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Aims: Tacrolimus-a widely used immunosuppressant to prevent allograft rejection after organ transplantation-is nephrotoxic, increasing the risk of kidney injury accompanied by kidney fibrosis. The mammalian target of rapamycin (mTOR) inhibitor, everolimus, is an immunosuppressant used together with tacrolimus. Although mTOR signaling inhibition has been demonstrated to exhibit antifibrotic effects, the efficacy of everolimus against tacrolimus-induced kidney fibrosis has not been explored. Therefore, we evaluated the protective effects of everolimus against tacrolimus-induced kidney fibrosis., Main Methods: To assess antifibrotic effect of everolimus against tacrolimus-induced kidney fibrosis, male Wistar rats were subcutaneously administered vehicle or tacrolimus (5 mg/kg per day) and/or everolimus (0.2 mg/kg per day) for 2 weeks after bilateral renal ischemia for 45 min. The antifibrotic effect of everolimus was also assessed using rat kidney fibroblast cell line (NRK-49F)., Key Findings: Tacrolimus administration increased predominant profibrotic cytokine transforming growth factor-β (TGF-β) and fibroblast activation marker α-smooth muscle actin (α-SMA) expression and promoted the infiltration of macrophages in the kidney cortex, resulting in renal interstitial fibrosis in rats. Tacrolimus increased serum creatinine, blood urea nitrogen, kidney injury molecule-1 (KIM-1), and kidney injuries, such as tubular dilation, vacuolization, and glomerular atrophy. Everolimus administration attenuated tacrolimus-induced kidney fibrosis and the associated abnormalities. Everolimus strongly suppressed TGF-β-induced kidney fibroblast activation and extracellular matrix protein expression by the mTOR signaling inhibition., Significance: We demonstrated that everolimus attenuates tacrolimus-induced renal interstitial fibrosis in rats. Owing to its protective effect against tacrolimus-induced kidney fibrosis, everolimus may be useful when used concomitantly with tacrolimus., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Dysferlin-deficient SJL mice are commonly used to study dysferlinopathy. We demonstrated that poloxamer 188 (P188), a membrane sealant, is effective in reducing the loss of muscle mass in SJL mice when administered using an osmotic pump for 6 weeks. We did not observe significant changes over a 2-week administration period, suggesting that longthier observation is necessary to determine the effectiveness of P188. We also examined exercise endurance in P188-administered SJL mice using a rolling cage. Phosphorylated p38 was found to be reduced in P188-administered SJL mice; additionally, using microarray analysis, we found diminished expression of atrogin-1, an E3 ubiquitin ligase, as the effector of muscular atrophy. Chronic infusion of P188 to dysferlin-deficient SJL mice reduced muscular atrophy, and administering p38 and atrogin-1 in the gastrocnemius muscle improved its motor function. These results provide a basis for potential treatments for dysferlin-deficient skeletal muscle fibers., (Copyright © 2011 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.)

Previously, we demonstrated that neuronal nitric oxide synthase (nNOS) is activated and promotes muscle atrophy in skeletal muscle during tail suspension, a model of unloading and denervation. Here, we examined patients with amyotrophic lateral sclerosis (ALS) and mutant (H46R) SOD1 transgenic (Tg) mice model using immunohistochemistry, Western blotting and real time PCR. We found cytoplasmic nNOS staining of angulated muscle fibers in patients with ALS. We also examined mutant SOD1 Tg mice and found cytoplasmic nNOS staining even before the onset of clinical muscle atrophy. In the Tg mice, nNOS was largely extracted with 100 mM NaCl and barely detected in the pellet fraction, suggesting fragile anchoring of nNOS to the sarcolemma. We also showed an elevated expression of atrogin-1, key molecules in muscle atrophy at the end stage. A common nNOS dislocation/atrogin-1/muscle atrophy pathway among tail suspension, denervation and ALS is suggested. nNOS modulation therapy may be beneficial in several types of muscle atrophy., (Copyright 2010 Elsevier B.V. All rights reserved.)

Hyposmia in Parkinson's disease (PD) was evaluated by using neuroimaging techniques. It is well known that olfactory impairments are one of the cardinal non-motor symptoms in PD. However, all smell tests used in previous studies depend on subjective answers by examinees and on sniffing of odorants, the latter of which may be impaired in PD as a consequence of motor impairments. We developed an fMRI system, which can visualize brain activation by olfactory stimuli during natural breathing. Although 7 age-matched controls demonstrated significant activations in various brain areas including precentral gyrus (BA6/6) and middle temporal gyrus (BA19/39) by the odorant stimuli, 9 patients with PD showed little activations by the same stimuli. These data suggest that the olfactory dysfunction in PD is not a simple reflection of impaired sniffing. Recent epidemiological studies demonstrate that the olfactory impairments may precede the onset of motor symptoms. Moreover, several pathological studies suggest that amygdala is one of the most frequently affected regions and is closely related to hyposmia in PD. Further brain imaging studies of hyposmia will shed light on the early pathological changes in PD.

There are two distinct phenotypes of multiple sclerosis (MS) in Asians, optic-spinal MS (OSMS) and conventional MS (CMS). In 2004, we performed the fourth nationwide epidemiological survey of MS. The epidemiological features were reported elsewhere; here we report the characteristic features of patients with each MS phenotype, classified according to the clinically estimated sites of involvement and MRI findings. Among 1493 MS patients collated, 57.7% were classified as having CMS and 16.5% were classified as having OSMS. Based on MRI findings, MS patients were further subdivided into those with OSMS with or without longitudinally extensive spinal cord lesions (LESCLs) and those with CMS with or without LESCLs. Although disease duration did not differ significantly among the four groups, EDSS scores were significantly higher in patients with LESCLs than in those without LESCLs, irrespective of OSMS or CMS phenotype. Similar trends were found for the frequencies of bilateral visual loss, transverse myelitis, and marked CSF pleocytosis and neutrophilia. Increased IgG index, brain lesions fulfilling the Barkhof criteria and secondary progression were more commonly found in CMS patients than in OSMS patients, while negative brain MRIs were more commonly encountered in OSMS patients than CMS patients, irrespective of the presence of LESCLs. These findings suggest that demographic features not only vary based on CMS or OSMS phenotype, but also with the presence or absence of LESCLs, and that nonetheless, these four phenotypes constitute a continuum.

CCR7 and its ligands CCL19 and CCL21 are a key chemokine system in T cell priming in secondary lymphoid organs, and are rarely expressed in normal muscle tissue. We immunohistochemically investigated the expression of this chemokine system in the muscles of seven patients with inclusion body myositis (IBM). In all cases, CCR7+ mononuclear cells infiltrated in the endomysium, preferentially surrounded and invaded nonnecrotic muscle fibers. Double immunostaining revealed that such CCR7+ mononuclear cells included BDCA-1+ myeloid dendritic cells as well as CD8+ cells, CD4+ cells and CD68+ macrophages. On the other hand, CCL19 was widely expressed on muscle fibers including those invaded by mononuclear cells. CCL19 was also expressed diffusely on endomysial mononuclear cells and endothelium of vessels. Immunoreactivities of CCL21 were detected on some muscle fibers and mononuclear cells. By RT-PCR analyses, mRNA of CCR7 was detected in all the patients and that of CCL19 and CCL21 was detected in six. These findings showed that the CCL19, CCL21/CCR7 chemokine system is expressed in muscles of IBM. The chemokine mediated attraction in dendritic and other immune cells and muscle cells may be crucial in sustained antigen presentation, T cell activation and immune attack to muscles in the pathogenesis of IBM.

