. Similarly, in vitro models, such as two-dimensional (2D) cell cultures, lack the intricate 3D organization and cellular interactions necessary to mimic real-world brain injuries. As a result, many promising therapies that show efficacy in preclinical models fail in human trials, underscoring the urgent need for more human-relevant TBI models.[24]

Advanced organ-on-a-chip, bioengineered scaffolds, and cerebral organoid-based models offer promising solutions by recapitulating key features of the human brain, including neuronal diversity, synaptic connectivity, and biomechanical properties. Human-derived iPSC models enable patient-specific investigations, allowing for personalized insights into injury mechanisms and therapeutic responses. Additionally, integrating computational modeling and artificial intelligence can enhance TBI research by predicting injury progression and optimizing treatment strategies. The development of more sophisticated, physiologically relevant models is critical for bridging the gap between basic research and clinical translation, ultimately leading to more effective neuroprotective and regenerative therapies for TBI patients.[25] [26]

 

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Brain Organoids

Brain organoids are 3D, self-organizing neural structures derived from human pluripotent stem cells (hPSCs) that recapitulate key aspects of brain development, cellular composition, and functional connectivity. These miniature brain-like models provide an unprecedented opportunity to study human neurodevelopment, disease mechanisms, and injury responses in a controlled laboratory setting. Unlike traditional 2D cell cultures, brain organoids exhibit complex cytoarchitecture, including cortical layering, neuronal differentiation, and synaptic activity, making them highly relevant for modeling neurological disorders and injuries such as TBI, stroke, and neurodegenerative diseases.

In the context of TBI research, brain organoids enable the investigation of secondary injury processes such as neuroinflammation, oxidative stress, and synaptic dysfunction, which are difficult to replicate in conventional animal models. Additionally, their human origin provides an advantage over rodent-based models, allowing for more accurate genetic and molecular studies that align with clinical conditions.

Advances in bioengineering, bioprinting, and microfluidic systems are further enhancing the physiological relevance of brain organoids, paving the way for personalized medicine approaches, drug screening, and regenerative therapies. While challenges such as vascularization and long-term maturation remain, brain organoids hold immense potential as a transformative tool in neuroscience, offering deeper insights into brain function and pathology.

 

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Types of Brain Organoids

Brain organoids are classified based on their developmental origin and the specific brain regions they model. These organoids replicate key structural and functional characteristics of different areas of the human brain, making them valuable tools for studying neurodevelopment, disease mechanisms, and brain injuries such as TBI, stroke, and neurodegenerative disorders. Over the past decade, 3D organoid technology has gained significant attention in the field of stem cell research. Organoids are small, laboratory-grown 3D cell structures derived from iPSCs or specific organ progenitor cells. Thanks to their ability to self-renew and self-organize, organoids can closely replicate several features of real organs as they develop in the body.[27] Researchers have effectively generated brain organoids that represent distinct areas of the brain, including the forebrain, midbrain, cerebellum, cortex, and hippocampus.[28] A variety of techniques are used to generate these organoids, with many focused on mimicking the natural development of the human brain.

The main types of brain organoids include:

• Cerebral organoids: these are the most widely used brain organoids, modeling the cerebral cortex with distinct neuronal layers, progenitor zones, and functional synaptic activity. They are crucial for studying cortical development, neurogenesis, and injuries affecting higher order cognitive functions. Brain organoids have been merged with vascular organoids to produce vascularized brain organoids. These combined models formed functional structures similar to the BBB and exhibited an increased number of neural progenitor cells (NPCs).[29] The brain's microvasculature releases important developmental molecules such as brain-derived neurotrophic factor (BDNF).[30] When brain organoids are grown alongside a vascular system, their development improves, supported by factors like BDNF that promote growth and maturation. These findings indicate that the presence of blood vessels plays a key role in regulating neural development.[31]

• Midbrain organoids: midbrain organoids, which replicate the midbrain region and contain dopaminergic neurons, have been extensively used for studying Parkinson's disease and neurodegenerative conditions. However, their application in TBI research is gaining increasing attention due to their ability to model region-specific neuronal responses to mechanical and biochemical insults.

