Genetic factors affecting survival in Japanese patients with sporadic amyotrophic lateral sclerosis: a genome-wide association study and verification in iPSC-derived motor neurons from patients.

BACKGROUND: Several genetic factors are associated with the pathogenesis of sporadic amyotrophic lateral sclerosis (ALS) and its phenotypes, such as disease progression. Here, in this study, we aimed to identify the genes that affect the survival of patients with sporadic ALS. METHODS: We enrolled 1076 Japanese patients with sporadic ALS with imputed genotype data of 7 908 526 variants. We used Cox proportional hazards regression analysis with an additive model adjusted for sex, age at onset and the first two principal components calculated from genotyped data to conduct a genome-wide association study. We further analysed messenger RNA (mRNA) and phenotype expression in motor neurons derived from induced pluripotent stem cells (iPSC-MNs) of patients with ALS. RESULTS: Three novel loci were significantly associated with the survival of patients with sporadic ALS-FGF1 at 5q31.3 (rs11738209, HR=2.36 (95% CI, 1.77 to 3.15), p=4.85×10-9), THSD7A at 7p21.3 (rs2354952, 1.38 (95% CI, 1.24 to 1.55), p=1.61×10-8) and LRP1 at 12q13.3 (rs60565245, 2.18 (95% CI, 1.66 to 2.86), p=2.35×10-8). FGF1 and THSD7A variants were associated with decreased mRNA expression of each gene in iPSC-MNs and reduced in vitro survival of iPSC-MNs obtained from patients with ALS. The iPSC-MN in vitro survival was reduced when the expression of FGF1 and THSD7A was partially disrupted. The rs60565245 was not associated with LRP1 mRNA expression. CONCLUSIONS: We identified three loci associated with the survival of patients with sporadic ALS, decreased mRNA expression of FGF1 and THSD7A and the viability of iPSC-MNs from patients. The iPSC-MN model reflects the association between patient prognosis and genotype and can contribute to target screening and validation for therapeutic intervention.

Multiple cerebral infarctions due to calcified amorphous tumor growing rapidly from an antecedent infection and decreasing rapidly by detachment of fibrin and antithrombotic drugs: a case report.

BACKGROUND: Calcified amorphous tumor (CAT) of the heart is a rare non-neoplastic intracardiac mass, a calcium deposition surrounded by amorphous fibrous tissue, and possibly causes cerebral embolism. Even rarer is CAT associated with infection, and no CAT with antecedent infection has been reported to our knowledge. In addition, although some CAT in patients on hemodialysis has been reported to grow rapidly, no case has been reported on CAT that grew and diminished rapidly in a short period of time. Here, we report the case of an 82-year-old Japanese woman with normal renal function who developed multiple cerebral infarctions due to CAT that grew rapidly, associated with inflammation from an antecedent infection, and diminished rapidly by detachment of fibrin on the mass surface and antithrombotic drugs. CASE PRESENTATION: The patient developed fever after dental treatment and found musical hallucination on the left ear worsened in degree and frequency. In a nearby clinic, she was treated with antibiotics, and her body temperature turned to normal in approximately 1 month. She presented to our hospital for workup on the worsened musical hallucination. Magnetic resonance imaging (MRI) showed multiple cerebral infarctions, and transthoracic echocardiography (TTE) revealed an immobile hyperechoic mass with an acoustic shadow arising from a posterior cusp of the mitral valve. CAT was suspected and treated with apixaban and aspirin. Follow-up MRI and TTE showed newly developed multiple cerebral infarctions and rapidly diminished CAT. Cardiac surgery was performed to resect the CAT. The pathological findings showed calcifications surrounded by amorphous fibrous tissue including fibrin, indicating CAT. The patient's symptoms improved and no cerebral infarctions recurred in 4 months follow-up. CONCLUSION: Inflammation from an antecedent infection can cause CAT to grow rapidly. Fibrous tissue including fibrin may attach to the surface of CAT, resulting in multiple cerebral infarctions. Fibrous tissue may detach and disappear by antithrombotic drugs, leading to a rapid diminishment of CAT in size.

