Takao Mitsui

Glucocorticoid Excess Induces Superoxide Production in Vascular Endothelial Cells and Elicits Vascular Endothelial Dysfunction

Abstract— Glucocorticoid (GC) excess often elicits serious adverse effects on the vascular system, such as hypertension and atherosclerosis, and effective prophylaxis for these complications is limited. We sought to reveal the mechanism underlying GC-induced vascular complications. Responses in forearm blood flow to reactive hyperemia in 20 GC-treated patients were significantly decreased to 43±8.9% (mean±SEM) from the values obtained before GC therapy (130±14%). An administration of vitamin C almost normalized blood flow responses. In human umbilical vein endothelial cells (HUVECs), production of hydrogen peroxide was increased up to 166.5±3.3% of control values by 10−7 mol/L dexamethasone (DEX) treatment (P <0.01). Concomitant with DEX-induced hydrogen peroxide production, intracellular amounts of peroxynitrite significantly increased and those of nitric oxide (NO) decreased, respectively (P <0.01). Immunoblotting analysis using anti-nitrotyrosine antibody showed that peroxynitrite formation was increased in DEX-treated HUVECs. Using inhibitors against metabolic pathways for generation of reactive oxygen species (ROS), we identified that the major production sources of ROS by DEX treatment were mitochondrial electron transport chain, NAD(P)H oxidase, and xanthine oxidase. These findings suggest that GC excess causes overproduction of ROS and thereby perturbs NO availability in the vascular endothelium, leading to vascular complications in patients with GC excess.

Adhalin gene mutations in patients with autosomal recessive childhood onset muscular dystrophy with adhalin deficiency.

Homozygous adhalin gene mutations were found in three patients from two consanguineous families with autosomal recessive childhood onset muscular dystrophy. Muscle biopsies from patients in each family showed complete absence of adhalin. Sequencing of adhalin cDNA prepared from skeletal muscle by reverse transcription PCR demonstrated a cytosine to thymidine substitution at nt 229 in the patient in family 1 and an adenine to guanine substitution at nt 410 and a 15-base insertion between nt 408 and 409 in the two patients in family 2. Sequencing of genomic DNA prepared from peripheral blood leukocytes by PCR confirmed these mutations. The parents in each family were found to be heterozygous for the respective mutations. These adhalin gene mutations are presumed to be responsible for the absence of adhalin in the skeletal muscle. Adhalin deficiency likely causes disruption of the muscle cell membrane, resulting in dystrophic changes in the skeletal muscle similar to dystrophin deficiency in Duchenne muscular dystrophy.

Oxidative damage to skeletal muscle DNA from patients with mitochondrial encephalomyopathies

To estimate the oxidative damage to skeletal muscle DNA in mitochondrial encephalomyopathies, we studied the amount of 8-hydroxy-deoxyguanosine (8-OH-dG) and the localization of superoxide dismutase (SOD) in the skeletal muscles of patients with progressive external ophthalmoplegia (PEO) or Kearns-Sayre syndrome (KSS). The molar ratio of 8-OH-dG/deoxyguanosine in skeletal muscle from PEO or KSS patients was significantly higher than the control value. The ratio from patients with polymyositis or Duchennes muscular dystrophy was not significantly elevated. Immunohistochemical staining for both Mn-SOD and Cu,Zn-SOD showed pronounced staining in the subsarcolemmal and intermyofibrillar regions of cytochrome-oxidase-negative ragged red fibers of KSS or PEO muscles. Our findings suggest that overproduction of 8-OH-dG and mitochondrial dysfunction with gene deletions are associated with each other in muscle cells of patients with PEO or KSS, and that free radicals may play an important role in the pathophysiology of mitochondrial encephalomyopathies.

