Severe nerve injuries can be treated with electrical stimulation and stem cell therapies, but little is known about the potential benefits of combining these two treatments. In an effort to investigate this combination, we conducted a study to evaluate the effectiveness of electrical stimulation and Schwann-like cell transplantation in female Wistar albino rats. Our study consisted of five groups of rats: a sham group, an injury group, an electrical stimulation group, a Schwann-like cell group, and a combination group. The experimental groups received electrical stimulation, Schwann-like cell transplantation, or both. The animals sciatic function index was evaluated during a 6-week recovery period, and nerve conduction velocity, wet muscle mass, and nerve tissues were also analyzed. The results of the study showed that all experimental groups had a faster functional recovery compared to the injury group, although the difference between groups was not statistically significant. Both the combination group and the Schwann-like cell transplantation group had a higher nerve conduction velocity compared to the other experimental groups. However, there was no significant difference between the combination and Schwann-like cell transplantation groups. Nonetheless, histological analysis showed a better axonal reorganization in the combination group. The study provides preliminary evidence of the potential benefits of combining electrical stimulation and Schwann-like cell transplantation in treating severe nerve injuries. However, further studies with larger sample sizes are needed to confirm these findings and optimize the treatment parameters.

Schwann cells (SCs) are essential for the regenerative processes of peripheral nerve injuries. However, their use in cell therapy is limited. In this context, several studies have demonstrated the ability of mesenchymal stem cells (MSCs) to transdifferentiate into Schwann-like cells (SLCs) using chemical protocols or co-culture with SCs. Here, we describe for the first time the in vitro transdifferentiation potential of MSCs derived from equine adipose tissue (AT) and equine bone marrow (BM) into SLCs using a practical method. In this study, the facial nerve of a horse was collected, cut into fragments, and incubated in cell culture medium for 48 h. This medium was used to transdifferentiate the MSCs into SLCs. Equine AT-MSCs and BM-MSCs were incubated with the induction medium for 5 days. After this period, the morphology, cell viability, metabolic activity, gene expression of glial markers glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), p75 and S100β, nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and glial cell-derived neurotrophic factor (GDNF), and the protein expression of S100 and GFAP were evaluated in undifferentiated and differentiated cells. The MSCs from the two sources incubated with the induction medium exhibited similar morphology to the SCs and maintained cell viability and metabolic activity. There was a significant increase in the gene expression of BDNF, GDNF, GFAP, MBP, p75, and S100β in equine AT-MSCs and GDNF, GFAP, MBP, p75, and S100β in equine BM-MSCs post-differentiation. Immunofluorescence analysis revealed GFAP expression in undifferentiated and differentiated cells, with a significant increase in the integrated pixel density in differentiated cells and S100 was only expressed in differentiated cells from both sources. These findings indicate that equine AT-MSCs and BM-MSCs have great transdifferentiation potential into SLCs using this method, and they represent a promising strategy for cell-based therapy for peripheral nerve regeneration in horses.

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Peripheral nerve injuries (PNIs) remain one of the great clinical challenges because of their considerable long-term disability potential. Postnatal neural crest-derived multipotent stem cells, including gingiva-derived mesenchymal stem cells (GMSCs), represent a promising source of seed cells for tissue engineering and regenerative therapy of various disorders, including PNIs. Here, we generated GMSC-repopulated nerve protectors and evaluated their therapeutic effects in a crush injury model of rat sciatic nerves.
GMSCs were mixed in methacrylated collagen and cultured for 48 h, allowing the conversion of GMSCs into Schwann-like cells (GiSCs). The phenotype of GiSCs was verified by fluorescence studies on the expression of Schwann cell markers. GMSCs encapsulated in the methacrylated 3D-collagen hydrogel were co-cultured with THP-1-derived macrophages, and the secretion of anti-inflammatory cytokine IL-10 or inflammatory cytokines TNF-α and IL-1β in the supernatant was determined by ELISA. In addition, GMSCs mixed in the methacrylated collagen were filled into a nerve protector made from the decellularized small intestine submucosal extracellular matrix (SIS-ECM) and cultured for 24 h, allowing the generation of functionalized nerve protectors repopulated with GiSCs. We implanted the nerve protector to wrap the injury site of rat sciatic nerves and performed functional and histological assessments 4 weeks post-surgery.
GMSCs encapsulated in the methacrylated 3D-collagen hydrogel were directly converted into Schwann-like cells (GiSCs) characterized by the expression of S-100β, p75NTR, BDNF, and GDNF. In vitro, co-culture of GMSCs encapsulated in the 3D-collagen hydrogel with macrophages remarkably increased the secretion of IL-10, an anti-inflammatory cytokine characteristic of pro-regenerative (M2) macrophages, but robustly reduced LPS-stimulated secretion of TNF-1α and IL-1β, two cytokines characteristic of pro-inflammatory (M1) macrophages. In addition, our results indicate that implantation of functionalized nerve protectors repopulated with GiSCs significantly accelerated functional recovery and axonal regeneration of crush-injured rat sciatic nerves accompanied by increased infiltration of pro-regenerative (M2) macrophages while a decreased infiltration of pro-inflammatory (M1) macrophages.
Collectively, these findings suggest that Schwann-like cells converted from GMSCs represent a promising source of supportive cells for regenerative therapy of PNI through their dual functions, neurotrophic effects, and immunomodulation of pro-inflammatory (M1)/pro-regenerative (M2) macrophages.