Maximum Walking Speed at Discharge Could Be a Prognostic Factor for Vascular Events in Patients With Mild Stroke: A Cohort Study.

OBJECTIVE: To identify the prognostic value of physical activity-related factors as well as known vascular risk factors for vascular events in mild ischemic stroke (MIS). DESIGN: Single-center prospective cohort study. SETTING: University hospital. PARTICIPANTS: Consecutive patients (N=255) (175 men, median age 70.0y) with acute ischemic stroke and transient ischemic attack (TIA) with modified Rankin scale scores ranging from 0 to 2 were enrolled in this study. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Enrolled patients were followed up for composite vascular events as primary outcomes up to 3 years postdischarge. Primary outcomes included stroke and cardiovascular death, hospitalization due to stroke or TIA recurrence, cardiovascular disease, and peripheral artery disease. During hospitalization, known vascular risk factors such as previous history of vascular events, stroke subtype, white matter lesions, and ankle-brachial index were assessed. Moreover, at the time of discharge, physical activity-related factors such as maximum walking speed (MWS), handgrip strength, knee extensor isometric muscle strength, anxiety, and depression were assessed as potential predictors. RESULTS: The Kaplan-Meier estimates of cumulative risk of composite vascular events at 1, 2, and 3 years were 9.6%, 14.4%, and 15.2%, respectively. After multivariate analysis, cerebral white matter lesions of periventricular hyperintensity (PVH) (grade=3; hazard ratio: 2.904; 95% confidence interval: 1.160 to 7.266; P=.023) and MWS (<1.45m/s; hazard ratio: 2.232; 95% confidence interval: 1.010 to 4.933; P=.047) were identified as significant independent predictors of composite vascular events. CONCLUSIONS: The results of this study indicate that MWS could be an independent prognostic factor for composite vascular events in MIS.

Loss of TDP-43 causes age-dependent progressive motor neuron degeneration.

Amyotrophic lateral sclerosis is a devastating, progressive neurodegenerative disease that affects upper and lower motor neurons. Although several genes are identified as the cause of familial cases, the pathogeneses of sporadic forms, which account for 90% of amyotrophic lateral sclerosis, have not been elucidated. Transactive response DNA-binding protein 43 a nuclear protein regulating RNA processing, redistributes to the cytoplasm and forms aggregates, which are the histopathological hallmark of sporadic amyotrophic lateral sclerosis, in affected motor neurons, suggesting that loss-of-function of transactive response DNA-binding protein 43 is one of the causes of the neurodegeneration. To test this hypothesis, we assessed the effects of knockout of transactive response DNA-binding protein 43 in mouse postnatal motor neurons using Cre/loxp system. These mice developed progressive weight loss and motor impairment around the age of 60 weeks, and exhibited degeneration of large motor axon, grouped atrophy of the skeletal muscle, and denervation in the neuromuscular junction. The spinal motor neurons lacking transactive response DNA-binding protein 43 were not affected for 1 year, but exhibited atrophy at the age of 100 weeks; whereas, extraocular motor neurons, that are essentially resistant in amyotrophic lateral sclerosis, remained preserved even at the age of 100 weeks. Additionally, ultra structural analysis revealed autolysosomes and autophagosomes in the cell bodies and axons of motor neurons of the 100-week-old knockout mice. In summary, the mice in which transactive response DNA-binding protein 43 was knocked-out specifically in postnatal motor neurons exhibited an age-dependent progressive motor dysfunction accompanied by neuropathological alterations, which are common to sporadic amyotrophic lateral sclerosis. These findings suggest that transactive response DNA-binding protein 43 plays an essential role in the long term maintenance of motor neurons and that loss-of-function of this protein seems to contribute to the pathogenesis of amyotrophic lateral sclerosis.