Increasing incidence of elderly onset patients with myasthenia gravis in a local area of Japan

Objective: As the number of elderly patients with myasthenia gravis (MG) has recently increased in Europe and the USA, a retrospective survey of Japanese MG patients was conducted in a single neurological centre over several decades. Methods: The study consisted of 112 consecutive MG patients with onset of the disease from 1971 to 2006 from an area of approximately 0.8 million inhabitants in Japan. Patients were classified into three subgroups according to age at onset: young onset (39 years old), middle aged onset (40–59 years old) and elderly onset (60 years old). The trends in incidence rate and clinical features were examined: disease severity, seropositivity for antiacetylcholine receptor antibody, occurrence of other autoimmune diseases, occurrence of thymoma and therapeutic response. Results: The onset adjusted age specific average annual incidence per 100 000 of the elderly onset MG patients increased 20-fold from 1981–1990 (0.06; 95% CI 0.00 to 0.36) to 2001–2006 (1.30; 95% CI 0.77 to 2.05). Clinical features of the elderly onset MG patients included low antiacetylcholine receptor antibody titres (mean 24.6 nmol/l), less frequent autoimmune overlaps (8.0%) and nearly no complete stable remission with or without thymectomy. Conclusion: The increasing incidence of elderly onset MG in Japanese patients similar to that reported in Caucasians has been confirmed. The clinical features suggest different immunological backgrounds between young onset and elderly onset MG patients, irrespective of the ethnic background.

Beneficial Effect of Steroid Pulse Therapy on Acute Viral Encephalitis

Corticosteroids are often used in the treatment of acute viral encephalitis, although the efficacy of corticosteroid therapy has not been proven. We examined the effects of high-dose corticosteroid therapy on acute viral encephalitis in 5 patients with progressive disturbances of consciousness. In 3 patients who were treated within 5 days after the onset of illness, pulse therapy dramatically reduced the degree of consciousness disturbance. They became alert within 24 h, and then neurological symptoms gradually improved. Corticosteroid therapy in the other 2 patients, in whom treatment was started more than 3 weeks after the onset of illness, was not as effective, but repeated therapy at 2-week intervals resulted in complete recovery. These findings suggest that high-dose corticosteroid therapy is effective, particularly for disturbances of consciousness, an important prognostic factor in acute viral encephalitis.

Preferential gray matter involvement in dengue myelitis

Dengue virus (DV) belongs to the family Flaviviridae , and its infection is characterized by dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. Some flaviviruses, such as Japanese encephalitis, St. Louis encephalitis, Murray Valley encephalitis, and West Nile viruses, have the common feature of neurovirulence in the human CNS, including poliomyelitis-like syndrome.1 We report a patient with dengue fever who developed gray matter myelitis. A 42-year-old Japanese man working in Indonesia was admitted to a local hospital because of fever (38° C). Three days after admission, he had weakness in both legs and retained urine, and he was transferred to Singapore General Hospital. On admission, he was febrile and had a generalized macular rash. There was flaccid paralysis of the lower limbs with absent knee and ankle jerks. Sensation to pinprick was decreased below T6 segment. Laboratory examination revealed thrombocytopenia. Results of CSF examination were as follows: cells, 201/mm3 (mononuclear cell dominant); glucose, 1.6 mmol/L; and protein, 100 …

UNDIMINISHED REGULATORY T CELLS IN THE THYMUS OF PATIENTS WITH MYASTHENIA GRAVIS

Objective: The thymus has been implicated as a possible site of origin that triggers autoimmunity in myasthenia gravis (MG). Although several groups have suggested that the decrease in the number of regulatory T (Treg) cells contributes to the onset of MG, the exact role of Treg cells in MG remains unclear. To address this point, we examined the number and distribution of Treg cells in a large number of patients with MG. Methods: Immunohistofluorescence analysis of Foxp3 along with CD4 and CD8 was performed in thymic sections of MG (+) (n = 24) and MG (−) patients (n = 27). Circulating CD4+CD25+ cells in the peripheral blood of patients with MG (n = 15) and age-matched healthy subjects (n = 15) were also analyzed. Results: Foxp3+CD4+CD8− cells were predominantly found in the thymic medulla and their number declined with age. There was no significant difference in the number or the distribution of Foxp3+CD4+CD8− cells in the thymus between MG (+) and MG (−) patients. The number of circulating CD4+CD25+ cells in the peripheral blood of patients with MG was not significantly altered compared to that in healthy subjects. Conclusion: The cellularity of Treg cells in the thymus and circulation is not diminished in patients with myasthenia gravis.

