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Although macrophages (MΦ) are known to play a central role in neuropathic pain, their contribution to cancer pain has not been established. Here we report that depletion of sciatic nerve resident MΦs (rMΦ) in mice attenuates mechanical/cold hypersensitivity and spontaneous pain evoked by intraplantar injection of melanoma or lung carcinoma cells. MΦ-colony stimulating factor (M-CSF) was upregulated in the sciatic nerve trunk and mediated cancer-evoked pain via rMΦ expansion, transient receptor potential ankyrin 1 (TRPA1) activation, and oxidative stress. Targeted deletion of Trpa1 revealed a key role for Schwann cell TRPA1 in sciatic nerve rMΦ expansion and pain-like behaviors. Depletion of rMΦs in a medial portion of the sciatic nerve prevented pain-like behaviors. Collectively, we identified a feed-forward pathway involving M-CSF, rMΦ, oxidative stress, and Schwann cell TRPA1 that operates throughout the nerve trunk to signal cancer-evoked pain. SIGNIFICANCE: Schwann cell TRPA1 sustains cancer pain through release of M-CSF and oxidative stress, which promote the expansion and the proalgesic actions of intraneural macrophages. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/12/3387/F1.large.jpg., (©2021 American Association for Cancer Research.)

Leprosy is a disease with a clinical spectrum of presentations that is also manifested in diverse histological features. At one pole, lepromatous lesions (L-pole) have phagocytic foamy macrophages heavily parasitized with freely multiplying intracellular Mycobacterium leprae . At the other pole, the presence of epithelioid giant cells and granulomatous formation in tuberculoid lesions (T-pole) lead to the control of M. leprae replication and the containment of its spread. The mechanism that triggers this polarization is unknown, but macrophages are central in this process. Over the past few years, leprosy has been studied using large scale techniques to shed light on the basic pathways that, upon infection, rewire the host cellular metabolism and gene expression. M. leprae is particularly peculiar as it invades Schwann cells in the nerves, reprogramming their gene expression leading to a stem-like cell phenotype. This modulatory behavior exerted by M. leprae is also observed in skin macrophages. Here, we used live M. leprae to infect (10:1 multiplicity of infection) monocyte-derived macrophages (MDMs) for 48 h and analyzed the whole gene expression profile using microarrays. In this model, we observe an intense upregulation of genes consistent with a cellular immune response, with enriched pathways including peptide and protein secretion, leukocyte activation, inflammation, and cellular divalent inorganic cation homeostasis. Among the most differentially expressed genes (DEGs) are CCL5/RANTES and CYP27B1 , and several members of the metallothionein and metalloproteinase families. This is consistent with a proinflammatory state that would resemble macrophage rewiring toward granulomatous formation observed at the T-pole. Furthermore, a comparison with a dataset retrieved from the Gene Expression Omnibus of M. leprae -infected Schwann cells (MOI 100:1) showed that the patterns among the DEGs are highly distinct, as the Schwann cells under these conditions had a scavenging and phagocytic gene profile similar to M2-like macrophages, with enriched pathways rearrangements in the cytoskeleton, lipid and cholesterol metabolism and upregulated genes including MVK , MSMO1 , and LACC1 / FAMIN . In summary, macrophages may have a central role in defining the paradigmatic cellular (T-pole) vs. humoral (L-pole) responses and it is likely that the multiplicity of infection and genetic polymorphisms in key genes are gearing this polarization., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Leal-Calvo, Martins, Bertoluci, Rosa, Camargo, Germano, Brito de Souza, Pereira Latini and Moraes.)

