[Severe consciousness disturbance after cord blood transplantation for relapsed T lymphoblastic lymphoma].

T-lymphoblastic leukemia/lymphoma (T-ALL/LBL) has a poor prognosis. Nelarabine has recently shown relatively good results in patients with relapsed or refractory T-ALL/LBL, but requires careful monitoring for neurological complications. A 50-year-old man with early recurrence of T-LBL after allogenic peripheral blood stem cell transplantation received nelarabine monotherapy and achieved complete remission after 1 cycle. He then received umbilical cord blood transplantation, and experienced sustained disturbance of consciousness. He later died of multiple organ failure, and autopsy suggested that nelarabine-induced leukoencephalopathy had caused the disturbance of consciousness. This case suggests that physicians should carefully monitor patients for neurological complications and consider imaging follow-up and consultation with a neurologist.

Evaluation of BCAS1-positive immature oligodendrocytes after cerebral ischemic stroke and SVD.

Ischemic cerebrovascular disease is an important cause of physical disability and dementia. Oligodendrocytes (OLGs), which differentiate from oligodendrocyte precursor cells (OPCs), are crucial for remyelination of the damaged brain and functional recovery. Breast carcinoma amplified sequence 1 (BCAS1) has recently been shown to be highly expressed in newly formed pre-myelinating oligodendrocytes (pre-mOLGs), while its expression level is reduced in mature OLGs. In this study, we analyzed BCAS1 expression by immunohistochemical analysis of human post-mortem brain tissue from six stroke patients (death within 2 months after stroke onset) and eight small vessel disease (SVD) patients. Control post-mortem brain tissue was from eight age-matched patients without any obvious central nervous system (CNS) pathology. The Olig2 expression in the area corresponding to the same section of the BCAS1-stained slice was analyzed to determine the total oligodendrocyte lineage. The percentage of differentiating OPCs in the oligodendrocyte lineage was calculated as the ratio of BCAS1+ to Olig2+ cells (BCAS1+/Olig2+). The stroke and SVD cases showed demyelination with decreased expression of myelin basic protein (MBP, a mature OLG marker). The stroke cases showed significantly increased numbers of early-stage BCAS1+ cells with an immature morphology and Olig2+ cells (pan-oligodendrocyte lineages) in the peri-infarct areas in both the cortex and white matter, but showed no increase in the number of late-stage BCAS1+ cells with a mature morphology. In contrast, the SVD cases showed no significant increase in Olig2+ and BCAS1+ cells. These results indicated that remyelination dysfunction could be attributed to insufficient maturation of OPCs in stroke and impaired recruitment of OPCs in SVD.

Utility of Skeletal Muscle CT in Diagnosing Spinal Muscular Atrophy Type 3 in a Patient Who Had Been Undiagnosed for 50 Years.

A 69-year-old woman presented with progressive limb weakness lasting 50 years. She denied any congenital disorders or a family history of neuromuscular disease. At ages 29, 46, and 58 years, she underwent hospitalization and evaluations including electromyogram (EMG) and muscle biopsy, but the results were inconclusive. As a result, she received a tentative diagnosis of myopathy of unknown etiology. However, at the age of 69 years, a computed tomography (CT) scan of her skeletal muscles revealed severe involvement of the triceps brachii, iliopsoas, and gastrocnemius muscles, along with preservation of the biceps brachii, gluteus maximus, and tibialis anterior muscles, which was consistent with spinal muscular atrophy (SMA). Finally, genetic testing revealed the deletion of the survival of the motor neuron 1 (SMN1) gene, confirming the diagnosis of SMA type 3. As our case suggests, SMA patients with prolonged disease duration could be underdiagnosed even after EMG and muscle biopsy. A skeletal CT scan could be useful for the diagnosis of SMA patients compared with MRI.

Efficacy of oligodendrocyte precursor cells as delivery vehicles for single-chain variable fragment to misfolded SOD1 in ALS rat model.

