Endoneurial Fibroblast Research Articles

A low dose of curcumin-PDA nanoparticles improves viability and proliferation in endoneurial fibroblasts and Schwann cell cultures

Curcumin is a polyphenol extracted from Curcuma longa’s roots. Low doses of curcumin are related to anti-inflammatory, antioxidant, and neuroprotective effects, while high doses are used for their lethality. This diversity of behaviors allows us to understand curcumin as a compound with hormetic action. Due to its strongly hydrophobic character, curcumin is often solubilized in organic compounds. In this way, we have recently reported the undesirable and occasionally irreversible effects of alcohol and DMSO on the viability of primary Schwann cell cultures. In this scenario, the use of nanoparticles as delivery systems has become a successful alternative strategy for these compounds. In the present work, we describe the structure of Polydopamine (PDA) nanoparticles, loaded with a low dose of curcumin (Curc-PDA) without the use of additional organic solvents. We analyzed the curcumin released, and we found two different forms of curcumin. Small increased cell viability and proliferation were observed in endoneurial fibroblast and Schwann cell primary cultures when Curc-PDA was steadily supplied for 5 days. The increased bioavailability of this natural compound and the impact on cells in culture not only confirm the properties of curcumin at very low doses but also provide a glimpse of a possible therapeutic alternative for PNS conditions in which SCs are involved.

Hedgehog signaling promotes endoneurial fibroblast migration and Vegf-A expression following facial nerve injury

BackgroundPeripheral nerve injuries are a common clinical problem which may result in permanent loss of motor or sensory function. A better understanding of the signaling pathways that lead to successful nerve regeneration may help in discovering new therapeutic targets. The Hedgehog (Hh) signaling pathway plays significant roles in nerve development and regeneration. In a mouse model of facial nerve injury, Hedgehog-responsive fibroblasts increase in number both at the site of injury and within the distal nerve. However, the role of these cells in facial nerve regeneration is not fully understood. We hypothesize that the Hh pathway plays an angiogenic and pro-migratory role following facial nerve injury. MethodsHedgehog pathway modulators were applied to murine endoneurial fibroblasts isolated from the murine facial nerve. The impact of pathway modulation on endoneurial fibroblast migration and cell proliferation was assessed. Gene expression changes of known Hedgehog target genes and the key angiogenic factor Vegf-A were determined by qPCR. In vivo, mice were treated with pathway agonist (SAG21k) and injured facial nerve specimens were analyzed via immunofluorescence and in situ hybridization. ResultsHedgehog pathway activation in facial nerve fibroblasts via SAG21k treatment increases Gli1 and Ptch1 expression, the rate of cellular migration, and Vegf-A expression in vitro. In vivo, expression of Gli1 and Vegf-A expression appears to increase after injury, particularly at the site of nerve injury and the distal nerve, as detected by immunofluorescence and in situ hybridization. Additionally, Gli1 transcripts co-localize with Vegf-A following transection injury to the facial nerve. DiscussionThese findings describe an angiogenic and pro-migratory role for the Hedgehog pathway mediated through effects on nerve fibroblasts. Given the critical role of Vegf-A in nerve regeneration, modulation of this pathway may represent a potential therapeutic target to improve facial nerve regeneration following injury.

Neuroinflammation in the peripheral nerve: Cause, modulator, or bystander in peripheral neuropathies?

The role of innate and adaptive inflammation as a primary driver or modifier of neuropathy in premorbidly normal nerves, and as a critical player in amplifying neuropathies of other known causes (e.g., genetic, metabolic) is incompletely understood and under‐researched, despite unmet clinical need. Also, cellular and humoral components of the adaptive and innate immune system are substantial disease modifying agents in the context of neuropathies and, at least in some neuropathies, there is an identified tight interrelationship between both compartments of the immune system. Additionally, the quadruple relationship between Schwann cell, axon, macrophage, and endoneurial fibroblast, with their diverse membrane bound and soluble signalling systems, forms a distinct focus for investigation in nerve diseases with inflammation secondary to Schwann cell mutations and possibly others. Identification of key immunological effector pathways that amplify neuropathic features and associated clinical symptomatology including pain should lead to realistic and timely possibilities for translatable therapeutic interventions using existing immunomodulators, alongside the development of novel therapeutic targets. GLIA 2016;64:475–486

A novel marker of Schwann cells and myelin of the peripheral nervous system.

