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The transplantation of rodent Schwann cells (SCs) provides anatomical and functional restitution in a variety of spinal cord injury (SCI) models, supporting the recent translation of SCs to phase 1 clinical trials for human SCI. Whereas human (Hu)SCs have been examined experimentally in a complete SCI transection paradigm, to date the reported behavior of SCs when transplanted after a clinically relevant contusive SCI has been restricted to the use of rodent SCs. Here, in a xenotransplant, contusive SCI paradigm, the survival, biodistribution, proliferation and tumorgenicity as well as host responses to HuSCs, cultured according to a protocol analogous to that developed for clinical application, were investigated. HuSCs persisted within the contused nude rat spinal cord through 6 months after transplantation (longest time examined), exhibited low cell proliferation, displayed no evidence of tumorigenicity and showed a restricted biodistribution to the lesion. Neuropathological examination of the CNS revealed no adverse effects of HuSCs. Animals exhibiting higher numbers of surviving HuSCs within the lesion showed greater volumes of preserved white matter and host rat SC and astrocyte ingress as well as axon ingrowth and myelination. These results demonstrate the safety of HuSCs when employed in a clinically relevant experimental SCI paradigm. Further, signs of a potentially positive influence of HuSC transplants on host tissue pathology were observed. These findings show that HuSCs exhibit a favorable toxicity profile for up to 6 months after transplantation into the contused rat spinal cord, an important outcome for FDA consideration of their use in human clinical trials., (© 2017 Wiley Periodicals, Inc.)

The elevation of intracellular cAMP synergistically enhances the neuregulin-dependent proliferation of cultured Schwann cells (SCs); however, the mechanism by which this occurs has not been completely defined. To better understand this mechanism, we investigated the effect of cAMP on the activation of the extracellular signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3-K)-Akt (PKB) pathways by heregulin, a member of the neuregulin family. Using primary cultures of adult SCs, we demonstrated that the adenylyl cyclase activator, forskolin, enhanced heregulin-dependent SC proliferation by reducing the time required for S-phase entry. When cAMP levels were increased, using either forskolin or a cell permeable analogue of cAMP, the heregulin-induced phosphorylation of ERK was converted from transient to sustained and the heregulin-induced phosphorylation of Akt was synergistically increased. Consistent with these observations, studies in which inhibitors of MEK, the upstream stimulating ERK kinase, and PI3-K were administered at different times following the onset of stimulation indicated that sustained high levels of both MEK/ERK and PI3-K/Akt activity before S-phase initiation were essential for S-phase entry. Overall, these novel results indicate that in neuregulin-stimulated SCs the activation of cAMP-mediated pathways accelerates G1-S progression by prolonging ERK activation and concurrently enhancing Akt activation.

Hypoxic preconditioning is protective in multiple models of injury and disease, but whether it is beneficial for cells transplanted into sites of spinal cord injury (SCI) is largely unexplored. In this study, we analyzed whether hypoxia-related preconditioning protected Schwann cells (SCs) transplanted into the contused thoracic rat spinal cord. Hypoxic preconditioning was induced in SCs prior to transplantation by exposure to either low oxygen (1% O 2 ) or pharmacological agents (deferoxamine or adaptaquin). All preconditioning approaches induced hypoxic adaptations, including increased expression of HIF-1α and its target genes. These adaptations, however, were transient and resolved within 24 h of transplantation. Pharmacological preconditioning attenuated spinal cord oxidative stress and enhanced transplant vascularization, but it did not improve either transplanted cell survival or recovery of sensory or motor function. Together, these experiments show that hypoxia-related preconditioning is ineffective at augmenting either cell survival or the functional outcomes of SC-SCI transplants. They also reveal that the benefits of hypoxia-related adaptations induced by preconditioning for cell transplant therapies are not universal., (© 2022 Wiley Periodicals LLC.)

Skin is an easily accessible tissue and a rich source of Schwann cells (SCs). Toward potential clinical application of autologous SC therapies, we aim to improve the reliability and specificity of our protocol to obtain SCs from small skin samples. As well, to explore potential functional distinctions between skin-derived SCs (Sk-SCs) and nerve-derived SCs (N-SCs), we used single-cell RNA-sequencing and a series of in vitro and in vivo assays. Our results showed that Sk-SCs expressed typical SC markers. Single-cell sequencing of Sk- and N-SCs revealed an overwhelming overlap in gene expression with the exception of HLA genes which were preferentially up-regulated in Sk-SCs. In vitro, both cell types exhibited similar levels of proliferation, migration, uptake of myelin debris and readily associated with neurites when co-cultured with human iPSC-induced motor neurons. Both exhibited ensheathment of multiple neurites and early phase of myelination, especially in N-SCs. Interestingly, dorsal root ganglion (DRG) neurite outgrowth assay showed substantially more complexed neurite outgrowth in DRGs exposed to Sk-SC conditioned media compared to those from N-SCs. Multiplex ELISA array revealed shared growth factor profiles, but Sk-SCs expressed a higher level of VEGF. Transplantation of Sk- and N-SCs into injured peripheral nerve in nude rats and NOD-SCID mice showed close association of both SCs to regenerating axons. Myelination of rodent axons was observed infrequently by N-SCs, but absent in Sk-SC xenografts. Overall, our results showed that Sk-SCs share near-identical properties to N-SCs but with subtle differences that could potentially enhance their therapeutic utility., (© 2022 Wiley Periodicals LLC.)