The interplay between immune cells and tumor cells determines the fate of tumorigenesis. Targeting the abnormal immune response of tumors has been recently achieved great success in some patients. Emerging evidence demonstrated the nervous system plays vital roles in immune regulation, but if the nervous system affects the immune-tumor response and the possible mechanism involved remain largely unexplored. Here, we report that Schwann cells, the major component of the peripheral nervous system (PNS), induce M2 polarization of macrophages by secreting cytokines and chemokines, and these polarized macrophages promote the proliferation of lung cancer cells. We cocultured peripheral blood mononuclear cells (PBMCs) with Schwann cells or treated PBMCs with the culture supernatant of Schwann cells. We found that both treatments induced M2 polarization of the macrophages in peripheral blood mononuclear cell cultures. We performed a bioinformatic analysis of the transcriptome of Schwann cells and analyzed cytokines and chemokines by ELISAs. We found that Schwann cells secreted high levels of CCL2, CXCL5, CXCL12, and CXCL8. CCL2 promotes the M2 polarization of macrophages. Furthermore, we isolated CD14-positive macrophages that were cocultured with the Schwann cells and treated A549 and H1299 lung cancer cells with these macrophages. We found that the Schwann cell-polarized macrophages increased the proliferation of the lung cancer cells. Our study sheds new light on the involvement of the PNS in the regulation of tumor progression via a "Schwann cell"-"immune cell"-"tumor cell" axis., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Chronic inflammatory demyelinating polyneuropathy (CIDP) and Guillain-Barre syndrome (GBS) are inflammatory neuropathies that affect humans and are characterized by peripheral nerve myelin destruction and macrophage-containing immune infiltrates. In contrast to the traditional view that the peripheral nerve is simply the target of autoimmunity, we report here that peripheral nerve Schwann cells exacerbate the autoimmune process through extracellular matrix (ECM) protein induction. In a spontaneous autoimmune peripheral polyneuropathy (SAPP) mouse model of inflammatory neuropathy and CIDP nerve biopsies, the ECM protein periostin (POSTN) was upregulated in affected sciatic nerves and was primarily expressed by Schwann cells. Postn deficiency delayed the onset and reduced the extent of neuropathy, as well as decreased the number of macrophages infiltrating the sciatic nerve. In an in vitro assay, POSTN promoted macrophage chemotaxis in an integrin-AM (ITGAM) and ITGAV-dependent manner. The PNS-infiltrating macrophages in SAPP-affected nerves were pathogenic, since depletion of macrophages protected against the development of neuropathy. Our findings show that Schwann cells promote macrophage infiltration by upregulating Postn and suggest that POSTN is a novel target for the treatment of macrophage-associated inflammatory neuropathies.

It is known that transient receptor potential ankyrin 1 (TRPA1) channels, expressed by nociceptors, contribute to neuropathic pain. Here we show that TRPA1 is also expressed in Schwann cells. We found that in mice with partial sciatic nerve ligation, TRPA1 silencing in nociceptors attenuated mechanical allodynia, without affecting macrophage infiltration and oxidative stress, whereas TRPA1 silencing in Schwann cells reduced both allodynia and neuroinflammation. Activation of Schwann cell TRPA1 evoked NADPH oxidase 1 (NOX1)-dependent H 2 O 2 release, and silencing or blocking Schwann cell NOX1 attenuated nerve injury-induced macrophage infiltration, oxidative stress and allodynia. Furthermore, the NOX2-dependent oxidative burst, produced by macrophages recruited to the perineural space activated the TRPA1-NOX1 pathway in Schwann cells, but not TRPA1 in nociceptors. Schwann cell TRPA1 generates a spatially constrained gradient of oxidative stress, which maintains macrophage infiltration to the injured nerve, and sends paracrine signals to activate TRPA1 of ensheathed nociceptors to sustain mechanical allodynia.