  • iPSC-derived brain organoids represent a promising platform for recapitulating protein expression patterns and physiological characteristics of the midbrain following trauma. Although numerous protocols exist for generating brain organoids, few studies have comprehensively examined the temporal dynamics of mature region-specific marker expression post-injury. To address this gap, recent advancements have enabled the differentiation of midbrain-specific organoids from iPSC lines using a matrix-free, bioreactor-based system, allowing for controlled evaluation of trauma-induced cellular and molecular changes.[32]

  • The characterization of midbrain organoids post-trauma includes immunofluorescence and immunohistochemistry analysis from early (day 7) to late (day 90) differentiation stages, alongside ultrastructural examination via electron microscopy and transcriptomic profiling through RNA sequencing.[33] This comprehensive approach provides critical insights into neuronal survival, inflammatory responses, and neurodegenerative cascades triggered by TBI-like conditions.

  • Beyond optimizing differentiation protocols, the identified midbrain-specific markers serve as key control parameters for investigating trauma-induced phenotypic alterations in organoids derived from patient iPSC lines with genetic predispositions to neurodegeneration. These models not only facilitate the study of TBI pathology in a human-relevant system but also provide a foundation for testing potential neuroprotective and regenerative therapies tailored for midbrain injury.

• Hippocampal organoids: hippocampal organoids are engineered to closely mimic the structure and function of the hippocampus, a critical brain region involved in learning and memory. These organoids serve as valuable models for studying neurodegenerative diseases like Alzheimer's disease, helping researchers investigate disease mechanisms and potential treatments. Additionally, hippocampal organoids provide important insights into how brain injuries affect memory formation and retrieval by allowing detailed examination of cellular and molecular changes after trauma.[34] Their ability to model both normal hippocampal function and pathological conditions makes them powerful tools in neuroscience research.

• Cerebellar organoids: cerebellar organoids are designed to closely resemble the cerebellum, containing key cell types such as Purkinje cells and granule neurons that are essential for motor coordination and balance. These organoids provide a valuable model for studying neurological conditions like ataxia and other movement disorders, as well as motor dysfunction following TBI.[35] By replicating the cellular architecture and function of the cerebellum, these models enable researchers to explore disease mechanisms and test potential therapies targeting cerebellar-related impairments.

• Thalamic organoids: thalamic organoids are developed to replicate the structure and function of the thalamus, an essential brain region that acts as a relay center for sensory processing and signal integration.[36] These organoids are particularly useful for studying sensory deficits that can occur after brain injuries as well as various neurodevelopmental disorders. By modeling the thalamus in vitro, researchers can better understand how disruptions in this area contribute to neurological dysfunction.

• Hypothalamic organoids: hypothalamic organoids contain hormone-producing neurons and are valuable for investigating the neuroendocrine system's regulation of bodily functions. They are especially important for studying metabolic disorders and hypothalamic dysfunctions, which affect processes such as appetite, temperature regulation, and stress responses.[37] These organoids provide a platform to explore the complex interactions between the brain and endocrine system in both health and disease.

 

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Generation of Brain Organoids for TBI Research

The development of brain organoids for TBI research has emerged as a promising approach to better understand injury mechanisms and explore potential therapeutic interventions. These 3D, self-organizing neural structures are generated from hPSCs, including embryonic stem cells and iPSCs. The organoid formation process involves directing neural differentiation under controlled conditions to mimic the development of specific brain regions affected by TBI.

The generation of TBI-specific brain organoids follows key steps:

• Stem cell differentiation: hPSCs are cultured in specialized media containing neurotrophic factors and small molecules that guide their differentiation into NPCs and subsequently into neurons, astrocytes, and other brain cell types.[38]

• Self-organization and maturation: the differentiating cells aggregate and develop into structured organoids, forming layers that resemble those found in the human brain, including the cortical plate, ventricular zone, and intermediate zone. Long-term culture allows for neuronal maturation, synaptogenesis, and functional activity, making them highly relevant for TBI studies.[28]

• Biomechanical injury induction: to model TBI in vitro, brain organoids are subjected to mechanical forces that mimic injury mechanisms seen in TBI, such as compression, shear stress, and cavitation-induced damage. These models enable researchers to study secondary injury processes, including neuroinflammation, oxidative stress, excitotoxicity, and BBB disruption.[39]