Polymyositis is an autoimmune disorder in which autoaggressive CD8(+) T cells are important in the pathogenesis. However, the mechanisms underlying sustained recruitment of these cells in the muscle tissue are still unknown. CCR7 and its ligands CCL19 and CCL21 are a chemokine system related to mononuclear cell migration and antigen presentation, and are suggested to play a key role in several autoimmune disorders. We investigated the expression of CCR7, CCL19 and CCL21 in frozen muscles of polymyositis. In immunohistochemistry, CCR7 was expressed mainly on mononuclear cells that infiltrated in the endomysium of polymyositis. 34.8+/-9.4% of endomysial mononuclear cells expressed CCR7. By double immunostaining, about 60% of endomysial CD8(+) T cells that surrounded the nonnecrotic muscle fibers coexpressed CCR7. Because most endomysial CD8(+) T cells expressed CD45RO, these were regarded as CD45RO(+)CCR7(+)CD8(+) T cells. On the other hand, CCL19 was expressed mainly on muscle fibers in proximity to CCR7(+) mononuclear cells, on the endothelium of the vessels and some mononuclear cells. CCL21 immunoreactivities were found on small numbers of mononuclear cells. In some cases, CCL21 immunoreactivities were also found on muscle fibers and the endothelium of vessels. In RT-PCR analysis, transcripts of CCR7 and CCL21 were detected in all the polymyositis muscles examined and that of CCL19 was detected in five out of seven polymyositis muscles. The CCL19,CCL21/CCR7 chemokine system is expressed in inflamed muscles of polymyositis and may be involved in the pathomechanism of polymyositis.

Accumulation of misfolded Cu/Zn superoxide dismutase (SOD1) occurs in patients with a subgroup of familial amyotrophic lateral sclerosis (fALS). To identify the conversion of SOD1 from a normally soluble form to insoluble aggregates, we investigated the change of SOD1 solubility with aging in fALS-linked H46R SOD1 transgenic mice. Mutant SOD1 specifically altered to insoluble forms, which were sequentially separated into Triton X-100-insoluble/sodium dodecyl sulfate (SDS)-soluble and SDS-insoluble/formic acid-soluble species. In spinal cords, the levels of SDS-dissociable soluble SOD1 monomers and SDS-stable soluble dimers were significantly elevated before motor dysfunction onset. In COS-7 cells expressing H46R SOD1, treatment with proteasome inhibitors recapitulated the alteration of SOD1 solubility in transgenic mice. In contrast, overexpression of Hsp70 reduced accumulation of mutant-specific insoluble SOD1. SDS-soluble low molecular weight species of H46R SOD1 may appear as early misfolded intermediates when their concentration exceeds the capacity of the proteasome and molecular chaperones.

The SJL mouse is a model for human dysferlinopathy (limb-girdle muscular dystrophy type 2B and Miyoshi myopathy). We used cDNA microarrays to compare the expression profiles of 10,012 genes in control and SJL quadriceps femoris muscles in order to find genes involved in the degeneration and regeneration process and in dysferlin's functional network. Many genes involved in the process of muscle regeneration are observed to be up-regulated in SJL mice, including cardiac ankyrin repeated protein (CARP), Neuraminidase 2, interleukin-6, insulin-like growth factor-2 and osteopontin. We found the upregulation of S100 calcium binding proteins, neural precursor cell expressed, developmentally down-regulated gene 4-like (NEDD4L) with C2 domain, and intracellular protein traffic associated proteins (Rab6 and Rab2). These proteins have the potential to interact with dysferlin. We must reveal some other molecules which may work with dysferlin in order to clarify the pathological network of dysferlinopathy. This process may lead to future improvements in the therapy for human dysferlinopathy.

We examined the effects of perindopril on the dopaminergic system in mice after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment. The mice received four intraperitoneal injections of MPTP at 1-h intervals. Administration of perindopril showed dose-dependent neuroprotective effects against striatal dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) depletion 3 days after MPTP treatment. Our immunohistochemical study showed that MPTP can severe damage in tyrosine hydroxylase (TH)-immunoreactive neurons after MPTP treatment. The administration of perindopril significantly attenuated MPTP-induced substantia nigra and striatal damage. The present study also showed that the immunoreactivity of parvalbumin (PV)- or neuronal nitric oxide synthase (nNOS)-positive cells in the substantia nigra was decreased 7 days after MPTP treatment, whereas no significant changes were observed in these cells of the striatum throughout the experiments. The administration of perindopril significantly attenuated MPTP-induced decrease of the PV- or nNOS-immunoreactivity in the nigral cells. In double-labeled immunostaining with anti-PV and anti-nNOS antibody, PV-immunoreactive cell bodies and fibers were not double-labeled for nNOS-immunoreactive cell bodies and fibers in both the striatum and substantia nigra after MPTP treatment. Furthermore, PV- or nNOS-immunoreactive cell bodies and fibers in both the striatum and substantia nigra were not double-labeled for TH-immunoreactive cell bodies and fibers. These results demonstrate that the ACE inhibitor perindopril has a dose-dependent protective effect against MPTP-induced striatal dopamine, DOPAC and HVA depletion in mice. The present study also demonstrates that perindopril is effective against MPTP-induced degeneration of the nigral neurons and interneurons. Furthermore, our immunohistochemical study suggests that PV-immunoreactive cells and nNOS-immunoreactive cells are different interneurons in both the striatum and substantia nigra. Thus, our results provide further evidence that the ACE inhibitor perindopril may offer a novel therapeutic strategy for Parkinson's disease (PD).

Background: Paraplegia is a serious complication of operations on the thoracic and thoracoabdominal aorta. To investigate the mechanism by which motor neurons are damaged during these operations, we have reported a rabbit model of spinal cord ischemia. We also tested whether a free radical scavenger MCI-186 that is useful for treating ischemic damage in the brain can protect against ischemic spinal cord damage., Methods: Fifteen minutes of ischemia was induced, then MCI-186 or vehicle was injected intravenously. Cell damage was analyzed by observing the function of the lower limbs and by counting the number of motor neurons. To investigate the mechanism by which MCI-186 prevents ischemic spinal cord damage, we observed the immunoreactivity of 8-hydroxy-2'-deoxyguanosine as an oxidative DNA damage marker and redox effector as a DNA repair marker., Results: In sham control, 8-hydroxy-2'-deoxyguanosine was not observed, and the nuclear expression of redox effector was observed. In vehicle injection group (group I), the nuclear expression of 8-hydroxy-2'-deoxyguanosine was observed at 1 and 2 days after reperfusion. The nuclear expression of redox effector was observed at 8 hours and 1 day, and disappeared at 2 days after transient ischemia. In MCI-186 injection group (group M), the nuclear expression of 8-hydroxy-2'-deoxyguanosine was not observed, and redox effector was observed at 8 hours and 1 and 2 days., Conclusions: These results suggest that redox effector decreased in motor neurons after transient ischemia and this reduction preceded oxidative DNA damage. MCI-186 works as a radical scavenger and reduced oxidative DNA damage, so redox effector did not disappear. MCI-186 could be a strong candidate for a use as a therapeutic agent in the treatment of ischemic spinal cord injury.

