X inactive specific transcript drives mitochondrial metabolic rewiring and neural stem cell fate after spinal cord injury

Abstract

Spinal cord injury triggers irreversible functional loss and metabolic collapse, where mitochondrial dysfunction creates a critical bottleneck for regenerative therapies. Zeng et al recently published a study in World Journal of Stem Cells identify the long noncoding RNA X-inactive specific transcript (XIST) as a master regulator of mitochondrial metabolism and neural stem cell fate through the insulin-like growth factor 2 mRNA-binding protein 2/carnitine palmitoyl transferase 1A axis. By stabilizing carnitine palmitoyl transferase 1A mRNA via insulin-like growth factor 2 mRNA-binding protein 2, XIST enhances fatty acid oxidation and oxidative phosphorylation, thereby driving neuronal differentiation while curbing reactive astrogliosis in murine models. Beyond metabolic control, XIST interfaces with chromatin remodeling and inflammatory signaling, although its multifaceted roles are highly context-dependent. Despite therapeutic promise, challenges regarding lineage-specific effects and the need for longitudinal in vivo validation persist. Future strategies integrating spatial transcriptomics and advanced delivery systems are essential to translate XIST-mediated mechanisms into clinical spinal cord injury repair.

Key Words: Carnitine palmitoyltransferase 1A; Insulin like growth factor 2 messenger RNA binding protein 2; Long noncoding RNA; Mitochondrial metabolism; Neural stem cells; Regenerative medicine; Spinal cord injury; X inactive specific transcript

Core Tip: This study highlights the pivotal role of the long noncoding RNA X inactive specific transcript as a metabolic-epigenetic regulator in spinal cord injury. By stabilizing carnitine palmitoyltransferase 1A messenger RNA through insulin like growth factor 2 messenger RNA binding protein 2, it enhances fatty acid oxidation and mitochondrial oxidative phosphorylation, promoting neural stem cell differentiation and functional recovery. The proposed framework integrates mitochondrial metabolism, lineage regulation, and regenerative strategies, providing a conceptual bridge from molecular mechanisms to translational medicine.

This editorial refers to “X inactive-specific transcript regulates mitochondrial function and neuronal differentiation of stem cells via IGF2BP2/CPT1A axis in models of spinal cord injury” by Zeng et al, 2025; https://doi.org/10.4252/wjsc.v17.i7.101929.

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INTRODUCTION

Recent studies have clearly identified X-inactive specific transcript (XIST) as a pivotal molecular “switch” that governs mitochondrial metabolism in neural stem cells (NSCs)[1,2]. Zeng et al[1] published a study in the recent issue of World Journal of Stem Cells demonstrated that XIST interacts with insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) to stabilize the mRNA of carnitine palmitoyl transferase 1A (Cpt1a), thereby markedly enhancing fatty acid oxidation (FAO) and mitochondrial oxidative phosphorylation (OXPHOS). This metabolic enhancement elevates ATP production and the oxygen consumption rate (OCR), promoting neuronal differentiation of NSCs while suppressing reactive astrogliosis. In mouse models of spinal cord injury (SCI), activation of this pathway coincides with reduced inflammation and improved locomotor recovery, whereas interference with IGF2BP2 or knockdown of Cpt1a reverses these effects - establishing the causal link and therapeutic potential of the XIST-IGF2BP2-CPT1A axis in energy metabolism and neural repair[1].

From a clinical standpoint, recovery of urinary, motor, and psychological functions in SCI patients critically depends on systematic rehabilitative care, underscoring that functional reconstruction requires not only neuronal regeneration but also metabolic support - consistent with the metabolic reprogramming paradigm centered on the XIST axis[3]. Moreover, independent studies have confirmed that IGF2BP2 directly binds and stabilizes multiple mitochondrial-related transcripts, including CPT1A and NADH dehydrogenase (ubiquinone) 1 alpha subcomplex subunit 2, further reinforcing the molecular validity and generalizability of this regulatory pathway[1,3,4].

