CRISPR-Based Therapy for Ischemic Stroke: A Narrative Review

Abstract

Ischemic stroke (IS) is one of the most common neurological diseases worldwide and is caused by the blockage of cerebral blood vessels, leading to reduced blood flow and neuronal damage. Given the limitations of existing treatments, CRISPR gene-editing technology has emerged as a promising strategy to precisely target the molecular pathways underlying IS pathophysiology. By enabling intervention in genes regulating inflammation, apoptosis, and repair, CRISPR enables more precise and effective therapies. Various CRISPR delivery systems, including viral vectors, nanocarriers, and extracellular vesicles, play crucial roles in the effective access of this tool to neural cells. Studies have shown that the use of CRISPR-Cas9 to modulate key pathogenic pathways, including those governing inflammation, oxidative stress, and cell death, can prevent neuronal damage and improve neurological function. Additionally, targeting ncRNAs and RNA methylation with CRISPR-based systems plays a role in regulating oxidative stress and stress granule formation. The use of CRISPR to modulate cell communication and organelle transfer and correct mitochondrial mutations has also been considered a neuroprotective mechanism. Despite persistent challenges in targeted and safe delivery, substantial preclinical advances, primarily in rodent models, underscore the potential for CRISPR-based therapies to transform future stroke treatment. These findings suggest that CRISPR-based strategies could evolve into precision neurotherapeutics that address root molecular pathologies, potentially complementing or surpassing current stroke interventions.

Introduction

Ischemic stroke (IS), a leading cause of mortality and disability worldwide, results from the occlusion of cerebral arteries, which reduces cerebral blood flow and causes neuronal damage (Campbell et al. 2019). IS accounts for approximately 85% of all stroke cases (Ding et al. 2022). The need for novel therapies is intensified by the increasing global burden of IS and the limitations of current treatments. Hypertension is recognized as the most important risk factor for this disease, accounting for nearly 54% of stroke cases. Between 1990 and 2015, the prevalence of hypertension increased from 442 to 874 million people, and the prevalence of IS is predicted to increase by 24.9% over the next two decades. Despite advances in reperfusion therapies, such as intravenous thrombolysis and mechanical thrombectomy, their utility is limited by narrow therapeutic windows and often incomplete recovery. Furthermore, the repeated failure of neuroprotective agents in clinical trials underscores the urgent need for novel therapeutic paradigms (Paul and Candelario-Jalil 2021).

Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-based gene-editing technologies have emerged as transformative approaches, enabling precise modulation of molecular pathways central to IS pathophysiology (Radenovic 2024; Bonowicz et al. 2025). CRISPR allows scientists to accurately and purposefully alter or correct specific genes or segments of genetic codes. Variants such as CRISPR-Cas9 and CRISPR-CasRx can target DNA and RNA, respectively, offering advanced tools for modifying or silencing molecules implicated in IS pathology (Li et al. 2023).

CRISPR-CasRx enables intervention at critical ischemic signaling nodes, including necroptosis, apoptosis, and mitochondrial dysfunction. Simultaneous silencing of mRNA via CRISPR-CasRx has been shown to reduce infarct volume, decrease edema, and improve sensorimotor functions in mice subjected to transient middle cerebral artery occlusion (tMCAO) (Song et al. 2024). Additionally, CRISPR-Cas9 has demonstrated efficacy in correcting mitochondrial mutations such as m.15059G > A in the MT-CYB gene and attenuating inflammatory responses in cell lines (Bonowicz et al. 2025), thereby addressing root molecular causes of ischemic injury.

This article comprehensively reviews the functional mechanisms, delivery strategies, and therapeutic potential of CRISPR technology for IS while also discussing key translational challenges and future directions.

Figure 1 provides a schematic overview of key CRISPR-based strategies for treating ischemic stroke.

Fig. 1.

Schematic overview of CRISPR-based therapeutic strategies in ischemic stroke. CRISPR-Cas9 enables DNA editing to correct pathogenic mutations, such as MT-CYB, and attenuate neuroinflammatory responses. CRISPR-CasRx targets and degrades specific mRNAs involved in deleterious pathways (e.g., necroptosis), thereby reducing neuronal damage and promoting functional recovery. The figure was created using Microsoft PowerPoint. MT-CYB, Mitochondrially Encoded cytochrome b; mRNA, Messenger Ribonucleic Acid

CRISPR Delivery Systems in Ischemic Stroke Therapy

Effective delivery of CRISPR components to the central nervous system (CNS) remains a critical hurdle. Recent advances in delivery platforms have enhanced the translational potential of CRISPR for IS (Wilbie et al. 2019; Lino et al. 2018). Below, some of these systems are introduced:

Viral Vectors

Viral vectors, notably adeno-associated viruses (AAVs) and lentiviruses (LVs), are widely used for CNS gene delivery because of their high transduction efficiency and stable expression. AAVs, such as AAV9, which exhibits strong neural tropism, are favored for their favorable safety profile and potential for cell type-specific targeting (Song et al. 2024). LVs, which are capable of genomic integration, enable long-term expression of CRISPR components, which is useful for studying chronic poststroke processes (Skukan et al. 2022).

As depicted in Fig. 2, AAV-mediated delivery of CRISPR-CasRx enables targeted degradation of NSF and RIPK1 mRNAs, thereby preventing necroptotic neuronal death and preserving function in ischemic models.

Fig. 2.

Schematic illustration of AAV9-mediated delivery of CRISPR-CasRx for neuroprotection in ischemic stroke. The diagram depicts the targeted knockdown of NSF and RIPK1 mRNAs within neurons and glial cells, leading to inhibition of necroptotic pathways, reduced cell death, and preservation of neuronal integrity following stroke. The figure was created using Microsoft PowerPoint. AAV9, adeno-associated virus serotype 9; CasRx, CRISPR-associated RNA-targeting protein; NSF, N-ethylmaleimide-sensitive fusion ATPase; RIPK1, receptor-interacting protein kinase 1

The broader application of viral vectors is illustrated in Fig. 3, which compares the AAV and LV platforms for delivering both the CRISPR-Cas9 and CRISPR-CasRx systems in the context of ischemic stroke gene therapy.

Fig. 3.