Superoxide dismutase1 (SOD 1) mutation is a leading cause of familial amyotrophic lateral sclerosis (ALS). Growing evidence suggests that antibody therapy against misfolded SOD1 protein can be therapeutic. However, the therapeutic effects are limited, partly because of the delivery system. Therefore, we investigated the efficacy of oligodendrocyte precursor cells (OPCs) as a drug delivery vehicle of single-chain variable fragments (scFv). Using a Borna disease virus vector that is pharmacologically removable and episomally replicable in the recipient cells, we successfully transformed wild-type OPCs to secrete scFv of a novel monoclonal antibody (D3-1), specific for misfolded SOD1. Single intrathecal injection of OPCs scFvD3-1, but not OPCs alone, significantly delayed disease onset and prolonged the lifespan of ALS rat models expressing SOD1 H46R . The effect of OPC scFvD3-1 surpassed that of a 1 month intrathecal infusion of full-length D3-1 antibody alone. scFv-secreting OPCs suppressed neuronal loss and gliosis, reduced levels of misfolded SOD1 in the spinal cord, and suppressed the transcription of inflammatory genes, including Olr1, an oxidized low-density lipoprotein receptor 1. The use of OPCs as a delivery vehicle for therapeutic antibodies is a new option for ALS in which misfolded protein and oligodendrocyte dysfunction are implicated in the pathogenesis.

Safety and tolerability of bosutinib in patients with amyotrophic lateral sclerosis (iDReAM study): A multicentre, open-label, dose-escalation phase 1 trial.

Background: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease caused by the loss of motor neurons, and development of effective medicines is urgently required. Induced pluripotent stem cell (iPSC)-based drug repurposing identified the Src/c-Abl inhibitor bosutinib, which is approved for the treatment of chronic myelogenous leukemia (CML), as a candidate for the molecular targeted therapy of ALS. Methods: An open-label, multicentre, dose-escalation phase 1 study using a 3 + 3 design was conducted in 4 hospitals in Japan to evaluate the safety and tolerability of bosutinib in patients with ALS. Furthermore, the exploratory efficacy was evaluated using Revised ALS Functional Rating Scale (ALSFRS-R), predictive biomarkers including plasma neurofilament light chain (NFL) were explored, and single-cell RNA sequencing of iPSC-derived motor neurons was conducted. Patients, whose total ALSFRS-R scores decreased by 1-3 points during the 12-week, received escalating doses starting from 100 mg quaque die (QD) up to 400 mg QD based on dose-limiting toxicity (DLT) occurrence, and all participants who received one dose of the study drug were included in the primary analysis. This trial is registered with ClinicalTrials.gov, NCT04744532, as Induced pluripotent stem cell-based Drug Repurposing for Amyotrophic Lateral Sclerosis Medicine (iDReAM) study. Findings: Between March 29, 2019 and May 7, 2021, 20 patients were enrolled, 13 of whom received bosutinib treatment and 12 were included in the safety and efficacy analyses. No DLTs were observed up to 300 mg QD, but DLTs were observed in 3/3 patients of the 400 mg QD cohort. In all patients receiving 100 mg-400 mg, the prevalent adverse events (AEs) were gastrointestinal AEs in 12 patients (92.3%), liver function related AEs in 7 patients (53.8%), and rash in 3 patients (23.1%). The safety profile was consistent with that known for CML treatment, and ALS-specific AEs were not observed. A subset of patients (5/9 patients) was found to respond well to bosutinib treatment over the 12-week treatment period. It was found that the treatment-responsive patients could be distinguished by their lower levels of plasma NFL. Furthermore, single-cell RNA sequencing of iPSC-derived motor neurons revealed the pathogenesis related molecular signature in patients with ALS showing responsiveness to bosutinib. Interpretation: This is the first trial of a Src/c-Abl inhibitor, bosutinib, for patients with ALS. The safety and tolerability of bosutinib up to 300 mg, not 400 mg, in ALS were described, and responsiveness of patients on motor function was observed. Since this was an open-label trial within a short period with a limited number of patients, further clinical trials will be required. Funding: AMED and iPS Cell Research Fund.