This mechanistic framework closely mirrors the mitochondrial pathological cascade that spans from the acute to chronic phases of SCI. The accumulation of reactive oxygen species (ROS), abnormal opening of the mitochondrial permeability transition pore, ATP depletion, and calcium imbalance together constitute a self-reinforcing loop of “energy failure-oxidative stress-cell death”. Multiple studies have indicated that initiating mitochondrial protection or antioxidant interventions within approximately 8 hours post-injury may represent a critical therapeutic window to avert the ensuing energy crisis[5].

Meanwhile, although NSC transplantation holds promise for reconstructing neural circuits and modulating immune responses, its reparative efficacy remains heavily contingent upon the energy supply and inflammatory status of the injured tissue. The heterogeneous outcomes reported across systematic reviews and translational studies further underscore this dependency, suggesting that metabolic reprogramming combined with cell-based therapy is emerging as a promising future direction[6,7].

At the molecular level, long noncoding RNAs (lncRNAs) serve as central hubs that integrate epigenetic, transcriptional, and metabolic signaling. XIST can reshape stem cell fate through multiple mechanisms, including chromatin remodeling, X-chromosome inactivation, RNA stability regulation, and microRNA sponging. In particular, IGF2BP2, functioning as an N6-methyladenosine (m⁶A) “reader”, broadly enhances the stability and translational efficiency of metabolism-related transcripts, thereby providing a molecular scaffold that bridges XIST-mediated epigenetic regulation with metabolic control. A growing body of evidence has demonstrated that the lncRNA-IGF2BP regulatory network plays a pivotal role in energy metabolism, lineage commitment, and cellular homeostasis[7-9].

Accordingly, a metabolic guidance strategy based on the XIST-IGF2BP2-CPT1A axis, by enhancing FAO and OXPHOS activity and restoring energy homeostasis, may optimize stem cell function both before and after NSC transplantation. Moreover, combining this approach with peroxisome proliferator-activated receptor-γ coactivator-1α-mediated mitochondrial biogenesis or antioxidant regimens could further amplify its reparative and regenerative benefits[10] (Table 1).

Table 1 Multidimensional regulatory landscape of X-inactive specific transcript in spinal cord injury: From mitochondrial metabolism to epigenetic and immune control.

Regulatory dimension	Key molecular partners/targets	Core mechanistic action	Functional impact and SCI outcome	Ref.
Epigenetic and chromatin	SAF-A/HNRNPU, PRC1/2, MED14	XIST coating and chromatin anchoring; recruitment of polycomb complexes; suppression of specific enhancers	Maintains XCI integrity and lineage stability in stem cells	Kolpa et al[14], 2016; Bousard et al[15], 2019; Richart et al[16], 2022
Post-transcriptional and metabolic (core axis)	IGF2BP2, CPT1A, NDUFA2	XIST recruits IGF2BP2 (m6A reader) to stabilize CPT1A and NDUFA2 mRNAs; markedly enhances FAO and OXPHOS	Promotes neuronal differentiation of NSCs; increases ATP production and OCR; improves locomotor recovery and reduces neuroinflammation	Zeng et al[1], 2025; Huang et al[4], 2022; Wang et al[11], 2021; Weng et al[17], 2022
Immune-metabolic and ceRNA	miR-124-3p, IRF1	XIST acts as a ceRNA sponge for miR-124-3p, modulating the Irf1 pathway and microglial polarization	Regulates M1/M2 macrophage balance; exerts context-dependent effects on the inflammatory microenvironment	Yang et al[18], 2023
Systemic and intercellular	miR-539-3p, ADAMTS5; MSC-derived exosomes	XIST modulates hepatic mitochondrial injury via Adamts5 axis; XIST-containing EVs facilitate stem-immune crosstalk	Sustains cross-organ energy homeostasis; drives pro-regenerative M2 phenotype through exosomal shuttling of regulatory RNAs	Wu et al[2], 2023; Karpenko[19], 2025; Phinney et al[20], 2015; Arabpour et al[21], 2021

