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The diagnosis of polyneuropathy (PNP) is based on the anamnesis and description of complaints of the patient and clinical findings. The type of distribution as well as known diseases and drug toxic factors can provide indications. Electromyography and electroneurography can be used to differentiate between axonal and demyelinating PNP. The laboratory examinations are initially directed towards frequent and treatable causes. These are then expanded depending on the suspected diagnosis. Analysis of cerebrospinal fluid (CSF) is facultative and should be carried out when there is a suspicion of a certain form of PNP with CSF findings indicative of the diagnosis. Nerve biopsy is indicated when the etiology of a severe or progressive PNP cannot be clarified by less invasive means and can have consequences for the treatment. A genetic investigation can be meaningful with a positive family anamnesis or with typical signs of hereditary PNP. Depending on the neuropathy and context, the diagnostic approach is structured differently. The special diagnostics for small fiber neuropathy and amyloid neuropathy as well as for diabetes and alcohol abuse are dealt with in detail in this article. Numerous cases of polyneuropathy remain unexplained and regularly have a favourable prognosis.

The diagnosis of polyneuropathy (PNP) is based on the anamnesis and description of complaints of the patient and clinical findings. The type of distribution as well as known diseases and drug toxic factors can provide indications. Electromyography and electroneurography can be used to differentiate between axonal and demyelinating PNP. The laboratory examinations are initially directed towards frequent and treatable causes. These are then expanded depending on the suspected diagnosis. Analysis of cerebrospinal fluid (CSF) is facultative and should be carried out when there is a suspicion of a certain form of PNP with CSF findings indicative of the diagnosis. Nerve biopsy is indicated when the etiology of a severe or progressive PNP cannot be clarified by less invasive means and can have consequences for the treatment. A genetic investigation can be meaningful with a positive family anamnesis or with typical signs of hereditary PNP. Depending on the neuropathy and context, the diagnostic approach is structured differently. The special diagnostics for small fiber neuropathy and amyloid neuropathy as well as for diabetes and alcohol abuse are dealt with in detail in this article. Numerous cases of polyneuropathy remain unexplained and regularly have a favourable prognosis.Die Diagnose einer Polyneuropathie (PNP) beruht auf der Anamnese und Beschwerdeschilderung des Patienten sowie dem klinischen Befund. Der Verteilungstyp sowie bekannte Erkrankungen und medikamentös-toxische Faktoren können richtungsweisend sein. Elektroneurographisch soll zwischen axonaler und demyelinisierender PNP unterschieden werden. Die Laboruntersuchungen richten sich zunächst auf häufige und behandelbare Ursachen. Je nach erarbeiteter Verdachtsdiagnose werden sie erweitert. Die Liquoranalytik ist fakultativ und sollte durchgeführt werden, wenn der Verdacht auf eine bestimmte PNP-Form mit diagnoseweisendem liquoranalytischem Befund besteht. Eine Nervenbiopsie ist indiziert, wenn bei hinreichend schwerer oder progredienter PNP die Ätiologie mit weniger invasiven Mitteln nicht geklärt werden kann und sich Konsequenzen fur die Behandlung ergeben konnen. Eine genetische Untersuchung kann bei positiver Familienanamnese für PNP oder bei typischen Zeichen einer hereditären PNP sinnvoll sein. Je nach Neuropathie und Kontext gestaltet sich das diagnostische Vorgehen anders. Auf die spezielle Diagnostik bei Small-fiber-Neuropathie und Amyloidneuropathie sowie bei Diabetes oder Alkoholmissbrauch wird im Beitrag genauer eingegangen. Zahlreiche Fälle einer PNP bleiben ungeklärt und haben regelmäßig eine günstige Prognose.

Background Studies of antigen presentation in retina using mice that expressed green fluorescent protein (GFP) from a transgenic CD11c promoter found that retinal GFP hi cells possessed antigen presentation function. Subsequent studies found that these high GFP hi cells preferentially localized to sites of retinal injury, consistent with their APC function. Interest in the roles of macrophages in degenerative CNS diseases led us to study the GFP hi cells in a retinal model of neurodegeneration. We asked if apoptotic cone photoreceptor cell death in Rpe65 −/− knockout mice induced the GFP hi cells, explored their relationship to resident microglia (MG), and tested their role in cone survival. Methods Rpe65 −/− mice were bred to CD11c GFP mice on the B6/J background. CD11c GFP Rpe65 −/− mice were also backcrossed to CX3CR1 YFP-creER ROSA DTA mice so that CX3CR1 + mononuclear cells could be depleted by Tamoxifen. Retinas were analyzed by immunohistochemistry, confocal microscopy, fluorescence fundoscopy and flow cytometry. Results Elevated numbers of GFP hi cells were concentrated in photoreceptor cell layers of CD11c GFP Rpe65 −/− mice coinciding with the peak of cone death at 2 to 4 weeks of age, and persisted for at least 14 months. After the initial wave of cone loss, a slow progressive loss of cones was found that continued to retain GFP hi cells in the outer retina. Sustained, four-week Tamoxifen depletions of the GFP hi cells and MG in Rpe65 −/− mice from day 13 to day 41, and from day 390 to day 420 promoted a small increase in cone survival. We found no evidence that the GFP hi cells were recruited from the circulation; all data pointed to a MG origin. MG and GFP hi cells were well segregated in the dystrophic retina; GFP hi cells were foremost in the photoreceptor cell layer, while MG were concentrated in the inner retina. Conclusions The expression of GFP on a subset of retinal mononuclear cells in CD11c GFP mice identified a distinct population of cells performing functions previously attributed to MG. Although GFP hi cells dominated the macrophage response to cone death in the photoreceptor cell layer, their ablation led to only an incremental increase in cone survival. The ability to identify, ablate, and isolate these cells will facilitate analysis of this activated, antigen-presenting subset of MG.