Schematic illustration of AAV and LV vector–mediated delivery of CRISPR-Cas9 (DNA-targeting) and CRISPR-CasRx (RNA-targeting) systems for neuroprotection in ischemic stroke. Cas9 induces genomic modifications to correct disease-associated mutations or modulate gene expression, whereas mediates sequence-specific mRNA degradation to transiently silence pathogenic transcripts such as Ripk1 or Nsf. The figure was created using Microsoft PowerPoint. AAV: adeno-associated virus; LV: lentivirus; Cas9: CRISPR-associated protein 9; CasRx: CRISPR-associated RfxCas13d; gRNA: guide RNA

Nonviral Carrier Nanoparticles

Nonviral nanoparticles offer advantages, including reduced immunogenicity and increased cargo capacity. Lipid nanoparticles (LNPs), polymeric nanoparticles (e.g., PLGA) (Do et al. 2021), and inorganic nanoparticles (e.g., calcium phosphate) have been employed. A notable example is intranasal delivery of dCas9-SunTag via functionalized calcium phosphate nanoparticles, which upregulated Sirt1 expression and improved outcomes in a mouse IS model without significant toxicity (Ryu et al. 2024). Magnetic nanoparticles (MNPs) can also increase delivery efficiency via magnetofection and enable real-time MRI tracking (Huang et al. 2021).

Extracellular Vesicles (EVs)

EVs, including exosomes, are natural lipid nanoparticles that exhibit low immunogenicity and an innate ability to cross the blood‒brain barrier (BBB). Engineered EVs can package and deliver CRISPR ribonucleoproteins or mRNAs, enabling efficient, nonimmunogenic transport across the BBB (Li et al. 2023; Wang et al. 2024a; Sokolov et al. 2019) (Fig. 4).

Fig. 4.

Engineered extracellular vesicl-mediated CRISPR/Cas9 delivery to the brain for targeted gene editing in neurological disorders. EVs are loaded with CRISPR/Cas9 ribonucleoprotein complexes (Cas9 protein and sgRNA). These modified EVs traverse the blood‒brain barrier BBB and selectively release their cargo into neurons, enabling accurate genome editing. The figure was created using Microsoft PowerPoint. CD9-HuR: Cluster of Differentiation 9/Human antigen R

Comparative Analysis and Strategic Selection

The choice of delivery system must align with therapeutic goals. For acute neuroprotection requiring transient gene modulation (e.g., silencing RIPK1), RNA-targeting CRISPR systems delivered via EVs or LNPs offer favorable safety profiles because of their self-limiting activity and lack of genomic integration. For strategies requiring sustained expression, such as chronic neuroregeneration, AAV vectors may be more appropriate, provided that immune responses are managed (Wilbie et al. 2019; Skukan et al. 2022; Wang et al. 2024a).

Therapeutic Mechanisms Based on CRISPR

CRISPR gene editing technology affects the pathophysiology of IS through various mechanisms. Below, we synthesize preclinical evidence under mechanistic headings, emphasizing the quality of evidence (in vitro, rodent, primate) and translational caveats.

Neuroinflammation and Cytokine Modulation

Ischemic stroke triggers the release of damage-associated molecular patterns (DAMPs) and activates microglia through Toll-like receptors (TLRs), NF-κB signaling, and downstream proinflammatory cytokines, including IL-1β, IL-6, and TNF-α. Conditional gene-editing platforms, such as NBS-CRISPR, which activates NF-κB, permit inflammation-restricted genome editing. Moreover, inhibition of MyD88 or proximal TLR signaling attenuates cytokine production in preclinical models (Lakhan et al. 2009; Simats et al. 2016; Bustamante et al. 2016).

In immunosuppressive environments, CRISPR-mediated activation of STAT3 has been shown to restore natural killer (NK) cell function, underscoring the context-dependent nature of immune modulation strategies. However, therapeutic modulation of innate immunity carries the risk of compromising host defense mechanisms; consequently, any anti-inflammatory intervention must be temporally controlled and reversible (Jin et al. 2018) (Fig. 5).

Fig. 5.

Schematic diagram of poststroke damaging pathways and potential intervention and potential CRISPR-based interventions. The illustration highlights key pathological processes such as ROS production, neuronal damage, and neuroinflammation, alongside targeted CRISPR strategies for neuroprotection. The figure was created using Microsoft PowerPoint. NBS-CRISPR, engineered CRISPR system for specific targeting; MYD88, Myeloid Differentiation Primary Response 88; ROS, Reactive Oxygen Species; STAT3, Signal Transducers and Activators of Transcription 3; PLA2g4A, Phospholipase A2 Group IV A; P65, RelA (an NF-κB subunit)

Oxidative Stress and Antioxidant Responses

Closely linked to neuroinflammation, oxidative stress also plays a central role in poststroke pathology and represents a critical focus for CRISPR-based interventions. Oxidative stress is a major driver of ischemic brain injury, where an imbalance between reactive oxygen species (ROS) production and clearance exacerbates neuronal damage. The Nrf2/Keap1 signaling axis serves as a master regulator of cellular antioxidant responses. CRISPR-Cas9 technology has been instrumental in functionally validating this pathway and identifying its upstream regulators. For example, Wang et al. (2024) used CRISPR-Cas9 to generate RNF13-knockout mice and demonstrated that RNF13 deficiency destabilizes the p62 protein, leading to impaired Nrf2/HO-1 signaling and exacerbated neuronal death after ischemic/reperfusion (I/R) injury (Wang et al. 2024a). This study genetically confirmed the neuroprotective role of the p62-mediated Nrf2 axis. Furthermore, the role of Nrf2/Keap1 as a key therapeutic target was underscored via the use of CRISPR-generated knockout models. Gao et al. (2020) employed Sirt3-KO mice created with CRISPR-Cas9 to demonstrate that the protective effect of the compound trilobatin against cerebral I/R injury is dependent on Sirt3, which concurrently modulates both the TLR4/NF-κB pathway and the Nrf2/Keap1 pathway (Gao et al. 2020). These studies exemplify how CRISPR is used not only to target the core antioxidant pathway directly but also to validate the specificity of pharmacological interventions that engage this axis. The goal of CRISPR-mediated modulation of pathways such as Nrf2/Keap1 and TLR4/NF-κB aligns with the broader therapeutic objective of other intervention strategies, including those employing natural compounds to rebalance the pro-oxidant/inflammatory state and enhance endogenous neuroprotection.

In addition to direct antioxidant pathway modulation, CRISPR/Cas9 has been employed to dissect the role of specific metabolic processes in oxidative stress. For example, glycogen metabolism in astrocytes critically influences postischemic neuroinflammation and oxidative balance. Knockout of the glycogen phosphorylase (GP) gene in astrocytes via CRISPR/Cas9 revealed that impaired glycogenolysis leads to increased ROS, reduced NADPH and glutathione production via the pentose phosphate pathway, and a shift toward a proinflammatory A1-like astrocyte phenotype. This study confirmed that glycogen breakdown normally suppresses the NF-κB pathway and activates the protective STAT3 pathway, highlighting astrocyte glycogen metabolism as a viable target for IS management (Guo et al. 2021).