Open-label study to evaluate the pharmacodynamics, clinical efficacy, and safety of meropenem for adult bacterial meningitis in Japan.

The aim of this study was to assess the efficacy, safety, and concentration of meropenem in cerebrospinal fluid when meropenem (2 g every 8 h) was administered to Japanese adult patients with bacterial meningitis. Five Japanese patients (mean age 60.6 years [range 35-71]) were enrolled. Infection with Streptococcus pneumoniae (three patients), Streptococcus salivarius (one patient), and Staphylococcus aureus (one patient) was confirmed by cerebrospinal fluid culture. Meropenem (2 g every 8 h) was administered to all five patients. Treatment duration ranged from 14 to 28 days (mean 22.6 days). All the patients were successfully treated. The concentration of meropenem in cerebrospinal fluid ranged from 0.27 to 6.40 µg/ml up to 8.47 h and was over 1 µg/ml 3 h after starting meropenem infusion. In each patient, the present study confirmed for the first time that the concentration of meropenem in cerebrospinal fluid exceeded the minimal inhibitory concentration for these pathogens. Eleven clinical and laboratory adverse events considered to be related to meropenem were observed in all patients, but no serious adverse event and no discontinuance of treatment due to adverse events occurred. Thus meropenem appeared to be a well-tolerated and effective agent for Japanese adult patients with bacterial meningitis. 2 g every 8 h of meropenem was delivered to CSF and its concentration was exceed in MICs for the detected pathogens.

Simple and efficient differentiation of human iPSCs into contractible skeletal muscles for muscular disease modeling.

Pathophysiological analysis and drug discovery targeting human diseases require disease models that suitably recapitulate patient pathology. Disease-specific human induced pluripotent stem cells (hiPSCs) differentiated into affected cell types can potentially recapitulate disease pathology more accurately than existing disease models. Such successful modeling of muscular diseases requires efficient differentiation of hiPSCs into skeletal muscles. hiPSCs transduced with doxycycline-inducible MYOD1 (MYOD1-hiPSCs) have been widely used; however, they require time- and labor-consuming clonal selection, and clonal variations must be overcome. Moreover, their functionality should be carefully examined. Here, we demonstrated that bulk MYOD1-hiPSCs established with puromycin selection rather than G418 selection showed rapid and highly efficient differentiation. Interestingly, bulk MYOD1-hiPSCs exhibited average differentiation properties of clonally established MYOD1-hiPSCs, suggesting that it is possible to minimize clonal variations. Moreover, disease-specific hiPSCs of spinal bulbar muscular atrophy (SBMA) could be efficiently differentiated via this method into skeletal muscle that showed disease phenotypes, suggesting the applicability of this method for disease analysis. Finally, three-dimensional muscle tissues were fabricated from bulk MYOD1-hiPSCs, which exhibited contractile force upon electrical stimulation, indicating their functionality. Thus, our bulk differentiation requires less time and labor than existing methods, efficiently generates contractible skeletal muscles, and may facilitate the generation of muscular disease models.

Oxidative stress induced by glutathione depletion reproduces pathological modifications of TDP-43 linked to TDP-43 proteinopathies.

TAR DNA-binding protein 43 (TDP-43) is a major component of ubiquitin-positive inclusion of TDP-43 proteinopathies including amyotrophic lateral sclerosis and frontotemporal lobar degeneration with ubiquitinated inclusions, which is now referred to as FTLD-TDP. TDP-43 in the aberrant inclusion is known to be hyperphosphorylated at C-terminal sites, to be truncated at the N-terminal region, and to re-distribute from nucleus to cytoplasm or neurite. The pathogenic role of these modifications, however, has not been clarified. Furthermore, there is no evidence about the initial cause of these modifications. Herein we show that ethacrynic acid (EA), which is able to increase cellular oxidative stress through glutathione depletion, induces TDP-43 C-terminal phosphorylation at serine 403/404 and 409/410, insolubilization, C-terminal fragmentation, and cytoplasmic distribution in NSC34 cells and primary cortical neurons. In the investigation using a nonphosphorylable mutant of TDP-43, there was no evidence that C-terminal phosphorylation of TDP-43 contributes to its solubility or distribution under EA induction. Our findings suggest that oxidative stress induced by glutathione depletion is associated with the process of the pathological TDP-43 modifications and provide new insight for TDP-43 proteinopathies.