• Integration with advanced technologies: approaches, such as microfluidics, organ-on-a-chip platforms, and bioprinting, enhance the physiological relevance of brain organoids by enabling the study of vascularization, immune responses, and neuronal network disruptions following TBI. Additionally, incorporating real-time imaging, electrophysiology, and single-cell transcriptomics allows for a deeper understanding of injury progression at both cellular and molecular levels.[40]

 

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Brain Organoid–Based TBI Models

Mechanical injury models: brain organoids are subjected to controlled mechanical forces, such as compression, shear stress, and cavitation-induced damage, to mimic primary injury mechanisms in TBI. These models allow researchers to study cellular and molecular responses following biomechanical trauma.[39] Mechanical injury models for TBI in brain organoids are designed to replicate the physical forces and damage associated with TBI ([Fig. 1]). Here are some common types:

Fig. 1 A schematic representation of various mechanical injury models. (A) Compression models, demonstrating localized tissue compression. (B) The scratch injury model, where mechanical abrasion is applied to induce damage. Stretchmodels, showcasing controlled tensile forces applied to cells or tissues. (C)

Compression models: these models use controlled compressive forces on brain organoids to simulate the effects of impact or pressure-related brain injuries. A previous study by Ramirez et al stands out as the first to use cerebral organoids to model TBI. Their innovative method, which involved penetrative techniques and gradual compression applied to both human cerebral organoids and mouse models, showed the underlying mechanisms of TBI.[41] Another study demonstrated a method to simulate mild and moderate TBI in human cerebral organoids by applying uniaxial compression at relevant strain rates. This approach allowed researchers to observe how TBI affects neuronal signaling and gene expression within the organoids.[42]

Impact injury models simulate blunt force trauma to the brain through direct mechanical impact. One common method involves weight-drop-based impactors, where a controlled weight-drop mechanism drives a stylus or flat impactor head onto tissue. These setups replicate focal contusion and compression injuries and are frequently used to study severe TBI, often with rodent tissue explants or brain slices. Alternatively, engineered 3D neural tissue models serve as valuable tools for trauma research.[43] These models compartmentalize cell bodies and axonal tracts, and show how injuries affect white matter. For studying injury effects at the single-cell level, researchers have also developed a microfluidic device that delivers up to 90% strain to individual cells using a nickel–iron armature, allowing for further culture or analysis afterward.[44] [45]

Stretch models: these models involve stretching brain organoids to mimic the tensile forces that occur during rapid acceleration or deceleration, as seen in many traumatic brain and spinal cord injuries. Stretch-induced injury models are among the most widely used in vitro systems for studying TBI. In these setups, surrogate tissues or cultured cells are placed on a flexible surface, often made of polydimethylsiloxane, which is then stretched using a hollow indenter or through pulses of compressed air or nitrogen gas.[46] These mechanical strain models have been thoroughly studied and are available commercially, typically as flexible silicone-based multi-well plates paired with validated systems that control and deliver injury.[47]

Scratch injury model: transection or scratch injury models are commonly used to study trauma-induced axon damage and evaluate treatments that promote axonal regeneration. While primary axotomy is rare in TBI, it is more common in spinal cord injury and can be replicated through transection, which also triggers secondary injury responses seen in vivo.[48] The scratch assay, a simple in vitro model, involves scraping neuron or astrocyte cultures to study cellular responses like astrocytic activation, important for healing. Another approach, the “brain-on-a-chip” model, uses 3D cell cultures in a microfluidic setup to mimic brain tissue responses. It is useful for drug screening, studying glial scar formation, and modeling disease processes, though delivering precise mechanical injury at the microscopic level remains a technical challenge.[49]

 

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Inflammation and Secondary Injury Models

Brain organoids can be used to simulate the secondary injury phase of TBI by inducing pro-inflammatory responses. This stage, which occurs after the initial trauma, involves complex biological processes such as glial activation, oxidative stress, excitotoxicity, and programmed cell death (apoptosis). Studying these processes is essential to understanding how ongoing inflammation contributes to long-term brain damage and functional decline in TBI patients.