We examined the neuroprotective effects of a novel astrocyte-modulating agent, (R)-(-)-2-propyloctanoic acid (ONO-2506), in a mouse model of Parkinson's disease. Male C57BL/6 mice received four intraperitoneal injections of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) (10 mg/kg) at 1-h intervals. Dopamine content in the striatum, measured with HPLC 3 days after MPTP injection, was reduced to 23% of control. But this dopamine depletion was dose-dependently prevented by repeated treatments with ONO-2506 (3, 10 and 30 mg/kg, i.p.) administered 1, 6, 24 and 48 h after MPTP injection (51% of control in 30 mg/kg group, p<0.01). ONO-2506 treatment (30 mg/kg) started after 6 h, followed by treatments at 24 and 48 h, also prevented the reduction of dopamine content (42% of control vs. 11% of control in the saline-treated group, p<0.01). We also performed immunohistochemistry for tyrosine hydroxylase (TH) and glial fibrillary acidic protein (GFAP). The MPTP injection resulted in a loss of TH-positive dopaminergic neurons (42% of control, p<0.01) in the substantia nigra after 7 days, but ONO-2506 treatment prevented this neuronal loss (70% of control, p<0.01). The MPTP injection led to reactive astrocytosis in the striatum after 7 days, but ONO-2506 induced earlier, moderate astrocytic activation after 3-7 days. These findings show that ONO-2506 protects dopaminergic neurons against MPTP neurotoxicity probably through facilitating astrocytic support for neuronal recovery from injury. Pharmacological modulation of astrocytes may offer a novel therapeutic strategy for Parkinson's disease.

Mutations in a Cu, Zn-superoxide dismutase (SOD1) cause motor neuron death in human familial amyotrophic lateral sclerosis (FALS) and its mouse model, suggesting that mutant SOD1 has a toxic effect on motor neurons. However, the question of how the toxic function is gained has not been answered. Here, we report that the mutant SOD1s linked to FALS, but not wild-type SOD1, aggregated in association with the endoplasmic reticulum (ER) and induced ER stress in the cDNA-transfected COS7 cells. These cells showed an aberrant intracellular localization of mitochondria and microtubules, which might lead to a functional disturbance of the cells. Motor neurons of the spinal cord in transgenic mice with a FALS-linked mutant SOD1 also showed a marked increase of GRP78/BiP, an ER-resident chaperone, just before the onset of motor symptoms. These data suggest that ER stress is involved in the pathogenesis of FALS with an SOD1 mutation.

Objective: The mechanism of spinal cord injury has been thought to be related to the vulnerability of spinal motor neuron cells against ischemia. However, the mechanisms of such vulnerability are not fully understood. We previously reported that spinal motor neurons may be lost by programmed cell death and thus now investigate a possible mechanism of neuronal death with immunohistochemical analysis for 8-hydroxy-2'-deoxyguanosine (8-OHdG) and redox factor-1 (Ref-1)., Methods: We used a rabbit spinal cord ischemia model with a balloon catheter. The spinal cord was removed at 8 hours, 1, 2, or 7 days after 15 minutes of transient ischemia, and histologic changes were studied with hematoxylin-eosin staining. Western blot analysis for Ref-1, temporal profiles of 8-OHdG and Ref-1 immunoreactivity, and double-label fluorescence immunocytochemical studies were performed., Results: Most motor neurons were preserved until 2 days but were selectively lost at 7 days of reperfusion. Western blot analysis of a sample from sham control spinal cord showed a characteristic 37-kDa band that was reduced after ischemia. Immunohistochemistry showed the nuclear expression of Ref-1 in motor neurons of control spinal cords, and immunoreactivity was decreased 1 day after ischemia. On the other hand, no nuclear expression was seen of 8-OHdG in motor neurons of control spinal cords, and immunoreactivity was increased 1 day after ischemia. Double-label fluorescence immunocytochemical study revealed that both 8-OHdG and Ref-1 were positive at 8 hours of reperfusion in the same motor neurons, which eventually die., Conclusion: These results suggest that Ref-1 decreased in motor neurons after transient spinal cord ischemia and that this reduction preceded oxidative DNA damage. The reduction of Ref-1 protein at the moderately late stage of reperfusion may be one of the factors responsible for the delay in neuronal death after spinal cord ischemia.

OX40 is selectively expressed on activated autoreactive memory T cells and these OX40+ lymphocytes may play a crucial role in autoimmune diseases. To determine whether OX40+ lymphocytes are involved in the pathomechanism of human inflammatory muscle diseases, we immunohistochemically examined the distribution of OX40+ cells in muscles from patients with polymyositis and granulomatous myopathy, and compared with that of cells bearing other activation markers, such as IL-2 receptor (IL-2R) and HLA-DR. In polymyositis, OX40+ mononuclear cells were found predominantly in the perivascular sites and to a lesser degree in the endomysium. Scanty IL-2R+ mononuclear cells were located only in the endomysium and HLA-DR was expressed on half of the mononuclear cells distributed diffusely in the perivascular sites and in the endomysium. Mononuclear cell infiltration in the perivascular sites was greater in the muscles in which OX40+ cells were present in the perivascular sites than in those without OX40+ cells in the perivascular sites (p<0.05). In granulomatous myopathy, OX40+ cells were detected in the centers of the granulomas. In contrast, IL-2R+ cells were present at the periphery of the granulomas and HLA-DR was detected on mononuclear cells throughout the granulomas. OX40+ mononuclear cells with specific distributions in muscles may be involved in the pathomechanism of polymyositis and granulomatous myopathy, and can be a candidate molecule of selective immunotherapy in these diseases.

The changes of regional cerebral blood flow (rCBF) in Parkinson's disease (PD) were investigated. Because of individual differences in brain volume and the extent of brain atrophy, previous functional imaging studies involved potential methodological difficulties. In this study, using the statistical parametric mapping technique, 99mTechnetium-labeled hexamethylpropyleneamineoxime brain single-photon emission computed tomography images from 18 patients with PD were transformed into standard brain-based stereotaxic coordinate spaces and then compared with such images for 11 control subjects matched for age and extent of brain atrophy. A rCBF decrement in the supplementary motor area (SMA) and such decrement in the dorsolateral prefrontal cortex (DLPFC) were observed in the summarized PD images as compared with controls (p<0.005). In a subgroup in the Hoehn-Yahr III/IV stage (11 cases), the rCBF decrement was demonstrated not only in the SMA, but also in the DLPFC and insular cortex (p<0.001). There was a correlation between the degree of the rCBF decrement in the DLPFC or the insular cortex and the score of the unified Parkinson's disease rating scale (p<0.05), while the rCBF decrement in the SMA showed no relationship with the severity of disease. The function of the SMA is closely associated with the nigro-striatal pathway and its impairment can explain the basic akinetic symptoms in PD, which are responsive to L-DOPA treatment. On the other hand, the DLPFC and insular cortex may play key roles in specific symptoms of impairment at advanced stages, such as impaired working memory, postural instability and autonomic dysfunction. We hypothesize that the impairment of the DLPFC and insular function is correlated with the progression of the disease and is related to DOPA-refractory symptoms, which are major problems in the care of patients with advanced PD.

We investigated possible motor effects of 7-nitroindazole (7-NI), an neuronal nitric oxide synthase (nNOS) inhibitor, and N(G)-nitro-L-arginine methyl ester (L-NAME), an non-selective NOS inhibitor in mice using catalepsy and pole tests in comparison with dopamine D(2) receptor antagonist, haloperidol. We also studied the change in dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) contents of these compounds. The administration of 7-NI and L-NAME (40-160 mg/kg, s.c.) dose-dependently induced motor deficit in both catalepsy and pole tests. The motor deficit induced by 7-NI was more pronounced than the one produced by L-NAME. In contrast, haloperidol showed a marked motor deficit in mice. Haloperidol showed a marked motor deficit as compared with 7-NI and L-NAME. For dopamine, DOPAC and HVA contents, haloperidol exhibited a significant decrease in dopamine content and a significant increase in DOPAC and HVA content in the striatum. In contrast, 7-NI showed a significant increase in the striatal dopamine content. However, 7-NI had no significant change in the striatal DOPAC and HVA contents. On the other hand, no significant change in the striatal dopamine, DOPAC and HVA contents was observed in L-NAME-treated mice. The present study also showed that the motor deficit induced by 7-NI or L-NAME was significantly attenuated by the treatment with L-arginine. These results demonstrate that NOS inhibitors as well as dopamine D(2) receptor antagonist haloperidol can induce motor deficit in mice. The present study also suggests that the mechanism in the motor deficit caused by NOS inhibitors may be different from that in the motor deficit induced by haloperidol. Furthermore, our findings suggest that nNOS may play some role in control of motor behavior.