Cellular markers are useful in the immunohistochemical studies of normal and pathological tissues. Herein, the development of a monoclonal antibody, Schwann/2E, which reacts with Schwann cells and myelin of the peripheral nervous system (PNS) is described. The Schwann/2E antibody was secreted by a hybridoma of mouse myeloma cells and mouse spleen cells that were immunized in vivo with a cytoskeletal fraction of the human spinal nerve. This antibody immunostained formalin-fixed, paraffin-embedded human, rat, and mouse tissue by the indirect immunoperoxidase method. Schwann cells and myelin of the PNS were intensely labeled by the Schwann/2E antibody. Both the nuclei and cytoplasm of the Schwann cells were labeled. As shown by a comparative light and an electron microscopic study, the Schwann/2E antibody immunoreacted with the Schwann cells that had myelinated axons, but not with those that had unmyelinated axons. The endoneurial fibroblast was not immunolabeled. This antibody slightly stained the endothelial cells of the lung and kidney. Myelin and oligodendroglia of the central nervous system did not react with the Schwann/2E antibody. The Schwann/2E antigen was stable in several histological fixatives. These results indicate that, under normal and pathological conditions, the Schwann/2E antibody could be a useful immunohistochemical marker of Schwann cells and myelin of the PNS.

Peripheral nerves in shiverer----dystrophic mouse chimeras: evidence that a non-Schwann cell component is required for axon ensheathment in vivo

Peripheral nerves in dystrophic mice express multiple axon ensheathment abnormalities. If an intrinsic deficiency expressed within the Schwann cells themselves were to account for this neuropathy, then such cells, existing in a chimera preparation, would be expected to express the same ensheathment abnormalities, whereas a coexisting population of non-dystrophic Schwann cells should not be similarly affected. The genotype of myelinated Schwann cells in shiverer----dystrophic chimera was established with immunocytochemical techniques. Shiverer myelin lacks the P1 component of myelin basic protein (MBP), whereas dystrophic myelin appears to contain normal levels of MBP. No correlation between the ensheathment characteristics of the chimera spinal roots and the genotype of the local Schwann cell population was found; both dystrophic Schwann cell populations expressing normalized ensheathment characteristics and shiverer Schwann cells failing to respond to the local presence of naked axons were observed. These results require that a defective extra Schwann cell component is involved in the pathogenesis of the dystrophic neuropathy. Moreover, the normal realization of that component appears to be a necessary prerequisite for shiverer Schwann cells to achieve full ensheathment competence. Although a definitive identification of the cell type(s) that expresses the dy gene locus has not been achieved in this chimera preparation, the observations are consistent with defective endoneurial fibroblast function.

The role of endoneurial fibroblasts in myelin degradation.

Counts of nuclei in electron micrographs of rat sciatic nerve show that approximately one third of the nuclear population is fibroblasts. Comparison of these figures with counts in teased fibers indicates that the majority of fibroblasts adheres tightly to myelinated nerve fibers. The early reactions and cellular transformations of the endoneurial fibroblast population upon intraneurial injections of india ink or of a purified preparation of rat sciatic myelin were studied for intervals of from 6 hours to 3 days after injection. There was phagocytosis of carbon or myelin by endoneurial fibroblasts as early as 6 hours after injection. A subsequent massive increase in phagocytic endoneurial cells appeared to develop from, and at the expense of, the normal endoneurial fibroblast population. The extent to which hematogenous monocytes also contributed to the macrophage population could not be assessed accurately by morphological criteria. Schwann cells never ingested carbon or myelin preparation during the period of investigation. Early invasion of the Schwann tube by macrophages from the endoneurial spaces could be demonstrated however. These macrophages were labeled by either carbon or myelin preparation which they had taken up from the endoneurial spaces before they entered the fibers through small gaps of the basal lamina, preferentially at the nodes of Ranvier. Carbon- or myelin-labeled hematogenous granulocytes entered nerve fibers in the same way, but the invasion of fibers by granulocytes was a transient early response limited to the first two days after injection. The data demonstrate the phagocytic capacities of endoneurial fibroblasts, their capacity for myelin degradation, and an invasion of Schwann tubes by elements of the endoneurial macrophage population during very early phases of fiber injury.