Peroxiredoxins are involved in the pathogenesis of multiple sclerosis and neuromyelitis optica spectrum disorder.

Peroxiredoxins (PRXs) are intracellular anti-oxidative enzymes but work as inflammatory amplifiers under the extracellular condition. To date, the function of PRXs in the pathogenesis of multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD) is not fully understood. The aim of this study was to investigate whether PRXs play a role in the pathogenesis of MS and NMOSD. We analyzed levels of PRXs (PRX1, PRX5 and PRX6) in the cerebrospinal fluid (CSF) and serum of 16 patients with MS, 16 patients with NMOSD and 15 patients with other neurological disorders (ONDs). We identified potential correlations between significantly elevated PRXs levels and the clinical variables in patients with MS and NMOSD. Additionally, pathological analyses of PRXs (PRX1-6) in the central nervous system (CNS) were performed using the experimental autoimmune encephalomyelitis (EAE), animal model of MS. We found that serum levels of PRX5 and PRX6 in patients with MS and NMOSD were higher compared with those in patients with ONDs (P < 0·05). Furthermore, high levels of PRX5 and PRX6 were partly associated with blood-brain barrier dysfunction and disease duration in NMOSD patients. No significant elevation was found in CSF PRXs levels of MS and NMOSD. Spinal cords from EAE mice showed remarkable PRX5 staining, especially in CD45+ infiltrating cells. In conclusion, PRX5 and PRX6 may play a role in the pathogeneses of MS and NMOSD.

Comparison of brain atrophy in patients with multiple sclerosis treated with first- versus second-generation disease modifying therapy without clinical relapse.

BACKGROUND AND PURPOSE: The silent progression of patients with multiple sclerosis (MS) has been reported. The aim of this study was to investigate the association between brain atrophy rates and disease-modifying drugs (DMDs) in patients with MS during their relapse-free period. METHODS: Patients with relapsing-remitting MS were classified into two groups on the basis of clinical records, i.e. a first-generation DMD group treated with interferon-beta-1a, interferon-beta-1b or glatiramer acetate and a second-generation DMD group treated with dimethyl fumarate, fingolimod or natalizumab. Brain volume was calculated with SPM12. RESULTS: A total of 45 patients with relapsing-remitting MS were enrolled in the first-generation (n = 22) or second-generation (n = 23) DMD group. The annualized relapse rate was lower in the first-generation than in the second-generation DMD group (median 0.26 vs. 0.59; P < 0.001). The annualized atrophy rate of the normalized brain volume was not different between the first- and second-generation DMD groups after analysis of covariance (median 0.13% vs. 0.59%; P = 0.17). CONCLUSIONS: The median annualized atrophy rate of normalized brain volume in the first-generation DMD group was similar to the previously reported annual brain atrophy rate of healthy controls, which may suggest that treatment with a first-generation DMD need not be changed when patients with MS are clinically inactive.

Recombinant thrombomodulin ameliorates experimental autoimmune encephalomyelitis by suppressing high mobility group box 1 and inflammatory cytokines.

Recombinant thrombomodulin (rTM) has pleiotrophic properties, including anti-coagulation and anti-inflammation; however, its effectiveness as a treatment for multiple sclerosis (MS) has not been evaluated fully. High mobility group box 1 (HMGB1) and proinflammatory cytokines, working as inflammatory mediators, are reportedly involved in the inflammatory pathogenesis of MS. The aim of this study was to determine whether rTM can be a potential therapeutic agent for experimental autoimmune encephalomyelitis (EAE). EAE mice received rTM treatment (1 mg or 0·1 mg/kg/day) from days 11 to 15 after immunization. The clinical variables, plasma levels of inflammatory cytokines and HMGB1 and pathological findings in EAE were evaluated. rTM administration ameliorated the clinical and pathological severity of EAE. An immunohistochemical study of the spinal cord showed weaker cytoplasmic HMGB1 staining in the rTM-treated EAE mice than in the untreated EAE mice. Plasma levels of inflammatory cytokines and HMGB1 were suppressed by rTM treatment. In conclusion, rTM down-regulated inflammatory mediators in the peripheral circulation and prevented HMGB1 release from nuclei in the central nervous system, suppressing EAE-related inflammation. rTM could have a novel therapeutic potential for patients with MS.

