Masatoshi Omoto

Sera from patients with multifocal motor neuropathy disrupt the blood-nerve barrier

Objective In multifocal motor neuropathy (MMN), the destruction of the blood-nerve barrier (BNB) has been considered to be the key step in the disease process. The purpose of the present study was to ascertain whether sera from patients with MMN can open the BNB, and which component of patient sera is the most important for this disruption. Methods We evaluated the effects of sera from patients with MMN, patients with amyotrophic lateral sclerosis, and control subjects on the expression of tight junction proteins and vascular cell adhesion molecule-1 (VCAM-1), and on the transendothelial electrical resistance (TEER) in human peripheral nerve microvascular endothelial cells (PnMECs). Results The sera from patients with MMN decreased the claudin-5 protein expression and the TEER in PnMECs. However, this effect was reversed after application of an anti-vascular endothelial growth factor (anti-VEGF) neutralising antibody. The VEGF secreted by PnMECs was significantly increased after exposure to the sera from patients with MMN. The sera from patients with MMN also increased the VCAM-1 protein expression by upregulating the nuclear factor kappa-B (NF-κB) signalling. The immunoglobulin G purified from MMN sera decreased the expression of claudin-5 and increased the VCAM-1 expression in PnMECs. Conclusions The sera from MMN patients may disrupt the BNB function via the autocrine secretion of VEGF in PnMECs, or the exposure to autoantibodies against PnMECs that are contained in the MMN sera. Autoantibodies against PnMECs in MMN sera may activate the BNB by upregulating the VCAM-1 expression, thereby allowing for the entry of a large number of circulating inflammatory cells into the peripheral nervous system.

NMO sera down-regulate AQP4 in human astrocyte and induce cytotoxicity independent of complement

Autoantibodies against astrocyte water channel aquaporin-4 (AQP4) are highly specific for neuromyelitis optica (NMO). However, the molecular mechanism of NMO still remains unclear. The purpose of this study was to identify the possible humoral mechanisms responsible for the occurrence of astrocytic damage. Human primary astrocytes (AST) were immortalized by retroviral vectors harboring temperature-sensitive SV40 T antigen gene and AQP4 cDNA (M23), designated as hAST-AQP4. The effects of NMO sera on the content and localization of AQP4, including cytotoxicity and astrocytic morphology, were evaluated. In addition, this study examined whether the amount and localization of AQP4 protein in astrocytes were influenced by direct contact with the immortalized human brain microvascular endothelial cell line, TY09. NMO sera alone induced cytotoxicity and addition of complement had a more harmful effect on hAST-AQP4. NMO sera also decreased AQP4 mRNA and protein. NMO sera alone up-regulated TNFα and IL-6 in astrocytes and co-incubation with anti-TNFα and anti-IL-6 neutralizing antibodies blocked both the cytotoxicity and reduction of AQP4 in astrocytes. In the experiment using the in vitro BBB models, AQP4 protein mainly localized at the astrocytic membrane after co-culture with TY09, in contact with TY09. The future elucidation of factors that up-regulate AQP4 in astrocytes presumably released by blood brain barrier forming endothelial cells and that block the production of inflammatory cytokines may therefore lead to the development of a novel therapeutic strategy.

Pathological study on amyloidosis in Cygnus olor (mute swan) and other waterfowl

Between 2004 and 2007, we examined a total of 70 waterfowl. Forty of 51 (78.4%) mute swans (Cygnus olor) had amyloidosis. Amyloid deposits were detected in the spleen of 39 of 49 birds (79.6%), liver of 37 of 47 birds (78.7%), intestine of 38 of 50 birds (76.0%), pancreas of 30 of 42 birds (71.4%), kidney of 32 of 47 birds (68.1%), thyroid gland of 20 of 30 birds (66.7%), heart of 26 of 49 birds (53.1%), and lung of 5 of 45 birds (11.1%). In some birds, there was a globular pattern of amyloid deposition or infiltration of foreign-body giant cells around amyloid nodules in the spleen. Immunostaining with anti-AA antibody and Western blotting revealed that all were cases of AA amyloidosis. In sections treated with potassium permanganate, which removes Congo red stain, the green refringence under polarized light had mostly vanished. However, staining was not completely eliminated in some areas. Electron microscopy confirmed that the star-shaped amyloid fibrils were 10 nm in diameter and lacked branching. We also demonstrated amyloid bundles and star-shaped amyloid fibrils. A high percentage (96.3%) of mute swans had an inflammatory condition known as bumblefoot. Swans are useful model for studies of animals that have high amounts of amyloid. This research may help in the elucidation of the mechanism of amyloidogenesis in humans, and more research regarding amyloidosis in birds that are consumed as food is necessary.