Purpose We previously demonstrated that passive transfer of lymphocytes from glaucomatous mice induces retinal ganglion cell (RGC) damage in recipient animals, suggesting a role for immune responses in the multifactorial pathophysiology of glaucoma. Here we evaluate whether absence of an adaptive immune response reduces RGC loss in glaucoma. Methods Elevated intraocular pressure (IOP) was induced in one eye of C57BL/6J (B6) or T- and B-cell-deficient Rag1-/- knockout mice. After 16 weeks RGC density was determined in both the induced and the normotensive contralateral eyes. Data were compared to mice having received injections of "empty" vector (controls). The number of extravascular CD3+ cells in the retinas was determined using FACS. Results Retinas of eyes with elevated IOP contain significantly more extravasated CD3+ cells than control retinas (46.0 vs. 27.1, P = 0.025). After 16 weeks of elevated IOP the average RGC density in B6 mice decreased by 20.7% (P = 1.9 × 10-4). In contrast, RGC loss in Rag1-/- eyes with elevated IOP was significantly lower (10.3%, P = 0.006 vs. B6). RGC loss was also observed in the contralateral eyes of B6 mice, despite the absence of elevated IOP in those eyes (10.1%; P = 0.008). In RAG1-/- loss in the contralateral eyes was minimal (3.1%) and significantly below that detected in B6 (P = 0.02). Conclusions Our findings demonstrate that T Rag1-/- mice are significantly protected from glaucomatous RGC loss. In this model, lymphocyte activity contributes to approximately half of all RGC loss in eyes with elevated IOP and to essentially all loss observed in normotensive contralateral eyes.

Using mice expressing green fluorescent protein (GFP) from a transgenic CD11c promoter we found that a controlled optic nerve crush (ONC) injury attracted GFPhi retinal myeloid cells to the dying retinal ganglion cells and their axons. However, the origin of these retinal myeloid cells was uncertain. In this study we use transgenic mice in conjunction with ONC, partial and full optic nerve transection (ONT), and parabiosis to determine the origin of injury induced retinal myeloid cells. Analysis of parabiotic mice and fate mapping showed that responding retinal myeloid cells were not derived from circulating macrophages and that GFPhi myeloid cells could be derived from GFPlo microglia. Comparison of optic nerve to retina following an ONC showed a much greater concentration of GFPhi cells and GFPlo microglia in the optic nerve. Optic nerve injury also induced Ki67+ cells in the optic nerve but not in the retina. Comparison of the retinal myeloid cell response after full versus partial ONT revealed fewer GFPhi cells and GFPlo microglia in the retina following a full ONT despite it being a more severe injury, suggesting that full transection of the optic nerve can block the migration of responding myeloid cells to the retina. Our results suggest that the optic nerve can be a reservoir for activated microglia and other retinal myeloid cells in the retina following optic nerve injury. Electronic supplementary material The online version of this article (10.1186/s40478-018-0571-8) contains supplementary material, which is available to authorized users.

© 2018 Taylor and Francis Group, LLC. In yeast, Tom22, the central component of the TOMM (translocase of outer mitochondrial membrane) receptor complex, is responsible for the recognition and translocation of synthesized mitochondrial precursor proteins, and its protein kinase CK2-dependent phosphorylation is mandatory for TOMM complex biogenesis and proper mitochondrial protein import. In mammals, the biological function of protein kinase CSNK2/CK2 remains vastly elusive and it is unknown whether CSNK2-dependent phosphorylation of TOMM protein subunits has a similar role as that in yeast. To address this issue, we used a skeletal muscle-specific Csnk2b/Ck2β-conditional knockout (cKO) mouse model. Phenotypically, these skeletal muscle Csnk2b cKO mice showed reduced muscle strength and abnormal metabolic activity of mainly oxidative muscle fibers, which point towards mitochondrial dysfunction. Enzymatically, active muscle lysates from skeletal muscle Csnk2b cKO mice phosphorylate murine TOMM22, the mammalian ortholog of yeast Tom22, to a lower extent than lysates prepared from controls. Mechanistically, CSNK2-mediated phosphorylation of TOMM22 changes its binding affinity for mitochondrial precursor proteins. However, in contrast to yeast, mitochondrial protein import seems not to be affected in vitro using mitochondria isolated from muscles of skeletal muscle Csnk2b cKO mice. PINK1, a mitochondrial health sensor that undergoes constitutive import under physiological conditions, accumulates within skeletal muscle Csnk2b cKO fibers and labels abnormal mitochondria for removal by mitophagy as demonstrated by the appearance of mitochondria-containing autophagosomes through electron microscopy. Mitophagy can be normalized by either introduction of a phosphomimetic TOMM22 mutant in cultured myotubes, or by in vivo electroporation of phosphomimetic Tomm22 into muscles of mice. Importantly, transfection of the phosphomimetic Tomm22 mutant in muscle cells with ablated Csnk2b restored their oxygen consumption rate comparable to wild-type levels. In sum, our data show that mammalian CSNK2-dependent phosphorylation of TOMM22 is a critical switch for mitophagy and reveal CSNK2-dependent physiological implications on metabolism, muscle integrity and behavior.