DOPAnization of tyrosine in α-synuclein by tyrosine hydroxylase leads to the formation of oligomers.

Parkinson's disease is a progressive neurodegenerative disorder characterized by the preferential loss of tyrosine hydroxylase (TH)-expressing dopaminergic neurons in the substantia nigra. Although the abnormal accumulation and aggregation of α-synuclein have been implicated in the pathogenesis of Parkinson's disease, the underlying mechanisms remain largely elusive. Here, we found that TH converts Tyr136 in α-synuclein into dihydroxyphenylalanine (DOPA; Y136DOPA) through mass spectrometric analysis. Y136DOPA modification was clearly detected by a specific antibody in the dopaminergic neurons of α-synuclein-overexpressing mice as well as human α-synucleinopathies. Furthermore, dopanized α-synuclein tended to form oligomers rather than large fibril aggregates and significantly enhanced neurotoxicity. Our findings suggest that the dopanization of α-synuclein by TH may contribute to oligomer and/or seed formation causing neurodegeneration with the potential to shed light on the pathogenesis of Parkinson's disease.

Conformational change of RNA-helicase DHX30 by ALS/FTD-linked FUS induces mitochondrial dysfunction and cytosolic aggregates.

Genetic mutations in fused in sarcoma (FUS) cause amyotrophic lateral sclerosis (ALS). Although mitochondrial dysfunction and stress granule have been crucially implicated in FUS proteinopathy, the molecular basis remains unclear. Here, we show that DHX30, a component of mitochondrial RNA granules required for mitochondrial ribosome assembly, interacts with FUS, and plays a crucial role in ALS-FUS. WT FUS did not affect mitochondrial localization of DHX30, but the mutant FUS lowered the signal of mitochondrial DHX30 and promoted the colocalization of cytosolic FUS aggregates and stress granule markers. The immunohistochemistry of the spinal cord from an ALS-FUS patient also confirmed the colocalization, and the immunoelectron microscope demonstrated decreased mitochondrial DHX30 signal in the spinal motor neurons. Subcellular fractionation by the detergent-solubility and density-gradient ultracentrifugation revealed that mutant FUS also promoted cytosolic mislocalization of DHX30 and aggregate formation. Interestingly, the mutant FUS disrupted the DHX30 conformation with aberrant disulfide formation, leading to impaired mitochondrial translation. Moreover, blue-native gel electrophoresis revealed an OXPHOS assembly defect caused by the FUS mutant, which was similar to that caused by DHX30 knockdown. Collectively, our study proposes DHX30 as a pivotal molecule in which disulfide-mediated conformational change mediates mitochondrial dysfunction and cytosolic aggregate formation in ALS-FUS.

Failure of DNA double-strand break repair by tau mediates Alzheimer's disease pathology in vitro.

DNA double-strand break (DSB) is the most severe form of DNA damage and accumulates with age, in which cytoskeletal proteins are polymerized to repair DSB in dividing cells. Since tau is a microtubule-associated protein, we investigate whether DSB is involved in tau pathologies in Alzheimer's disease (AD). First, immunohistochemistry reveals the frequent coexistence of DSB and phosphorylated tau in the cortex of AD patients. In vitro studies using primary mouse cortical neurons show that non-p-tau accumulates perinuclearly together with the tubulin after DSB induction with etoposide, followed by the accumulation of phosphorylated tau. Moreover, the knockdown of endogenous tau exacerbates DSB in neurons, suggesting the protective role of tau on DNA repair. Interestingly, synergistic exposure of neurons to microtubule disassembly and the DSB strikingly augments aberrant p-tau aggregation and apoptosis. These data suggest that DSB plays a pivotal role in AD-tau pathology and that the failure of DSB repair leads to tauopathy.

White matter edematous change with moderate vascular lesions in pretreated acute stage of leukoencephalopathy with cerebral amyloid angiopathy.