SCI: Spinal cord injury; SAF-A: Scaffold attachment factor A; HNRNPU: Heterogeneous nuclear ribonucleoprotein U; PRC1: Polycom repressive complex 1; PRC2: Polycom repressive complex 2; MED14: Mediator complex subunit 14; XIST: X-inactive specific transcript; XCI: X-chromosome inactivation; IGF2BP2: Insulin-like growth factor 2 mRNA-binding protein 2; CPT1A: Carnitine palmitoyl transferase 1A; NDUFA2: NADH ubiquinone oxidoreductase subunit A2; m⁶A: N6-methyladenosine; FAO: Fatty acid oxidation; OXPHOS: Oxidative phosphorylation; NSCs: Neural stem cells; ATP: Adenosine triphosphate; OCR: Oxygen consumption rate; IRF1: Interferon regulatory factor 1; ceRNA: Competing endogenous RNA; ADAMTS5: A disintegrin and metalloproteinase with thrombospondin motifs 5; MSCs: Mesenchymal stromal/stem cells; EVs: Extracellular vesicles; M1: Classically activated (pro-inflammatory) macrophage phenotype; M2: Alternatively activated (anti-inflammatory/pro-regenerative) macrophage phenotype.

THE XIST-IGF2BP2-CPT1A AXIS: A “METABOLISM-TO-FATE” SWITCH IN NSCS

The latest in vivo and in vitro evidence demonstrates that XIST, by recruiting or depending on the m⁶A reader protein IGF2BP2, selectively stabilizes CPT1A transcripts, thereby enhancing FAO and coupling it with mitochondrial OXPHOS to increase ATP production and OCR. This metabolic gain translates into enhanced neuronal lineage differentiation and suppressed astrocytic differentiation of NSCs, accompanied by reduced inflammation and improved locomotor recovery in mouse models of SCI. Disruption of Igf2bp2 or knockdown of Cpt1a reverses these effects, indicating a clear causal chain and the therapeutic feasibility of targeting this axis[1].

IGF2BP2: The post-transcriptional “molecular scaffold” of XIST

IGF2BP2 possesses dual functionality as both an RNA-binding protein and an m⁶A “reader”, capable of recognizing m⁶A-modified sites and enhancing the stability, intracellular localization, and translational efficiency of its target mRNAs. Through this mechanism, IGF2BP2 effectively projects the epigenetic regulatory signals of XIST onto metabolic pathways. Robust evidence supports the role of this protein family in immunometabolism and lipid metabolic reprogramming - for instance, by modulating OXPHOS/FAO preference through nodes such as peroxisome proliferator activated receptor γ and tuberous sclerosis 1. Additional findings indicate that IGF2BP2 facilitates its binding to Cpt1a mRNA and upregulates Cpt1a expression, further substantiating the molecular plausibility of the XIST-IGF2BP2-CPT1A axis[11].

CPT1A: The rate-limiting gatekeeper of FAO in the nervous system and its “double-edged” nature

CPT1A serves as the rate-limiting enzyme governing the entry of long-chain fatty acids into mitochondria for β-oxidation. At the glial level, FAO has been shown to organize and sustain the structure of mitochondrial respiratory supercomplexes, generating moderate levels of signaling ROS that support cognitive homeostasis. Disruption of CPT1A in astrocytes impairs respiratory chain assembly and energetic coupling, which in turn alters the functional phenotype of co-cultured neurons - highlighting the critical importance of maintaining FAO for neural network stability[12].

However, in pathological contexts, pharmacologic inhibition of CPT1 (e.g., etomoxir) remains controversial due to its off-target and toxicological issues, including fatty acid mimetic properties and mitochondrial/cytotoxic effects. Although such agents effectively suppress FAO, they cannot be regarded as specific probes for CPT1A activity. Therefore, any intervention targeting CPT1A must carefully define dosage, timing, and cell-type specificity to avoid adverse consequences such as energy crises or immune-metabolic dysregulation[13].

Conceptual framework and translational implications

Collectively, current evidence positions the XIST-IGF2BP2-CPT1A axis as a pivotal pathway bridging metabolic reprogramming and cell fate determination. Through IGF2BP2-dependent stabilization of Cpt1a transcripts, XIST enhances FAO and mitochondrial OXPHOS activity, thereby increasing cellular energy supply, promoting neuronal differentiation of NSCs, and suppressing astrocytic lineage commitment. Within the context of mitochondrial dysfunction that pervades the acute, subacute, and chronic stages of SCI, this axis provides a novel metabolic intervention paradigm.