Subinsular vascular lesions: an analysis of 119 consecutive autopsied brains.

The insula of Reil constitutes a functionally intriguing complex of the brain related to multifunctional activities. We examined the subinsular region in 119 consecutively autopsied patients, as T2 hyperintense lesions are frequently observed in magnetic resonance diagnosis of this region. The patients were admitted in neurology wards and were diagnosed as having cerebrovascular disease in 55 patients (46%), other neurological diseases in 57 patients (48%) and non-neurological diseases in seven patients (6%). Demyelination of the white matter was semi-quantified as a fiber density score (percent stained area/total area) with computer-assisted image analysis on Klüver-Barrera-stained sections. Astrogliosis was assessed by immunohistochemistry for glial fibrillary acidic protein. The lesion analysis showed a dilated perivascular space in 29 patients (24%), demyelination (fiber density score less than the mean - 1 SD) in 27 patients (23%), slit-shaped lesion in six patients (5%), lacunar infarction in one patient (1%) and cerebral hemorrhage in one patient (1%). A histologic-radiologic comparison in two patients with subcortical ischemic vascular dementia showed correspondence between subinsular hyperintensities, and demyelination, gliosis and a dilated perivascular space. These results indicate that subinsular lesions rarely signifies focal vascular lesions, and are consisted of demyelination, gliosis and a dilated perivascular space.

Astroglial expression of ceramide in Alzheimer's disease brains: a role during neuronal apoptosis.

Accumulating evidences indicate that ceramide is closely involved in apoptotic cell death in neurodegenerative disorders and aging. We examined ceramide levels in the cerebrospinal fluid (CSF) or brain tissues from patients with neurodegenerative disorders and the mechanism of how intra- and extracellular ceramide was regulated during neuronal apoptosis. We screened the ceramide levels in the CSF of patients with neurodegenerative disorders, and found that ceramide was significantly increased in patients with Alzheimer's disease (AD) than in patients with age-matched amyotrophic lateral sclerosis (ALS) and other neurological controls. With immunohistochemistry in AD brains, ceramide was aberrantly expressed in astroglia in the frontal cortices, but not detected in ALS and control brains. To explore for the regulation of ceramide in astroglia in Alzheimer's disease brains, we examined the metabolism of ceramide during neuronal apoptosis. In retinoic acid (RA)-induced neuronal apoptosis, RA slightly increased de novo synthesis of ceramide, but interestingly, RA dramatically inhibited conversion of [14C] ceramide to glucosylceramide (GlcCer), suggesting that the increase of ceramide mass is mainly due to inhibition of the ceramide-metabolizing enzyme GlcCer synthase. In addition, a significant increase of the [14C] ceramide level in the culture medium was detected by chasing and turnover experiments without alteration of extracellular [14C] sphingomyelin levels. A 2.5-fold increase of ceramide mass in the supernatant was also detected after 48 h of treatment with RA. These results suggest a regulatory mechanism of intracellular ceramide through inhibition of GlcCer synthase and a possible role of ceramide as an extracellular/intercellular mediator for neuronal apoptosis. The increased ceramide level in the CSF from AD patients, which may be derived from astroglia, raises a possibility of neuronal apoptosis by the response to intercellular ceramide in AD.

Stimulation of angiotensinogen production in primary cultures of rat hepatocytes by glucocorticoid, cyclic adenosine 3',5'-monophosphate, and interleukin-6.