A 79-year-old man presented with subacute onset of dementia. Brain magnetic resonance imaging revealed leukoencephalopathy in the posterior lobes with presence of microbleeds. Although clinical manifestation suggested a diagnosis of leukoencephalopathy associated with cerebral amyloid angiopathy (CAA), the patient died of sudden rupture of an aneurysm of the thoracic aorta two months after the onset of dementia. Autopsy revealed pathological features of advanced-stage Alzheimer's disease. Immunohistochemistry for amyloid-ß revealed CAA mainly affecting arteries but not capillaries. Klüver-Barrera staining revealed white matter edema predominantly in the occipital lobes without ischemic changes. Perivascular cuffing was found to be sparse, but there was no evidence of angiitis. Pathological findings suggest that leukoencephalopathy was caused by the disruption of the blood-brain barrier rather than ischemia. Because the present patient died before immunotherapy, his neuropathological findings could reflect the pathomechanism of the acute stage of leukoencephalopathy with CAA.

An autopsy case of adult-onset neuronal intranuclear inclusion disease with perivascular preservation in cerebral white matter.

Neuronal intranuclear inclusion disease (NIID) is a neurodegenerative disease characterized by appearance of eosinophilic hyaline intranuclear inclusions. While the main symptoms of adult-onset NIID are dementia or limb weakness, some patients present with encephalitic episodes and transient neurological symptoms. The pathophysiology of these acute, transient symptoms, however, remains unknown. Here, we describe an autopsy case of adult-onset NIID with progressive dementia and transient hemiparesis. The patient was a 70-year-old man without a relevant family history, and initially presented with progressive dementia. He then exhibited transient left hemiparesis at 75 years of age and died of ureteral cancer at 77 years of age. Neuropathological examination revealed the presence of multiple areas of focal spongiosis in the subcortical white matter and patchy myelin pallor of the white matter, as in previous reports. However, perivascular areas were preserved even in the damaged white matter. In addition, dense glial fibrillary acidic protein (GFAP)-immunoreactive astrocytic processes were observed in these areas. [Correction added on 23 January 2022, after first online publication: the preceding sentence has been corrected to improve readability.] GFAP immunohistochemistry revealed decreased density and morphological abnormalities of astrocytes in the affected white matter. These pathological findings might reflect blood-brain barrier impairment and dysregulation of blood flow, which may be related to the pathophysiology of the acute, transient symptoms observed in NIID.

Insertable inductively coupled volumetric coils for MR microscopy in a human 7T MR system.

PURPOSE: To demonstrate the capability of insertable inductively coupled volumetric coils for MR microscopy in a human 7T MR system. METHODS: Insertable inductively coupled volume coils with diameters of 26 and 64 mm (D26 and D64 coils) targeted for monkey and mouse brain specimen sizes were designed and fabricated. These coils were placed inside the imaging volume of a transmit/receive knee coil without wired connections to the main system. Signal-to-noise ratio (SNR) evaluations were conducted with and without the insertable coils, and the g-factor maps of parallel imaging (PI) were also calculated for the D64 coil. Brain specimens were imaged using 3D T2∗ -weighted images with spatial resolution of isotropic 50 and 160 µm using D26 and D64 coils, respectively. RESULTS: Relative average (SD) SNRs compared with knee coil alone were 12.54 (0.30) and 2.37 (0.05) at the center for the D26 and D64 coils, respectively. The mean g-factors of PI with the D64 coil for the factor of 2 were less than 1.1 in the left-right and anterior-posterior directions, and around 1.5 in the superior-inferior direction or when the PI factor of 3 was used. Acceleration in two directions showed lower g-factors but suffered from intrinsic low SNR. Representative T2∗ -weighted images of the specimen showed structural details. CONCLUSION: Inductively coupled small diameter coils insertable to the knee coil demonstrated high SNR and modest PI capability. The concept was successfully used to visualize fine structures of the brain specimen. The insertable coils are easy to handle and enable MR microscopy in a human whole-body 7T MRI system.

The stimulator of interferon genes (STING) pathway is upregulated in striatal astrocytes of patients with multiple system atrophy.