Functional association between nicotinic acetylcholine receptor and sarcomeric proteins via actin and desmin filaments

By affinity chromatography utilizing α‐cobrotoxin from digitonin‐solubilized fractions of rabbit skeletal muscle, we found that many proteins are associated with the nicotinic acetylcholine receptor (AChR). In addition to the proteins we previously reported to bind to AChR (including dystrophin‐dystrophin‐associated protein (DAP) complex, utrophin, rapsyn, and actin; Mitsui et al. [1996] Biochem. Biophys. Res. Commun.224:802–807), α‐actinin, desmin, myosin, tropomyosin, troponin T, and titin are also identified to be associated with AChR. Alkaline treatment or Triton X‐100 solubilization released dystrophin‐DAP complex, utrophin, and rapsyn from the AChR fraction, while actin and desmin remained associated. These findings demonstrate that AChR is supported primarily by a submembranous organization of actin and desmin filaments, and is linked to sarcomeric proteins via these filaments. To further investigate whether the association has any functional role, we studied the effect of acetylcoline on ATPase activity of the AChR fraction. Acetylcholine (0.5–4 μM) significantly activated Mg2+‐ATPase activity of digitonin‐solubilized AChR fraction (P < 0.05). Furthermore, we found that desmin as well as actin activated myosin Mg2+‐ATPase activity. From these findings, it is suggested that desmin and actin form a submembranous organization in the postsynaptic region, and function as mediators of excitation of AChR to the sarcomeric contraction system. J. Cell. Biochem. 77:584–595, 2000.

Solitary plasmacytoma with VEGF overproduction: Report of a patient with polyneuropathy

Solitary plasmacytoma frequently involves polyneuropathy.1 Although several possible mechanisms, such as immunologic reaction of M proteins2 and neurotoxic substances, have been suggested, the pathogenesis of polyneuropathy has not yet been elucidated. Recently, we saw a patient with polyneuropathy and a solitary plasmacytoma. Her plasma vascular endothelial growth factor (VEGF) level was markedly increased, and rapidly decreased after resection of the tumor, followed by neurologic improvement. A 62-year-old Japanese woman was admitted to our hospital in May 1999 because of gait disturbance. The results of physical examination were unremarkable except for bilateral pretibial edema. Neurologic findings showed prominent muscle wasting and weakness in distal limbs and stoking and glove sensory disturbance. Deep tendon reflexes were absent or diminished in all four limbs. Laboratory studies revealed normal values of fasting blood sugar, vitamins, and thyroid function. M proteins were not detected by immunoelectrophoresis or immunofixation. Antinuclear antibody and anti-ganglioside antibodies were not detected. Her plasma VEGF level …

Complicated paraneoplastic neurological syndromes: a report of two patients with small cell or non-small cell lung cancer

Paraneoplastic neurological syndromes are frequently associated in patients with small cell lung cancer (SCLC) and antineuronal antibodies are involved in the autoimmune mechanism. Multiple syndromes are sometimes complicated in a single patient with SCLC. However, little is known about non-SCLC-associated neurological manifestations. We report two patients with complicated paraneoplastic neurological syndromes. Patient 1 showed paraneoplastic limbic encephalitis (PLE), paraneoplastic sensory neuropathy (PSN) and Lambert-Eaton myasthenic syndrome (LEMS) associated with SCLC. Patient 2 developed opsoclonus-ataxia and probable PLE associated with non-SCLC. Analysis of various antineuronal antibodies revealed that anti-Hu and P/Q-type voltage-gated calcium channel (VGCC) antibodies were positive in Patient 1 but any antibodies were not in Patient 2. Brain MRI demonstrated high intensity signals in temporal lobes particularly on fluid-attenuated inversion recovery (FLAIR) or diffusion-weighted images. These findings suggest that complicated paraneoplastic neurological syndromes occur in non-SCLC as well as SCLC and that unidentified antineuronal autoantibodies may underlie the pathophysiology.