Extrathymically derived regulatory T cells (iTregs) protect against autoimmunity to tissue-specific antigens (Ag). However, whether Ag-specific iTreg generation and function is limited to secondary lymphoid tissue or whether it can occur within the tissue-specific, local environment of the cognate Ag remains unresolved. Mice expressing beta-galactosidase (βgal) on a retina specific promoter (βgal mice) in conjunction with mice expressing green fluorescent protein (GFP) and diphtheria toxin (DTx) receptor (DTR) under control of the FoxP3 promoter, and βgal-specific T cell receptor transgenic (BG2) mice were used to examine this question. Local depletion (ocular DTx), but not systemic depletion (intra-peritoneal DTx), of βgal-specific iTregs enhanced experimental autoimmune uveoretinitis induced by activated βgal-specific effector T cells. Injections of small amounts of βgal into the anterior chamber of the eye produced similar numbers of βgal-specific iTregs in the retina whether or not the mouse was depleted of pre-existing, circulating Tregs. Together these results suggest that protection from tissue-specific autoimmunity depends on the function of local Ag-specific iTregs, and that the retina is capable of local, “on-demand” iTreg generation that is independent of circulating Tregs.

Phosphoprotein 8-50, also termed growth-associated protein GAP43, is a membrane-bound phosphoprotein expressed in neurons. It is particularly abundant during periods of axonal outgrowth in development and regeneration of the central and peripheral nervous systems. Recently the expression of phosphoprotein 8-50 in regenerating muscle fibers was reported and it was assumed that phosphoprotein 8-50, in muscles, plays a role in the determination of the growth morphology of regenerating muscle fibers. Thus, phosphoprotein 8-50 no longer can be regarded as a neuron-specific molecule. In this paper we study the subcellular localization of phosphoprotein 8-50 in regenerating human skeletal muscle fibers by electron immunohistochemistry. Phosphoprotein 8-50 immunoreactivity is randomly distributed over the nuclear and perinuclear area of regenerating muscle fibers. Previously, by light-microscopy phosphoprotein 8-50 immunoreactivity was demonstrated on the inner face of the sarcolemma in hypotrophic type 1 fibers in congenital fiber type disproportion. It is this distribution of phosphoprotein 8-50 in developmentally disordered myocytes in particular which allows an analogy to the corresponding results found for growing axons. But we did not find subsarcolemmal expression of 8-50 in regenerating muscle fibers. Probably this indicates that there is a difference in 8-50 expression between regeneration of muscle fibers and developmentally retarded/immature myofibers. The presented data suggest that phosphoprotein 8-50 is inserted only stage-dependent in the extending sarcolemma of the growing muscle fiber. Analogously to the nervous system, phosphoprotein 8-50 may serve a local function involving transmembrane signalling by means of calmodulin binding ofphosphoprotein 8-50 and/or phosphoprotein 8-50 phosphorylation and dephosphorylation. [Neural Res 1998; 20: 360-364].

Interest in the identities, properties, functions, and origins of local APC in CNS tissues is growing. We recently reported that dendritic cells (DC) distinct from microglia were present in quiescent retina and rapidly responded to injured neurons. In this study, the disease-promoting and regulatory contributions of these APC in experimental autoimmune uveoretinitis (EAU) were examined. Local delivery of purified, exogenous DC or monocytes from bone marrow substantially increased the incidence and severity of EAU induced by adoptive transfer of activated, autoreactive CD4 or CD8 T cells that was limited to the manipulated eye. In vitro assays of APC activity of DC from quiescent retina showed that they promoted generation of Foxp3+ T cells and inhibited activation of naive T cells by splenic DC and Ag. Conversely, in vitro assays of DC purified from injured retina demonstrated an enhanced ability to activate T cells and reduced induction of Foxp3+ T cells. These findings were supported by the observation that in situ activation of DC before adoptive transfer of β-galactosidase–specific T cells dramatically increased severity and incidence of EAU. Recruitment of T cells into retina by local delivery of Ag in vivo showed that quiescent retina promoted development of parenchymal Foxp3+ T cells, but assays of preinjured retina did not. Together, these results demonstrated that local conditions in the retina determined APC function and affected the pathogenesis of EAU by both CD4 and CD8 T cells.

Regulatory T cells (Tregs) are generated to antigens (Ag) found in the retina. Some Tregs are the result of ectopic expression of the retinal Ags in the thymus, where developing T cells are committed to enter the regulatory lineage. However, the generation of retinal Ag-specific Tregs independent of the thymus was uncertain. Our studies show that Tregs can be generated from mature, peripheral T cells based on exposure to retinal Ags. These peripherally induced Tregs limited immune responses and experimental autoimmune disease induced by retinal Ags and thus constitute a crucial component of retinal immune privilege.