Multiple system atrophy (MSA) is a progressive neurodegenerative disorder characterized by the accumulation of pathogenic phosphorylated α-synuclein in oligodendrocytes. In brains affected by MSA, severe astrogliosis is also observed, but its precise role in MSA pathogenesis remains largely unknown. Recently, the stimulator of interferon genes (STING) pathway and type I interferons, its downstream molecules, have been reported to be involved in the neurodegenerative process and to be activated in astrocytes. This study aimed to investigate the role of the STING pathway in the pathogenesis of MSA using postmortem brains. Samples used for immunohistochemical analysis included 6 cases of MSA parkinsonism type (MSA-P), 6 cases of MSA cerebellar type (MSA-C), and 7 age-matched controls. In MSA-P cases, astrocytes immunopositive for STING and TANK-binding kinase 1 (TBK1), its downstream molecule, were abundantly observed in the putamen and the substantia nigra. Moreover, these molecules colocalized with glial fibrillary acidic protein (GFAP) in reactive astrocytes, and the density of STING-positive astrocytes correlated with that of GFAP-positive reactive astrocytes in the brains of patients with MSA-P. These results suggest that the upregulated expression of STING pathway-related proteins in astrocytes and the subsequent inflammation may contribute to the pathogenesis in MSA-P and could provide novel therapeutic targets for the treatment of MSA.

Monoclonal gammopathy of renal significance (MGRS)-related AL amyloidosis complicated by amyloid myopathy: a case report.

BACKGROUND: Lately, monoclonal gammopathy of renal significance (MGRS) has been defined as a group of renal disorders that are strongly associated with monoclonal protein, including amyloid immunoglobulin light chain (AL) amyloidosis. Amyloid myopathy is rare (1.5% of all patients with amyloidosis) and the prognosis is poor. Furthermore, only approximately 20% of patients with amyloid myopathy are reported to have renal involvement, indicating a lack of data in the literature. CASE PRESENTATION: Here, we report a rare case of MGRS-related AL amyloidosis complicated by amyloid myopathy that presented with muscle weakness in the upper and lower limbs, neck and fingers, and nephrotic syndrome. Blood, urine, and bone marrow examination revealed monoclonal gammopathy of undetermined significance (MGUS) (Bence Jones protein-lambda). Muscle biopsy of the vastus lateralis muscle demonstrated amyloid proteins in the sarcolemma and in the blood vessel walls on Congo red staining, suggesting amyloid myopathy, and tiny inclusions in fibers on modified Gomori trichrome stain. Although we thought they were reminiscent of nemaline bodies, we could not confirm the nature of this structure. Renal biopsy demonstrated amyloid proteins in the mesangial region, part of the capillary walls, and the blood vessel walls on direct fast scarlet staining. As these amyloid proteins were positive for p-component staining and negative for amyloid A staining, ß2-microglobulin, and pre-albumin, and as lambda light chains were positive in the mesangial region, we diagnosed the patient with MGRS-related AL amyloidosis. Although he was treated with melphalan and dexamethasone, his symptoms did not improve. CONCLUSIONS: AL amyloidosis involving the kidneys and muscles has a poor prognosis, and a delayed diagnosis of amyloid myopathy is common because of its rarity and frequent misdiagnosis, which increases organ function deterioration. Therefore, early detection, therapeutic intervention, and careful follow-up are crucial.

Japanese Familial Cases of Frontotemporal Dementia and Parkinsonism with N279K Tau Gene Mutation.