Other key players in oxidative stress and apoptosis, such as caspase-3 and NOX4, have also been successfully targeted with CRISPR/Cas9 to mitigate poststroke brain injury (Katta et al. 2023).

CRISPR technology has also elucidated the role of mitophagy, the selective clearance of damaged mitochondria, in combating oxidative stress. Research using CRISPR/Cas9 to knockout tissue plasminogen activator (tPA) in neurons demonstrated that tPA deficiency exacerbates I/R injury by impairing FUNDC1-dependent mitophagy, leading to mitochondrial dysfunction and apoptosis. Conversely, tPA treatment enhances mitophagy, stabilizes mitochondria, and improves neuronal survival via AMPK/FUNDC1 signaling (Cai et al. 2021).

Additional CRISPR-based studies have identified novel targets within inflammatory signaling cascades that intersect with oxidative stress. For example, knockout of CRACR2A attenuated stroke-induced inflammation and injury by impairing neutrophil adhesion and migration (Lee et al. 2025). Conversely, knockout of Tollip, a negative regulator of TLR pathways, surprisingly confers protection against I/R injury by enhancing Akt signaling, revealing the complex context-dependent roles of innate immune modulators (Li et al. 2015).

Prevention of Apoptosis and Necroptosis

In addition to oxidative damage, programmed cell death mechanisms, such as apoptosis and necroptosis, represent key therapeutic targets to prevent secondary neuronal loss after IS. Apoptosis and necroptosis are central mechanisms of secondary neuronal loss, vascular inflammation, and impaired tissue regeneration following IS (Wang et al. 2024b). CRISPR-based interventions targeting these pathways have demonstrated robust neuroprotective effects. Transcriptional activation of SIRT1 via the CRISPR/dCas9-SunTag system in a mouse pMCAO model resulted in several-fold increases in Sirt1 mRNA and antiapoptotic Bcl2, concomitant with downregulation of proapoptotic Bax and PARP. These molecular changes are correlated with reduced brain edema and enhanced survival, all without permanent genomic modifications (Ryu et al. 2024). Similarly, CRISPR-Cas9–mediated BCL-2 overexpression in a tMCAO model suppressed caspase-3 activity and prevented neuronal loss, shifting the cellular balance toward survival (Liu et al. 2024).

Effective neuroprotection extends beyond single-gene modulation. Multiplex regulation via dCas9-engineered astrocytes simultaneously upregulated anti-inflammatory (IL-38) and antiapoptotic genes while repressing glial scar–associated genes (LZK, MST1), promoted VEGF-driven angiogenesis, attenuated inflammation, inhibited apoptosis, and facilitated neural stem cell migration (Melika 2024). Additionally, PGC-1α in microglia confers broad protection: its CRISPR-Cas9–mediated upregulation enhances mitochondrial function, activates ULK1-dependent autophagy and mitophagy, and suppresses NLRP3 inflammasome activity and proinflammatory cytokine release; conversely, PGC-1α deletion exacerbates poststroke injury (Han et al. 2021).

Necroptosis, driven by the RIPK1/RIPK3/MLKL axis, amplifies inflammatory neuronal death. CRISPR-CasRx–mediated knockdown of Ripk1 and NSF mRNAs delivered via AAV9 to neurons and glia in tMCAO models significantly reduced infarct volume, edema, and necrosis while improving neurological function. This RNA-targeting strategy offers high precision, tunability, and a favorable safety profile owing to its transient, nonintegrating nature, highlighting its promise for clinical translation (Song et al. 2024).

Blood–Brain Barrier (BBB) Integrity and Repair

In addition to preventing cell death, preserving BBB integrity is essential for limiting edema, inflammatory infiltration, and secondary damage. The importance of maintaining BBB integrity to improve neurological outcomes after IS is emphasized. Given advancements in genome editing, the use of CRISPR/Cas9 as an effective tool to target molecular pathways involved in BBB damage is proposed. The evidence presented below shows that targeted genetic interventions restore and maintain the integrity of this critical barrier and prevent secondary complications caused by barrier leakage. As demonstrated in a previous study, inhibition of Sema4D/PlexinB1 signaling through CRISPR/Cas9 gene editing of PlexinB1 led to a significant reduction in BBB permeability, improved neurological function, and decreased infarct volume after IS. Sema4D, which is increased in perivascular astrocytes after stroke, binds to PlexinB1 on pericytes, resulting in disruption of BBB integrity. This causes the release of proinflammatory factors and the expression of MMP-9, which degrades soluble proteins. Additionally, Sema4D induces an inflammatory CD11b-positive phenotype in pericytes and inhibits mRNA degradation by AUF1, thereby exacerbating inflammation (Zhou et al. 2018). In parallel with these findings, Xu and colleagues (2021) reported that deletion of the AIM2 gene via CRISPR/Cas9 improved BBB integrity and reduced ischemic brain injury in an experimental stroke model. In this study, AIM2⁻/⁻ mice subjected to MCAO had smaller infarct volumes, better neurological function, and reduced BBB damage. In these mice, soluble proteins such as zonula occludens-1 (ZO-1) and occludin, which play important roles in maintaining BBB integrity, were preserved, whereas neutrophil infiltration and ICAM-1 expression were significantly reduced. Mechanistically, AIM2 deletion led to decreased phosphorylation of STAT3, a central pathway involved in disruption of BBB integrity (Xu et al. 2021). The results of this study introduce the AIM2 gene as a therapeutic target in IS; thus, the use of gene deletion techniques with CRISPR is a suitable method for reducing BBB damage and improving clinical outcomes in stroke patients.

On the basis of this body of evidence, it has become clear that targeting genes involved in maintaining barrier structures is not limited to inflammatory signaling but also includes the repair and regeneration of structural proteins. In this context, more recent studies have shown that disruption of the tight junction protein claudin-5 (CLDN5) plays a key role in BBB leakage during IS (Winkler et al. 2021). Therefore, the use of CRISPR technology to restore CLDN5 expression is proposed to reduce vasogenic edema and improve vascular function. This approach not only reduces barrier leakage but also has the potential to restore vascular barrier function after ischemic injury (Xiang et al. 2025; Rust et al. 2025).