We measured the central motor conduction time (CMCT), central sensory conduction time (CSCT), F wave and mean F wave/M wave amplitude ratio in patients with HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP) and controls. CMCTs in upper (U) and lower (L) limbs were significantly prolonged in HAM/TSP. L-CSCT was significantly prolonged in HAM/TSP, but U-CSCT in HAM/TSP and controls were not significantly different. CMCT and CSCT were significantly correlated in HAM/TSP. U-CMCT, but not L-CMCT, correlated with the clinical severity of HAM/TSP. Although F wave conduction velocity and its occurrence were normal in HAM/TSP, U- and L-mean F wave/M wave amplitude ratio tended to be higher in HAM/TSP, and the L-mean F wave/M wave amplitude ratio was significantly correlated with the L- and thoracic CMCT. These findings demonstrate that the prolongation of CMCT sensitively reflects the extension of the lesions and the disinhibition to the anterior horn cells in HAM/TSP.

Mammalian sialidases have been reported to give a great influence on a number of cellular functions including cell differentiation and cell growth by removal of sialic acids from glycoproteins and gangliosides. To understand the roles of the sialidases during development, we investigated expression pattern of three types of sialidase in developing rat brain and liver. For this purpose we cloned a new membrane-associated sialidase cDNA from rat brain. The cDNA encodes 418 amino acids containing three ASP-boxes characteristic of sialidases and the major transcript of 3.5 kb is highly expressed in brain and cardiac muscle but low in liver. Competitive polymerase chain reaction methods were developed to evaluate the mRNA level together with activity assays in comparison with cytosolic and lysosomal sialidases previously obtained. The results indicate that the expression of individual sialidase genes is spatiotemporally controlled with distinct roles in determining the concentration and components of sialo-glycoconjugates during development., (Copyright 2001 Academic Press.)

Autosomal dominant cerebeller ataxias (ADCAs) are a heterogeneous group of neurodegenerative disorders that differ in both the clinical manifestations and modes of inheritance. At present, eight different genes causing ADCAs have been found: spinocerebeller ataxia type 1 (SCA1), SCA2, SCA3/Machado-Joseph disease (MJD), SCA6, SCA7, SCA8, SCA12 and dentatorubropallidoluysian atrophy (DRPLA). The relative prevalence of each mutation varies according to race and native place. We studied 117 unrelated ADCA families that originated from the Tohoku District in the northernmost part of Honshu Island in Japan (mainly Miyagi Prefecture in the central part of Tohoku District). The SCA1 mutation was the most frequent among the known disorders (24.8% of all such families). The relative prevalence of SCA1 in the Tohoku District is very high compared with the values already reported from other regions in the world. Because the population of this area had seldom moved, the alleles with SCA1 mutations (including alleles with an intermediate CAG repeat number) are assumed to have been present in this area for a long time.

Quantitative receptor autoradiography was used to examine the sequential patterns of changes in dopaminergic and glutamatergic receptors in the brain of rats lesioned with 6-hydroxydopamine. The animals were unilaterally lesioned in the medial forebrain bundle and the brains were analyzed at 1, 2, 4 and 8 weeks of postlesion. Degeneration of the nigrostriatal pathway caused a significant increase in dopamine D(2) receptors in the ipsilateral striatum from 1 to 8 weeks of postlesion. In the ipsilateral substantia nigra (SN), a significant decrease in dopamine D(2) receptors was also observed from 1 to 8 weeks of postlesion. On the other hand, dopamine D(1) receptors were increased in the ipsilateral ventromedial striatum from 2 to 4 weeks of postlesion. In the ipsilateral SN, a transient increase in dopamine D(1) receptors was observed only 1 week after lesioning. However, other regions in both ipsilateral and contralateral sides showed no significant change in dopamine D(1) and D(2) receptors during postlesion except for a transient change in a few regions. N-Methyl-D-aspartate (NMDA) receptors showed no significant changes in all brain regions studied during the postlesion. In contrast, a transient increase in excitatory amino acid transport sites was observed only in the frontal cortex and ventromedial striatum of the ipsilateral side at 2 weeks of postlesion. However, glycine receptors showed a significant change in any brain areas of both ipsilateral and contralateral sides after lesioning. The change in the brain areas of contralateral side was more pronounced than that of ipsilateral side for glycine receptors. In addition, dopamine uptake sites showed a severe damage in the ipsilateral striatum from 1 to 8 weeks after lesioning. In the contralateral side, in contrast, no significant change in dopamine uptake sites was found in the striatum during the postlesion. These results indicate that unilateral injection of 6-hydroxydopamine in the medial forebrain bundle can cause a significant increase in dopamine D(1) and D(2) receptors in the striatum. The increase in dopamine D(2) receptors was more pronounced than that in dopamine D(1) receptors in the striatum after 6-hydroxydopamine treatment. In contrast, dopamine uptake sites showed a severe damage in the striatum during the postlesion. Furthermore, our results support the existence of dopamine D(2) receptors on the neurons of SN, but not dopamine D(1) receptors. For glutamatergic receptor system, the present study suggests that the changes in glycine receptors may be more susceptible to degeneration of nigrostriatal pathway than NMDA receptors and excitatory amino acid transport sites. Thus, our findings are of interest in relation of degeneration of the nigrostriatal pathway that occurs in Parkinson's disease

We report the clinical and MRI findings of two patients with familial spinal arachnoiditis. Although their initial symptoms were various, they both showed spastic paraparesis and sensory disturbance below the thoracic level. Cytokines and WBC in the CSF were studied, but they were not elevated at all. The spinal magnetic resonance images of each showed extensive arachnoiditis and a cystic structure. The other impressive features included: (i) an enhancement within the thickened arachnoid and an adhesion between the spinal cord and the dura mater, (ii) deformation of the thoracic cord where the arachnoid adhered, and (iii) secondary syrinx formation. Laminectomy may have an adverse outcome for such patients.

Bilirubin, an efficient antioxidant, is the end product of the heme cleavage pathway, which is catalyzed by heme oxygenase (HO) and biliverdin reductase. Although an inducible form of HO is overexpressed in the Alzheimer's disease (AD) brain, it has not been determined whether bilirubin metabolism is actually activated or not. In this study, we measured CSF-bilirubins and their derivatives using an enzyme-linked immunosorbent assay with two kinds of anti-bilirubin monoclonal antibodies, designated 24G7 and 5M2. In AD patients, the levels of CSF bilirubin derivatives increased significantly compared with those of controls. This increase was not due to the increased permeability of the blood-brain barrier, because the levels of unconjugated bilirubin were not different between AD and controls. These data may reflect the increase of degraded bilirubin metabolites in the AD brain derived from the scavenging reaction against chronic oxidative stress.