BACKGROUND: Mutations in the tau gene linked to chromosome 17 cause frontotemporal dementia and parkinsonism (FTDP-17). OBJECTIVE: This study presents 3 Japanese familial cases diagnosed with N279K tau gene mutation, including 1 autopsy-confirmed case. METHODS: We compared the clinical presentations, cognitive functions, and images between the 3 familial cases diagnosed with N279K mutation. RESULTS: All 3 patients presented symptoms in their early 40s. One patient showed severe cognitive dysfunction and died in his sixth year after onset. The remaining 2 cases presented with parkinsonism-dominant clinical features. Among the 2 cases, 1 presented the characteristic symptoms of progressive supranuclear palsy. The pathological features of the dementia-dominant case showed frontal and temporal lobe-dominant neuronal loss and gliosis. Tau-positive neuronal and glial inclusions were found throughout. Further, tufted astrocytes and globose tangles were present whereas there were no Pick bodies and astrocytic plaques, compatible with pathology-confirmed frontotemporal lobar degeneration (FTLD) -tau subtypes. CONCLUSIONS: Patients with FTDP-17 can be classified into the following 2 major groups: dementia and parkinsonism-plus predominant phenotypes. Among our 3 cases, 1 showed dementia predominance whereas the other 2 showed parkinsonism predominance. Mutations in the microtubule-associated protein tau (MAPT) present with several pathological features. Clinically, our case presented a behavioral variant frontotemporal dementia (bvFTD). However, morphologically, the observed glial and neuronal pathology met the criteria for progressive supranuclear palsy (PSP). This study highlights the clinical heterogeneity within and between families with same MAPT mutation. Few pathologically confirmed PSP cases have been reported with mutations in MAPT.

Optineurin defects cause TDP43-pathology with autophagic vacuolar formation.

We previously showed that optineurin (OPTN) mutations lead to the development of amyotrophic lateral sclerosis. The association between OPTN mutations and the pathogenesis of amyotrophic lateral sclerosis remains unclear. To investigate the mechanism underlying its pathogenesis, we generated Optn knockout mice. We evaluated histopathological observations of these mice and compared with those of OPTN- amyotrophic lateral sclerosis cases to investigate the mechanism underlying the pathogenesis of amyotrophic lateral sclerosis caused by OPTN mutations. The Optn (-/-) mice presented neuronal autophagic vacuoles immunopositive for charged multivesicular body protein 2b, one of the hallmarks of granulovacuolar degenerations, in the cytoplasm of spinal cord motor neurons at the age of 8 months and the OPTN- amyotrophic lateral sclerosis case with homozygous Q398X mutation. In addition, Optn (-/-) mice showed TAR-DNA binding protein 43/sequestosome1/p62 -positive cytoplasmic inclusions and the clearance of nuclear TAR-DNA binding protein 43. The axonal degeneration of the sciatic nerves was observed in Optn (-/-) mice. However, we could not observe significant differences in survival time, body weight, and motor functions, at 24 months. Our findings suggest that homozygous OPTN deletion or mutations might result in autophagic dysfunction and TAR-DNA binding protein 43 mislocalization, thereby leading to neurodegeneration of motor neurons. These findings indicate that the Optn (-/-) mice recapitulate both common and specific pathogenesis of amyotrophic lateral sclerosis associated with autophagic abnormalities. Optn (-/-) mice could serve as a mouse model for the development of therapeutic strategies.

BCAS1-positive immature oligodendrocytes are affected by the α-synuclein-induced pathology of multiple system atrophy.

Multiple system atrophy (MSA) is pathologically characterized by the presence of fibrillar α-synuclein-immunoreactive inclusions in oligodendrocytes. Although the myelinating process of oligodendrocytes can be observed in adult human brains, little is known regarding the presence of α-synuclein pathology in immature oligodendrocytes and how their maturation and myelination are affected in MSA brains. Recently, breast carcinoma amplified sequence 1 (BCAS1) has been found to be specifically expressed in immature oligodendrocytes undergoing maturation and myelination. Here, we analyzed the altered dynamics of oligodendroglial maturation in both MSA brains and primary oligodendroglial cell cultures which were incubated with α-synuclein pre-formed fibrils. The numbers of BCAS1-expressing oligodendrocytes that displayed a matured morphology negatively correlated with the density of pathological inclusions in MSA brains but not with that in Parkinson's disease and diffuse Lewy body disease. In addition, a portion of the BCAS1-expressing oligodendrocyte population showed cytoplasmic inclusions, which were labeled with antibodies against phosphorylated α-synuclein and cleaved caspase-9. Further in vitro examination indicated that the α-synuclein pre-formed fibrils induced cytoplasmic inclusions in the majority of BCAS1-expressing oligodendrocytes. In contrast, the majority of BCAS1-non-expressing mature oligodendrocytes did not develop inclusions on day 4 after maturation induction. Furthermore, exposure of α-synuclein pre-formed fibrils in the BCAS1-positive phase caused a reduction in oligodendroglial cell viability. Our results indicated that oligodendroglial maturation and myelination are impaired in the BCAS1-positive phase of MSA brains, which may lead to the insufficient replacement of defective oligodendrocytes. In vitro, the high susceptibility of BCAS1-expressing primary oligodendrocytes to the extracellular α-synuclein pre-formed fibrils suggests the involvement of insufficient oligodendroglial maturation in MSA disease progression and support the hypothesis that the BCAS1-positive oligodendrocyte lineage cells are prone to take up aggregated α-synuclein in vivo.