Dermatomyositis and vasculitic neuropathies are disorders with immune mediated ischemic injuries. Cellular responses to hypoxia include the hypoxia-inducible factor-1 (HIF-1)-induced transcription of genes involved in angiogenesis. To study their possible roles in those disorders, the immunohistochemical expression of HIF-1alpha, HIF-1beta, HIF-2alpha, vascular endothelial growth factor (VEGF), VEGF-receptor (VEGF-R) and erythropoietin-receptor was investigated. Cases of normal nerves, diabetic neuropathy, normal muscles, polymyositis and inclusion-body-myositis served as controls. The latter were chosen because they represent comparable inflammatory disorders, however, in these ischemia/hypoxia is not supposed to play such a prominent pathogenetic role. Hypoxia-related proteins were not detected in normal controls. In polymyositis and inclusion-body-myositis, there was VEGF-R-expression in muscle fibers and HIF-2alpha reactivity in endothelial cells. In dermatomyositis, HIF-1alpha and HIF-1beta were found in endothelial cells, whereas HIF-2alpha, erythropoietin-receptor, VEGF and VEGF-R additionally were observed in muscle fibers. In vasculitic and diabetic neuropathies, a variable focal expression of hypoxia-inducible factors, VEGF, VEGF-R and erythropoietin-receptor was seen in vessels. These observations suggest that the upregulation of hypoxia-related proteins may represent an adaptation mechanism of neuromuscular tissues to immune mediated deprivation of the blood supply.

The contribution of peripheral expression of tissue-specific CNS Ags to the generation of tolerance is uncertain. To study this question, we examined mice transgenic (Tg) for expression of β-galactosidase (βgal) on the retinal photoreceptor cell arrestin promoter, in conjunction with TCR Tg mice producing CD4+ T cells specific for βgal (βgalTCR). Several strategies were used to test the hypothesis that βgal expressed in the retina supported thymus-independent tolerance and regulatory T cell development. Retinal expression generated an immunoregulatory response that depressed development of immune responses to βgal following systemic immunization with βgal. This regulation was transferable to naive mice by CD3+4+25+ T cells from naive retinal βgal+ donors. Experiments that removed the βgal+ retina by enucleation showed that subsequent development of a regulatory response was lost. Adoptive transfer of CD25− βgalTCR T cells into retinal βgal Tg mice on the Rag−/− background led to regulatory activity that limited lymphopenia-induced proliferation of βgalTCR T cells in mice with retinal expression of βgal and inhibited the ear-swelling assay for delayed type hypersensitivity. These results show that retinal expression of very small amounts of a tissue-specific Ag can generate tolerance that includes regulatory T cells.

Objective: To describe the clinical, electrophysiologic and morphologic features of a Costa Rican family with an autosomal dominant inherited Charcot–Marie–Tooth (CMT) neuropathy. Methods: The field study took place in Costa Rica, Central America. Seven patients underwent neurological examinations and standard electrodiagnostic tests, and a sural nerve biopsy was taken from one patient. Fifteen family members were screened for gene defects associated with CMT disease. Results: Characteristic features of this family were a late age of onset (35–56 years), positive sensory symptoms and muscle cramps. Based on electrodiagnostic and morphologic data, the patients were classified as having a CMT2 neuropathy. The CMT1A duplication/HNPP deletion and point mutations in genes PMP22, MPZ, Cx32 and EGR2 implicated in the most common types of CMT disease were excluded. Subsequently, almost all known CMT loci were excluded by linkage analysis. Discussion: Features of this family were a late age of onset and positive sensory symptoms. This new autosomal dominant CMT neuropathy is associated with an unknown gene defect. [Neurol Res 2009; 31: 283–288] Objetivo: Describir las características clínicas, electrofisiológicas y morfológicas de una familia costarricense con una neuropatía de Charcot-Marie-Tooth (CMT) hereditaria autosómica dominante. Métodos: El estudio de campo se llevó a cabo en Costa Rica, Centroamérica. Siete pacientes se sometieron a exámenes neurológicos y pruebas de electrodiagnóstico estándar, ya un paciente se le tomó una biopsia del nervio sural. Quince miembros de la familia fueron evaluados por defectos genéticos asociados con la enfermedad de CMT. Resultados: Los rasgos característicos de esta familia fueron una edad de inicio tardía (35-56 años), síntomas sensoriales positivos y calambres musculares. Sobre la base de los datos morfológicos y de electrodiagnóstico, los pacientes se clasificaron con una neuropatía CMT2. Se excluyeron la duplicación de CMT1A/deleción HNPP y las mutaciones puntuales en los genes PMP22, MPZ, Cx32 y EGR2 implicados en los tipos más comunes de enfermedad de CMT. Posteriormente, casi todos los loci CMT conocidos fueron excluidos por análisis de ligamiento. Discusión: Las características de esta familia fueron una edad de inicio tardía y síntomas sensoriales positivos. Esta nueva neuropatía de CMT autosómica dominante está asociada con un defecto genético desconocido. [NeurolRes 2009; 31: 283–288] Universidad Nacional, Costa Rica. Escuela de Ciencias Biológicas