The effects of hormones and cytokines on angiotensinogen production were studied in primary cultured rat hepatocytes. The basal secretion of angiotensinogen decreased during culture. The addition of dexamethasone and (Bu)2cAMP completely prevented this decrease. Angiotensinogen secretion by freshly plated hepatocytes was slightly increased in response to dexamethasone, but after 24 h in culture, hepatocytes no longer responded to dexamethasone alone. When hepatocytes were treated with (Bu)2cAMP, glucagon, or forskolin, angiotensinogen secretion increased in response to dexamethasone in a concentration-dependent manner. 17 beta-Estradiol and T3 failed to stimulate angiotensinogen secretion in either the presence or absence of (Bu)2cAMP. Interleukin-6 (IL-6) exhibited a stimulatory activity on angiotensinogen secretion, which was dependent on the presence of dexamethasone, whereas IL-1 and tumor necrosis factor had no effect in either the presence or absence of dexamethasone and/or (Bu)2cAMP. Unlike primary cultured hepatocytes, angiotensinogen secretion by rat hepatoma H4IIEC3 cells increased in response to dexamethasone alone. This increase was not enhanced by (Bu)2cAMP, but was enhanced by IL-6. Thus, in primary cultures of rat hepatocytes, neither glucocorticoid, cAMP, nor IL-6 alone stimulated angiotensinogen production, but a combination of glucocorticoid and cAMP or of glucocorticoid and IL-6 exhibited a stimulatory activity on angiotensinogen production. These results suggest that angiotensinogen production in the liver is synergistically regulated by these factors, whereas the hepatoma cell line H4IIEC3 lacks the regulatory mechanism of cAMP on glucocorticoid-induced angiotensinogen production.

Angiotensinogen production by rat hepatoma cells is stimulated by B cell stimulatory factor 2/interleukin-6.

Angiotensinogen has been identified as one of the acute-phase reactants. In vitro studies were carried out using the Reuber H35 hepatoma cell line to identify the species of cytokines contributing to the increased synthesis of angiotensinogen in the liver. Angiotensinogen secretion by H35 cells was maximally increased 4-fold by the addition of 10(-7) M dexamethasone. Under this condition, angiotensinogen secretion was further stimulated by B cell stimulatory factor 2/interleukin-6 (IL-6, 50 U/ml), but not by interleukin-1 or interferon-alpha. In the absence of glucocorticoid, IL-6 did not affect angiotensinogen secretion by H35 cells, indicating that the presence of glucocorticoid is required for the stimulatory activity of IL-6. These results suggest that IL-6 is a mediator responsible for the increased synthesis of angiotensinogen in the liver during acute inflammation.

Location of forelimb motoneurons in the Japanese toad (Bufo japonicus): a horseradish peroxidase study.

To label the spinal motoneurons innervating the forelimb muscles of the Japanese toad, horseradish peroxidase (HRP) was injected into these muscles or applied to the cut end of the brachial nerves (N. radialis and N. ulnaris). Spatial distribution of the HRP-labeled motoneurons was reconstructed from serial frontal sections of the spinal cord and their location was examined. Motoneurons innervating forelimb muscles were distributed in the lateral cell column from segment 3 to segment 5 of the ipsilateral brachial spinal cord. In the transverse plane of the spinal cord, motoneurons innervating the medial forearm muscles (innervated by N. ulnaris) were located in the more medial part of the lateral cell column, whereas those innervating the lateral forearm muscles and the upper arm muscle (innervated by N. radialis) were located in the more lateral part of the lateral cell column. Along the longitudinal axis of the spinal cord, motoneurons innervating the more anterior (flexor side) forearm muscles were located in the more rostral part of the spinal cord, whereas those innervating the more posterior (extensor side) forearm muscles were located in the more caudal part of the spinal cord. Thus, motoneurons innervating forearm muscles were well organized somatotopically not only in the transverse plane, but also along the longitudinal axis of the spinal cord. Such a somatotopic organization of motoneurons along the longitudinal axis could also be regarded as a functional one; the flexor motoneurons were located rostrally to the extensor motoneurons.

The coagulation-fibrinolysis system in patients with leukoaraiosis and Binswanger disease.