Purpose: In an attempt to study whether ischemic spinal cord expresses a foreign gene in vivo, a replication-defective adenoviral vector containing the Escherichia coli lacZ gene was directly injected into the ischemic spinal cord of rabbits, and temporal and spatial profiles of the exogenous gene expression were compared with that of the control spinal cord., Methods: Thirty-nine Japanese domesticated white rabbits weighing 2 to 3 kg were used in this study and were divided into two subgroups, a 15-minute ischemia group and a sham control group. The adenoviral vector was directly injected into lumbar spinal cord by a needle from dorsal spine just after the infrarenal aortic occlusion in the case of ischemia. Animals were allowed to recover at ambient temperature and were killed at 1, 2, 4, and 7 days after reperfusion (n = 3 at each time point)., Results: In the control rabbit, adenoviral vector was transferred into the spinal cord, and the lacZ gene was expressed at dorsal astroglia and anterior motor neurons at 1 to 7 days of reperfusion. After 15 minutes of ischemia, the lacZ gene was expressed at 2 and 4 days of reperfusion in dorsal astroglia and anterior motor neurons, which were positive for Fas antigen., Conclusion: This result suggests that it is possible to transfer and express the lacZ gene in ischemic motor neurons, which eventually show apoptotic change with induction of Fas antigen, and also suggests a great potential of gene therapy for paraplegic patients in the future.

In rare cases, Miller Fisher syndrome (MFS) has been known to recur. However, clinical features of recurrent MFS have not been well analyzed, and the precipitating factors relating to recurrence remain unknown. From 1981 to 1996, we examined four patients with recurrent MFS among 28 Japanese MFS patients. In the four patients, the recurrent episodes occurred after long asymptomatic intervals, ranging from 2.5 to 12.5 years. The clinical and laboratory features of recurrent episodes were similar either to those of the initial episodes or to those of the 24 non-recurrent patients. Of the two patients tested for serum IgG anti-GQ1b antibody, both were positive. Serological HLA typing showed that all recurrent patients were both HLA-Cw3 and -DR2 positive. However, out of 13 non-recurrent patients examined, six had HLA-Cw3, and four had HLA-DR2. The frequency of HLA-DR2 among the recurrent patients was significantly higher than among healthy controls (corrected P = 0.038), and was also higher than among the non-recurrent patients but not significantly. These findings suggest that recurrent MFS is clinically the same as typical MFS and that HLA-DR2 is possibly associated with recurrence.

We investigated the sequential patterns of changes in dopamine uptake sites, D1 and D2 receptors in the brain of animals lesioned with 6-hydroxydopamine using quantitative receptor autoradiography. The rats were unilaterally lesioned in the medial forebrain bundle and the brains were analyzed at 1, 2, 4 and 8 weeks postlesion. Degeneration of the nigrostriatal pathway caused a significant loss of dopamine uptake sites in the ipsilateral striatum, substantia nigra (SN) and ventral tegmental area (VTA) in the lesioned animals. Dopamine D1 receptors were significantly increased in the ventromedial part of striatum of the ipsilateral side from 2 to 4 weeks postlesion. In the ipsilateral SN, a transient increase in dopamine D1 receptors was observed only 1 week after lesioning. However, the frontal cortex, parietal cortex and dorsolateral part of the striatum showed no significant change in dopamine D1 receptors throughout the experiments. On the other hand, dopamine D2 receptors were decreased increased in the ipsilateral SN and VTA from 1 week to 8 weeks postlesion. In the ipsilateral striatum, dopamine D2 receptors were increased in the dorsolateral part from 2 weeks to 8 weeks and in the ventromedial part from 2 weeks to 4 weeks. However, the frontal cortex and parietal cortex showed no significant change in dopamine D2 receptors during postlesion. In the contralateral side, most of regions examined showed no significant change in dopamine uptake sites, dopamine D1 receptors and dopamine D2 receptors during postlesion except for a transient change in a few regions. These results demonstrate that 6-hydroxydopamine can cause a severe functional damage in dopamine uptake sites in the striatum, SN and VTA. Our findings also suggest that the up-regulation in dopamine D2 receptors is more pronounced than that in dopamine D1 receptors in the brain after 6-hydroxydopamine treatment. Furthermore, our results support the existence of dopamine D2 receptors on the neurons of SN and VTA. Thus, our findings provide insights into the pathogenesis of Parkinson's disease.

Although perinuclear anti-neutrophil cytoplasmic antibodies (pANCA) are associated with vasculitic neuropathy, their association with central nervous system (CNS) disorders has not been studied except for one report on optic-spinal type of multiple sclerosis associated with serum pANCA. We examined pANCA in sera from 98 patients with various CNS disorders, such as 58 MS, 17 myelitis, 12 HTLV-1 associated myelopathy, and 11 other CNS diseases using indirect immunofluorescence methods. The results showed serum pANCA to be positive in five patients with a peculiar type of myelitis, including two with MS and three with etiology unknown myelitis. All of these ANCA-positive patients were women and had acute or subacute myelopathy with various severities. MRI revealed segmental swelling of the spinal cord with T2 hyperintensity in the acute stage of the disease. Marked pleocytosis (227.8+/-101/mm3) and elevated protein level (128.8+/-52 mg/dl) in CSF were noted. Four of the patients had anti-nuclear antibodies and two had previous histories of symptoms suggesting autoimmune disorders. In a search for target antigens of pANCA, myeloperoxidase reactivity was found in the sera from two myelitis patients. Clinical and laboratory features of myelitis patients with pANCA in the present study are different from those of typical MS patients. Further study will be needed to delineate the role of pANCA in the pathogenesis of a specific type of myelitis.

We analyzed 13 patients with spinocerebellar ataxia type 2 (SCA2) in seven unrelated families who live in Gunma Prefecture, Japan (population approx. 2,000,000), and documented the clinical and molecular properties correlated with the CAG repeat expansion. Twelve of the 13 patients and one presymptomatic female were genetically examined, and the CAG repeat number of the expanded and normal alleles was 40.8+/-4.8 (mean+/-S.D., n=13) and 22+/-0 (n=13), respectively. The repeat size of the expanded alleles was inversely correlated with the patients' age at onset. Paternal anticipation was observed, accompanied by an increase of the CAG repeat size. The patients presented here were clinically characterized by a relatively higher frequency of slow saccades, hyporeflexia, hypotonia, and tremor. A number of peaks in the expanded allele on polyacrylamide gel electrophoresis showed the presence of cell mosaicism in SCA2 as well. In Gunma Prefecture, SCA2, Machado-Joseph disease and spinocerebellar ataxia type 6 are almost equally present and at higher frequencies than spinocerebellar ataxia type 1 and hereditary dentatorubropallidoluysian atrophy, which are rare. Thus, the difference of frequency of autosomal dominant spinocerebellar ataxias may be present in Japan.

We assayed IL-6 in 105 cerebrospinal fluid (CSF) samples from patients with ALS, MS, HTLV-1 associated myelopathy (HAM), and controls. There was considerable overlap in IL-6 levels in all patient groups. The mean IL-6 in 27 patients with ALS was significantly higher than in 21 patients in the other neurological disease (OND) group (P=0.0075). There were no significant differences in MS or HAM and the OND control group. Overall, CSF IL-6 correlated with protein concentration but not with percentage IgG or IgG-albumin index. Patients with CSF oligoclonal bands were no more likely to have detectable IL-6 than patients without oligoclonal bands. Similarly, IL-6 did not correlate with clinical disease activity in MS when subgroups of patients were compared or when an individual patient was followed over time. The elevated IL-6 in ALS may reflect an ongoing humoral immune response, or IL-6 may be non-specifically expressed in these patients as a putative neurotrophic factor in response to nerve cell degeneration.