Peripheral nerve injury (PNI) is a relatively frequent type of trauma that results in the suffering of many patients worldwide every year. Schwann cells (SCs) are expected to be applied in cell therapy because of their ability to promote peripheral nerve regeneration. However, the lack of clinically renewable sources of SCs hinders the application of SC-based therapies. Adipose-derived stem cells (ADSCs) have generated great interest in recent years because of their multipotency and ease of harvest, and they have already been verified to differentiate into Schwann-like cells (SLCs) in vitro. However, the efficiency of differentiation and the functions of SLCs remain unsatisfactory. We newly generated three-dimensional (3D) SLC spheroids from ADSCs using a modified protocol with human recombinant peptide (RCP) petaloid μ-piece. Morphological analysis, gene expression analysis by qRT-PCR, ELISA measurement of the secretion capabilities of neurotrophic factors, and neurite formation assay were performed to evaluate the functions of these 3D SLCs in vitro. Motor function recovery was measured in a sciatic nerve injury mouse model to analyze the nerve regeneration-promoting effect of 3D SLCs in vivo. The differentiation efficiency and the secretion of neurotrophic factors were enhanced in 3D SLCs compared with conventional SLCs. 3D SLCs could more effectively promote neurite growth and longer neurite extension in a neuron-like SH-SY5Y model. Additionally, 3D SLCs had a better therapeutic effect on nerve regeneration after transplantation into the sciatic nerve injury mouse model. These findings demonstrated that the potential of ADSC-derived SLCs to promote nerve regeneration could be significantly increased using our modified differentiation protocol and by assembling cells into a 3D sphere conformation. Therefore, these cells have great potential and can be used in the clinical treatment of PNI.

BACKGROUND: The iatrogenic effects of repairing peripheral nerve injuries (PNIs) with autografts (AGTs) encouraged the present study to involve a new approach consisting of grafting xenogeneic prerecellularized allogeneic cells instead of AGTs.
METHODS: We compared sheep\'s AGT regenerative and functional capacity with decellularized human nerves prerecellularized with allogeneic Schwann-like cell xenografts (onwards called xenografts). Mesenchymal stem cells were isolated from ovine adipose tissue and induced in vitro to differentiate into Schwann-like cells (SLCs). Xenografts were grafted in ovine sciatic nerves. Left sciatic nerves (20 mm) were excised from 10 sheep. Then, five sheep were grafted with 20 mm xenografts, and five were reimplanted with their nerve segment rotated 180° (AGT).
RESULTS: All sheep treated with xenografts or AGT progressively recovered the strength, movement, and coordination of their intervened limb, which was still partial when the study was finished at sixth month postsurgery. At this time, numerous intrafascicular axons were observed in the distal and proximal graft extremes of both xenografts or AGTs, and submaximal nerve electrical conduction was observed. The xenografts and AGT-affected muscles appeared partially stunted.
CONCLUSIONS: Xenografts and AGT were equally efficacious in starting PNI repair and justified further studies using longer observation times. The hallmarks from this study are that human xenogeneic acellular scaffolds were recellularized with allogenic SCL and were not rejected by the nonhuman receptors but were also as functional as AGT within a relatively short time postsurgery. Thus, this innovative approach promises to be more practical and accessible than AGT or allogenic allografts and safer than AGT for PNI repair.

Peripheral nerve injury (PNI) after pelvic surgery is a common issue with a significant impact on patients. Autologous nerve grafting is the gold standard treatment for PNI, but this technique cannot be applied to fine nerve fibers in the pelvis. Schwann-like cell (SLC) differentiation is a novel therapeutic strategy for this clinical condition. However, the efficiency of SLC differentiation remains unsatisfactory. We modified an SLC differentiation protocol using adipose-derived stem cells (ADSCs) and folic acid. Morphology, gene expression and secretion of neurotrophic factors were examined to assess the differentiation quality and phenotypic characteristics. Our new modified protocol effectively induced a Schwann cell (SC) phenotype in ADSCs as assessed by morphology and expression of SC markers [S100 calcium-binding protein B (S100B), P < 0.01 ; p75 neurotrophic receptor (p75NTR), P < 0.05]. SLCs produced by the new protocol displayed a repair phenotype with decreased expression of ERBB2 and early growth response protein 2 (EGR2) / KROX20 (P < 0.01). Furthermore, our new protocol enhanced both mRNA expression and secretion of nerve growth factors by SLCs (P < 0.01). This protocol enhanced the SC characteristics and functions of ADSC-derived SLCs. This promising protocol requires further research and may contribute to SC-based nerve regeneration. J. Med. Invest. 68 : 347-353, August, 2021.