Accumulation of HAX-1 and PARL in brainstem- and cortical-type Lewy bodies in Parkinson's disease and dementia with Lewy bodies.

HS1-associated protein X-1 (HAX-1) and presenilin-associated rhomboid-like protein (PALR) were reported to play an important role in the activation of HtrA2/Omi, which is also designated PARK13, in the mitochondria. To elucidate the role of HAX-1 and PARL in patients with Parkinson's disease (PD) and dementia with Lewy bodies (DLB), we performed immunohistochemical studies on HtrA2/Omi, HAX-1 and PARL using autopsied brains from 8 normal subjects, 10 patients with PD and 5 patients with DLB. In accordance with our previous report, brainstem-type and cortical Lewy bodies were strongly immunopositive for HtrA2/Omi. In the normal brains, HAX-1 and PARL immunoreactivities were observed in various types of neurons in the cerebral cortex, midbrain, and upper pons. HAX-1 and PARL immunoreactivities were also observed in the remaining neurons, and brainstem-type and cortical Lewy bodies were intensely immunoreactive for HAX-1 and PARL. Both immunoreactivities were localized to the halo or core of brainstem-type Lewy bodies. Our results suggest that brainstem-type and cortical Lewy bodies may contain HAX-1 and PARL as well as HtrA2/Omi, and that these proteins may partially contribute to the formation of Lewy bodies and may be associated with the pathogenesis of PD and DLB.

Linear Polyubiquitin Chain Modification of TDP-43-Positive Neuronal Cytoplasmic Inclusions in Amyotrophic Lateral Sclerosis.

Neuronal cytoplasmic inclusions (NCIs) containing TAR DNA-binding protein of 43 kDa (TDP-43) are pathological hallmarks of amyotrophic lateral sclerosis (ALS) and are known to be ubiquitinated. Eight linkage types of polyubiquitin chains have been reported, each type of chain exerting different intracellular actions. The linkage type of polyubiquitin chain involved in the formation of NCIs in sporadic ALS (sALS), however, has not yet been elucidated. We performed immunohistochemical study of the spinal cords of 12 patients with sALS and on those of 6 control subjects. Virtually all ubiquitinated NCIs were immunolabeled with lysine 48-linked polyubiquitin chain (K48-Ub). Although the majority of NCIs were triple-immunoreactive for K48-Ub, linear polyubiquitin chain (L-Ub), and lysine 63-linked polyubiquitin chain (K63-Ub), thin parts of K48-Ub-immunopositive NCIs were not labeled for K63-Ub or L-Ub. We also detected HOIP and SHARPIN, components of linear ubiquitin chain assembly complex, colocalizing with L-Ub on NCIs. Moreover, the immunosignal of optineurin, an autophagy receptor working with L-Ub, and that of activated NF-κB p65, were observed to be colocalizing with L-Ub on certain parts of NCIs. The L-Ub modification of TDP-43-positive NCIs may function as an inducer of autophagic clearance of NCIs, neuroinflammation, and neurodegeneration in sALS.