Recurring Weakness in Rhabdomyolysis Following Pfizer-BioNTech Coronavirus Disease 2019 mRNA Vaccination.

Rhabdomyolysis is a well-known clinical syndrome of muscle injury. Rhabdomyolysis following coronavirus disease 2019 (COVID-19) vaccination has recently been reported. The patients' weakness gradually subsided and did not recur. Rhabdomyolysis associated with COVID-19 vaccination has not been assessed by repeated magnetic resonance imaging (MRI) within a short time. We report a rare case of an older woman who developed recurring weakness with rhabdomyolysis after COVID-19 vaccination. A 76-year-old woman presented with myalgia 2 days after receiving a third dose of the COVID-19 vaccine. A physical examination showed weakness of the bilateral iliopsoas muscles. Her creatine kinase concentration was 9816 U/L. MRI showed hyperintensity of multiple limb muscles. She was treated with intravenous normal saline. Her symptoms disappeared within 3 days. However, MRI on day 4 of hospitalization showed exacerbation of the hyperintensity in the left upper limb muscles. On day 5 of hospitalization, weakness of the left supraspinatus and deltoid muscles appeared. MRI on day 8 of hospitalization showed attenuation of the hyperintensity in all muscles. Her weakness and elevated creatine kinase concentration disappeared by day 10. Repeated MRI over a short time may be useful to predict potential weakness and monitor the course of COVID-19 vaccine-induced rhabdomyolysis.

A 47-Year-Old Japanese Woman with Symptoms of Increased Salty and Reduced Sweet Taste Perception Preceding a Diagnosis of Thymoma-Associated Myasthenia Gravis.

BACKGROUND Myasthenia gravis (MG) is an autoimmune neuromuscular disorder, which is often accompanied by various complications. Partial dysgeusia is an uncommon nonmotor symptom of MG, and dysgeusia preceding typical MG symptoms is rare. Although ageusia and hypogeusia have been reported in patients with MG, increased perception of taste has not been reported. CASE REPORT A 47-year-old Japanese woman presented with a reduced perception of sweet taste and an increased perception of salty taste. Meanwhile, she was diagnosed with thymoma-associated generalized MG and underwent extended thymectomy. Three months later, her anti-acetylcholine receptor (AChR) antibody (Ab) titer increased to 70 nmol/L, when she had completely lost perception of sweet taste and had developed a markedly increased perception of salty taste. Prednisolone and tacrolimus were then added to the medication, and her partial dysgeusia gradually improved. As the AChR Ab titer decreased, disturbance of sweet taste resolved, although a slight decrease persisted. The increased perception of salty taste returned to normal. CONCLUSIONS This is a rare case of a patient with MG who developed an increased salty taste perception with a reduced sweet taste perception 3 months before the onset of her motor symptoms. We suggest that MG should be considered as a differential diagnosis in patients with partial dysgeusia but no motor symptoms.

Subcortical infarction in a young adult with Hunter syndrome.