A regulatory strategy centered on XIST-IGF2BP2-CPT1A could be synergistically integrated with peroxisome proliferator-activated receptor-γ coactivator-1α-mediated mitochondrial biogenesis, antioxidant and anti-inflammatory therapies, and NSC transplantation, collectively achieving energy rebalance in the microenvironment, precise lineage guidance, and comprehensive neural functional recovery. Future studies should emphasize temporal alignment between interventions and disease stages, clarify cell type-specific responses, and optimize dosage and delivery modalities. In addition, reliance on nonspecific inhibitors such as etomoxir as causal probes should be avoided to ensure mechanistic validity and translational safety[1] (Figure 1).

Figure 1 X-inactive specific transcript-insulin-like growth factor 2 mRNA-binding protein 2-carnitine palmitoyl transferase 1A-driven bioenergetic rewiring links neural stem cell fate to multi-scale recovery after spinal cord injury. A: Spinal cord injury triggers a hostile microenvironment around the lesion core and penumbra, featuring reactive oxygen species burst, Ca²⁺ overload, mitochondrial permeability transition pore opening, ATP depletion, mitochondrial membrane damage, and inflammatory cytokines, forming an “energy failure → oxidative stress → cell death” loop within an early mitochondrial protection window; B: In neural stem cells, X-inactive specific transcriptional cooperates with insulin-like growth factor 2 mRNA-binding protein 2 to promote carnitine palmitoyl transferase 1A (CPT1A) mRNA stabilization/enhanced translation, increasing CPT1A at the outer mitochondrial membrane and driving fatty-acid import, fatty acid oxidation, and oxidative phosphorylation/electron transport chain activity, thereby restoring ATP production and oxygen consumption rate; C: This metabolism-to-fate switch favors neuronal lineage commitment (doublecortin/neuronal nuclei ↑) and suppresses reactive astrogliosis (glial fibrillary acidic protein ↓), supporting circuit reconstruction and reduced glial scarring; D: A context-dependent immunometabolic layer highlights microglial polarization regulated via a competing endogenous RNA pathway (X-inactive specific transcript-miR-124-3p-interferon regulatory factor 1) and a multi-layer regulatory module including chromatin/epigenetic control (scaffold attachment factor A/heterogeneous nuclear ribonucleoprotein U, polycomb repressive complex 1/2, trimethylation of histone H3 lysine 27, mediator complex subunit 14) and post-transcriptional N6-methyladenosine-linked regulation; E: System/functional outcomes include reduced inflammation, diminished cavity/scar, improved axonal continuity/remyelination (myelin basic protein), enhanced locomotor recovery (Basso-Beattie-Bresnahan locomotor rating scale-Basso Mouse Scale), and improved autonomic function; F: Translational modules summarize delivery options (adeno-associated virus, engineered exosomes/extracellular vesicles, hydrogel scaffolds, neural stem cell transplantation) and rational combination add-ons (e.g., peroxisome proliferator-activated receptor gamma coactivator 1-alpha; elamipretide), with caution that CPT1A/fatty acid oxidation targeting requires dose-timing-cell specificity and avoidance of nonspecific CPT1 inhibition/off-target systemic metabolic effects. Solid arrows denote activation (→) and blunt lines denote inhibition (⊣). SCI: Spinal cord injury; ROS: Reactive oxygen species; mPTP: Mitochondrial permeability transition pore; ATP: Adenosine triphosphate; XIST: X-inactive specific transcript; NSC: Neural stem cell; CPT1A: Carnitine palmitoyltransferase 1A; IGF2BP2: Insulin-like growth factor 2 mRNA-binding protein 2; OMM: Outer mitochondrial membrane; FAO: Fatty acid oxidation; OXPHOS: Oxidative phosphorylation; ETC: Electron transport chain; OCR: Oxygen consumption rate; DCX: Doublecortin; NeuN: Neuronal nuclei; GFAP: Glial fibrillary acidic protein; SAF-A: Scaffold attachment factor A; hnRNPU: Heterogeneous nuclear ribonucleoprotein U; PRC1/PRC2: Polycomb repressive complex 1/2; H3K27me3: Trimethylation of histone H3 lysine 27; m⁶A: N6-methyladenosine; MED14: Mediator complex subunit 14; ceRNA: Competing endogenous RNA; IRF1: Interferon regulatory factor 1; MBP: Myelin basic protein; BBB: Basso-Beattie-Bresnahan locomotor rating scale; BMS: Basso Mouse Scale; AAV: Adeno-associated virus; EVs: Extracellular vesicles; PGC-1α: Peroxisome proliferator-activated receptor gamma coactivator 1-alpha; SS-31: Elamipretide.