Cellular Reprogramming and Neuronal Replacement

Neural regeneration is hindered by glial scar formation, which arises from reactive astrogliosis and the disruption of cellular equilibrium following ischemic stroke. Consequently, alongside vital acute neuroprotection, long-term recovery necessitates regenerative approaches, such as the conversion of glial cells into neurons (Safina and Embree 2022; Cekanaviciute and Buckwalter 2016). Recent advances have focused on direct glia-to-neuron reprogramming as a regenerative strategy, bypassing the need for exogenous cell transplantation. This approach employs neurogenic transcription factors, such as NeuroD1, Ascl1, and Neurogenin2, to convert reactive astrocytes into functional neurons and thereby restore neural circuitry (Greșiță et al. 2025). Notably, AAV-mediated delivery of NeuroD1 in nonhuman primates with an IS increased neuronal density and preserved parvalbumin-positive interneurons in cortical areas, demonstrating translational potential (Ge et al. 2020).

CRISPR-Cas9 further refines this paradigm by enabling the targeted activation of neuronal genes or the silencing of astrocytic identity genes. For example, CRISPR-Cas9–mediated knockdown of the RNA-binding protein PTBP1 via specific single-guide RNAs (sgRNAs) has significantly improved in vivo neural reprogramming efficiency (Zhou et al. 2022; Ma et al. 2019). However, conversion rates remain inconsistent across brain regions and injury models, limiting the therapeutic scale of neuronal replacement.

Although conceptually compelling, the clinical translation of glia-to-neuron reprogramming faces substantial challenges. Key concerns include suboptimal in vivo conversion efficiency (Ge et al. 2020; Bocchi et al. 2022), uncertain long-term survival and functional integration of converted neurons into existing networks, and the fidelity of region- and neurotransmitter-specific phenotypic acquisition in the chronically injured brain. Additionally, incomplete or aberrant reprogramming may yield dysfunctional intermediates or unintended proliferative states, underscoring the need for fail-safe mechanisms (Rivetti Di Val Cervo et al. 2017). Realizing the therapeutic promise of this strategy will require advanced delivery platforms for spatial precision, lineage-tracing tools for fate mapping, and rigorous functional assessments in chronic IS models (Li et al. 2025; Peng et al. 2022).

However, critical questions regarding the long-term durability, safety, and functional synaptic integration of CRISPR-reprogrammed neurons remain largely unanswered in current preclinical studies. The field urgently requires longitudinal fate-mapping studies in higher-order stroke models to assess the stability of the converted phenotype and potential oncogenic risks. Furthermore, the functional efficacy of these new neurons within rebuilt neural circuits needs rigorous validation through advanced behavioral and electrophysiological assays. Future research directions should prioritize the development of controllable and reversible CRISPR systems combined with spatial‒temporal delivery platforms to mitigate risks and increase the precision of in vivo reprogramming for clinical translation.

Modulating Cellular Crosstalk, Organelle Transfer, and Synaptic Plasticity

Increasing Intercellular Support via Mitochondrial Transfer

Enhancing intercellular support mechanisms, particularly astrocyte-mediated transfer of functional organelles, such as mitochondria, to injured neurons after ischemic stroke, offers a complementary therapeutic approach. CRISPR enables specific modulation of key genes involved in this process. For example, CRISPR/Cas9-mediated upregulation of CD38, a calcium-dependent signaling molecule, in astrocytes facilitated cyclic ADP ribose-dependent mitochondrial release, increased neuronal ATP levels, and activated key survival pathways, including the phosphorylation of Akt (pAkt) and the upregulation of the antiapoptotic protein Bcl-xL. These molecular changes collectively enhance neuronal survival and improve outcomes both in in vitro oxygen‒glucose deprivation conditions and in vivo after stroke (Hayakawa et al. 2016). Conversely, CD38 suppression impaired mitochondrial transfer and recovery (Fig. 6).

Fig. 6.

CRISPR-mediated enhancement of astrocyte-to-neuron mitochondrial transfer to promote neuronal survival following ischemic stroke. This diagram depicts how CRISPR/Cas9 upregulation of CD38 in astrocytes facilitates the release and transfer of functional mitochondria (Mito) to stressed neurons, bolstering cellular bioenergetics (elevated adenosine triphosphate, ATP) and activating pro-survival signaling pathways such as pAkt and B-cell lymphoma-extra large (Bcl-xL). The figure was created using Microsoft PowerPoint. CRISPR, Clustered Regularly Interspaced Short Palindromic Repeats; Cas9, CRISPR-associated protein 9; CD38, cluster of differentiation 38; Mito, mitochondria; ATP, adenosine triphosphate; pAkt, phosphorylated protein kinase B; Bcl-xL, B-cell lymphoma-extra large

Nevertheless, the path from experimental success to clinical application is fraught with complexities. The therapeutic potential of enhancing mitochondrial transfer is tempered by practical and biological considerations. The efficiency of intercellular organelle delivery in vivo is difficult to quantify and likely represents a fraction of the total cellular need in a widespread ischemic lesion. Sustaining the metabolic function of transferred mitochondria in a hostile postischemic environment characterized by oxidative stress and calcium dysregulation remains a significant challenge (Meng et al. 2025). There is also a potential risk of transferring dysfunctional or heteroplasmic mitochondria, which could exacerbate neuronal metabolic stress rather than alleviate it (Bustamante-Barrientos et al. 2023). Moreover, the long-term fate and replicative capacity of donated mitochondria within postmitotic neurons are unknown, raising questions about the durability of this intervention (Tripathi and Ben-Shachar 2024). Therefore, despite being a powerful neuroprotective concept, clinical translation will require advances in controlling the quality, quantity, and specificity of mitochondrial delivery, coupled with long-term safety assessments.

Promoting Neuronal Regeneration and Synaptic Rewiring

Concurrent with cellular support, promoting endogenous repair and synaptic plasticity is essential for functional recovery. A central mediator is brain-derived neurotrophic factor (BDNF), whose levels decline after stroke, exacerbating neurodegeneration (Rhee and Shih 2021). CRISPR-based strategies to upregulate BDNF enhance neuronal survival and synaptic plasticity by activating key pathways such as the MAPK/ERK and PI3K/Akt pathways, leading to synaptic reorganization and motor function restoration (Toader et al. 2025). Similarly, the modulation of microRNAs (miRNAs) via CRISPR fine-tunes poststroke repair (Bindal et al. 2025).