BACKGROUND: Hypercoagulability is observed in vascular dementia, including Binswanger disease. However, the correlation between hypercoagulability, leukoaraiosis, and dementia remains unclear. OBJECTIVE: To examine how activation of the coagulation fibrinolysis correlates with leukoaraiosis and dementia. PATIENTS AND METHODS: Thrombin-antithrombin complex (TAT), prothrombin fragment(1 + 2) (F1 + 2) and cross-linked D-dimer (XDP) were measured consecutively in 18 subjects without dementia and with leukoaraiosis, and in 29 subjects with subcortical vascular dementia and severe leukoaraiosis (Binswanger disease) at either stable or deteriorating stages. They were compared with 19 patients with old lacunar infarctions and 24 patients with other neurological diseases. We also examined the indices of cognitive impairment and brain atrophy. In each group, the ventricular area-cranial space area ratio was measured by an image analyzer. RESULTS: Patients with Binswanger disease who were exclusively at deteriorating stages showed increased TAT and XDP levels and an increased ventricular area-cranial space area ratio, as compared with the patients with other neurological diseases (P<.001). The index of cognitive impairment in patients at a deteriorating stage showed a decreasing trend vs that of patients in the stable stage. Among the variables that were significantly associated with a hypercoagulable condition (ie, age, scores on Mini-Mental State Examination or the Hasegawa Dementia Rating Scale, Revised [MMSE/HDRS], white matter lesions, ventricular area-cranial space area ratio, and C-reactive protein), age (odds ratio [OR], 2.82) and MMSE/HDSR scores (OR, 0.43) survived as predictors for coagulation activation, and C-reactive protein survived for fibrinolysis activation (OR, 4.63) in multivariate analysis. CONCLUSION: Hypercoagulability in a subgroup of patients with Binswanger disease and with more severe cognitive impairment and brain atrophy does not support a triggering role for a coagulation-fibrinolysis system, although it may contribute to worsening of neurological deficits.

Reduction of lipofuscin by centrophenoxine and chlorpromazine in the neurons of rat cerebral hemisphere in primary culture.

Cells from neonatal rat cerebral hemispheres were dispersed by trypsin and cultured for 32 days. Histochemical, fluorescence, and electron microscopic analyses demonstrated that lipofuscin pigments increased in neuronal and non-neuronal cells in primary culture according to the lapse of time. When centrophenoxine (10(-4) or 5 X 10(-4) M) or chlorpromazine (10(-6) or 10(-5) M) was added to the medium, the accumulation of lipofuscin pigments in neurons was significantly reduced. However, the effects of these agents were not detected in non-neuronal cells.

Interleukin-6 as a mediator responsible for inflammation-induced increase in plasma angiotensinogen.

The concentration of plasma angiotensinogen increases upon induction of inflammation. Studies were carried out using serum samples collected from mice and rats after injection of lipopolysaccharide (LPS) to determine whether interleukin-6 (IL-6) is a mediator responsible for the inflammation-induced increase of angiotensinogen synthesis in liver cells. Serum collected from mice or rats 2 and 4 hr after injection of LPS contained a factor that stimulated [35S]methionine incorporation into angiotensinogen newly synthesized by rat hepatoma H4IIEC3 (H4) cells. Assay of IL-6 using an IL-6-dependent murine hybridoma, MH60.BSF2 cells, showed the presence of IL-6-like activity in sera of mice or rats 2 and 4 hr after injection of LPS. Anti-mouse IL-6 monoclonal antibody completely inhibited not only the IL-6-like activity present in LPS-treated mouse serum but also the ability of the serum to stimulate angiotensinogen synthesis of H4 cells. These results suggest that increased synthesis of angiotensinogen in the liver after induction of inflammation is mediated by IL-6, a cytokine important in immune reactions and the hepatic acute-phase response.

A novel cell type in the parathyroid glands of spontaneously hypertensive rats.