BEYOND CPT1A: THE MULTILAYERED REGULATORY LANDSCAPE AND CONTEXT DEPENDENCE OF XIST

Beyond the IGF2BP2 → CPT1A → FAO/OXPHOS metabolic axis, XIST influences SCI pathology and stem cell fate through multiple additional layers of regulation.

Chromatin/epigenetic layer (X-chromosome inactivation and chromatin braking)

Acting in cis, XIST coats the chromosome and interacts with the nuclear-matrix anchoring protein scaffold attachment factor A/heterogeneous nuclear ribonucleoprotein U, positioning itself on chromatin to provide both a “scaffold” and spatial coordinates for the delivery of silencing complexes and modifying enzymes. This process is a core step in X-chromosome inactivation. Studies indicate that the requirement for scaffold attachment factor A in XIST-chromatin anchoring varies by cellular context, yet its role as an anchoring factor is consistently supported[14]. With respect to silencing effectors, recruitment of polycomb repressive complex 1/2 (PRC1/PRC2) and transcriptional repression can be partially decoupled in time and pathway: XIST can first establish baseline silencing, while the stable occupancy of PRC1/2 and deposition of trimethylation of histone H3 lysine 27 consolidate the silent state and promote chromatin folding - providing evidence for a hierarchical “silence first, consolidate later” sequence[15]. Notably, loss of XIST not only weakens silencing but also triggers reactivation at specific loci and aberrant enhancer activity, implicating Mediator components (e.g., mediator complex subunit 14) and other transcriptional co-regulators; functionally, this manifests as impaired stem cell differentiation and lineage skewing - suggesting that XIST possesses dual properties as both a silencer and an enhancer/mediator brake[16].

Post-transcriptional layer (RNA-binding protein/m⁶A bridge to metabolism)

At the post-transcriptional level, XIST cooperates with RNA-binding proteins, most prominently the m⁶A reader IGF2BP2. Members of the IGF2BP family recognize m⁶A-modified sites and broadly enhance the stability and translational efficiency of their target mRNAs, thereby transmitting lncRNA-derived regulatory signals downstream to metabolic and stress-response transcripts such as CPT1A and mitochondrial respiratory chain subunits. This “m⁶A reader-stabilizer” paradigm has been validated across multiple biological systems, expanding the conceptual framework for how XIST → IGF2BP2 → metabolic gene expression may operate mechanistically[17].

Competing endogenous RNA/immune layer (miR-124-3p/interferon regulatory factor 1 axis and microglial polarization)

At the inflammatory and immune level, XIST can function as a competing endogenous RNA sponge to modulate the miR-124-3p/interferon regulatory factor 1 axis, thereby influencing microglial polarization and the broader cytokine network. Evidence from mouse and cellular models shows that inhibition of XIST promotes M2 polarization, yet paradoxically exacerbates the SCI phenotype (Heliyon, 2023) - illustrating the paradox that improved immune phenotype ≠ functional recovery[18]. This finding reflects the complex interplay among temporal windows, polarization dynamics, and metabolic energy states following SCI. The same study also demonstrated the converse effect - XIST upregulation → M1 polarization → aggravated inflammation - highlighting the context dependence of XIST regulation: Depending on timing, injury severity, and microenvironmental metabolic status, the overall outcomes of modulating XIST may be diametrically opposed[18].