In addition to promoting neuronal survival and angiogenesis, a paramount goal of poststroke therapy is the restoration of functional neural circuitry through synaptic plasticity and the dynamic strengthening, weakening, and formation of synaptic connections. Disruptions in synaptic plasticity are a hallmark of neurological deficits following stroke (Singh et al. 2024). Therefore, CRISPR-based strategies that modulate genes such as BDNF or miR-195 hold promise not only as neuroprotectants but also as enablers of synaptic reorganization. By creating a molecular environment conducive to dendritic growth, spinogenesis, and long-term potentiation, these interventions aim to facilitate the inherent capacity of the brain for functional rewiring, which is essential for meaningful motor and cognitive recovery.

Targeting Non-coding RNAs and RNA Methylation via CRISPR for Neuroprotection in Ischemic Stroke

Epigenetic and posttranscriptional regulation by noncoding RNAs (ncRNAs) and RNA methylation offers another layer of precision intervention in IS pathophysiology. ncRNAs, particularly long noncoding RNAs (lncRNAs), and RNA modifications perform significant regulatory functions in the pathophysiology of neurological diseases, including stroke (Si et al. 2020; Mehta et al. 2015). These elements, although lacking protein-coding capacity, serve as key modulators of inflammatory processes, apoptosis, oxidative stress, and cellular repair mechanisms. The advent of CRISPR/Cas9 technology has enabled the manipulation of these targets to elucidate their functions and therapeutic potential.

Targeting lncRNAs

FosDT is a brain-specific lncRNA that is rapidly upregulated following IS and contributes significantly to poststroke brain injury (Mehta et al. 2015). To investigate its functional role, a research group employed CRISPR-Cas9 to generate FosDT knockout rats. Compared with wild-type controls, these animals exhibited normal growth, development, and brain structure but demonstrated markedly improved sensorimotor function and reduced cerebral damage after transient IS. Transcriptomic analysis revealed that FosDT deletion led to the downregulation of genes associated with neuroinflammation, apoptosis, mitochondrial dysfunction, and oxidative stress, underscoring its pleiotropic role in postischemic pathology (Mehta et al. 2021). This work identified lncRNAs such as FosDT as promising therapeutic targets. Complete functional silencing of noncoding genes remains challenging, as small indels do not always abolish lncRNA activity; complementary approaches, including CRISPR–Cas9–mediated large deletions through homologous recombination, are necessary for effective ablation.

CRISPR-Display for lncRNA Targeting and Functional Analysis

To specifically dissect lncRNA function, innovative tools such as CRISPR-Display have been developed (Shechner et al. 2015). This method utilizes a catalytic dCas9 protein fused to gRNAs that incorporate additional RNA segments. These segments can harbor functional RNA domains or entire lncRNA sequences, allowing their targeted recruitment to specific genomic loci. CRISP-Disp thus provides a unique platform to study the cis- and trans-regulatory roles of lncRNAs in neurological diseases by enabling locus-specific manipulation without permanent genomic alteration, offering insights into their mechanisms in conditions such as cerebral ischemia.

Modifying m⁶A RNA Methylation via CRISPR

N6-methyladenosine (m⁶A) is a prevalent RNA modification that influences mRNA stability, processing, and translation and plays a crucial role in cellular stress responses. Methyltransferase-like 3 (METTL3) is a central writer enzyme involved in m⁶A deposition. In the context of IS, research has identified a protective axis involving METTL3, the microRNA miR-335, and the transcription factor ELK-related factor 1 (Erf1). Under ischemic conditions, METTL3-mediated m⁶A methylation promotes the processing of pri-miR-335 to its mature form. miR-335, in turn, targets and downregulates Erf1 mRNA, a process that facilitates the formation of stress granules, cytoprotective assemblies that help neurons manage translational arrest during acute stress (Si et al. 2020; Wang et al. 2020; Lu et al. 2003; Xu et al. 2024) CRISPR-Cas9 knockout of METTL3 in neuronal PC12 cells subjected to oxygen‒glucose deprivation/reperfusion (OGD/R) disrupted this axis, reducing m⁶A levels on pri-miR-335, diminishing mature miR-335 production, and impairing stress granule formation. Consequently, these cells exhibited increased susceptibility to death. Re-expression of METTL3 rescued these phenotypes, confirming the role of this pathway in neuronal survival (Si et al. 2020). This example provides a clear mechanistic link between CRISPR-mediated editing of an RNA methylation enzyme, the regulation of a specific ncRNA (miR-335), and a functional outcome (stress granule formation) directly tied to managing oxidative and proteotoxic stress postischemia.

Targeting Regulatory Genetic Variants in Non-coding Regions

In addition to canonical ncRNAs, noncoding regulatory elements are also amenable to CRISPR intervention. For example, a genetic variant (rs17114036) located in a flow-sensitive enhancer within intron 5 of the PLPP3 gene has been associated with protection against atherosclerosis. Using CRISPR-Cas9 to delete this 66-bp enhancer region in human aortic endothelial cells resulted in reduced PLPP3 expression and heightened inflammatory responses. Complementary CRISPR interference (CRISPRi) silencing of this enhancer further established its causal role in regulating PLPP3, illustrating how CRISPR tools can dissect the function of noncoding regulatory variants in vascular pathophysiology relevant to stroke risk (Krause et al. 2018). Collectively, these studies highlight the potential of CRISPR for mechanistic dissection. However, this precision reveals a critical challenge: neuroprotective efficacy is often pathway, and context specific, limiting the generalizability of individual findings. For example, while strategies enhancing antioxidant responses (e.g., via Nrf2) or inhibiting specific inflammatory nodes show promise, their net benefit may be counteracted in the multifaceted pathology of human stroke, where compensatory pathways are activated. This mechanistic siloing, compounded by the predominant use of homogeneous, young rodent models lacking prevalent comorbidities such as diabetes or sustained hypertension, creates a formidable efficacy gap between controlled preclinical settings and the clinical reality of stroke patients (Li et al. 2025; Bustamante-Barrientos et al. 2023).

Collectively, these strategies converge on a modular, targetable network of ischemic injury pathways. An integrated overview is provided in Fig. 7, illustrating how CRISPR platforms can be deployed to simultaneously counteract neuroinflammation, oxidative stress, cell death, and impaired regeneration.

Fig. 7.

