Objective. Our previous study demonstrated that the transcription factor, Krüppel-like Factor 7 (KLF7), stimulates axon regeneration following peripheral nerve injury. In the present study, we used a gene therapy approach to overexpress KLF7 in bone marrow-derived stem/stromal cells (BMSCs) as support cells, combined with acellular nerve allografts (ANAs) and determined the potential therapeutic efficacy of a KLF7-transfected BMSC nerve graft transplantation in a rodent model for sciatic nerve injury and repair. Approach. We efficiently transfected BMSCs with adeno-associated virus (AAV)-KLF7, which were then seeded in ANAs for bridging sciatic nerve defects. Main results. KLF7 overexpression promotes proliferation, survival, and Schwann-like cell differentiation of BMSCs in vitro. In vivo, KLF7 overexpression promotes transplanted BMSCs survival and myelinated fiber regeneration in regenerating ANAs; however, KLF7 did not improve Schwann-like cell differentiation of BMSCs within in the nerve grafts. KLF7-BMSCs significantly upregulated expression and secretion of neurotrophic factors by BMSCs, including nerve growth factor, ciliary neurotrophic factor, brain-derived neurotrophic factor, and glial cell line-derived neurotrophic factor in regenerating ANA. KLF7-BMSCs also improved motor axon regeneration, and subsequent neuromuscular innervation and prevention of muscle atrophy. These benefits were associated with increased motor functional recovery of regenerating ANAs. Significance. Our findings suggest that KLF7-BMSCs promoted peripheral nerve axon regeneration and myelination, and ultimately, motor functional recovery. The mechanism of KLF7 action may be related to its ability to enhance transplanted BMSCs survival and secrete neurotrophic factors rather than Schwann-like cell differentiation. This study provides novel foundational data connecting the benefits of KLF7 in neural injury and repair to BMSC biology and function, and demonstrates a potential combination approach for the treatment of injured peripheral nerve via nerve graft transplant.

Previous animal studies have demonstrated that the flavonoid small-molecule TrkB agonist, 7, 8-dihydroxyflavone (DHF), promotes axon regeneration in transected peripheral nerves. In the present study, we investigated the combined effects of 7, 8-DHF treatment and bone marrow-derived stem/stromal cells (BMSCs) engraftment into acellular nerve allografts (ANAs) and explore relevant mechanisms that may be involved. Our results show that TrkB and downstream ERK1/2 phosphorylation are increased upon 7, 8-DHF treatment compared to the negative control group. Also, 7, 8-DHF promotes proliferation, survival, and Schwann-like cell differentiation of BMSCs in vitro. While selective ERK1/2 inhibitor U0126 suppressed the effect of upregulation of ERK1/2 phosphorylation and decreased cell proliferation, survival, and Schwann-like cell differentiation partially induced by 7, 8-DHF. In vivo, 7, 8-DHF promotes survival of transplanted BMSCs and upregulates axonal growth and myelination in regenerating ANAs. 7, 8-DHF+BMSCs also improved motor endplate density of target musculature. These benefits were associated with increased motor functional recovery. 7, 8-DHF+BMSCs significantly upregulated TrkB and ERK1/2 phosphorylation expression in regenerating ANA, and increased TrkB expression in the lumbar spinal cord. The mechanism of 7, 8-DHF action may be related to its ability to upregulate TrkB signaling, and downstream activation of survival signaling molecules ERK1/2 in the regenerating ANAs and spinal cord and improved survival of transplanted BMSCs. This study provides novel foundational data connecting the benefits of 7, 8-DHF treatment in neural injury and repair to BMSCs biology and function and demonstrates a potential combination approach for the treatment of injured peripheral nerve via nerve graft transplant.