Cross-tissue mitochondrial network (hepatic example)

The impact of XIST on mitochondrial homeostasis is not confined to the nervous system. In models of hepatic fibrosis, the XISt/miR-539-3p/ADAMTS5 axis has been shown to induce mitochondrial injury in hepatocytes and promote hepatic stellate cell activation, suggesting that XIST exerts a cross-tissue influence on energy homeostasis and mitochondrial quality control. This finding provides additional evidence supporting XIST as an epigenetic-transcriptional-mitochondrial nexus[2].

Intercellular communication and stem-immune coregulation: The expanding regulatory horizon of XIST

Beyond its established cell-autonomous functions, the regulatory influence of XIST likely transcends intracellular boundaries, positioning it as a pivotal mediator in the sophisticated intercellular dialogue required for spinal cord regeneration and inflammatory modulation. Emerging conceptual frameworks suggest that XIST may participate in a “stem system” evolved to meet the demands of tissue homeostasis, where neural and mesenchymal stem cells utilize extracellular vesicles (EVs) or exosomes to shuttle regulatory RNAs to the immune microenvironment[19]. Specifically, mesenchymal stem cell-derived exosomes have been shown to outsource metabolic challenges such as mitophagy and transfer specific lncRNAs or microRNAs to immune cells, thereby driving macrophage polarization toward a pro-regenerative M2 phenotype and alleviating chronic inflammation[20,21]. In the context of SCI, this EV-mediated crosstalk facilitates a coregulation hub between stem cells, macrophages, and regulatory T cells, which is essential for proper tissue repair and functional reconstruction[22,23]. Furthermore, evidence indicates that such exosomal shuttling of RNAs can activate downstream regenerative pathways, such as the phosphatidylinositol 3-kinase/protein kinase B axis, to relieve tissue damage and promote cellular survival[24,25].

Integrative perspective and translational implications

Collectively, current evidence positions XIST as a multidimensional regulatory hub spanning epigenetic, transcriptional, metabolic, and immunological layers in SCI. In NSCs, XIST stabilizes metabolism-related transcripts via IGF2BP2, enhancing OXPHOS and mitochondrial function, thereby promoting neuronal differentiation. Conversely, in microglial cells, XIST amplifies pro-inflammatory signaling through the miR-124-3p/interferon regulatory factor 1 axis; yet, its inhibition at certain stages may paradoxically impair overall recovery due to insufficient energy supply, inefficient debris clearance, and disrupted glia-neuron interactions, underscoring a strong cell type- and time-dependent complexity.

Future therapeutic strategies should achieve precise stratification and metabolic coupling across distinct cellular lineages: Enhancing FAO and OXPHOS activity before and after NSC transplantation to optimize the energetic microenvironment, while integrating anti-inflammatory and antioxidant regimens. In microglia, regulation should be reversible, low-dose, and temporally phased to preserve essential clearance and repair functions.

Furthermore, multi-omics validation - including chromatin immunoprecipitation sequencing, an assay for transposase-accessible chromatin using sequencing, RNA immunoprecipitation, and m⁶A-CLIP - is warranted to systematically delineate the causal chain linking XIST, RNA-binding proteins, and metabolic target genes, with behavioral and bioenergetic outcomes incorporated as terminal endpoints. Overall, this framework positions XIST as a master integrator of chromatin regulation, post-transcriptional amplification, and immunometabolic coordination, thereby laying a theoretical and experimental foundation for precise, cell-specific, and temporally controllable interventions in SCI repair (Table 1).