To evaluate the clinical characteristics of Machado-Joseph disease (MJD) with reference to CAG repeat length and disease duration, we analyzed neurologic findings in 108 patients from 84 families. The majority of MJD patients presented with an ataxic gait as the initial symptom. Dysarthria and nystagmus were observed from an early stage. Bulging eyes, muscle atrophy and bradykinesia developed later. Patients with a shorter CAG repeat length or later onset had more frequent involvement of proprioceptive sensory deficit. Incidence of abnormal reflexes, tones, and proprioceptive sensation was not associated with disease duration, but with CAG repeat length. Based on these results, we propose a new clinical classification: type A (juvenile type), with hyperreflexia and dystonia, but without a proprioceptive sensory deficit; type C (adult type), with hyporeflexia and a proprioceptive sensory deficit, but without dystonia; and type B (intermediate type), the remaining patients with a mixed presentation.

Dopamine D1 and D2 receptors and uptake sites were studied in the gerbil hippocampus, parietal cortex and thalamus 1 h to 7 days after 10 min of cerebral ischemia using the occlusion of bilateral common carotid arteries. [3H]SCH23390 ([N-methyl-3H]R[+]-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-7-ol-be nzazepine) and [3H]mazindol were used as markers of dopamine D1 receptors and uptake sites, respectively. [3H]Nemonapride was used to label dopamine D2 receptors. No obvious alteration in [3H]SCH23390 and [3H]mazindol binding was found in the hippocampus up to 48 h after ischemia. These bindings showed a significant reduction in the hippocampus after 7 days of recirculation. In contrast, [3H]nemonapride binding was unaffected in the hippocampus during the recirculation periods. The parietal cortex and thalamus also exhibited no significant changes in [3H]SCH23390, [3H]nemonapride and [3H]mazindol binding after ischemia. MAP2 (microtubule-associated protein 2) immunoreactivity was unchanged in all regions up to 48 h after ischemia. Thereafter, a marked loss of MAP2-immunoreactive neurons was observed in the hippocampal CA1 and CA3 neurons 7 days after recirculation. These findings were consistent with histological observations with cresyl violet staining. Our results demonstrate that dopamine D1 receptors and dopamine uptake sites in the hippocampus are susceptible to cerebral ischemia, whereas dopamine D2 receptors in this region are particularly resistant. Furthermore, these findings suggest that dopamine transmission may not be major factor in producing ischemic hippocampal damage.

A case of central nervous system (CNS) leukemia with normal bone marrow, associated with a novel chromosomal abnormality, is described. A 58 year-old woman complained of hearing disturbance, severe headache and vomiting, and showed signs of meningeal irritation, as well as papilledema and bilateral dysacusis. Immature atypical cells were found in the cerebrospinal fluid (CSF) with elevated pressure, pleocytosis, increased protein and decreased glucose levels. She was diagnosed as having neoplastic meningitis. In spite of intensive investigations, including bone marrow puncture, malignancies were not found in organs other than intra-cranial site. The symptoms and CSF findings were temporarily improved with chemotherapy and irradiation, but she relapsed into neoplastic meningitis. The anaplastic cells in CSF were positive with CD45 by immunocytochemistry, and were positive by peroxidase staining. Thus, the anaplastic cells were considered to be myelocytic leukemic cells. Chromosomal analysis showed that these leukemic cells had a novel chromosomal abnormality: 46XX, 4q+, 10q-, 16q-. There has been no report of leukemic meningitis without bone marrow abnormalities. It is possible that this peculiar abnormal chromosome is related to the primary infiltration of the central nervous system. With this novel chromosomal abnormality, this case is important for considering the mechanism of primary leukemic meningitis.

McLeod syndrome is a rare X-linked disorder involving neurological defects and acanthocytosis. We examined the XK gene in three patients with neuroacanthocytosis, one of whom had cardiomyopathy, and his symptoms were very similar to those of McLeod syndrome. We found two new transversions (C to G at codon 204 and G to C at codon 205) in exon 3 in all those cases. However, the transversion at codon 205 was found in all 70 Japanese normal subjects and four non-Japanese (two Caucasian males, one Chinese female and one Micronesian female) and that at codon 204 was also detected in all 14 normal Japanese males and the four non-Japanese. These findings suggest that they are not the cause of McLeod syndrome, but normal polymorphisms which have not been reported. Moreover, there is a possibility that patients with neuroacanthocytosis similar to McLeod syndrome exist without the XK gene abnormalities.

We studied the age-related alterations of dopaminergic receptors in the brain of Fisher 344 rats with various age (3 weeks and 6, 12, 18 and 24 months) using in vitro receptor autoradiography. [3H]SCH 23390, [3H]spiperone and [3H]nemonapride, and [3H]mazindol were used to label dopamine D1 receptors, dopamine D2 receptors and dopamine uptake sites, respectively. In immature rats (3 weeks old), [3H]SCH 23390 binding showed a significant increase in most brain regions compared to adult animals (6 months old), whereas [3H]spiperone and [3H]nemonapride bindings showed no significant alteration in any brain areas. In contrast, [3H]mazindol binding showed a significant decline in most brain regions. On the other hand, the age-related alterations in [3H]SCH 23390 binding were not observed in any brain regions. [3H]Spiperone and [3H]nemonapride bindings also showed no significant alteration in the brain during aging, except for a transient alteration in [3H]spiperone binding in the nucleus accumbens and hippocampus of 12 months old rats. However, [3H]mazindol binding showed a significant reduction in most brain areas of 12 months old rats. Thereafter, the age-related reduction in [3H]mazindol binding was observed in most brain regions of 18 and 24 months old rats. The results demonstrate that dopamine uptake sites are more susceptible to the aging process than both dopamine D1 and D2 receptors. Furthermore, our results suggest that dopaminergic receptors and dopamine uptake sites may develop with different patterns and speeds after birth. Our studies may provide valuable information concerning the effect of aging on dopaminergic systems.

We immunohistochemically examined biopsied muscles from nine untreated patients with polymyositis (PM) and five patients with other neuromuscular diseases (ONMD), using monoclonal antibodies to tumor necrosis factor-alpha (TNF-alpha) and lymphoid surface markers. In muscles of three patients with PM, we observed many TNF-alpha positive macrophages and lymphocytes in endomysium and around vessels in the muscles. By contrast, there were few, weakly TNF-alpha stained cells in muscles of three patients with ONMD. The ratio of TNF-alpha-positive cells to the muscle fibers and the ratio of TNF-alpha-positive cells to the mononuclear cells were significantly higher in PM compared with ONMD. In addition, we observed atrophic muscle fibers more frequently in TNF-alpha-positive muscles than TNF-alpha-negative ones. We conclude that, at least, in a part of PM patients, TNF-alpha produced locally may contribute to the pathogenesis of PM.

L-3,4-Dihydroxyphenylalanine (L-DOPA), the precursor of dopamine, and bromocriptine, a dopamine D2 receptor agonist, were investigated in haloperidol-induced motor impairments in mice using both catalepsy and pole tests. In catalepsy test, subcutaneous treatment with haloperidol (0.125, 0.25 and 0.5 mg/kg) caused a cataleptic effect in mice in a dose-dependent manner. This cataleptic effect was evident upto 7 hr after haloperidol treatment. In pole test, haloperidol (0.125, 0.25 and 0.5 mg/kg) produced the prolongation of Tturn and TLA as a marker of bradykinesia in mice and the prolongation lasted at least 7 hr after haloperidol treatment. Intraperitoneal co-pretreatment with L-DOPA (400 mg/kg) + carbidopa (10 mg/kg) in mice decreased the catalepsy induced by haloperidol at a dose of 0.125 mg/kg, while co-pretreatment with L-DOPA (200 and 400 mg/kg) + carbidopa (10 mg/kg) dose-dependently decreased the haloperidol (0.125 mg/kg)-induced bradykinesia. The effect of LDOPA + carbidopa in pole test was more pronounced than that in catalepsy test. Intraperitoneal pretreatment with bromocriptine (2 and 4 mg/kg) in mice reduced the catalepsy and bradykinesia produced by haloperidol at a dose of 0.125 mg/kg. The effect of bromocriptine in pole test was relatively similar to that in catalepsy test. Also, co-pretreatment with LDOPA (400 mg/kg) + carbidopa (10 mg/kg) and pretreatment with bromocriptine (2 and 4 mg/kg) significantly decreased the catalepsy induced by haloperidol at a higher dose of 0.5 mg/kg. These results indicate that co-administration with L-DOPA + carbidopa and single treatment with bromocriptine can decrease haloperidol-induced catalepsy and bradykinesia in mice. Furthermore, our study suggests that pole test as well as catalepsy test is of value in the screening of drugs against neuroleptic-induced motor deficits.