Recently, we described a circulating hypertensive factor, present in the plasma of spontaneously hypertensive rats (SHR). This factor seems to be produced by the parathyroid gland but is not identical to parathyroid hormone (PTH). In view of these findings, we attempted to search for histological differences in parathyroid glands between SHR and normotensive Wistar-Kyoto (WKY) rats by light and electron microscopy. Novel cells, distinct from normal chief cells, were frequently found in parathyroid glands of SHR rats, whereas they were scarcely observed in WKY rats. Our findings suggest that the novel cells are involved in the development of hypertension in SHR rats.

Changes in activity of the renin-angiotensin system of the rat by induction of acute inflammation.

Angiotensinogen is the precursor of biologically active peptide angiotensin II and its hepatic synthesis is increased by the induction of acute inflammation. Studies were carried out to know whether the rise in plasma angiotensinogen is actually involved in the activity of the renin-angiotensin system during acute inflammation. The plasma level of angiotensinogen in rats was increased to 2.5 times the normal level 16 h after the induction of acute inflammation by administration of lipopolysaccharide (LPS). The plasma renin concentration (PRC) was decreased to about 40% of the normal level concomitantly with a reduction of plasma renin activity (PRA) at 4 h after LPS administration. In contrast, 16 h after LPS injection, when plasma angiotensinogen showed a high level and PRC had recovered to the normal range, PRA was increased to 1.7 times the normal level. These results indicate that acute inflammation induced by LPS causes a biphasic change in the generation of angiotensin I, i.e., an early decrease depending upon the reduction of PRC and later increase depending upon elevation of the angiotensinogen concentration.

Restenosis after successful emergency coronary angioplasty for acute myocardial infarction: comparison with elective angioplasty.

We investigated the initial and late restenosis rate after successful emergency coronary angioplasty in 64 patients with acute myocardial infarction, and compared these results with those of 100 patients (110 lesions) who had successful angioplasty on an elective basis. The majority of the baseline clinical and angiographic variables were similar in the myocardial infarction and elective groups. The restenosis rate at 1 month was high in patients undergoing emergency angioplasty for acute myocardial infarction (23 vs. 12%). At 3-6 months, the angiographic restenosis rate was low for the infarction group (26 vs. 37%). The overall restenosis rate was similar in the infarction and elective groups (39 vs. 40%). Lesion regression after coronary angioplasty was more frequent in the infarction than in the elective angioplasty group (27 vs. 14%, P < 0.05). These findings suggest that considering the high restenosis rate at 1 month and the lower, but still 20% or more, rate at 3-6 months, a follow-up angiography should be performed both prior to discharge and at 3-6 months after the procedure, in patients with acute myocardial infarction.

Progression of coronary artery disease in patients receiving percutaneous transluminal coronary angioplasty: angiographic comparison of angioplasty and non-angioplasty sites.

Long-term follow-up of coronary artery stenosis was carried out in 81 patients undergoing successful percutaneous transluminal coronary angioplasty (PTCA). Progression of coronary stenosis was observed in 38 patients with 52 sites, and regression was seen in 23 patients with 23 sites during 2 years or more of follow-up. Progressive change occurred in three sites of 97 PTCA lesions and 49 of 1,090 non-PTCA lesions (no significant difference). On the other hand, regressive change was observed in 19 sites of PTCA lesions and four of non-PTCA lesions (p < 0.05). The percentage of narrowing in PTCA lesions was 28.0 +/- 14.4% at 3-6 months after PTCA, and this improved to 22.8 +/- 15.8% at late study angiography. Coronary score changes were -0.16 +/- 0.54 in PTCA lesions, and 0.08 +/- 0.46 in non-PTCA lesions (p < 0.01). We concluded that the progression of obstructive coronary artery disease in sites with previous PTCA is less than in those with non-PTCA.

Effects of an antithrombin drug in patients with subacute exacerbations of Binswanger disease.

The blood coagulation system has been shown to be activated in subacute exacerbations of Binswanger disease (BD). In our previous study, the antithrombin drug argatroban t ameliorated the neurological exacerbations in a BD patient with antiphospholipid antibody syndrome. We have further examined the therapeutic efficacy of argatroban in 3 BD patients with subacute exacerbations, but without any immune-mediated prothrombotic complications. In 1 out of these 4 patients, treatment with sodium ozagrel, an antiplatelet drug was applied, but was ineffective. In all patients, argatroban treatment reduced the levels of the hemostatic markers, with a corresponding improvement in cognitive dysfunction and gait disorders. These results suggest that the antithrombin effect is true also for BD patients not compromised by the immune-mediated prothrombotic condition.