Although peripheral nerves are capable of regeneration, advanced age decreases the potential for functional recovery after injury. The cellular mechanisms for this are not currently understood. Here, we performed sciatic nerve grafting with young (2 months old) and aged (18 months old) Brown-Norway male rats, in which 1 cm nerve grafts from young or aged rats were sutured into nerves of young or aged rats. Axons were allowed to regenerate until the nerve grafts and distal nerves were harvested at 1, 3, and 7 days and 2 and 6 weeks. At 6 weeks, our data suggested that young nerve grafts supported regeneration better than aged nerve grafts. In addition, myelin debris clearance was inhibited in young nerves when grafted into aged rats, but clearance was faster when aged nerves were grafted into young rats. Further analysis revealed that aged macrophages have delayed migration into injured nerve, and macrophages and Schwann cells from aged rats were less phagocytic for myelin debris in vitro. To understand these impairments, expression levels of pro- and anti-inflammatory cytokines were analyzed at 1 day after injury. Based on these levels, there was not a clear polarization to either an M1 or M2 phenotype; however, expression levels of IL-6, IL-10, CCL2 (MCP1), and Arg-1 were decreased in aged nerves. Taken together, both macrophages and Schwann cells had attenuated responses to nerve injury in aged rats, leading to inefficient clearance of debris and impaired axonal regeneration., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Leprosy is a persistent infectious disease caused by Mycobacterium leprae that still affects over 200,000 new patients annually. The host genetic background is an important risk factor for leprosy susceptibility and the PARK2 gene is a replicated leprosy susceptibility candidate gene. The protein product of PARK2, Parkin, is an E3 ubiquitin ligase that is involved in the development of various forms of Parkinsonism. The human macrophage is both a natural host cell of M. leprae as well as a primary mediator of natural immune defenses, in part by secreting important pro-inflammatory cytokines and chemokines. Here, we report that down-regulation of Parkin in THP-1 macrophages, human monocyte-derived macrophages and human Schwann cells resulted in a consistent and specific decrease in interleukin-6 (IL-6) and monocyte chemoattractant protein 1 (MCP-1/CCL2) production in response to mycobacteria or LPS. Interestingly, production of IL-6 at 6 hours by THP-1 cells stimulated with live M. leprae and M. bovis BCG was dependent on pretreatment with 1,25-dihydroxyvitamin D(3) (VD). Parkin knockdown in VD-treated cells blocked IL-6 induction by mycobacteria. However, IκB-α phosphorylation and levels of IκB-ξ, a nuclear protein required for IL-6 expression, were not affected by Parkin silencing. Phosphorylation of MAPK ERK1/2 and p38 was unaffected by Parkin silencing while JNK activation was promoted but did not explain the altered cytokine production. In a final set of experiments we found that genetic risk factors of leprosy located in the PARK2 promoter region were significantly correlated with M. leprae sonicate triggered CCL2 and IL6 transcript levels in whole blood assays. These results associated genetically controlled changes in the production of MCP-1/CCL2 and IL-6 with known leprosy susceptibility factors.

Peripheral nerve repair across long gaps remains clinically challenging despite progress made with autograft transplantation. While scaffolds that present trophic factors and extracellular matrix molecules have been designed, matching the performance of autograft-induced repair has been challenging. In this study, we explored the effect of cytokine mediated 'biasing' of macrophage phenotypes on Schwann cell (SC) migration and axonal regeneration in vitro and in vivo. Macrophage phenotype was successfully modulated by local delivery of either Interferon-gamma (IFN-γ) or Interleukin-4 (IL-4) within polymeric nerve guidance channels, polarizing them toward pro-inflammatory (M1) or pro-healing (M2a and M2c) phenotypes, respectively. The initial polarization of macrophages to M2a and M2c phenotype results in enhanced SC infiltration and substantially faster axonal growth in a critically-sized rat sciatic nerve gap model (15 mm). The ratio of pro-healing to pro-inflammatory population of macrophages (CD206+/CCR7+), defined as regenerative bias, demonstrates a linear relationship with the number of axons at the distal end of the nerve scaffolds. The present results clearly suggest that rather than the extent of macrophage presence, their specific phenotype at the site of injury regulates the regenerative outcomes., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Human apolipoprotein E (apoE) is a 34.2kDa glycosylated protein with three isoforms (apoE2, apoE3 and apoE4). Experimental autoimmune neuritis (EAN), an animal model for human Guillain-Barré syndrome, is an immune-mediated experimental disorder of the peripheral nervous system (PNS). Increased susceptibility to EAN in apoE deficient mice has been previously found. To elucidate the isoform-dependent effects of apoE on EAN, we used human apoE2, E3 and E4 transgenic mice (Tg) immunized with P0 peptide 180-199, as well as T cell proliferation test, macrophage and Schwann cell (SC) cultures to investigate the effects of apoE isoforms on the functions of T cells, macrophages and SCs both under naïve conditions and in EAN. Clinical signs of EAN were most severe in wild type (WT) C57BL/6 mice and apoE4 Tg mice, followed by apoE2 Tg mice and apoE3 Tg mice (WT≈E4>E2>E3, p<0.01). At the nadir of EAN, spleen weight and lymphocyte proliferation were in line with the clinical severity of the disease. Proliferation tests of purified T cells from naive mice stimulated with phytohemagglutinin or interleukin-12 showed isoform-specific differences (WT≈E4>E3≈E2, p<0.01). Macrophages from both naïve and EAN mice produced nitric oxide upon inflammatory stimulation with lipopolysaccharide, interferon-γ, polyinosinic:polycytidylic acid or combinations thereof, in an isoform-dependent manner (WT≈E4>E2>E3, p<0.01). Generalized intervention with 1400W, a specific inducible nitric oxide synthase inhibitor, significantly suppressed the clinical course of EAN in apoE2, E3 and E4 Tg mice and in WT mice. During the recovery stage of disease, the highest expression of CD178 (FasL) on SCs was found in apoE3 Tg mice. Our data support an isoform-dependent effect of apoE on EAN. This might be due to the isoform-specific effects of apoE on functions of T cells, macrophages and SCs, which contribute to the distinct clinical courses of EAN. ApoE3 might not only inhibit the onset and suppress the clinical severity of EAN, but also enhance the termination of immune responses in the PNS., (Copyright © 2011 Elsevier Inc. All rights reserved.)