To study retinal immunity in a defined system, a CD4+ TCR transgenic mouse line (βgalTCR) specific for β-galactosidase (βgal) was created and used with transgenic mice that expressed βgal in retinal photoreceptor cells (arrβgal mice). Adoptive transfer of resting βgalTCR T cells, whether naive or Ag-experienced, into arrβgal mice did not induce retinal autoimmune disease (experimental autoimmune uveoretinitis, EAU) and gave no evidence of Ag recognition. Generation of βgalTCR T cells in arrβgal mice by use of bone marrow grafts, or double-transgenic mice, also gave no retinal disease or signs of Ag recognition. Arrβgal mice were also resistant to EAU induction by adoptive transfer of in vitro-activated βgalTCR T cells, even though the T cells were pathogenic if the βgal was expressed elsewhere. In vitro manipulations to increase T cell pathogenicity before transfer did not result in EAU. The only strategy that induced a high frequency of severe EAU was transfer of naive, CD25-depleted, βgalTCR T cells into lymphopenic arrβgal recipients, implicating regulatory T cells in the T cell inoculum, as well as in the recipients, in the resistance to EAU. Surprisingly, activation of the CD25-depleted βgalTCR T cells before transfer into the lymphopenic recipients reduced EAU. Taken together, the results suggest that endogenous regulatory mechanisms, as well as peripheral induction of regulatory T cells, play a role in the protection from EAU.

It is well known that immunoproteasome generates peptides for MHC Class I occupancy and recognition by cytotoxic T lymphocytes (CTL). The present study focused on evidence for alternative roles for immunoproteasome. Retina and brain were analyzed for expression of immunoproteasome subunits using immunohistochemistry and western blotting under normal conditions and after injury/stress induced by CTL attack on glia (brain) or neurons (retina). Normal retina expressed substantial levels of immunoproteasome in glia, neurons, and retinal pigment epithelium. The basal level of immunoproteasome in retina was two-fold higher than in brain; CTL-induced retinal injury further up-regulated immunoproteasome expression. Immunoproteasome up-regulation was also observed in injured brain and corresponded with expression in Purkinje cells, microglia, astrocytes, and oligodendrocytes. These results suggest that the normal environment of the retina is sufficiently challenging to require on-going expression of immunoproteasome. Further, immunoproteasome up-regulation with retinal and brain injury implies a role in neuronal protection and/or repair of damage.

Motoneurons release the heparansulfate proteoglycan agrin and thereby activate the muscle-specific receptor tyrosine kinase (MuSK), which is the main organizer of subsynaptic specializations at the neuromuscular junction. Recently, we showed that (1) the protein kinase CK2 interacts with the intracellular region of MuSK; (2) the CK2 protein is enriched and co-localized with MuSK at postsynaptic specializations; (3) CK2-mediated phosphorylation of serine residues within a specific MuSK epitope, named the kinase insert, regulates acetylcholine receptor (AChR) clustering; (4) muscle-specific CK2beta knockout mice develop a myasthenic phenotype due to impaired muscle endplate structure and function (see Genes Dev 20(13):1800-1816, 2006). Here, we investigated for the first time if CK2 is modulated in biopsies from human patients. To this end, we measured transcript amounts of the subunits CK2alpha and CK2beta and determined holoenzyme CK2 activity in 34 muscle biopsies of human patients with different muscle pathologies.

Background: Thymic expression of a photoreceptor cell antigen, interphotoreceptor retinoid-binding protein, is known to generate regulatory T cells (Treg) that prevent spontaneous autoimmune disease of the retina. However, the contribution of other endogenous, tissue-specific antigens (Ags) expressed in the retina to the generation of Treg is uncertain. Methods: Transgenic mice that express β-galactosidase (β-gal) in photoreceptor cells, together with β-gal-specific T cell receptor transgenic mice, were used to study the induction of Treg in vivo. Results: Transgenic expression of β-gal on the arrestin promoter led to a spontaneous immunoregulatory response that inhibited the development of immune responses to β-gal. The regulation was transferred by CD3+4+25+ Treg. Several strategies were then used to show that β-gal expressed in the retina supported spontaneous, thymus-independent Treg development. The endogenous Treg also differed from the Treg induced by Ag inoculation into the anterior chamber of the eye. Conclusion: These results demonstrate that retinal expression of very small amounts of a tissue-specific Ag can generate Treg in the periphery.

To explore pathways to neuron loss in inflammatory diseases, transgenic mice expressing beta-galactosidase (beta-gal) in either astrocytes or photoreceptor cells, or both, were inoculated with activated, beta-gal-specific cytotoxic CD8 T lymphocytes (CTLs). beta-gal-positive astrocytes in brain were rapidly attacked, with particular damage in cerebellum. Substantial loss of cerebellar granule cells was found, even though these neurons did not express beta-gal. The small number of beta-gal-positive retinal astrocytes present in these mice was also rapidly destroyed by transferred CTLs, but without detectable consequences for retinal neurons. However, in mice with photoreceptor cell-specific beta-gal expression, near-total destruction of photoreceptor cells was produced by CTL transfer. Attack on photoreceptor cells displayed minimal inflammation, and onset was a week later than onset of astrocyte-directed disease. CTL transfer into F1 mice expressing beta-gal in both astrocytes and photoreceptor cells confirmed that pathogenesis directed against antigen expressed in glia versus neurons proceeded in two distinct, independent phases. beta-gal-positive retinal astrocytes were severely affected by 5 days post-transfer, followed by rapid resolution. Photoreceptor cells in the same retina were unaffected until 12 days post-transfer. The susceptibility of photoreceptor cells was not enhanced by the prior CTL attack on beta-gal-expressing retinal astrocytes. The results demonstrate that extensive bystander killing of neurons can occur in vivo as a result of direct CTL attack on surrounding astrocytes. Antigen-expressing retinal neurons were also efficiently killed by CTLs, but by a mechanism that was substantially delayed and dissociated from the killing of retinal astrocytes.