THERAPEUTIC POTENTIAL AND TRANSLATIONAL CONSIDERATIONS

A metabolic intervention strategy centered on the XIST-IGF2BP2-CPT1A axis requires suitable delivery vehicles and combinatorial approaches as a foundation. In terms of delivery systems, several complementary platforms have demonstrated potential for translating the XIST-IGF2BP2-CPT1A-centered metabolic intervention into clinical practice. Viral vectors, particularly adeno-associated viruses, have accumulated extensive experience in central nervous system delivery and clinical trials; however, high-dose immune responses, hepatotoxicity, and neurotoxicity require rigorous evaluation and dose-engineering optimization. Adeno-associated viruses remain most suitable for in vivo long-term overexpression or knockdown validation and for exploring clinically scalable production routes[26]. Exosomes provide additional advantages, including biocompatibility, ability to cross the blood-spinal cord barrier, and engineering versatility. They can encapsulate lncRNAs, siRNAs, or antisense oligonucleotides and have already shown therapeutic efficacy in SCI regeneration and synaptic plasticity; when combined with biomaterials, their local retention and controlled release at the lesion site are markedly enhanced[27]. Furthermore, hydrogels and porous scaffolds can serve as co-delivery platforms for NSCs and exosomes, offering mechanical support, cavity bridging, and sustained release of microenvironmental cues. These biomaterials represent a critical interface integrating gene-, cell-, and drug-based interventions into a cohesive “regenerative engineering” framework[28].

Regarding intervention modes, based on the evidence by Zeng et al[1] (i.e., XIST → IGF2BP2 → CPT1A stabilization → enhanced FAO/OXPHOS → promoted neuronal differentiation with reduced inflammation and improved motor scores), the primary strategy should involve upregulating XIST or enhancing CPT1A activity/expression within NSCs or their transplant microenvironment. Systemic or ectopic modulation must be approached cautiously, with careful attention to timing and targeting specificity. Supporting studies indicate that Cpt1a overexpression restores mitochondrial homeostasis and energy metabolism to promote tissue repair, providing rationale for “XIST + metabolic enhancement” as a combinatorial approach[29].

Nevertheless, FAO modulation safety remains a major translational bottleneck. Astrocytes rely on FAO to maintain respiratory supercomplex architecture and signaling ROS levels that support central nervous system network function. Crude inhibition or imbalance may disrupt these processes. Clinical cases of hepatotoxicity and neurotoxicity linked to Cpt1 inhibitors (e.g., perhexiline) highlight the necessity of establishing metabolic and organ toxicity monitoring and drug-withdrawal thresholds[12].

Potential combination strategies include XIST gene modulation co-delivered via exosome- or hydrogel-based platforms, together with mitochondria-targeted therapeutics such as SS-31 (elamipretide) to stabilize membranes and protect mitochondrial integrity. Moreover, pairing these interventions with neuromodulatory techniques (e.g., optogenetic or electrical stimulation) could further promote circuit reconstruction and functional recovery[30].

In summary, XIST should not be applied in isolation, but rather integrated into a multimodal SCI repair framework that combines gene therapy, stem cell transplantation, biomaterials, and neural modulation. Such an integrated approach - grounded in precise delivery and rigorous risk management - will be essential to advance clinical translation[31].

UNRESOLVED ISSUES & RESEARCH AGENDA

Precision histomorphology and cell-lineage resolution

A critical limitation in existing evidence, including the study by Zeng et al[1], is the insufficient depth of histomorphological characterization. Robust validation of NSC fate necessitates the use of specialized and specific parameters rather than relying on generalized pan-neuronal markers. Future studies must prioritize confocal co-localization analysis - such as the overlapping of BrdU/Edu with mature neuronal markers (e.g., neuronal nuclei or microtubule-associated protein 2) to confirm terminal differentiation, and synaptic proteins (e.g., synaptophysin or postsynaptic density protein-95) to verify functional integration into neural circuits[7]. Furthermore, the XIST-IGF2BP2-CPT1A axis likely exerts distinct regulatory effects across cellular sub-populations (e.g., inhibitory vs excitatory neurons). Integrating single-nucleus RNA sequencing with spatial transcriptomics at multiple post-injury stages is essential to construct a high-resolution cell-space-time atlas for XIST, thereby clarifying its lineage-specific roles that remain obscured in bulk tissue analyses[32].