In the present study, we investigated the role of the macrophage inflammatory protein-1alpha (MIP-1alpha) in the pathogenesis of neuropathic pain following partial sciatic nerve ligation (PSL) in mice. MIP-1alpha mRNA and its protein were dramatically up-regulated after PSL, and MIP-1alpha was localized on macrophages and Schwann cells in the injured sciatic nerve (SCN). PSL-induced long-lasting tactile allodynia and thermal hyperalgesia were prevented by the perineural injection of anti-MIP-1alpha (2ng). Intraneural (20ng) and perineural (100ng) injection of recombinant MIP-1alpha elicited tactile allodynia and thermal hyperalgesia in sham-operated limb. MIP-1alpha receptors (CCR1 and CCR5) mRNA and their proteins were also up-regulated in the SCN after PSL, and were localized on macrophages and Schwann cells. PSL-induced tactile allodynia was attenuated by perineural injection (0.2nmol) of siRNA against CCR1 and CCR5. On the other hand, PSL-induced thermal hyperalgesia was prevented by siRNA against CCR5, but not CCR1. Interleukin-1beta (IL-1beta) mRNA and its precursor protein in macrophages and Schwann cells were also up-regulated in the SCN after PSL, and PSL-induced neuropathic pain was prevented by the perineural injection of anti-IL-1beta (2ng). PSL-induced IL-1beta up-regulation was suppressed by anti-MIP-1alpha and siRNA against CCR1 and CCR5. Perineural injection of nicotine (20nmol), a macrophage suppressor, prevented PSL-induced neuropathic pain and suppressed MIP-1alpha and IL-1beta expressions. In conclusion, we propose a novel critical molecule MIP-1alpha derived from macrophages and Schwann cells that appears to play a crucial role in the development of neuropathic pain induced by PSL., (Copyright 2010 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.)

In this article we first discuss the factors that regulate macrophage recruitment, activation, and myelin phagocytosis during Wallerian degeneration and some of the factors involved in the termination of inflammation at the end of the period of Wallerian degeneration after peripheral nerve injuries. In particular, we deal with the early events that trigger chemokine and cytokine expression; the role of phospholipase A(2) in initiating the breakdown of compact myelin, and chemokine, cytokine expression; and the role of MCP-1, MIP-1alpha, and IL-1beta in macrophage recruitment and myelin phagocytosis. We also discuss how inflammation may be switched off and the recently identified role of the Nogo receptor on activated macrophages in the clearance of these cells from the injured nerve. In the second half of the article we focus on the role of certain Schwann cell borne cytokines and chemokines, such as M-CSF and MCP-1 as well as intracellular signaling that regulate their expression in animal models of inherited demyelinating disease. Additionally, we present the preservation of sensory nerves fibers from macrophage attack in these animal models as a challenging paradigm for the development of putative treatment approaches. Finally, we also discuss the similarities and differences in these Schwann cell-macrophage responses in injury-induced Wallerian degeneration and inherited demyelinating diseases. Knowledge of the molecular mechanisms underlying Schwann cell-macrophage interaction under pathological conditions is an important prerequisite to develop effective treatment strategies for various peripheral nerve disorders.

Macrophages are critically involved in the pathogenesis of genetically caused demyelination, as it occurs in models for inherited demyelinating neuropathies. It is presently unknown which factors link the Schwann cell-based myelin mutation to the activation of endoneurial macrophages. Here we identified the chemokine monocyte chemoattractant protein-1 (MCP-1) as a first and crucial factor upregulated in Schwann cells of mice heterozygously deficient for the myelin protein zero. The chemokine could be identified as an important mediator of macrophage immigration into peripheral nerves. Furthermore, a 50% reduction of chemokine expression by crossbreeding with MCP-1-deficient mice reduced the increase in macrophage numbers in the mutant nerves and lead to a robust amelioration of pathology. Surprisingly, the complete absence of MCP-1 aggravated the disease. Our findings show that reducing but not eliminating chemokine expression can rescue genetically caused demyelination that may be an interesting target in treating demyelinating diseases of the peripheral nervous system.