We measured 6-[18F]fluorodopa (FDOPA) uptake in the caudate nucleus and the putamen of 20 patients with early and late onset of Parkinson's disease (EOPD and LOPD) and 20 normal control subjects using positron emission tomography. The mean influx rate constant values (Ki) were significantly reduced in the caudate nucleus and the putamen of the patients with EOPD and LOPD compared with age-matched control groups (p < 0.01), respectively. There were significant negative correlations between Ki values in the caudate nucleus (r = -0.67, p = 0.0024) and the putamen (r = -0.67, p = 0.0014), and duration of disease in the LOPD group compared with the EOPD group. Similar negative relationships between Ki values and clinical stages by Hoehn and Yahr and degrees of main clinical symptoms (bradykinesia, tremor and rigidity) were more markedly seen in the LOPD group than in the EOPD group. The present results suggest that the function of presynaptic dopaminergic terminals correlates well with clinical disease severity and degrees of main symptoms in the LOPD group, but not in the EOPD group. We speculate that compensatory up-regulatory function in the postsynaptic dopaminergic receptors may modify disease severity and the degrees of main clinical symptoms of EOPD.

Defects in immune surveillance mechanisms may allow increased replication of human T-lymphotropic virus type I (HTLV-I) in peripheral blood mononuclear cells in HTLV-I associated myelopathy/tropical spastic paraparesis (HAM/TSP). To explore this possibility, antibody-dependent cell-mediated cytotoxicity (ADCC) against HTLV-I infected cells in HAM/TSP and in asymptomatic HTLV-I-seropositive carriers (SPC), was studied. ADCC activity was significantly depressed in HAM/TSP compared to SPC subjects (p < 0.025) and was due to specific reduction in ADCC effector activity. On the other hand, there was no difference in antibody component of anti-HTLV-I ADCC (ADCC-Ab) activities between HAM/TSP and SPC, although patients with more severe forms of disease tended to have higher anti-HTLV-I ADCC-Ab activity. In HAM/TSP, depressed ADCC activity against HTLV-I due to reduced ADCC-effector activity may allow increased replication of HTLV-I which may implicate the development of inflammatory neuropathologic lesions of HAM/TSP.

Presenilin 1 (PS-1) is the main causal gene of familial Alzheimer's disease. In this report, we describe the abnormal behavior of PS-1 in gel electrophoresis in the presence of SDS. Freshly in vitro synthesized PS-1 was identified as a single molecule with the molecular size of 43,000 on SDS gels but was found to disappear after incubation at 37 degrees C for 24 hr due to the formation of aggregates. Intermediate aggregates with M(r) 74,000 and 100,000 were formed before the final aggregate which was retained at the top of the gel. Thus the amount of 43,000-protein species of PS-1 was found to decrease on gels with a concomitant increase in the amount of 74,000/100,000 proteins. Similar abnormality was seen in PS-1 expressed in COS cells transfected with PS-1 cDNA. Moreover, cellular PS-1 was strongly suggested to be cleaved into the fragments with M(r) approximately 20,000 in COS and CHO cells. Fragmentation of cellular PS-1 was not affected by the missense point mutation of Ala260Val on PS-1 which was identified in a pedigree with familial Alzheimer's disease.

We measured cerebral glucose utilization and fluorodopa metabolism in the brain of patients with corticobasal degeneration using position emission tomography. The clinical pictures are distinctive, comprising features referable to both cerebral cortical and basal ganglionic dysfunctions. Brain images of glucose metabolism can demonstrate specific abnormalities with a marked asymmetry in the parietal cortex (the primary motor and sensory cortex and the lateral parietal cortex), the thalamus, the caudate nucleus and the putamen of the dominantly affected hemisphere related to clinical symptoms in six patients. [18F]dopa uptake also reduced in an asymmetric pattern, both the caudate nucleus and the putamen in four patients. This unique combination study measuring both cerebral glucose utilization and fluorodopa metabolism in the nigrostriatal system can provide efficient information about the dysfunctions which are correlated with individual clinical symptoms.

We report clinical characteristics of familial amyotrophic lateral sclerosis (FALS) with 4 different missense point mutations in exons 2, 4, and 5 of the Cu/Zn superoxide dismutase (SOD) gene, that result in amino acid substitutions of histidine46 by arginine (H46R), leucine84 by valine (L84V), isoleucine104 by phenylalanine (I104F), and valine148 by isoleucine (V148I), in 5 Japanese families. Although features of progressive neurogenic muscular atrophy were common in patients of these families, patients of each family showed characteristic clinical features. FALS patients with the H46R mutation showed a benign clinical course and stereotype progression of muscular weakness and atrophy beginning from the legs. In FALS with the L84V mutation, while the clinical course of the disease was similar, the age at onset was younger in men than women. The patients with I104F showed wide ranges of age at onset and duration with ophthalmoparesis and sensory involvement in one patient. Those with the V148I mutation showed younger age at onset and variable first symptoms within the family. Although lower motor sign was evident in all cases, hyperreflexia varied from 0 to 100% among patients with the different mutations, and the Babinski sign was not observed in any case. Bulbar palsy was frequent with I104F, but not with H46R. SOD activity of the red blood cells was severely reduced with I104F and V148I, but was slightly reduced with H46R. These results suggest that familial ALS with different mutations of the Cu/Zn SOD gene each showed clinical characteristics, and that genetic mutations and clinical features are well correlated in familial ALS.

There are currently some types of autosomal dominant cerebellar ataxias such as Machado-Joseph disease (MJD), spinocerebellar ataxia types 1-5 (SCA1-5), or hereditary dentatorubropallidoluysian atrophy. It is very important for these ataxias to be clinically differentiated, but that is sometimes difficult. In particular, the differential diagnosis between MJD and SCA1 is thought to be the most difficult. Recently, both MJD and SCA1 have been proven to be related to expansions of CAG trinucleotide in their causative genes. In this study, 20 cases of MJD in 13 unrelated Japanese families were genetically and clinically examined in comparison with 20 cases of age at onset- and duration-matched Japanese SCA1. The CAG repeat number of expanded MJD and SCA1 alleles was 72.2 +/- 3.1 (mean +/- SD, n = 20) and 47.3 +/- 4.4 (n = 20), respectively, and each repeat size was inversely correlated with age at onset in both MJD and SCA1. The repeat number in leukocytes increased from parents to children with acceleration of age at onset (anticipation) in MJD. In MJD, the number of CAG repeats in the expanded allele was lower in sperm than that of leukocytes, but was more in SCA1. However, the number of peaks in the expanded allele was greater in sperm than in leukocytes in both MJD and SCA1 (increased mosaicism level). MJD was clinically characterized by a relatively higher frequency of ocular signs such as eyelid retraction, bulging eyes, ophthalmoparesis, and nystagmus, spasticity in lower limbs, and sensory and urinary disturbances in contrast to the SCA1 patients except for slow eye movement. These results indicate that the expanded CAG repeat and clinical features are correlated in both MJD and SCA1, and MJD can be differentiated from SCA1 by clinical characteristics mentioned above as well as DNA analysis.