Various models within this framework allow researchers to examine specific aspects of the inflammatory response. Neuroinflammation models focus on the role of microglia and astrocytes, which are the brain's resident immune cells. While short-term inflammation can help with tissue repair, prolonged or chronic inflammation often results in further damage to neurons and worsened outcomes. Cytokine and chemokine pathway models are used to explore how immune signaling molecules are released after injury. These molecules help recruit other immune cells to the injury site, but if not properly regulated, they can lead to excessive inflammation and secondary injury.[50] [51]

Another key area is oxidative stress, which is modeled by studying the production of reactive oxygen species and free radicals after TBI. These harmful molecules can damage DNA, proteins, and cell membranes, amplifying the injury. In addition, BBB disruption models help understand how the protective barrier around the brain becomes compromised after TBI, allowing harmful substances to enter and trigger further inflammation. Lastly, neurogenic inflammation models investigate the role of neuropeptides like substance P in increasing the inflammatory response, contributing to prolonged damage.[52]

Together, these organoid-based models provide a comprehensive and human-relevant way to study the secondary injury mechanisms in TBI, especially those driven by inflammation. This knowledge is vital for developing therapies that can reduce long-term damage and improve recovery after brain injury.

 

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Hypoxia and Ischemic Injury Models

TBI frequently disrupts blood flow and oxygen supply to the brain, resulting in vascular dysfunction and low oxygen levels (hypoxia). These conditions can result in the death of brain cells (neurons) and disrupt the brain's normal metabolism. To study these effects in the laboratory, researchers use brain organoids and simulate oxygen and glucose deprivation, mimicking the conditions of ischemia. This helps in understanding how the brain is damaged by lack of oxygen and in testing treatments that might protect brain cells.

Several methods are used to explore hypoxia-related injury. Controlled hypoxia models reduce the amount of oxygen in the environment to mimic low-oxygen conditions, helping researchers observe how the brain responds to hypoxia alone. Ischemic stroke models, like the middle cerebral artery occlusion technique in animals, are used to block blood flow to certain parts of the brain, replicating the effects of stroke and showing how this interacts with TBI.[53] Another approach is the fluid percussion injury combined with hypoxia, where physical brain trauma is followed by induced oxygen deprivation. This simulates real-life situations where a patient suffers a head injury along with breathing difficulties.

In laboratory settings, in vitro hypoxia models use brain organoids or cultured brain cells exposed to low oxygen to study responses like inflammation, oxidative stress, and cell death. Finally, BBB disruption models explore how hypoxia and ischemia damage the protective barrier around the brain. This allows harmful substances to enter, causing more injury and inflammation. Together, these models help researchers understand the complex effects of hypoxia in TBI and develop strategies to reduce brain damage.[54]

 

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Microfluidic and Organ-on-a-Chip Models

New bioengineering techniques now make it possible to combine brain organoids with microfluidic devices, allowing scientists to carefully control injury conditions and observe how brain cells respond in real time. These integrated systems help create more realistic models of the human brain and are especially useful for high-throughput testing of potential treatments for neurological conditions such as TBI. Microfluidic and organ-on-a-chip technologies are cutting-edge platforms that replicate how human organs function at a miniature scale. These systems use tiny channels and chambers to simulate the movement of fluids and the exchange of nutrients, mimicking the natural environment of human tissues more closely than traditional laboratory setups.[55]

By combining this technology with tissue engineering, researchers can create dynamic and controlled environments that simulate specific organ functions, study disease processes, and test how drugs behave in the body. Organ-on-a-chip models take this a step further by using multiple cell types and engineered materials to recreate the structure and activity of entire organs, such as the brain, liver, or lungs.[56] These advanced models are transforming biomedical research because they provide real-time data, reduce the need for animal testing, and offer more accurate predictions of how the human body might react to treatments. As a result, they are increasingly used in drug discovery, toxicity screening, disease modeling, and personalized medicine.[57]

 

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Brain Organoid-TBI Models and Their Applications

Brain organoid-TBI models represent an approach to studying the complex mechanisms of brain injury, overcoming the limitations of traditional in vivo (animal) and in vitro (2D cell culture) models. These 3D self-organizing neural structures, derived from hPSCs, offer a physiologically relevant system to investigate TBI pathophysiology, neuroinflammation, and potential therapeutic interventions.