INTRODUCTION: Hunter syndrome (mucopolysaccharidosis type II, MPS II) is an X-linked lysosomal storage disease caused by deficiency of iduronate-2-sulfatase. Recently, stroke caused by embolization with Hunter syndrome has been reported. Here, we report the case of a 23-year-old Japanese man with Hunter syndrome who developed subcortical infarction by the mechanism similar to branch atheromatous disease (BAD). CASE PRESENTATION: He had been treated with idursulfase supplementation. He presented with left-sided weakness and conjugate eye deviation to the right, and was diagnosed with branch atheromatous disease affecting the right corona radiata, based on MRI findings. The patient was treated with argatroban and aspirin. Magnetic resonance angiography demonstrated no evidence of luminal narrowing of the cerebral arteries. T1-sampling perfection with application-optimized contrasts by using different flip angle evolutions (SPACE) imaging revealed thickened middle cerebral artery. The patient had markedly low flow-mediated vasodilation, suggesting impaired vasodilation in response to nitric monoxide. CONCLUSION: The arterial wall thickening and impaired vasodilation in the cerebral arteries related to subcortical infarction. We should clarify the mechanism of cerebral infarction in Hunter syndrome patients.

Antiparkinsonian drugs as potent contributors to nocturnal sleep in patients with Parkinson's disease.

OBJECTIVE: To clarify whether antiparkinsonian drugs contribute to nocturnal sleep disturbances in patients with Parkinson's disease (PD). BACKGROUND: Although the major antiparkinsonian drugs L-dopa and dopamine agonists (DAs) have been found to affect sleep, little is known about the effects of specific drugs on sleep in PD patients. METHODS: The study participants consisted of 112 PD patients (median age 72.5 years [inter-quartile range: IQR 65-79]; mean disease duration 8.44 years [standard deviation: 7.33]; median Hoehn and Yahr stage 3 [IQR 2-3.75]) taking one of three types of non-ergot extended-release DAs (rotigotine 32; pramipexole 44; ropinirole 36) with or without L-dopa (median daily total dosage of antiparkinsonian drugs 525.5 mg [IQR 376.25-658] levodopa equivalent dose [LED]). Participants were assessed using the PD Sleep Scale-2 (PDSS-2). RESULTS: For the whole PD patient cohort, the PDSS-2 sleep disturbance domain score and the scores for item 1 assessing sleep quality and item 8 assessing nocturia were positively correlated with daily total dosage of antiparkinsonian drugs and dosage of L-dopa, but not with the dosage of DAs. Sub-analysis according to DA treatment revealed that DA dosage was not correlated with item 1 or 8 score in any of the subgroups. The LED ratio of rotigotine to the total dosage of antiparkinsonian drugs was inversely correlated with the item 1 score. CONCLUSIONS: These data suggest that antiparkinsonian drugs, in particular L-dopa, adversely affect nocturnal sleep in PD patients, especially in terms of sleep quality and nocturia. Thus, adjusting both the total dosage of antiparkinsonian drugs and the dose-ratio of L-dopa might be key actions for alleviating poor sleep quality in patients with PD. Among DAs, we found a clear positive correlation between the dose-ratio of rotigotine and sleep quality. Thus, partial L-dopa replacement with rotigotine could improve sleep quality in patients with PD.

Dorfin ameliorates phenotypes in a transgenic mouse model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that is characterized by progressive motor neuron degeneration and leads to death within a few years of diagnosis. One of the pathogenic mechanisms of ALS is proposed to be a dysfunction in the protein quality-control machinery. Dorfin has been identified as a ubiquitin ligase (E3) that recognizes and ubiquitinates mutant SOD1 proteins, thereby accelerating their degradation and reducing their cellular toxicity. We examined the effects of human Dorfin overexpression in G93A mutant SOD1 transgenic mice, a mouse model of familial ALS. In addition to causing a decrease in the amount of mutant SOD1 protein in the spinal cord, Dorfin overexpression ameliorated neurological phenotypes and motor neuron degeneration. Our results indicate that Dorfin overexpression or the activation or induction of E3 may be a therapeutic avenue for mutant SOD1-associated ALS.

Aberrant Expression of Nodal and Paranodal Molecules in Neuropathy Associated With IgM Monoclonal Gammopathy With Anti-Myelin-Associated Glycoprotein Antibodies.