BACKGROUND: Acellular nerve allografts (ANA) recellularized with mesenchymal stem cells (MSC) or Schwann cells (SC) are, at present, a therapeutic option for peripheral nerve injuries (PNI). This study aimed to evaluate the regenerative and functional capacity of a recellularized allograft (RA) compared with autograft nerve reconstruction in PNI.
METHODS: Fourteen ovines were randomly included in two groups (n=7). A peroneal nerve gap 30 mm in length was excised, and nerve repair was performed by the transplantation of either an autograft or a recellularized allograft with SC-like cells. Evaluations included a histomorphological analysis of the ANA, MSC pre differentiated into SC-like cells, at one year follow-up functional limb recovery (support and gait), and nerve regeneration using neurophysiological tests and histomorphometric analysis. All evaluations were compared with the contralateral hindlimb as the control.
RESULTS: The nerve allograft was successfully decellularized and more than 70% of MSC were pre differentiated into SC-like cells. Functional assessment in both treated groups improved similarly over time (p <0.05). Neurophysiological results (latency, amplitude, and conduction velocity) also improved in both treated groups at twelve months. Histological results demonstrated a less organized arrangement of nerve fibers (p <0.05) with an active remyelination process (p <0.05) in both treated groups compared with controls at twelve months.
CONCLUSIONS: ANA recellularized with SC-like cells proved to be a successful treatment for nerve gaps. Motor recovery and nerve regeneration were satisfactorily achieved in both graft groups compared with their contralateral nontreated nerves. This approach could be useful for the clinical therapy of PNI.

Vascular network reconstruction plays a pivotal role in the axonal regeneration and nerve function recovery after peripheral nerve injury. Increasing evidence indicates that Schwann cells (SCs) can promote nerve function repair, and the beneficial effects attributed to SCs therapy may exert their therapeutic effects through paracrine mechanisms. Recently, the previous research of our group demonstrated the promising neuroregenerative capacity of Schwann-like cells (SCLCs) derived from differentiated human embryonic stem cell-derived neural stem cells (hESC-NSCs) in vitro. Herein, the effects of SC-like cell conditioned medium (SCLC-CM) on angiogenesis and nerve regeneration were further explored. The assays were performed to show the pro-angiogenic effects of SCLC-CM, such as promoted endothelial cell proliferation, migration and tube formation in vitro. In addition, Sprague-Dawley rats were treated with SCLC-CM after sciatic nerve crush injury, SCLC-CM was conducive for the recovery of sciatic nerve function, which was mainly manifested in the SFI increase, the wet weight ratio of gastrocnemius muscle, as well as the number and thickness of myelin. The SCLC-CM treatment reduced the Evans blue leakage and increased the expression of CD34 microvessels. Furthermore, SCLC-CM upregulated the expressions of p-Akt and p-mTOR in endothelial cells. In conclusion, SCLC-CM promotes angiogenesis and nerve regeneration, it is expected to become a new treatment strategy for peripheral nerve injury.

Schwann cells (SCs) combined with acellular nerve allografts (ANAs) effectively promote the regeneration and repair of peripheral nerves, but the exact mechanism has not been fully elucidated. However, the disadvantages of SCs include their limited source and slow rate of expansion in vitro. Previous studies have found that adipose-derived stem cells have the ability to differentiate into Schwann-like cells. Therefore, we speculated that Schwann-like cells combined with ANAs could profoundly facilitate nerve regeneration and repair. The aim of the present study was to investigate the cellular and molecular mechanisms of regeneration and repair. In this study, tissue-engineered nerves were first constructed by adipose-derived Schwann-like cells and ANAs to bridge missing sciatic nerves. Then, the rats were randomly divided into five groups (n = 12 per group): a Control group; a Model group; an ADSC group; an SC-L group; and a DMEM group. Twelve weeks postsurgery, behavioral function tests and molecular biological techniques were used to evaluate the function of regenerated nerves and the relevant molecular mechanisms after sciatic nerve injury (SNI). The results showed that adipose-derived Schwann-like cells combined with ANAs markedly promoted sciatic nerve regeneration and repair. These findings also demonstrated that the expression of neurotrophic factors (NFs) was increased, and the expression of Janus activated kinase2 (JAK2)/P-JAK2, signal transducer and activator of transcription-3 (STAT3)/P-STAT3 was decreased in the spinal cord after SNI. Therefore, these results suggested that highly expressed NFs in the spinal cord could promote nerve regeneration and repair by inhibiting activation of the JAK2/STAT3 signaling pathway.