Background We previously reported that the peripheral regulatory T cells (pTregs) generated ‘on-demand’ in the retina were crucial to retinal immune privilege, and in vitro analysis of retinal dendritic cells (DC) showed they possessed antigen presenting cell (APC) activity that promoted development of the Tregs and effector T cells (Teffs). Here, we expanded these findings by examining whether locally generated, locally acting pTregs were protective against spontaneous autoimmunity and autoimmunity mediated by interphotoreceptor retinoid-binding protein (IRBP). We also examined the APC capacity of retinal DC in vivo. Methods Transgenic (Tg) mice expressing diphtheria toxin receptor (DTR) and/or green fluorescent protein (GFP) under control of the endogenous FoxP3 promoter (GFP only in FG mice, GFP and DTR in FDG mice) or the CD11c promoter (GFP and DTR in CDG mice) were used in conjunction with Tg mice expressing beta-galactosidase (βgal) as retinal neo-self antigen and βgal-specific TCR Tg mice (BG2). Retinal T cell responses were assayed by flow cytometry and retinal autoimmune disease assessed by histological examination. Results Local depletion of the Tregs enhanced actively induced experimental autoimmune uveoretinitis to the highly expressed retinal self-antigen IRBP in FDG mice and spontaneous autoimmunity in βgal-FDG-BG2 mice, but not in mice lacking autoreactive T cells or their target antigen in the retina. The presence of retinal βgal downregulated the generation of antigen-specific Teffs and pTregs within the retina in response to local βgal challenge. Retinal DC depletion prevented generation of Tregs and Teffs within retina after βgal injection. Microglia remaining after DC depletion did not make up for loss of DC-dependent antigen presentation. Conclusions Our results suggest that local retinal Tregs protect against spontaneous organ-specific autoimmunity and that T cell responses within the retina require the presence of local DC.

Oxidative stress and nuclear factor-kappaB (NF-kappaB) activation are linked to the pathogenesis of many metabolic, degenerative, and chronic inflammatory diseases. Activation of the receptor for advanced glycation end products (RAGE) by its specific ligand N(epsilon)-carboxymethyllysine (CML) results in the activation of NF-kappaB and the production of proinflammatory cytokines. To determine whether engagement of RAGE contributes to the pathogenesis of inflammatory myopathies, we performed immunohistochemical studies on the presence of CML-modified proteins, RAGE and activated NF-kappaB in muscle biopsies of patients with polymyositis (PM, n=10), dermatomyositis (DM, n=10), limb girdle muscular dystrophy (LGMD, n=10) and in 10 controls with normal muscle biopsy results. In inflammatory myopathies CML, RAGE and NF-kappaB were detected in mononuclear cells and in regenerating muscle fibers. CML, NF-kappaB and, to a lesser extent, RAGE were also found in degenerating muscle fibers, but colocalization of CML, RAGE and NF-kappaB was only seen in infiltrating mononuclear cells and regenerating muscle fibers. Immunofluorescence double labeling demonstrated an expression of CML, RAGE and NF-kappaB in CD4-, CD8-, CD22- and CD68-positive mononuclear cells. Western blot analysis showed an increased immunoreactivity for CML-modified proteins in PM and DM. In LGMD, CML, RAGE and NF-kappaB were found in regenerating muscle fibers and less frequently in degenerating muscle fibers, and with lower staining intensities than in inflammatory myopathies. Our data suggests that the CML-RAGE-NF-kappaB pathway is an evident proinflammatory pathomechanism in mononuclear effector cells in PM and DM. RAGE-mediated NF-kappaB activation may be involved in muscle fiber regeneration in inflammatory myopathies and LGMD.

ZusammenfassungMyalgien können als unspezifisches Symptom bei einer Vielzahl neurologischer und anderer Erkrankungen auftreten. Häufig sind diese Schmerzen nicht durch eine primäre Schädigung des Skelettmuskels selbst verursacht, sondern Begleitsymptome unterschiedlicher Erkrankungen. Primäre muskuläre Ursachen müssen jedoch bedacht werden, um entsprechend zu beraten und eine angemessene Therapie anbieten zu können.In der vorliegenden Arbeit werden Myopathien und primär nicht muskuläre Erkrankungen im Hinblick auf die Differenzialdiagnose von Muskelschmerzen vorgestellt.

ZusammenfassungLokalisierte oder generalisierte Schmerzen der Skelettmuskulatur sind häufige Leit- oder Begleitsymptome von Erkrankungen aus verschiedenen medizinischen Fachdisziplinen. Die ätiologische Abklärung von Myalgien erfolgt anhand des Beschwerdebildes, des klinischen Untersuchungsbefundes und gegebenenfalls weiterführender Untersuchungen. Die vorliegende Arbeit stellt alle derzeitigen diagnostischen Möglichkeiten vor sowie einen Algorithmus für die zielgerichtete Diagnostik bei Myalgien.