The purpose of this study was to investigate quantitatively the longitudinal temporal, spatial changes of the tumor necrosis factor-alpha (TNF) and interleukin-10 (IL-10) immunopositive cells during Wallerian degeneration and the following regeneration after crush injury in rat sciatic nerve using immunohistochemistry and enzyme linked immunosorbent assay (ELISA). The number of TNF-immunopositive cells reached its peak and increased significantly in all the segments distal to the crush site 3 days after injury. On Day 7, TNF-immunopositive cells decreased in all the segments distal to the crush site, and a significant decrease was observed 14 days after injury. From Day 21 to Day 56, there were no significant differences in the numbers of TNF-immunopositive cells. The average size of TNF immunopositive cells became significantly larger with degeneration. The number of IL-10-immunopositive cells decreases significantly 1 day after crush injury. IL-10-immunopositive cells increased on Day 3, returning to control levels. Seven days after injury, a significant increase in the number of IL-10-immunopositive cells was observed. There was also no significant difference in the number of IL-10-immunopositive cells beyond Day 14 except for a part of distal segments. The number of IL-10-immunopositive cells showed no significant differences in all the segments on Day 56. The protein levels of IL-10 measured by ELISA were similar to the result of immunohistochemistry. These results suggest that the significant change in IL-10 occurred prior to the significant change in TNF and that IL-10 may be the key to the change in TNF.

We report a role for Nogo receptors (NgRs) in macrophage efflux from sites of inflammation in peripheral nerve. Increasing numbers of macrophages in crushed rat sciatic nerves express NgR1 and NgR2 on the cell surface in the first week after injury. These macrophages show reduced binding to myelin and MAG in vitro, which is reversed by NgR siRNA knockdown and by inhibiting Rho-associated kinase. Fourteen days after sciatic nerve crush, regenerating nerves with newly synthesized myelin have fewer macrophages than cut/ligated nerves that lack axons and myelin. Almost all macrophages in the cut/ligated nerves lie within the Schwann cell basal lamina, while in the crushed regenerating nerves the majority migrate out. Furthermore, crush-injured nerves of NgR1- and MAG-deficient mice and Y-27632-treated rats show impaired macrophage efflux from Schwann cell basal lamina containing myelinated axons. These data have implications for the resolution of inflammation in peripheral nerve and CNS pathologies.

Matrix metalloproteinase-9 (MMP-9) is an extracellular protease that is induced hours after injury to peripheral nerve. This study shows that MMP-9 gene deletion and neutralization with MMP-9 antibody reduce macrophage content in injured wild-type nerves. In mice with delayed Wallerian degeneration (WldS), MMP-9 and tumor necrosis factor alpha (TNFalpha) decline in association with the reduced macrophage recruitment to injured nerve that characterizes this strain of mice. We further determined that TNFalpha acts as an MMP-9 inducer by establishing increased MMP-9 levels after TNFalpha injection in rat sciatic nerve in vivo and primary Schwann cells in vitro. We found reduced MMP-9 expression in crushed TNFalpha knockout nerves that was rescued with exogenous TNFalpha. Finally, local application of MMP-9 on TNFalpha-/- nerves increased macrophage recruitment to the lesion. These data suggest that TNFalpha lies upstream of MMP-9 in the pathway of macrophage recruitment to injured peripheral nerve.

The lower regeneration potential reported for C57BL/6J mice strain after peripheral nerve lesion may result from alterations in crucial events during Wallerian degeneration. We analysed neurofilament and myelin breakdown, macrophage recruitment, NADPH-diaphorase reaction and inducible nitric oxide synthase (iNOS) expression in transected sciatic nerves of C57BL/6J and A/J mice. The neurofilament volume density was lower in C57BL/6J strain mice at 1 and 3 days after lesion, and later equalled the density observed in A/J. C57BL/6J mice presented a high number of cells containing myelin debris, 3 and 5 days after the lesion. In both strains iNOS immunoreactivity was intense in macrophages and less evident in Schwann cells. However, a delay in macrophage recruitment and a lower percentage of iNOS-expressing macrophages on the third day were observed in C57BL/6J mice. NADPH-diaphorase reaction disclosed a similar pattern for both strains until the seventh day. However, at 5 days, cells with slender processes involving ellipsoid segments showed a well-defined cytoplasmic labelling in C57BL/6J whereas in A/J most of these cells exhibited a more granular and disperse labelling. We propose that these differences between A/J and C57BL/6J strains during Wallerian degeneration may be implicated in the lower regeneration potential observed in the latter.