Spinal cord injury (SCI) can result in severe motor and sensory deficits, for which currently no effective cure exists. The pathological process underlying this injury is extremely complex and involves many cell types in the central nervous system. In this study, we have uncovered a novel function for macrophage G protein-coupled receptor kinase-interactor 1 (GIT1) in promoting remyelination and functional repair after SCI. Using GIT1 flox/flox Lyz2-Cre (GIT1 CKO) mice, we identified that GIT1 deficiency in macrophages led to an increased generation of tumor necrosis factor-alpha (TNFα), reduced proportion of mature oligodendrocytes (mOLs), impaired remyelination, and compromised functional recovery in vivo. These effects in GIT1 CKO mice were reversed with the administration of soluble TNF inhibitor. Moreover, bone marrow transplantation from GIT1 CWT mice reversed adverse outcomes in GIT1 CKO mice, further indicating the role of macrophage GIT1 in modulating spinal cord injury repair. Our in vitro experiments showed that macrophage GIT1 plays a critical role in secreting TNFα and influences the differentiation of oligodendrocyte precursor cells (OPCs) after stimulation with myelin debris. Collectively, our data uncovered a new role of macrophage GIT1 in regulating the transformation of OPCs into mOLs, essential for functional remyelination after SCI, suggesting that macrophage GIT1 could be a promising treatment target of SCI., (© 2024 Wiley Periodicals LLC.)

Schwann cells are critical for the proper development and function of the peripheral nervous system (PNS), where they form a collaborative relationship with axons. Past studies highlighted that a pair of proteins called the prohibitins play major roles in Schwann cell biology. Prohibitins are ubiquitously expressed and versatile proteins. We have previously shown that while prohibitins play a crucial role in Schwann cell mitochondria for long-term myelin maintenance and axon health, they may also be present at the Schwann cell-axon interface during development. Here, we expand on this, showing that drug-mediated modulation of prohibitins in vitro disrupts myelination and confirming that Schwann cell-specific ablation of prohibitin 2 (Phb2) in vivo results in severe defects in radial sorting and myelination. We show in vivo that Phb2-null Schwann cells cannot effectively proliferate and the transcription factors EGR2 (KROX20), POU3F1 (OCT6), and POU3F2 (BRN2), necessary for proper Schwann cell maturation, are dysregulated. Schwann cell-specific deletion of Jun, a transcription factor associated with negative regulation of myelination, confers partial rescue of the developmental defect seen in mice lacking Schwann cell Phb2. Finally, we identify a pool of candidate PHB2 interactors that change their interaction with PHB2 depending on neuronal signals, and thus are potential mediators of PHB2-associated developmental defects. This work develops our understanding of Schwann cell biology, revealing that Phb2 may modulate the timely expression of transcription factors necessary for proper PNS development, and proposing candidates that may play a role in PHB2-mediated integration of axon signals in the Schwann cell., (© 2024 Wiley Periodicals LLC.)

Schwann cell (SC) grafts promote axon regeneration in the injured spinal cord, but transplant efficacy is diminished by a high death rate in the first 2-3 days postimplantation. Both hypoxic preconditioning and pharmacological induction of the cellular hypoxic response can drive cellular adaptations and improve transplant survival in a number of disease/injury models. Hypoxia-inducible factor 1 alpha (HIF-1α), a regulator of the cellular response to hypoxia, is implicated in preconditioning-associated protection. HIF-1α cellular levels are regulated by the HIF-prolyl hydroxylases (HIF-PHDs). Pharmacological inhibition of the HIF-PHDs mimics hypoxic preconditioning and provides a method to induce adaptive hypoxic responses without direct exposure to hypoxia. In this study, we show that hypoxia-mimetics, deferoxamine (DFO) and adaptaquin (AQ), enhance HIF-1α stability and HIF-1α target gene expression. Expression profiling of hypoxia-related genes demonstrates that HIF-dependent and HIF-independent expression changes occur. Analyses of transcription factor binding sites identify several candidate transcriptional co-regulators that vary in SCs along with HIF-1α. Using an in vitro model system, we show that hypoxia-mimetics are potent blockers of oxidative stress-induced death in SCs. In contrast, traditional hypoxic preconditioning was not protective. The robust protection induced by pharmacological preconditioning, particularly with DFO, indicates that pharmacological induction of hypoxic adaptations could be useful for promoting transplanted SC survival. These agents may also be more broadly useful for protecting SCs, as oxidative stress is a major pathway that drives cellular damage in the context of neurological injury and disease, including demyelinating diseases and peripheral neuropathies., (© 2021 Wiley Periodicals LLC.)