The pathogenesis of the inflammatory myopathies is still unclear, making their treatment largely empirical. Improved understanding of the molecular mechanisms of inflammatory muscle injury may, however, lead to the development of more specific immunotherapies. To elucidate a possible pathogenic contribution of calcium-binding proteins such as the annexins, we immunohistochemically investigated muscle biopsy specimens from patients with dermatomyositis (10 cases), polymyositis (9 cases), and inclusion-body myositis (4 cases), compared to control cases comprising sarcoid myopathy (3 cases), Duchenne muscular dystrophy (DMD; 4 cases), and normal muscle (3 cases). We found expression of annexins A1, A2, A4, and A6 in the vascular endothelium of all cases. Myofibers expressed annexins A5, A6, and A7 diffusely and weakly in the cytosol, whereas annexins A5 and A7 were also particularly localized to the sarcolemma. In the inflammatory myopathies, in areas of myonecrosis in DMD, and in granulomatous lesions of sarcoid myopathy, reactivity of annexins A1, A2, A4, A5, and A6 was observed in macrophages and T-lymphocytes. Whereas the latter annexins appear to be nonspecific indicators of activation, annexin A1 upregulation may represent endogenous anti-inflammatory mechanisms that merit further investigation.

Binding of ligands to the receptor for advanced glycation endproducts (RAGE) results in activation of the proinflammatory transcription factor nuclear factor-kappaB (NF-kappaB) and subsequent expression of NF-kappaB-regulated cytokines. In order to determine whether engagement of RAGE contributes to the pathogenesis of vasculitic neuropathy, we studied the presence of the RAGE ligand N(epsilon)-(carboxymethyl)lysine (CML), the receptor itself, NF-kappaB, and interleukin-6 (IL-6) in sural nerve biopsies of 12 patients with vasculitic neuropathies and 12 controls. In the patients, CML, RAGE, NF-kappaB, and IL-6 were localized in mononuclear cells, epineurial and endoneurial vessels and the perineurium. CML, RAGE, NF-kappaB, and IL-6 were expressed by CD4(+), CD8(+), and CD68(+) cells invading the nerves. Controls showed only weak staining. These data suggest that the RAGE pathway plays a critical proinflammatory role in vasculitic neuropathy.

Prostaglandins (PGs) are critical mediators of physiologic processes and inflammation. They are produced by two different isoforms of the cyclooxygenase (COX) enzyme, namely COX-1 and COX-2. In particular COX-2 was demonstrated to be crucial for PG-synthesis in inflammation. Recently, inhibition of COX-2 was shown to prevent the loss of motor neurons in a model of amyotrophic lateral sclerosis (ALS). Furthermore, spinal COX-2 expression was shown to be increased in transgenic mice that produce an ALS-like syndrome. Therefore, we investigated the expression of COX-1 and COX-2 in the spinal cord of seven human sporadic ALS patients by means of immunohistochemistry. Specimens from seven patients without any neurological disease served as controls. COX-2 expression was dramatically increased in the spinal cord of patients with ALS. Its protein was found in motor neurons, interneurons and glial cells. Statistical analysis showed a significantly higher expression of COX-2 in ALS for both neurons and glia. In contrast, COX-1 expression was predominantly confined to microglia and no apparent difference was detected between controls and ALS. In addition, we studied the concentration of prostaglandin E2 (PG E2) as a marker for COX activity in the cerebrospinal fluid of nine patients diagnosed for ALS and compared the results with those from nine patients without motor neuron disease. PG E2 levels were markedly increased in ALS cases (45.8 +/- 35.1 pg/mL) compared to the non-ALS specimens (15.8 +/- 3.7 pg/mL). The results of our study corroborate a potential role for COX-2 in the pathogenesis of motor neuron death in ALS. Selective COX-2 inhibition might therefore offer a new possibility in the treatment of human ALS. However, to determine the exact role of COX-2 in human ALS will require further research.

Vasculitic neuropathies are immune mediated diseases of the peripheral nervous system, in which inflammation of the blood vessels causes damage to the nerves. We distinguish neuropathies associated with primary and secondary systemic vasculitis, with rheumatic diseases, with malignant disorders, drug-induced vasculitis and the non-systemic vasculitic neuropathies (NSVN). The typical clinical picture consists in an asymmetric or multifocal, painful sensorimotor neuropathy with an acute, sub-acute or chronic course and acute relapses. Neurophysiology reveals an active, asymmetric, axonal sensorimotor neuropathy. The disorders usually respond to immunosuppressive treatment. A diagnosis of definite vasculitis can be made with evidence of vasculitis in a biopsy specimen. The absence of positive morphological evidence, however, does not exclude the diagnosis. There is no single laboratory test that can prove or exclude vasculitis, in NSVN even an elaborate panel of blood tests can show normal findings. Systemic vasculitis has an incidence of 4/100000 per year and, untreated, has a poor prognosis, which is greatly improved by the use of immunosuppressive treatment. The prognosis of NSVN is generally better, although many patients need long term immunosuppression. Current treatment recommendations for vasculitic neuropathies are presented.