Autoimmune demyelinating neuropathies, like Guillain-Barré syndrome (GBS) and chronic inflammatory demyelinating polyneuropathy (CIDP), are characterized by local inflammation and demyelination of peripheral nerves. The therapeutic response to plasma exchange and IVIg points to an important role for humoral factors in the pathogenesis of these autoimmune disorders of the peripheral nervous system (PNS). The similarity of the human disease with its animal model disease experimental autoimmune neuritis (EAN), and the activation of T-cell and macrophage derived cytokines all indicate that cellular immunity is also of relevance. Many of the basic principles in autoimmune inflammatory neuropathies have been learned from studies in EAN which can now be linked to the human disorders. This article summarizes current aspects of cellular immunity in autoimmune demyelinating disorders of the PNS. Better understanding of these cellular immunoregulatory mechanisms may help to develop new therapeutic strategies.

The literature concerning Schwann cells (SCs) and macrophages in myelin phagocytosis during Wallerian degeneration is reviewed. SCs carry out the first step in the removal of myelin by segmenting myelin and then incorporating the degraded myelin. The recruited macrophages then join in the myelin-phagocytosis event, appearing to make full use of their original phagocyte abilities until the end of myelin clearance. The molecular mechanisms of the two cells underlying myelin phagocytosis are thought to be different; myelin phagocytosis by SCs being lectin-mediated, i.e., opsonin-independent, whereas that of macrophages is mainly opsonin-dependent. It is important to note that SCs and macrophages cooperatively accomplish myelin phagocytosis., (Copyright 2002 Wiley-Liss, Inc.)

An association between macrophages and remyelination efficiency has been observed in a variety of different models of CNS demyelination. In order to test whether this association is causal or coincidental, we have examined the effects of macrophage depletion on the rate of remyelination of lysolecithin-induced demyelination in the spinal cord of young adult female rats. Macrophage depletion was achieved by reducing the monocyte contribution to the macrophages within the lesion using the clodronate-liposome technique. This technique not only resulted in a decrease in Ox-42-positive cells in the spleen of treated animals but also in the levels of macrophage scavenger receptor type B mRNA expression within the demyelinating lesion. In animals treated with clodronate-liposomes throughout the remyelination process, there was a significant decrease in the extent of oligodendrocyte remyelination at 3 weeks after lesion induction, but no effect on Schwann cell remyelination. If macrophage depletion was delayed until the second half of the remyelination phase, then there was no effect on the repair outcome, implying that macrophages are required for the early stages of CNS remyelination. The results of this study indicate that the macrophage response is an important component of successful CNS remyelination and that approaches to the treatment of demyelinating disease based on inhibition of the inflammatory response may also impair regenerative events that follow demyelination., (Copyright 2001 Wiley-Liss, Inc.)

Peripheral nerve injury is followed by Wallerian degeneration which is characterized by cellular and molecular events that turn the degenerating nerve into a tissue that supports nerve regeneration. One of these is the removal, by phagocytosis, of myelin that contains molecules which inhibit regeneration. We have recently documented that the scavenger macrophage and Schwann cells express the galactose-specific lectin MAC-2 which is significant to myelin phagocytosis. In the present study we provide evidence for a mechanism leading to the augmented expression of cell surface MAC-2. Nerve lesion causes noneuronal cells, primarily fibroblasts, to produce the cytokine granulocyte macrophage-colony stimulating factor (GM-CSF). In turn, GM-CSF induces Schwann cells and macrophages to up-regulate surface expression of MAC-2. The proposed mechanism is based on the following novel observations. GM-CSF mRNA was detected by PCR in in vitro and in vivo degenerating nerves, but not in intact nerves. The GM-CSF molecule was detected by ELISA in medium conditioned by in vitro and in vivo degenerating peripheral nerves as of the 4th h after injury. GM-CSF activity was demonstrated by two independent bioassays, and repressed by activity blocking antibodies. Significant levels of GM-CSF were produced by nerve derived fibroblasts, but neither by Schwann cells nor by nerve derived macrophages. Mouse rGM-CSF enhanced MAC-2 production in nerve explants, and up-regulated cell surface expression of MAC-2 by Schwann cells and macrophages. Interleukin-1 beta up-regulated GM-CSF production thus suggesting that injury induced GM-CSF production may be mediated by interleukin-1 beta. Our findings highlight the fact that fibroblasts, by producing GM-CSF and thereby affecting macrophage and Schwann function, play a significant role in the cascade of molecular events and cellular interactions of Wallerian degeneration.