CRISPR-based mechanisms target pathogenic pathways activated after ischemic stroke to improve neuronal function and facilitate brain tissue regeneration. CRISPR systems such as Cas9 and CasRx enable modulation of injury pathways, including neuroinflammation (via TLR4/NF-κB and JAK-STAT3 signaling), oxidative stress (via Nrf2/Keap1), apoptosis/necroptosis, and excitotoxicity. By intervening at these molecular nodes, CRISPR strategies aim to enhance neuronal survival, promote neuroprotection, and support structural and functional recovery of the ischemic brain. The figure was created using Microsoft PowerPoint. CRISPR: Clustered Regularly Interspaced Short Palindromic Repeats. Cas9: CRISPR-associated protein 9; JAK-STAT3: Janus Kinas-Signal Transducer and Activator of Transcription 3; Nrf2/keap1: Nuclear factor erythroid 2-related factor 2/Kelch-like ECH-associated protein 1; TLR4/NF-kB: Toll-Like Receptor 4/Nuclear Factor kappa-light-chain-enhancer of activated B cells

Technical Considerations and Comparative Advantages of RNA-Targeting CRISPR Systems

Following the discussion of therapeutic applications, this section presents a technical and comparative analysis of RNA-targeting and epigenome-modifying CRISPR tools. The focus is on the unique mechanistic attributes that differentiate systems such as Cas13 and dCas9 from conventional DNA-editing Cas9, with direct implications for their strategic application in IS therapy.

Mechanistic Principles: DNA vs RNA Targeting

The fundamental distinction lies in the target molecule. CRISPR-Cas9 induces double-strand breaks in DNA, leading to permanent alterations via nonhomologous end joining or homology-directed repair (Bonowicz et al. 2025). In contrast, CRISPR-Cas13 (e.g., CasRx) and CRISPR-dCas9-based systems (e.g., CRISPRi/a) operate without permanent DNA damage (Li et al. 2023). Cas13 directly cleaves target mRNA, causing transient knockdown, which is subject to cellular RNA turnover rates. dCas9, when fused to effector domains such as KRAB (for CRISPRi), modulates gene expression epigenetically by altering chromatin states or recruiting transcriptional machinery, effects that are often reversible upon cessation of expression (Zhou et al. 2022; Yang and Patel 2024).

Pharmacokinetic and Pharmacodynamic Implications

This mechanistic difference translates to distinct pharmacokinetic/pharmacodynamic profiles critical for clinical translation. RNA-targeting and epigenetic systems offer a self-limiting therapeutic window. Their activity is constrained by the half-life of the Cas protein/gRNA complex and the target RNA itself, reducing the risk of long-term overdosing or persistent off-target effects. This is particularly advantageous for modulating acute, transient pathological processes in stroke, such as the early inflammatory burst or necroptotic signaling (Song et al. 2024). Conversely, DNA-editing strategies such as Cas9 are better suited for correcting stable genetic defects or enabling long-term expression of therapeutic genes, bearing a different risk‒benefit calculus involving potential genotoxicity (Krause et al. 2018; Roy et al. 2022; Guo et al. 2023).

Design and Specificity Considerations

Achieving high specificity requires different strategies for each system. For DNA-editing Cas9, off-target effects arise from sgRNA tolerance to DNA mismatches; mitigation strategies include the use of high-fidelity Cas9 variants or paired nickases (Roy et al. 2022; Guo et al. 2023). For RNA-targeting Cas13, specificity challenges relate to collateral RNAse activity and seed-sequence interactions; engineered high-fidelity variants address this (Chen et al. 2025; Han et al. 2017). For CRISPRi/a, off-target effects stem from dCas9 binding specificity and the pleiotropic effects of epigenetic modifiers; optimal sgRNA design to avoid regulatory regions is key (Duke et al. 2020; Gilbert et al. 2014).

Integration with Therapeutic Goals: A Decision Framework

The choice among these systems should be guided by the specific therapeutic objective. For rapid, acute neuroprotection, such as silencing Ripk1 postischemia, Cas13 systems are ideal because of rapid mRNA degradation and transient action (Song et al. 2024; Tian et al. 2025). For sustained but reversible modulation, CRISPRi offers tunable, durable yet potentially reversible repression (Bendixen et al. 2023). For permanent correction or sustained expression, such as the repair of a mitochondrial mutation or continuous BDNF production, CRISPR-Cas9 (with HDR) or CRISPRa may be needed, with acknowledgment of the need for long-term safety monitoring (Bonowicz et al. 2025; Toader et al. 2025).

Figure 8 summarizes these RNA/epigenome-targeting tools and their therapeutic contexts.

Fig. 8.

Comparative overview of RNA-targeting and epigenome-modifying CRISPR systems for neuroprotection in ischemic stroke. The diagram illustrates three primary CRISPR-based platforms for modulating gene expression without altering genomic DNA sequence, highlighting their distinct mechanisms, advantages, and therapeutic contexts. The figure was created using Microsoft PowerPoint. IL-6: Interleukin-6; BAX: Bcl2-associated X protein; hfCas13x: high-fidelity/CRISPR-associated protein 13; cx43: Connexin 43; kRAB: Kruppel-associated box; IL-38: Interleukin-38; BRAG-1: Brain-Related Guanine Nucleotide Exchange Factor 1; MST-I: Mammalian Sterile 20-like kinase 1; LZK: Leucin zipper-Bearing kinase

The therapeutic potential of CRISPR in ischemic stroke can be further contextualized through an integrative framework that connects molecular targets to functional outcomes. As illustrated in Fig. 9, CRISPR systems simultaneously modulate multiple interconnected pathological pathways. This coordinated approach addresses neuroinflammation, oxidative stress, programmed cell death, and BBB disruption more comprehensively than single-pathway interventions do, leading to synergistic neuroprotection.

Fig. 9.

Integrative framework linking CRISPR-targeted pathways to functional recovery in ischemic stroke. This figure synthesizes how CRISPR systems modulate key pathological processes and maps these interventions to corresponding functional outcomes, including neuronal survival, reduced infarct volume, preserved BBB integrity, and improved neurological function. The figure was created using Microsoft PowerPoint. BBB: Blood–Brain Barrier; CRISPR: Clustered Regularly Interspaced Short Palindromic Repeats; Cas9: CRISPR-associated protein 9; CasRx: CRISPR-associated RfxCas13d; ROS: Reactive Oxygen Species; TLR4: Toll-Like Receptor 4; NF-κB: Nuclear Factor kappa-light-chain-enhancer of activated B cells; Nrf2: Nuclear factor erythroid 2–related factor 2; Keap1: Kelch-like ECH-associated protein 1; JAK: Janus Kinase; STAT3: Signal Transducer and Activator of Transcription 3; RIPK1: Receptor-Interacting Protein Kinase 1; MLKL: Mixed Lineage Kinase Domain-Like Protein; BDNF: Brain-Derived Neurotrophic Factor; VEGF: Vascular Endothelial Growth Factor; MMP-9: Matrix Metallopeptidase 9; IL-1β: Interleukin-1 beta; TNF-α: Tumor Necrosis Factor-alpha; SIRT1: Sirtuin 1