To clarify the pathogenesis of anti-myelin-associated glycoprotein (MAG) antibody neuropathy associated with IgM monoclonal gammopathy (anti-MAG neuropathy), sural nerve biopsy specimens from 15 patients were investigated. Sodium channels, potassium channels, contactin-associated protein 1 (Caspr1), contactin 1, and neurofascin were evaluated by immunofluorescence in teased-fiber preparations. Immunoreactivity to the pan-sodium channel in both anti-MAG neuropathy patients and in normal controls was concentrated at the node of Ranvier unless there was demyelination, which was defined as the widening of the node of Ranvier. However, this immunoreactivity became weak or disappeared as demyelination progressed. In contrast, KCNQ2 immunostaining was nearly absent even in the absence of demyelination. The lengths of Caspr1, contactin 1, and pan-neurofascin immunostaining sites at the paranode were significantly increased compared with those of normal controls despite the absence of demyelination. The length of paranodal neurofascin staining correlated with the anti-MAG antibody titer, nerve conduction indices, the frequency of de/remyelination in teased-fiber preparations, and the frequency of widely spaced myelin (p < 0.05, p < 0.05, p < 0.01, and <0.05, respectively). These findings suggest that nodal and paranodal molecular alterations occur in early stages preceding the morphological changes associated with demyelination in anti-MAG neuropathy.

Motor Improvement-Related Regional Cerebral Blood Flow Changes in Parkinson's Disease in Response to Antiparkinsonian Drugs.

Little is known about the relationship between regional cerebral blood flow (rCBF) change and clinical improvement in patients with Parkinson's disease (PD). Single-photon emission computed tomography (SPECT) measurement of cerebral blood flow allows evaluation of temporal changes in brain function, and using SPECT, we aimed to identify motor improvement-related rCBF changes in response to the administration of antiparkinsonian drugs. Thirty PD patients (16 without dementia; 14 with dementia) were scanned with technetium-99m labeled ethyl cysteinate dimer SPECT and were rated with the Movement Disorder Society-Unified Parkinson's Disease Rating Scale part III, both before and after a single administration of antiparkinsonian drugs. The SPECT data were processed using Statistical Parametric Mapping 2, the easy Z-score Imaging System, and voxel-based Stereotactic Extraction Estimation. The rCBF responses in the deep brain structures after administration of antiparkinsonian drugs tended to be larger than those in cortical areas. Among these deep brain structures, the rCBF increases in the substantia nigra (SN), lateral geniculate (LG) body, and medial geniculate (MG) body correlated with drug efficacy (p < 0.05, respectively). A subgroup analysis revealed that the motor improvement-related rCBF change in the MG was statistically significant, irrespective of cognitive function, but the significant changes in the LG and SN were not found in subjects with dementia. In conclusion, our SPECT study clearly exhibited drug-driven rCBF changes in PD patients, and we newly identified motor improvement-related rCBF changes in the LG and MG. These results suggest that rCBF changes in these regions could be considered as candidates for clinical indicators for objective evaluation of disease progression. Furthermore, functional studies focusing on the LG and MG, especially in relation to therapies using audio-visual stimuli, may bring some new clues to explain the pathophysiology of PD.

[Exploration of pathogenesis-associated molecules and development of disease models for sporadic ALS].

The mechanism underlying the characteristic selective motor neuron degeneration in amyotrophic lateral sclerosis (ALS) has remained elusive. Modest advances in this research field have been achieved by the identification of copper/zinc superoxide dismutase 1 (SOD1) as one of the causative genes for rare familial ALS and by the development and analysis of mutant SOD1 transgenic animal models. However, in sporadic ALS (SALS) with many more patients, causative or critical genes situated upstream of the disease pathway have not yet been elucidated and no available disease models have been established. We have been working on screening these genes employing and combining several new technologies such as cDNA microarray, molecular indexing, and laser capture microdissection. Many of the resultant genes are of intense interest and may provide a powerful tool for determining the molecular mechanisms of SALS. Of these, in this paper, we will focus on Dorfin, a RING finger-type E3 ubiquitin ligase and dynactin1, a major component of dynein/dynactin complex that is important for retrograde axonal transport. We are now challenging creation of the disease models by simulating the gene expression changes specifically observed in SALS patients.