The presence of peripheral myelinating cells in the central nervous system (CNS) has gained the neurobiologist attention over the years. Despite the confirmed presence of Schwann cells in the CNS in pathological conditions, and the long list of their beneficial effects on central remyelination, the cues that impede or allow Schwann cells to successfully conquer and remyelinate central axons remain partially undiscovered. A better knowledge of these factors stands out as crucial to foresee a rational therapeutic approach for the use of Schwann cells in CNS repair. Here, we review the diverse origins of Schwann cells into the CNS, both peripheral and central, as well as the CNS components that inhibit Schwann survival and migration into the central parenchyma. Namely, we analyze the astrocyte- and the myelin-derived components that restrict Schwann cells into the CNS. Finally, we highlight the unveiled mode of invasion of these peripheral cells through the central environment, using blood vessels as scaffolds to pave their ways toward demyelinated lesions. In short, this review presents the so far uncovered knowledge of this complex CNS-peripheral nervous system (PNS) relationship., (© 2020 Wiley Periodicals LLC.)

Human Schwann cells (hSCs) can be isolated directly from peripheral nerve and cultured using methods similar to those used for SCs from other species. Yet, important interspecies differences are revealed when the primary or expanded hSCs are compared to their nonhuman counterparts. This review addresses the special properties of nerve-derived hSCs that have resulted to date from both in vitro studies and in vivo research on cell transplantation in animal models and human subjects. A consensus has yet to emerge about the essential attributes of cultured normal hSCs. Thus, an emphasis is placed on the importance of validating hSC cultures by means of purity, identity, and biological activity to reliably use them as in vitro models of the SC phenotype and cell therapy products for injury repair. Combining traditional immunological methods, high-resolution omics approaches, and assorted cell-based assays is so far the best approach to unequivocally identify hSC populations obtained by direct isolation or derivation from stem cells. Special considerations are required to understand and manage the variability and heterogeneity proper of donor batches, as well as to evaluate risk factors. This is particularly important if the intended use of the hSCs is implantation in the human body, diagnosis of disease, or drug testing aimed at targeting any aspect of SC function in human patients. To conclude, in view of their unique properties, new concepts and methods are needed to better understand the biology of hSCs and exploit their full potential in basic science and regenerative medicine., (© 2020 Wiley Periodicals, Inc.)

Oligodendrocyte progenitor cells (OPCs) are the most proliferative and dispersed population of progenitor cells in the adult central nervous system, which allows these cells to rapidly respond to damage. Oligodendrocytes and myelin are lost after traumatic spinal cord injury (SCI), compromising efficient conduction and, potentially, the long-term health of axons. In response, OPCs proliferate and then differentiate into new oligodendrocytes and Schwann cells to remyelinate axons. This culminates in highly efficient remyelination following experimental SCI in which nearly all intact demyelinated axons are remyelinated in rodent models. However, myelin regeneration comprises only one role of OPCs following SCI. OPCs contribute to scar formation after SCI and restrict the regeneration of injured axons. Moreover, OPCs alter their gene expression following demyelination, express cytokines and perpetuate the immune response. Here, we review the functional contribution of myelin regeneration and other recently uncovered roles of OPCs and their progeny to repair following SCI., (© 2019 Wiley Periodicals, Inc.)

Common in vitro models used to study the mechanisms regulating myelination rely on co-cultures of oligodendrocyte precursor cells (OPCs) and neurons. In such models, myelination occurs in an environment that does not fully reflect cell-cell interactions and environmental cues present in vivo. To avoid these limitations while specifically manipulating oligodendroglial cells, we developed a reliable ex vivo model of myelination by seeding OPCs on cerebellar slices, deprived of their endogenous oligodendrocytes. We showed that exogenous OPCs seeded on unmyelinated cerebella, efficiently differentiate and form compact myelin. Spectral confocal reflectance microscopy and electron microscopy analysis revealed that the density of compacted myelin sheaths highly increases all along the culture. Importantly, we defined the appropriate culture time frame to study OPC differentiation and myelination, using accurate quantification resources we generated. Thus, this model is a powerful tool to study the cellular and molecular mechanisms of OPC differentiation and myelination. Moreover, it is suitable for the development and validation of new therapies for myelin-related disorders such as multiple sclerosis and psychiatric diseases., (© 2021 Wiley Periodicals LLC.)