ZusammenfassungUnter vaskulitischen Neuropathien versteht man immunvermittelte Erkrankungen des peripheren Nervensystems, bei denen die Schädigung der Nerven durch eine Entzündung der Blutgefäße bedingt ist. Unterschieden werden vaskulitische Neuropathien bei primären und sekundären systemischen Vaskulitiden, bei entzündlichrheumatischer Systemerkrankung, bei malignen Erkrankungen, medikamenteninduzierten Vaskulitiden und die nicht-systemische vaskulitische Neuropathie (NSVN). Das typische klinische Erscheinungsbild einer vaskulitischen Neuropathie ist eine asymmetrische oder multifokale schmerzhafte sensomotorische Neuropathie mit akutem/subakutem oder chronischem Verlauf mit schubförmigen Verschlechterungen. Charakteristisch sind weiterhin der elektrophysiologische Nachweis einer aktiven asymmetrischen axonalen sensomotorischen Neuropathie und die Besserung durch immunsuppressive Therapie. Eine gesicherte Vaskulitis liegt vor, wird ein positiver Biopsiebefund erhoben; das Fehlen eindeutiger morphologischer Befunde schließt sie jedoch nicht aus. Es gibt keinen Labortest, der eine Vaskulitis definitiv nachweist oder ausschließt, so ist auch bei ausschließlich unauffälligen Ergebnissen von durchaus umfangreichen Blutuntersuchungen das Vorliegen einer NSVN möglich. Systemische Vaskulitiden treten mit einer Inzidenz von mindestens 4/100000 jährlich auf und haben unbehandelt eine schlechte Prognose, die durch immunsuppressive Therapie entscheidend verbessert wird. Bei der NSVN ist die Prognose deutlich besser, eine längerfristige immunsuppressive Therapie ist auch hier häufig erforderlich.

The iron Lewis acid catalyst, [(η5-C5H5)Fe(CO)2(THF)]+[BF4]−, has previously been found to catalyze a growing array of synthetic transformations. Building upon these precedents, the utility of the catalyst has now been extended to catalytic reaction involving olefins and (N-(p-tosyl)imino)phenyliodinane, PhI=NTs, which serves as a nitrenoid precursor, to produce aziridines with yields up to 85%.

We studied the occurrence of Ne-(carboxymethyl)lysine (CML), the major in vivo Maillard product in nerve and muscle in various inflammatory and noninflammatory diseases using a previously characterised specific CML antiserum. We found an increased CML staining in the perineurium of biopsies from B12-deficient, diabetic and vasculitic neuropathy and to a lesser extent, in alcoholic neuropathy and in mononuclear inflammatory cells. In diabetic neuropathy CML, the receptor for advanced glycation endproducts (RAGE) and nuclear factor kappa B (activated NFκB) were colocalised indicating possible interaction. In myositis, intracellular CML depositions were observed in infiltrating monocytes, myoblasts and necrotic fibers and Western blot analysis showed a marked increase of CML-modified proteins. To evaluate the impact of inflammation-associated cells on CML formation, we designed in vitro experiments using granulocytes and the monocytic cell line Mono-Mac6 (MM6). Oxidative burst of zymosan-stimulated granulocytes led to CML modification of extracellular proteins within 60–120 min. Predifferentiated MM6 cells, exerting NADPH oxidase activity but lacking myeloperoxidase activity, showed CML modification of distinct intracellular proteins within 1–3 days upon stimulation. Our findings suggest that CML modification occurs in vivo and in vitro when oxidative stress from e.g. inflammatory cells is present, indicating that CML may represent a marker for oxidative stress.

In amyotrophic lateral sclerosis (ALS) patients, hyperintense signals at the subcortical precentral gyrus in brain fluid attenuated inversion recovery (FLAIR) MR images have been found more frequently than in controls. Quantitative analysis has revealed a significant increase of the FLAIR-magnetic resonance imaging (MRI) signal at the subcortical precentral gyrus of ALS patients compared to healthy controls. In addition, hypointense signals at the rim of the precentral gyrus in FLAIR and T2-weighted images have been shown in ALS patients. In 17 ALS patients, we evaluated hyperintense signals in T2-, T1-, proton density-weighted and FLAIR MR images, and hypointense signals in T2-weighted and FLAIR images 15.7+/-3.0 months after the initial examination by visual scoring. In FLAIR images, a quantitative analysis was added. The visual scores of hyperintense signals along the corticospinal tract did not change significantly in all sequences. However, the quantitative evaluation of FLAIR images revealed a significant increase of the signal intensity at the subcortical precentral gyrus (p

Amyotrophic lateral sclerosis is a rapidly progressive and fatal disease which has no known cure and limited symptomatic treatment. While coping strategies in more common diseases are widely assessed, coping is poorly understood in ALS.We examined 41 ALS patients using a standardised interview, a validated coping self-rating questionnaire and a self-rating depression scale. The evaluation was repeated after six months."Loss of speech", "loss of mobility" and "the poor prognosis" were the most frequent answers in the standardised interview to questions regarding the worst aspect of the disease. Pain was seldom mentioned. "Family members" were most helpful in coping with the disease, followed by "unspecific mechanisms" and "technical aids". None of our patients expressed a wish for assisted suicide. In comparison with other fatal diseases, patients with ALS had similar rankings in the coping mechanism of "rumination", but lower rankings in "search for social integration", "defence of fear", "search for information and communication". In contrast,* "search for hold in the religion" was of high importance for our ALS patients. In the follow-up examination the importance of "search for information and communication" increased.The results emphasise the importance of "loss of speech" and the importance of the caring family as well as the availability of technical aids in ALS. Coping in ALS seems to be based mainly on "rumination" and *"hold in the religion", but the increasing importance of "search for information" indicates that the sustained offer of information is essential.