Colocalization of Apolipoprotein AI in Various Kinds of Systemic Amyloidosis

Apolipoprotein AI (apoAI), a major component of high-density lipoproteins, is one of the major amyloid fibril proteins and a minor constituent of the senile plaques observed in Alzheimers disease. We examined colocalization of apoAI in various kinds of systemic amyloidosis in this study. Forty-three of 48 formalin-fixed paraffin-embedded heart specimens with various forms of systemic amyloidosis reacted immunohistochemically with anti-human apoAI antibody. ApoAI was also detected in water-extracted amyloid material by immunoblotting. In addition, we observed colocalization of apoAI and murine amyloid A (AA) amyloidosis in human apoAI transgenic mice. This is the first report of colocalization of apoAI with amyloid deposits in various forms of human systemic amyloidosis and murine AA amyloidosis in human apoAI transgenic mice. ApoAI may not always be a major component of amyloid fibrils, even when it is present in systemic amyloid deposits.

Rituximab-associated progressive multifocal leukoencephalopathy derived from non-Hodgkin lymphoma: neuropathological findings and results of mefloquine treatment.

A 66-year-old man with non-Hodgkin lymphoma (NHL) developed progressive multifocal leukoencephalopathy (PML) after undergoing chemotherapy including rituximab. Although the administration of mefloquine at a dose of 500 mg weekly temporarily led to a dramatic decrease in the copy number of JC Virus DNA in the cerebrospinal fluid, the patients symptoms gradually worsened. The CD4(+) T count remained continuously low, at least until approximately five months after the last cycle of chemotherapy. A postmortem examination performed 10 months after the onset of PML disclosed a severe condition associated with rituximab-treated PML originating from NHL and a high mefloquine concentration in the brain. The accumulation of further data regarding mefloquine treatment in PML cases may help to elucidate the optimal dosage and time window for effectively treating PML.

Inactivation of amyloid-enhancing factor (AEF) : study on experimental murine AA amyloidosis

It is known that amyloid-enhancing factor (AEF) shortens the preamyloid phase in experimentally induced AA amyloidosis in mice. Because it is reported that AEF serves as both a nidus and a template for amyloid formation, AA amyloidosis may have transmissibility by a prion-like mechanism. It has been shown that amyloid fibrils also have AEF activity, and amyloid fibrils with AEF activity were named fibril-amyloid enhancing factor (F-AEF). In this study, we investigated methods to inactivate the AEF activity. AEF was extracted from the thyroid gland obtained at autopsy of a patient with AA amyloidosis. Before injection into mice, AEF was treated with several methods for inactivation. Of all the tested treatments, 1 N NaOH, 0.1 N NaOH, and autoclaving consistently demonstrated complete inactivation of AEF. Heat treatment led to incomplete inactivation, but 0.01 N NaOH, 0.001 N NaOH, pepsin, trypsin, pronase, and proteinase K treatment had no effect on AEF activity. By analysis with transmission electron microscopy, the AEF preparation contains amyloid fibrils, and a change of ultrastructure was shown after 1 N NaOH, 0.1 N NaOH, and autoclaving treatment. Furthermore, immunoblotting of AEF with antihuman AA antibody revealed that the protein band was scarcely found after autoclaving, 1 N NaOH, and 0.1 N NaOH treatment. Our results suggest that, similar to Creutzfeldt–Jakob disease (CJD), amyloidosis may require chemical or autoclaving decontamination.

Autosomal dominant tauopathy with parkinsonism and central hypoventilation

Frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17T) can show various clinical phenotypes.1 We describe Japanese siblings with the intronic 10 + 14 splice site mutation of the microtubule-associated protein tau (MAPT) gene, showing parkinsonism, depression, weight loss, and central hypoventilation with reduced serotonin concentration suggested by low 5-hydroxyindole acetic acid (5-HIAA) in CSF. These clinical and biochemical features are just shared by Perry syndrome,2–4 although neither DCTN1 gene mutation nor TDP-43 proteinopathy was found. ### Case reports. A Japanese woman (III-5) (figure, A) developed clumsiness, tremor of the upper limbs, appetite loss, and apathy at age 44. She had lost 14 kilograms of body weight during 10 months. Mask-like face, hypophonic voice, bradykinesia, and muscle rigidity with predominance on the right side were observed. Deep tendon reflexes were hyperactive, especially in the right extremities with the Babinski sign. Despite levodopa treatment, her parkinsonism progressed and memory loss, disorientation, and cyanosis became apparent. Blood tests, EKG, and chest X-ray were not remarkable. Arterial blood gases showed reduced oxygen (60.3 Torr) and increased carbon dioxide (55.5 Torr). CSF analysis showed reduced 5-HIAA (5.6 ng/mL [normal 28.5 ± 7.2 ng/mL]). Brain MRI demonstrated mild atrophy in the brainstem tegmentum. Polysomnography revealed central hypoventilation with hypoxia. Although memory loss, disorientation, and …