The expression of HLA-DR and the macrophage marker CD 68 was studied in 44 sural nerve biopsies from patients with inflammatory and non-inflammatory neuropathies and controls using immunohistochemistry on non-osmicated semithin sections, a technique that has not been used before in such a biopsy study. Most HLA-DR-immunoreactive (ir) cells were fibroblasts, macrophages or perineural cells, some were perivascular and endothelial cells, and only few were Schwann cells. Counts of immunoreactive cells revealed (a) increased HLA-DR expression in severe as compared to less severe neuropathies and to controls, (b) no correlation between the numbers of HLA-DR-ir cells and CD 68-ir macrophages, and (c) no close correlation between diagnostic groups and the number of HLA-DR-ir cells, but higher numbers in inflammatory neuropathies. We conclude that endoneurial fibroblasts and macrophages as antigen-presenting cells may be mediators in various peripheral nerve diseases, not only in inflammatory disorders.

A unique pathogenetic process for onion-bulb (Ob) formation is disclosed with disclosed with immunohistochemistry and electron microscopy. Biopsy of a swollen segment of tibial nerve from a 42 year-old white female histologically demonstrated diffuse and angiocentric lymphocytic infiltrate in both endo- and perineurium with occasional lymphofollicular formation. Extensive Ob formation of nerve fibers was most striking with or without associated lymphocytes. Axis-cylinders were intact in the majority of Ob. Immunocytochemically, Ob are composed of alternately laminated leaflets of Schwann cells (S100+) and mononuclear macrophage (HAM56+/LeuMl+/Muramidase+) processes but no perineurial (EMA+) cells. Immunohistochemical evidence of antigen presentation (HLA-DR/LN3+/Ia+) was confined to macrophages. Electron microscopy insinuates that intricate interactions between macrophages and Schwann cells exists. Putative inhibition of remyelination along with proliferation of Schwann cells most probably is secondary to the effects of macrophages secretory products. No direct participation of B or T lymphocytes was detected in Ob. Thus, modified macrophages may emit a factor for concomitantly promoting proliferation of Schwann cells and an enzyme for myelin breakdown. In addition, only a few macrophages could be detected in some Ob and could be easily overlooked or misinterpreted as "vacuolated fibroblasts", if no immunohistochemical correlation is made, as modified macrophages making the external leaflets of Ob are more vacuolated.

This study was designed to identify which cells express major histocompatibility complex class II (Ia) antigen in experimental autoimmune neuritis and may therefore be antigen presenters. Serial 1-micron-thick cryosections of ventral roots of animals with experimental autoimmune neuritis were labeled with Ox6 antibody against rat Ia, the ED1 antibody to identify monocytes/macrophages and an antiserum against S100, a marker for Schwann cells. Ia-positive cells were predominantly present before overt clinical signs and demyelination (day 12). At later stages when many axons were demyelinated, their number was markedly reduced. Few Ia-positive cells that had extending long processes, which over some distance were in immediate contact with several myelin sheaths, were scattered in normal-appearing nerve roots at these later time points. Most of the Ia-positive cells could be identified as ED1-positive lean monocytes/macrophages, but in contrast most phagocytic macrophages in advanced stages of myelin degradation no longer expressed Ia. Ia-positive structures were invariably negative for S100 at early and late stages of experimental autoimmune neuritis, indicating that Schwann cells did not express identifiable Ia antigen. These findings contrast with reports of expression of major histocompatibility complex class II antigens by Schwann cells in human neuropathies. Furthermore they do not support the notion that aberrant Ia expression by Schwann cells plays a major pathogenic role in experimental autoimmune disease of the peripheral nervous system.