Extended longitudinal observation in chronic SCI models

Most current investigations focus on the acute or subacute phases of SCI, yet the clinically relevant processes - such as scar remodeling, cavity stabilization, and long-term functional reconstruction - predominantly evolve during the chronic stage. To enhance translational relevance, research should extend follow-up periods to 12-24 weeks using standardized compression or contusion models. This longitudinal approach should integrate behavioral trajectories (e.g., gait analysis and grid walking) with magnetic resonance imaging/diffusion tensor imaging-based structure-function correlations to ensure that short-term metabolic gains translate into sustained neurological recovery[33].

Metabolic safety and scalable delivery platforms

Targeting CPT1A requires a rigorous evaluation of its biphasic effects, as its systemic or non-specific inhibition may trigger adverse metabolic reprogramming in astrocytes. Future strategies should employ cell-type-specific Cpt1a conditional knockout models to precisely define the safety window. Simultaneously, clinical translation relies on developing scalable delivery systems, such as engineered exosomes or EVs integrated into bio-responsive hydrogel scaffolds, to optimize local retention and the synergistic reconstruction of the injured microenvironment[12,34]. Collectively, these priorities establish a roadmap for the clinical realization of XIST-centered repair strategies (Table 2).

Table 2 Prioritized research agenda and experimental strategies for X-chromosome inactivation-centered metabolic interventions in spinal cord injury.

Research priority	Current gap (critique of evidence)	Suggested experimental approach (specific parameters)	Expected outcome	Ref.
Precision histomorphology and lineage specificity	Insufficiency of specialized histological characterization; lack of specific parameters to confirm NSC terminal differentiation	Confocal co-localization analysis (e.g., BrdU/NeuN for neurogenesis; synaptophysin for synaptic integration); integration of snRNA-seq and spatial transcriptomics	High-resolution mapping of XIST-mediated lineage commitment and functional integration across neural/glial subpopulations	Li et al[7], 2024; Zhang et al[32], 2024
Long-term structural and functional stability	Absence of 12-24 week chronic-phase data; limited correlation between short-term metabolic gains and longitudinal functional trajectories	Standardized chronic injury paradigms (compression/contusion); 12-24 weeks follow-up; multiparametric MRI/DTI coupled with composite behavioral metrics (gait analysis, grid walking)	Verification of sustained regenerative benefits, scar-stabilizing effects, and long-term structure-function correlations in chronic SCI	Rosenzweig et al[35], 2018; Chen et al[33], 2023
Metabolic safety and targeted delivery systems	Systemic metabolic risks of non-specific CPT1A intervention; demand for scalable, CNS-oriented and bio-responsive delivery platforms	Cell-type-specific Cpt1a cKO or knock-in models; engineered exosomes (EVs) or LNPs integrated into hydrogel scaffolds	Establishment of an optimized therapeutic window and clinically feasible “gene-cell-vector” platforms for spinal cord repair	Morant-Ferrando et al[12], 2023; Williams et al[34], 2025

NSC: Neural stem cell; BrdU: 5-bromo-2’-deoxyuridine; NeuN: Neuronal nuclei; snRNA-seq: Single-nucleus RNA sequencing; XIST: X-inactive specific transcript; MRI: Magnetic resonance imaging; DTI: Diffusion tensor imaging; SCI: Spinal cord injury; CPT1A: Carnitine palmitoyl transferase 1A; CNS: Central nervous system; cKO: Conditional knockout; EVs: Extracellular vesicles; LNPs: Lipid nanoparticles.

CONCLUSION

As a dual hub connecting metabolic regulation and epigenetic control, XIST stabilizes CPT1A via IGF2BP2 in NSCs, thereby enhancing FAO and OXPHOS, promoting neuronal differentiation, and alleviating inflammation. CPT1A thus emerges as a pivotal effector and therapeutic target within this regulatory cascade. For clinical translation, careful evaluation of cell-type specificity, systemic metabolic compensation, dose-timing relationships, and long-term safety is essential. Looking ahead, translational progress will rely on multidisciplinary integration, combining XIST modulation with exosome- or hydrogel-based delivery, NSC engineering, and neuromodulatory approaches, alongside single-cell and spatial omics, lineage tracing, and mitochondrial imaging. These efforts should be coupled with the development of companion biomarkers and stratified intervention strategies to accelerate the clinical realization of XIST-centered spinal cord repair.