Safety Considerations, Translational Barriers, and Mitigation Strategies

The clinical translation of CRISPR-based therapies for stroke demands a rigorous, multifaceted approach to safety, extending beyond foundational concerns to address specific translational hurdles. Paramount among these is the risk of off-target editing. For DNA-editing systems such as CRISPR-Cas9, unintended cleavages at genomic sites with sequence homology to the sgRNA can lead to deleterious mutations. Mitigation strategies are evolving and include the use of engineered high-fidelity Cas9 variants with enhanced specificity, paired nickases that generate staggered cuts to reduce off-target effects, and sophisticated bioinformatic algorithms for optimal sgRNA design to minimize homology with nontarget sites (Roy et al. 2022; Guo et al. 2023). For RNA-targeting systems (e.g., Cas13 and CasRx), off-target binding to untranslated RNAs remains a concern, although their transient activity and the absence of permanent genomic alterations offer a distinct safety advantage for applications requiring reversible gene modulation (Gilbert et al. 2014; Abudayyeh et al. 2017).

Immunogenicity presents a second major barrier. Immune reactions can be directed against both the bacterial-derived caspase proteins and the viral capsids used in many delivery systems, potentially reducing therapeutic efficacy and causing adverse inflammatory responses. Preclinical immunogenicity profiling in relevant models is therefore critical. The choice of delivery vector itself is a key safety determinant; nonintegrating vectors (e.g., adeno-associated viruses, AAVs) or transient nonviral carriers pose a lower risk of insertional mutagenesis than do integrated lentiviral vectors, although the latter can be engineered to be integration deficient (Skukan et al. 2022).

This preclinical optimism is tempered by fundamental translational gaps. Many studies report near-complete neuroprotection in rodents, a magnitude of effect rarely translatable to humans and potentially indicative of publication bias toward positive outcomes. The leap from controlled, short-term models to chronic stroke recovery in a neuroanatomically complex human brain remains monumental. Successful translation will require validation in higher-order primates and models incorporating aging, atherosclerosis, and diabetes to better approximate the clinical reality (Li et al. 2025; Bustamante-Barrientos et al. 2023).

Further translational challenges include ensuring targeted biodistribution to the central nervous system while minimizing exposure to peripheral organs (Wilbie et al. 2019; Lino et al. 2018; Wang et al. 2023). To address these barriers systematically and derisk the path to the clinic, we recommend the implementation of standardized preclinical test batteries. These should encompass utilizing cell type-specific panels and transcriptome-wide screens (e.g., CIRCLE-seq and GUIDE-seq for DNA; RIP-seq for RNA) to identify and quantify unintended editing events. Evaluating innate and adaptive immune responses to CRISPR components and delivery vehicles in immunocompetent animal models. The use of sensitive tracking methods in large-animal models to define pharmacokinetics and tissue tropism. Proactive dialog with regulatory agencies to align preclinical safety packages with expectations for first-in-human trials. Specifically, for nonviral platforms such as EVs and LNPs, key hurdles include batch-to-batch variability, controlledloading efficiency of CRISPR cargo, and achieving targeted delivery to specific brain cell types beyond passive BBB crossing. The standardization of production and rigorous characterization are paramount for clinical translation. Additionally, the ethical and regulatory landscape requires careful navigation, especially for strategies involving permanent genomic edits. A clear distinction must be maintained between somatic cell editing (the therapeutic goal) and germline modifications. Transparent informed consent processes, equitable access considerations, and robust, phased clinical trial designs are imperative to ensure the responsible development of these transformative therapies.

Conclusion

This narrative review synthesizes the rapidly evolving landscape of CRISPR-based gene editing as a transformative strategy against IS. The evidence compiled herein demonstrates that CRISPR technology, through systems such as Cas9 and CasRx, enables unprecedented precision in targeting the molecular underpinnings of stroke injury. Its applications span critical pathways, effectively modulating neuroinflammation, counteracting oxidative stress, inhibiting programmed cell death, preserving BBB integrity, and even promoting neural regeneration through cellular reprogramming. These multifaceted interventions, facilitated by advances in delivery vectors such as adeno-associated viruses and engineered nanoparticles, have consistently shown promise in improving structural and functional outcomes in preclinical models, positioning CRISPR as a cornerstone of next-generation neurotherapeutics.

The integration of CRISPR with broader therapeutic paradigms may unlock further potential. In addition to standalone genetic interventions, emerging research highlights the importance of supporting cellular homeostasis and metabolic resilience for optimal recovery. For example, nutraceuticals and lifestyle modifications that modulate autophagy, mitigate oxidative stress, and enhance neuroplasticity could act synergistically with precision gene-editing strategies. This combined approach, which combines targeted genomic modulation with systemic support for cellular health, may offer a more holistic and robust framework for addressing both the acute and chronic sequelae of IS. Nevertheless, this promising trajectory is balanced by substantial translational challenges. The field must move beyond proof-of-concept studies in idealized models and rigorously address inconsistent reproducibility, the long-term safety of genomic edits, and the development of robust biomarkers to monitor efficacy and off-target effects in humans. Future research prioritizing these critical gaps will determine whether CRISPR-based strategies can evolve from potent preclinical tools into viable clinical neurotherapeutics.

A comparative overview of key CRISPR-based approaches alongside conventional therapies is presented in Table 1. This table synthesizes critical parameters, including therapeutic mechanisms, delivery systems, model outcomes, advantages, and limitations, highlighting the distinct value proposition of gene-editing strategies.

Table 1.