Brain organoid models are advanced laboratory-grown mini-brains developed from human stem cells that offer a powerful platform to study the human brain, especially in the context of TBI. TBI occurs when a sudden physical impact or force causes damage to the brain, triggering a series of complex biological responses. These include neuronal injury, glial cell activation, synaptic dysfunction, and, in severe cases, progressive neurodegeneration. Brain organoids mimic the structure and function of the human brain more closely than traditional models and thus provide a unique opportunity to study these changes at the cellular and molecular level in a human-relevant context.[39]

One of the major advantages of using brain organoid models is their utility in drug screening. Organoids allow researchers to test the safety and effectiveness of potential therapeutic agents, such as neuroprotective drugs, anti-inflammatory compounds, and regenerative molecules. Because they are derived from human cells, organoids offer a more accurate and predictive platform for assessing how a human brain might respond to different treatments, which can accelerate the process of drug discovery and reduce reliance on animal models.[58]

Additionally, brain organoid-TBI models are increasingly being used in the field of regenerative medicine. They support the development of novel treatment approaches including stem cell therapies, gene editing techniques such as CRISPR, and bioengineered scaffolds that can potentially repair damaged neural tissue. These tools can be tested on organoid systems to understand their safety, integration, and effectiveness in promoting neural repair and functional recovery following brain injury.[59]

Overall, brain organoid models are transforming the way scientists approach TBI research. They provide a highly relevant system to study injury mechanisms, identify and test new drugs, and explore advanced therapies for brain repair, with the ultimate goal of improving outcomes for TBI patients.

 

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Key Insights from Organoid-Based TBI Models

• Neuronal and axonal damage: organoids subjected to mechanical trauma exhibit axonal fragmentation, cytoskeletal disruption, and neuronal apoptosis, similar to DAI observed in TBI patients. Immunostaining for β-III tubulin, MAP2, and NF-L confirms axonal integrity loss.[60]

• Neuroinflammatory responses: post-injury, brain organoids demonstrate activation of astrocytes (GFAP), microglia (Iba1), and cytokine secretion (interleukin [IL]-6, tumor necrosis factor-α, IL-1β). Transcriptomic profiling reveals upregulation of inflammatory pathways, akin to secondary injury in TBI.[61]

• BBB dysfunction and vascular damage: although traditional brain organoids lack vasculature, integration with endothelial cells and pericytes has enabled the modeling of BBB disruption, a key feature of TBI. Organoids co-cultured with vascular components exhibit tight junction breakdown (ZO-1, occludin loss) and increased permeability, mimicking real-world pathology.[62]

• Therapeutic screening and drug discovery: organoid-based platforms facilitate high-throughput drug screening for TBI. Compounds targeting oxidative stress (such as N-acetylcysteine), inflammation, and excitotoxicity have been tested with promising neuroprotective effects.[63]

 

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Future Prospects

Brain organoid models hold immense potential for transforming TBI research, offering a physiologically relevant platform to study injury mechanisms, neuroinflammation, and therapeutic responses. Future advancements will focus on improving vascularization and immune system integration within organoids to better mimic the human brain microenvironment post-TBI.[64] The development of mechanically induced TBI models in organoids will enhance our ability to replicate primary and secondary injury cascades, allowing for more precise investigations of cellular damage, synaptic dysfunction, and glial responses. Additionally, integrating organ-on-a-chip technologies and bioprinting techniques will enable controlled injury paradigms and high-throughput drug screening.[65] As patient-derived brain organoids become more refined, they could serve as personalized models for testing neuroprotective and regenerative therapies, bridging the gap between preclinical research and clinical applications. Moving forward, brain organoids will play a critical role in deciphering TBI pathophysiology, evaluating targeted treatments, and advancing precision medicine for brain trauma recovery.

 

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Conclusion

Brain organoid models represent a groundbreaking advancement in TBI research, contributing a 3D, human-relevant platform to study injury mechanisms, neuroinflammation, and potential therapeutic strategies. While current models still face challenges in vascularization, immune system integration, and long-term maturation, ongoing advancements in bioprinting, organ-on-a-chip technologies, and patient-derived organoids will significantly enhance their translational potential. By allowing precise modeling of primary and secondary injury cascades, organoids can bridge the gap between preclinical research and clinical applications, accelerating the discovery of effective neuroprotective and regenerative therapies. As the field progresses, brain organoids will play an increasingly crucial role in personalized medicine, drug screening, and mechanistic studies, ultimately improving outcomes for TBI patients and shaping the future of neurotrauma research.

 

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Conflict of Interest

None declared.

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Publication History

Article published online:
19 August 2025