Gene expressions specifically detected in motor neurons (dynactin 1, early growth response 3, acetyl-CoA transporter, death receptor 5, and cyclin C) differentially correlate to pathologic markers in sporadic amyotrophic lateral sclerosis.

In a differential gene expression profile, we showed previously that dynactin 1 (DCTN1), early growth response 3 (EGR3), acetyl-CoA transporter (ACATN), death receptor 5 (DR5), and cyclin C (CCNC) were prominently up- or downregulated in motor neurons of sporadic amyotrophic lateral sclerosis (ALS). In the present study, we examined the correlation between the expression levels of these genes and the levels of pathologic markers for motor neuron degeneration (i.e. cytoplasmic accumulation of phosphorylated neurofilament H [pNF-H] and ubiquitylated protein) and the numbers of residual motor neurons in 20 autopsies of patients with sporadic ALS. DCTN1 and EGR3 were widely downregulated, and the changes in gene expression were correlated to the number of residual motor neurons. In particular, DCTN1 was markedly downregulated in most residual motor neurons before the accumulation of pNF-H, even in cases with well-preserved motor neuron populations. ACATN, DR5, and CCNC were upregulated in subpopulations of residual motor neurons, and their expression levels were well correlated with the levels of pNF-H accumulation and the number of residual motor neurons. The expressions of DCTN1, EGR3, ACATN, and DR5 were all markedly altered before ubiquitylated protein accumulation. DCTN1 downregulation appears to be an early event before the appearance of neurodegeneration markers, whereas upregulations of DR5 and CCNC are relatively later phenomena associated with pathologic markers and leading to neuronal death. The sequence of motor neuron-specific gene expression changes in sporadic ALS can be beneficial information in developing appropriate therapeutic strategies for neurodegeneration.

Gene expression profiling toward understanding of ALS pathogenesis.

Although more than 130 years have gone by since the first description in 1869 by Jean-Martin Charcot, the mechanism underlying the characteristic selective motor neuron degeneration in amyotrophic lateral sclerosis (ALS) has remained elusive. Modest advances in this research field have been achieved by the identification of copper/zinc superoxide dismutase 1 (SOD1) as one of the causative genes for rare familial ALS (FALS) and by the development and analysis of mutant SOD1 transgenic mouse models. However, in sporadic ALS (SALS) with many more patients, causative or critical genes situated upstream of the disease pathway have not yet been elucidated and no available disease models have been established. To approach genes causative or critical for ALS, gene expression profiling in tissues primarily affected by the disease has represented an attractive research strategy. We have been working on screening these genes employing and combining several new technologies such as cDNA microarray, molecular indexing, and laser capture microdissection. Many of the resultant genes are of intense interest and may provide a powerful tool for determining the molecular mechanisms of ALS. However, we have barely arrived at the starting point and are confronting an enormous number of genes whose roles remain undetermined. Challenging tasks lie ahead of us such as identifying which genes are really causative for ALS and developing a disease model of SALS with due consideration for the expression changes in those genes.

[Dyskinesia-hyperpyrexia syndrome in a patient with Parkinson's disease: a case report].