Neuropathy in chronic graft‐versus‐host disease caused by Donor T cells

Chronic graft-versus-host disease (cGVHD) affects diverse organs, including peripheral nerves. Previous cases of neuropathy accompanied by cGVHD were diagnosed primarily based on electrophysiological studies and showed a favorable response to intravenous immunoglobulins. A case demonstrating infiltration of T cells in the sural nerve has been reported, but it remained unclear whether such neuropathy was caused by cGVHD or was coincidentally coexistent with other autoimmune diseases, including chronic inflammatory demyelinating polyneuropathy (CIDP). We describe direct invasion of the peripheral nervous system by the donor’s T cells in a recipient. A 29-year-old woman, after being diagnosed with acute lymphocytic leukemia (ALL), underwent a gendermismatched allogeneic bone marrow transplantation from her HLA-identical brother. Five months later, she developed cGVHD manifested by hyperkeratotic plaques of mouth ulceration and skin eruption, fulfilling the criteria for GVHD. She was treated with cyclosporine and prednisolone, after which all findings gradually lessened. Seven months later, she felt leg weakness and also noted numbness of all extremities, alopecia, dry eyes, and mouth ulcers. The neurological examination revealed distal-dominant muscle weakness in all limbs with areflexia. Tactile and pain sensations were diminished in all 4 limbs. Cerebrospinal fluid (CSF) analysis showed 13 mononuclear cells per microliter and elevated protein (264 mg/dl). Electromyography (EMG) revealed reduction of motor and sensory action potential amplitudes, decreased conduction velocities, and prolonged latencies with conduction block and abnormal temporal dispersion. The findings were consistent with a diagnosis of demyelinating neuropathy. After intravenous immunoglobulin treatment (400 mg/kg/day for 5 consecutive days), limb weakness gradually improved, and numbness of her extremities resolved. Thereafter, her demyelinating neuropathy relapsed 3 times, and she was treated with intravenous immunoglobulin, resulting in partial remission. After a fourth deterioration of the neurological and electrophysiological abnormalities, she was admitted to our department for a nerve biopsy. Sural nerve specimens showed a severe loss of large fibers and scattered thinly myelinated fibers. The teased fiber analysis showed segmental demyelination and irregular internode length. According to Dyck’s classification, a demyelinating process was seen in 8% of the fibers. Immunohistochemically, scattered CD4þ and CD8þ T cells were observed in the nerve bundles (Fig. 1A and B). The density of lymphocytes in the endoneurial space was 3.1/mm. Fluorescence in situ hybridization (FISH) showed that 15% of the infiltrating T cells carried both Xand Y-chromosomes (Fig. 1C and D), whereas others had two X-chromosomes. These data conclusively demonstrate that the infiltrating T cells were of donor origin, that is, from her brother. Periodic administration of immunoglobulin was effective in maintaining remission of the patient’s neuropathic deficits for 1 year thereafter. Gender-mismatched transplantation allowed us to distinguish the separation of donor and recipient cells using FISH, leading to the diagnosis of cGVHD-related neuropathy, and not CIDP. Arthritis with synovitis that developed in cGVHD after gender-mismatched transplantation for acute myelogenous leukemia has been reported. As in our patient, FISH analysis identified the donor origin of the inflammatory infiltrating T cells within the synovial tissues and confirmed the diagnosis of cGVHD.

Immunohistochemical analysis of laminin components in the blood–nerve barrier and blood–brain barrier

The blood–brain barrier (BBB) and blood–nerve barrier (BNB) play crucial roles in the maintenance of the central and peripheral nervous system. It had previously been thought that the BNB was leaky compared with the BBB, but several recent reports showed that the BNB has almost the same properties as a barrier system as the BBB, suggesting that the specific structural component of the basement membrane (especially laminin) in the BNB acts as a distinct barrier. Although the role of the laminin isoforms has been linked to BBB, the laminins in the BNB still remain unknown. Here, we aim to show the laminin components in the BNB and BBB.

Neuromyelitis optica shows hypermetabolism in 18-fluorodeoxyglucose positron emission tomography

Neuromyelitis optica (NMO) is an idiopathic severe necrotic inflammatory disease that mainly causes optic neuritis and myelopathy. Although it usually shows typical magnetic resonance imaging (MRI), the spinal lesions in NMO might sometimes mimic neoplasms or multiple sclerosis (MS) plaques. We present two NMO patients who showed marked hypermetabolism using 18‐fluorodeoxyglucose (FDG) positron emission tomography (PET) imaging at spinal cord lesions, corresponding to the gadolinium‐enhanced site. As MS plaques typically show hypometabolism, FDG‐PET imaging might be a useful and specific tool for the differential diagnosis of MS and NMO. Furthermore, NMO lesions should not be misdiagnosed as neoplasms simply because of the presence of hypermetabolism in FDG‐PET imaging. (Clin. Exp. Neuroimmunol. doi: 10.1111/j.1759‐1961.2011.00026.x, 2012)