Comparative overview of CRISPR-based and traditional therapies in ischemic stroke

Cat	Therapy/approach	Target/pathway	Delivery	Model system	Outcomes	Advantages	Limitations	Ethical considerations	References
Gene Editing	CasRx (targeting Ripk1/Nsf)	Modulates necroptosis via RIPK1/RIPK3/MLKL pathway	AAV9	Rodent (tMCAO), Primate	↓ Infarct volume (~ 14%); ↑ sensorimotor function	RNA-specific; durable	Delivery optimization; immunogenicity	Consent for permanent edit	Song et al. (2024)
	dCas9 (SIRT1 activation)	SIRT1- mediated neuroprotection	Intranasal NP	Mouse (pMCAO)	↓ Edema, ↑ survival (safe)	Non-viral; BBB bypass	Long-term safety unknown	Off-target risk	Ryu et al. (2024)
	NeuroD1-mediated astrocyte-to-neuron conversion	Neuronal regeneration	AAV	Rhesus Macaque	~ 90% conversion; tissue preserve	Regenerative	Scaling to human brain untested	Cellular reprogramming ethics	Ge et al. (2020)
Conventional Therapies	tPA (thrombolysis)	Clot lysis	i.v	Clinical	Effective if ≤ 4.5 h	Fast reperfusion	Narrow window; hemorrhage risk	Consent under pressure	Paul and Candelario-Jalil (2021)
	Neuroprotectants	↓Excitotoxicity & inflammation	Oral/intravenous	Preclinical & clinical trials	Limited human efficacy	Multi-target potential	Limited clinical applicability	Risk–benefit uncertainty	Paul and Candelario-Jalil (2021)
	MSCs	Paracrine signaling; anti-inflammatory	i.v./Local	Animal models, Phase I/II trials	Neurological improvement	Immunomodulatory; pro-repair	Limited human evidence	Stem-cell oversight	Do et al. (2021)
Delivery	AAV9	CNS gene delivery	Viral (i.v.)	Rodent/primate	Sustained expression	High CNS tropism	Immunogenicity; cargo limit	Viral vector regulation	Skukan et al. (2022); Wang et al. (2023)
	dCas9/CaP/PEI-PEG-bHb NPs	BBB bypass for gene modulation	Intranasal	Mouse (pMCAO)	Upregulated Sirt1; ↓ edema; ↑ survival	Biocompatible; avoids viral risks	Scalability/stability issues	Long-term safety monitoring	Ryu et al. (2024)
	LNPs	Transient mRNA/editor delivery	Systemic	Experimental	Short-lived; ↓ off-target	Safe, transien; tunable	Low brain penetration without targeting ligands	Germ-line editing concern	Salman et al. (2022)
Genetic Targets	Ripk1	Necroptosis master regulator	AAV9	Rodent	↓ infarct & edema	Central to cell death pathway	Possible off-target cleavage	Precision medicine ethics	Song et al. (2024)
	Nsf	Vesicle trafficking /synaptic function	AAV9	Rodent	↑ Neuro recovery	Synergy with Ripk1	Dual-targeting complexity	Consent for combo engineering	Song et al. (2024)
	Sirt1	Anti-apoptotic/anti-inflammatory	NPs	Mouse	↑ Survival; ↓ apoptosis	Broad neuroprotection	Expression duration unclear	Access equity	Ryu et al. (2024)
	PCSK9 (theoretical)	LDL-C lowering (preventive)	–	–	May reduce recurrent risk	Preventive strategy	Needs validation in stroke models	Cost/access barriers	Bonowicz et al. (2025)
Emerging	Engineered EVs	CRISPR cargo delivery	Local/ i.v	Preclinical	Rapid, reversible modulation potential	Low immunogenicity; natural carrier	Feasibility during thrombectomy unclear	EV biologic regulation	Li et al. (2023); Wang et al. (2024a); Chen et al. (2025)
	Magnetic EVs + CRISPR	Magnetically-guided delivery	Local	Conceptual	Site-specific targeting	Enhanced spatial precision	Early-stage development	Long-term biosafety unknown	Li et al. (2023); Huang et al. (2021); Wang et al. (2024a)
	iPSC-Neurons + CRISPR	Regeneration and circuit repair	Transplant	Preclinical	Functional recovery in rodent models	One-shot durable effect	Tumorigenicity risk; scalability	Dual oversight (stem cell + gene edit)	Song et al. (2024); Ge et al. (2020)
Long-Term Outcomes	CasRx (Ripk1/Nsf)	Sustained neuroprotection	AAV9	Rodent	Behavioral benefit (25 days)	One-shot durable effect	Long-term safety data limited	edit dilemma	Song et al. (2024)
	NeuroD1-mediated AtN conversion	Neuronal replacement & integration	AAV	Primate	↑ Neuron density (1 year preserv.)	Regenerative potential	Human translation needed	Circuit rewiring ethics	Greșiță et al. (2025); Ge et al. (2020)
Societal Aspects	Access & cost	–	–	–	High cost may limit availability	Technological advancement	Need for equitable policies	Need for equitable health policy	Bonowicz et al. (2025); Melika (2024)
	Public Perception	–	–	–	Hope vs. fear of designer therapies	Engages public in ethics	Misinformation/skepticism	Autonomy vs. safety balance	Bonowicz et al. (2025); Melika (2024)
	Regulatory Framework	–	–	–	Standards evolving rapidly	Ensures safety	Lengthy, costly approval	Harmonization across countries	Bonowicz et al. (2025); Skukan et al. (2022); Salman et al. (2022)

CRISPR: Clustered Regularly Interspaced Short Palindromic Repeats, CasRx: CRISPR-associated protein RfxCas13d, RIPK1: Receptor-Interacting Protein Kinase 1, RIPK3: Receptor-Interacting Protein Kinase 3, MLKL: Mixed Lineage Kinase Domain-Like Protein, AAV9: Adeno-Associated Virus serotype 9, tMCAO: transient Middle Cerebral Artery Occlusion, i.v: Intravenous, pMCAO: permanent Middle Cerebral Artery Occlusion, dCas9: Dead Cas9 (catalytically inactive Cas9 protein), SIRT1: Sirtuin 1 (NAD-dependent deacetylase sirtuin-1), CaP: Calcium Phosphate, PEI-PEG-bHb: Polyethylenimine-Polyethylene Glycol-Bovine Hemoglobin, BBB: Blood‒Brain Barrier, MCT1: Monocarboxylate Transporter 1, NeuroD1: Neurogenic Differentiation Factor 1, AtN: Astrocyte-to-Neuron, MSCs: Mesenchymal Stem Cells, tPA: Tissue Plasminogen Activator, LNPs: Lipid Nanoparticles, PCSK9: Proprotein Convertase Subtilisin/Kexin Type 9, LDL: Low-Density Lipoprotein, EVs: Extracellular Vesicles, iPSC: Induced Pluripotent Stem Cell

Author Contributions

F.A. conceived the original idea and drafted the initial version of the manuscript. S.GH. designed the figures and was responsible for the overall formatting. Both authors collaborated closely throughout the revision process to refine and finalize the paper.

Funding

This research was conducted without any external financial support. All associated costs were borne independently by the authors.

Data Availability

No datasets were generated or analysed during the current study.

Declarations

Conflict of interest

The authors affirm that they have no competing interests and that the study adheres.