Non-physiological, excessive dopaminergic stimulation can cause dyskinesia-hyperpyrexia syndrome (DHS), which was initially reported by Gil-Navarro and Grandas in 2010. A 70-years-old woman with a 13-years history of Parkinson's disease (PD) was hospitalized due to difficulty walking, despite being treated with levodopa/carbidopa (600 mg/day), immediate-release pramipexole (3 mg/day), and selegiline (5 mg/day). Immediate-release pramipexole was changed to extended-release pramipexole without changing the dose or levodopa equivalent dose (LED). The patient's adherence to drugs was good. The parkinsonism gradually improved and the patient was discharged. One month later, the patient developed severe generalized athetotic dyskinesia with visual hallucinations and hyperpyrexia that lasted for a week, and she was readmitted to hospital. On admission, the patient was conscious but slightly disoriented. Body temperature was 40.3°C with hyperhidrosis. Leukocyte count in the peripheral blood was 1.78×10(4)/ml and serum creatine kinase was >3×10(4) U/l. Chest survey, whole-body computed tomography, and cranial magnetic resonance imaging showed no abnormalities. The patient was diagnosed with DHS and treated by tapering the oral administration of dopaminergic drugs, including extended-release pramipexole. Her clinical condition recovered without dyskinesia, and serum creatine kinase level swiftly normalized. DHS and resemblant conditions are reported to occur in long-term PD patients with motor complications. In advanced stage PD, loss of dopaminergic neurons impairs the dopamine holding capacity of the striatum and exogenous dopaminergic drugs can result in uncontrollable and excessive fluctuations in dopamine concentration. Our case recommends caution when switching to long-acting dopaminergic drugs, even if the dose is unchanged, could lead to excessive dopaminergic stimulation. This case highlights the importance of considering both the LED and the duration of action of dopaminergic drugs when adjusting medication.

Differentially expressed genes in sporadic amyotrophic lateral sclerosis spinal cords--screening by molecular indexing and subsequent cDNA microarray analysis.

To analyze the genes related to the pathophysiology of sporadic amyotrophic lateral sclerosis (SALS) we performed gene profiling of SALS spinal cords using molecular indexing combined with cDNA microarray. Eighty-four fragments were cloned in the first screening procedure with molecular indexing. Subsequent quantitative microarray screening revealed 11 genes which were differentially expressed in SALS. Real-time RT-PCR verified that the expression level of the following six genes was altered in SALS: dorfin, metallothionein-3, 30 kDa TATA-binding protein-associated factor, neugrin, ubiquitin-like protein 5 and macrophage-inhibiting factor-related protein-8. These results indicated that genes associated with the ubiquitin-proteasome system, oxidative toxicity, transcription, neuronal differentiation and inflammation might be involved in the pathogenesis of SALS.

Hsp70 and Hsp40 improve neurite outgrowth and suppress intracytoplasmic aggregate formation in cultured neuronal cells expressing mutant SOD1.

Mutations of the superoxide dismutase 1 (SOD1) gene cause familial amyotrophic lateral sclerosis (FALS). Intracytoplasmic aggregate formation consisting of mutant SOD1 is the histological hallmark of FALS. Since a previous report revealed that Hsp70 reduced aggregate formation and cell death in a cell model of FALS, here we examined the combined effects of Hsp70 and its cofactor, Hsp40, on a cell model of FALS. The combination of Hsp70 and Hsp40 reduced intracytoplasmic aggregates and markedly improved neurite outgrowth. They also prevented cell death to a relatively lesser extent. Neurite outgrowth was recognized almost exclusively in the cells without intracytoplasmic aggregates. Hsp70 and Hsp40 were upregulated in cells expressing mutant SOD1, and were colocalized with intracytoplasmic aggregates of mutant SOD1. These findings suggest that heat shock proteins (HSPs) promote neurite outgrowth by suppressing intracytoplasmic aggregate formation and restoring cellular dysfunctions. This is the first demonstration that overexpression of HSPs improved neurite outgrowth as it suppressed intracytoplasmic aggregate formation and cell death in a cultured neuronal cell model of FALS. These findings may provide a basis for the utilization of HSPs in developing a treatment for FALS.