Objective. Acellular nerve allograft (ANA) is an effective surgical approach used to bridge the sciatic nerve gap. The molecular regulators of post-surgical recovery are not well-known. Here, we explored the effect of transgenic Schwann cells (SCs) overexpressing POU domain class 6, transcription factor 1 (POU6F1) on sciatic nerve regeneration within ANAs. We explored the functions of POU6F1 in nerve regeneration by using a cell model of H₂O₂-induced SCs injury and transplanting SCs overexpressing POU6F1 into ANA to repair sciatic nerve gaps. Approach. Using RNA-seq, Protein–Protein Interaction network analysis, gene ontology enrichment, and Kyoto Encyclopedia of Genes and Genomes pathway analysis, we identified a highly and differentially expressed transcription factor, POU6F1, following ANA treatment of sciatic nerve gap. Expressing a high degree of connectivity, POU6F1 was predicted to play a role in peripheral nervous system myelination. Main results. To test the role of POU6F1 in nerve regeneration after ANA, we infected SCs with adeno-associated virus—POU6F1, demonstrating that POU6F1 overexpression promotes proliferation, anti-apoptosis, and migration of SCs in vitro. We also found that POU6F1 significantly upregulated JNK1/2 and c-Jun phosphorylation and that selective JNK1/2 inhibition attenuated the effects of POU6F1 on proliferation, survival, migration, and JNK1/2 and c-Jun phosphorylation. The direct interaction of POU6F1 and activated JNK1/2 was subsequently confirmed by co-immunoprecipitation. In rat sciatic nerve injury model with a 10 mm gap, we confirmed the pattern of POU6F1 upregulation and co-localization with transplanted SCs. ANAs loaded with POU6F1-overexpressing SCs demonstrated the enhanced survival of transplanted SCs, axonal regeneration, myelination, and functional motor recovery compared to the ANA group loaded by SCs-only in line with in vitro findings. Significance. This study identifies POU6F1 as a novel regulator of post-injury sciatic nerve repair, acting through JNK/c-Jun signaling in SCs to optimize therapeutic outcomes in the ANA surgical approach.

The landscape of therapeutic deep brain stimulation (DBS) for locomotor function recovery is rapidly evolving. This review provides an overview of electrical neuromodulation effects on spinal cord injury (SCI), focusing on DBS for motor functional recovery in human and animal models. We highlight research providing insight into underlying cellular and molecular mechanisms. A literature review via Web of Science and PubMed databases from 1990 to May 29, 2024, reveals a growing body of evidence for therapeutic DBS in SCI recovery. Advances in techniques like optogenetics and whole-brain tractogram have helped elucidate DBS mechanisms. Neuronal targets sites for SCI functional recovery include the mesencephalic locomotor region (MLR), cuneiform nucleus (CNF), and nucleus raphe magnus (NRG), with pedunculopontine nucleus (PPN), periaqueductal gray (PAG), and nucleus ventroposterolateral thalami (VPL) for post-injury functional recovery treatment. Radiologically guided DBS optimization and combination therapy with classical rehabilitation have become an effective therapeutic method, though ongoing interventional trials are needed to enhance understanding and validate DBS efficacy in SCI. On the pre-clinical front, standardization of pre-clinical approaches are essential to enhance the quality of evidence on DBS safety and efficacy. Mapping brain targets and optimizing DBS protocols, aided by combined DBS and medical imaging, are critical endeavors. Overall, DBS holds promise for neurological and functional recovery after SCI, akin to other electrical stimulation approaches.

Spinal cord injury (SCI) causes permanent motor deficits primarily due to neuronal apoptosis and inadequate axonal regeneration, with the c-Jun N-terminal kinase (JNK)/c-Jun pathway playing a dual role in neurodegeneration and regeneration. Here, we identified Nyap1 as a spatiotemporally limited modulator that rearranges JNK signaling to shift its function from degeneration to regeneration. Time-resolved transcriptomic profiling showed Nyap1 was dynamically upregulated as a hub gene during the acute-to-subacute phase of SCI. Nyap1 was associated with altered subcellular distribution of p-JNK, with increased cytoplasmic retention, with overexpression enriching p-JNK in the cytosol and limiting nuclear translocation-sufficient to drive regenerative transcription without inducing apoptosis. This interaction was associated with modulation of apoptosis-related and regeneration-related signaling, including changes in the Bcl-2/Bax ratio, caspase-3 activation, c-Jun phosphorylation, and GAP43 expression, with these effects abrogated by JNK inhibition. In vivo, Nyap1 overexpression preserved thoracic spinal motor neurons, promoted descending propriospinal tract (DPST) axonal regeneration, enhanced lumbar motor neuron dendritic arborization and synaptic input, and reduced lesion cavity volume. These structural improvements were accompanied by better gait coordination, neuromuscular reinnervation, and translesional motor conduction. Our findings support a model in which Nyap1 is associated with spatial redistribution of p-JNK-related signaling, linking neuronal survival with circuit-level plasticity after SCI, highlighting its potential as a therapeutic candidate for CNS trauma.