Bromocriptine, a dopamine D2 receptor agonist, has widely been used for patients with Parkinson's disease. In this study, we examined its neuroprotective effects against neuronal damage in the CA1 subfield of the hippocampus following experimental cerebral ischemia in the Mongolian gerbil. Forebrain ischemia was induced by occlusion of bilateral common carotid arteries for 3 min. Bromocriptine, at a dose of 0.3 or 3 mg/kg, was injected i.p. 30 min before the onset of ischemia. Histopathological observations showed that neuronal damage to hippocampal CA1 neurons, which was seen 7 days after ischemia in vehicle-treated animals, was prevented by bromocriptine treatment. Immunohistochemical staining for copper/zinc superoxide dismutase and manganese superoxide dismutase decreased markedly in the CA1 neurons of vehicle-treated animals 2 days after ischemia when histological neuronal destruction was not yet seen, but was well preserved in bromocriptine-treated animals. The present findings show that bromocriptine protects against ischemia-induced neuronal damage, and that the mechanism of the neuroprotection may relate to the preservation of SODs. Bromocriptine, which was recently shown to be a potent free radical scavenger, may have a potent neuroprotective action against disorders including ischemic stroke.

Amyotrophic lateral sclerosis (ALS) is a progressive and fatal neurological disorder that results in relentless damage to the motor neuron system. Although about 5-10% of cases are familial, the pathophysiologic process of ALS remains unknown. We identified a novel point mutation A to G in exon 2 of the Cu/Zn SOD gene, resulting in an amino acid substitution of histidine46 by arginine (H46R), in two Japanese familial ALS (FALS) families. The segregations of the mutation were evident. The enzymatic activities of Cu/Zn SOD of peripheral red blood cell lysate were reduced to about 80% in the affected members, compared with other non-affected family members. The patients in these families are clinically characterized by relative late onset, initial involvement in lower extremities, relative rare impairment of bulbar muscles and much slow progression of muscular weakness and atrophy, compared with other Japanese FALS cases who have no mutation in the Cu/Zn SOD gene. These findings suggest that the H46R mutation in Cu/Zn SOD gene is highly related to this unique subtype of FALS.

Thirty-one HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients and 34 asymptomatic carriers were studied in order to investigate the presence of anti-lymphocyte antibodies in their sera. 32% of the patients produced allolymphocytotoxic antibodies reactive to peripheral blood lymphocytes (mainly T cells) as well as to B cells, which belonged to the IgM class, reacted under warm and cold conditions, and showed specificity to multiple HLA antigens, especially HLA A2, A26, A33, B7, B27, B35, B48, B61 and Cw3. Moreover, their reactivities could be absorbed by pooled platelets. Therefore, it may be suggested that either anti-HLA class I antibodies or antibodies cross-reactive to HLA were present. On Western blot analysis, the HAM/TSP cytotoxic sera reacted to the 110 kDa molecules of lymphocyte lysates. In the case of carriers, weak cytotoxic antibodies were detectable in 14% of the sera. No multispecificity to HLA class I antigens was observed and there was no reactivity to the 110 kDa band. The production of allolymphocytotoxic antibodies in HAM/TSP patients may therefore be explained by the presence of allogeneic-like stimuli in this disease. HTLV-I infected cells, expressing altered HLA antigens could possibly account for this stimulation and also contribute to the alteration of immune functions in HAM/TSP, and thus play an important role in its pathogenesis.

We describe a 21-year-old man presenting with proximal muscle weakness associated with hypernatremia. His manifestations other than muscle weakness included dry skin, loss of axillary and pubic hair, decreased libido and loss of thirst sensation. His serum sodium level was elevated to 169-171 mEq./l but all other electrolytes were normal. In addition, serum CK was elevated and an EMG study showed myogenic changes. Endocrinological studies revealed hypothalamic hypopituitarism, while MRI revealed a suprasellar mass. A partial correction of hypernatremia led to an immediate recovery of the muscle weakness as well as a normalization of both the serum CK level and EMG findings, suggesting a direct association between the muscle weakness and hypernatremia. The phosphocreatine/inorganic phosphorus (PCr/Pi) ratios in the resting calf muscle, obtained using 31P magnetic resonance spectroscopy (MRS), were very low during the state of muscle weakness, while they returned to nearly normal values after clinical improvement, suggesting that the muscle weakness in hypernatremic state was caused by a depletion of the intramuscular energy stores, probably due to an overworking Na-K pump to correct the intracellular electrolyte imbalance.

We found that the hemicellulose extracted from rice bran fiber (RBF) increased the peripheral blood leukocytes in rats. Wistar male rats were fed a 10% RBF or various proportions of hemicellulose diet for 2 weeks. The body weight and relative tissue weight were not affected by the RBF or hemicellulose dietary. The number of peripheral blood leukocytes and lymphocytes was significantly increased in rats fed a 10% hemicellulose diet than that of rats fed a control diet (p < 0.01). The proportion of each lymphocyte subset was not significantly changed, but the ratio of CD4 positive cells to CD8 positive cells was relatively increased.

To determine the functional diversity of thymic lymphoid cells in patients with myasthenia gravis (MG), we evaluated mixed lymphocyte reactions (MLR) using thymus cells in 7 MG patients and 8 controls. In the MLR, we used thymus cells as responder cells and mitomycin C-treated peripheral non-T cells as stimulator cells. In an autologous MLR test, a low proliferative response was observed in both the MG patients and controls. In an allogeneic MLR test, in which thymus cells were co-cultured with allogeneic non-T cells, the thymus cells from MG patients showed an increased proliferative response to stimulator cells, whether they were from MG patients or the controls. However, thymus cells from the controls showed a low proliferative response to any allogeneic stimulator cells. The enhanced allo-reactivity of thymus cells from MG patients thus suggests that there is an increase in the number of functionally mature T lineage cells in the MG thymus.

We studied the relationship between antibody titers to recombinant HTLV-I p40tax protein and gag-env hybrid protein in serum (by an enzyme-linked immunosorbent assay) and HTLV-I proviral DNA load in peripheral blood mononuclear cells (by a quantitative polymerase chain reaction method) in 18 patients with HTLV-I-associated myelopathy (HAM)/tropical spastic paraparesis (TSP), 17 HTLV-I carriers without HAM/TSP and 16 HTLV-I uninfected controls. The IgG and IgA antibody titers to either of the proteins correlated significantly with the HTLV-I pX (coding p40tax protein) and pol DNA amounts in HTLV-I infected subjects. HAM/TSP patients had significantly higher titers of IgG and IgA antibodies to the HTLV-I proteins than did the HTLV-I carriers without HAM/TSP. While the IgM antibodies to the HTLV-I proteins were found in only 6% of HTLV-I carriers without HAM/TSP, they were found in 40% of HAM/TSP patients, especially those having both a high HTLV-I proviral DNA load and high titers of the IgG and IgA antibodies. HAM/TSP patients with the IgM antibodies had a tendency to deteriorate more frequently on the Kurtzke's disability status scale and magnetic resonance imaging of the brain (leukoencephalopathy) than did those without in the two-year follow-up. Thus, the presence of IgM antibody and high titers of IgG and IgA antibodies to the HTLV-I proteins, together with the increased HTLV-I proviral DNA load, appears to distinguish HAM/TSP patients from HTLV-I carriers without HAM/TSP.