Acute-phase response of angiotensinogen in rat adjuvant arthritis.

The plasma level of angiotensinogen during the chronic phase of inflammation was studied for comparison with those of other acute-phase reactants in rat adjuvant arthritis. In response to a single injection of Freund's complete adjuvant, this level exhibited a transient increase during the first 24 h. By contrast, increased levels of plasma T-kininogen and alpha 2-macroglobulin, typical acute-phase reactions in the rat, were maintained during the 4-week experimental period. These results suggest that the hepatic synthesis of angiotensinogen is stimulated only in the early phase of chronic inflammation, and therefore that the mechanism underlying the acute-phase response of angiotensinogen is distinct from those currently suggested for other acute-phase reactants.

[A case of multiple cerebral arterial thrombosis due to congenital protein C deficiency].

We report a 49-year-old man who had right hemiparesis and motor aphasia. A computed tomography revealed hypodense areas in the left frontal subcortex. A cerebral angiography demonstrated occlusion of the left distal internal carotid artery and both anterior cerebral arteries, as well as stenosis of the left internal carotid artery at the cervical portion. The second angiogram obtained a month later showed no changes. The diagnosis of atherothrombotic cerebral infarction was established on the basis of clinical profile and angiographic findings. Protein C activity and antigen levels were reduced to approximately one half of the normal level in the patient and his brother. The patient had no other risk factors for stroke. Protein C deficiency has been considered one of the risk factors for thrombotic diseases. Venous thrombosis is the most common clinical manifestation, whereas arterial thrombosis is relatively rare. It is generally believed that arterial ischemic stroke associated with protein C deficiency occurs with embolic mechanism, and atherothrombotic infarction is extremely rare. This is the first report suggesting the possibility that protein C deficiency can cause cerebral thrombosis.

[A case of life-threatening ventricular arrhythmias probably due to psychotropic drugs].

We present a case provoked life-threatening ventricular arrhythmias probably due to psychotropic drugs. The patient was a 55-year-old man who had previously twice operations of aortic valve replacement (AVR). The signs of cardiac failure were recurrently appeared from the end of 1991, and he had received promethazine and sulpiride for his depressive state. From cardiac catheterization, we planned his third AVR. The electrocardiographic (ECG) QTc interval was prolonged to 0.48 seconds on this admission. In March 1992 syncopal attack appeared suddenly, and his monitor ECG revealed frequent polymorphous ventricular tachycardia (VT) and Torsade de Pointes (Tdp). These arrhythmias stopped by emergent cardiac pacing. After discontinuing these psychotropic drugs, no ventricular arrhythmias appeared. Since the patient complained severe insomnia one month before operation, the diminished dose of psychotropic drugs (promethazine and levomepromazine) was readministered. Ten days after the operation, syncopal attack reappeared and his ECG recorded frequent VT and Tdp. During both syncopal attacks his serum potassium and magnesium were within normal limits. Two days later, he died from multi-organ failure. We concluded that life-threatening arrhythmias such as VT and Tdp might develop under the administration of mild psychotropic drugs (promethazine or levomepromazine), therefore, must better take a care of ECG changes in cases of using any psychotropic drugs.

[Two cases of acute myocardial infarction with simultaneous occlusions of two main coronary branches].

We report two rare cases of acute myocardial infarction with simultaneous occlusion of two main coronary branches. Case 1 was a 66-year-old man. Emergent CAG demonstrated total occlusions of RCA-segment 1 and LCX-segment 13. The patient underwent a direct PTCA of the RCA occlusion and an intracoronary infusion of urokinase. Angiography before discharge revealed a 25% stenosis of the RCA and a 75% stenosis of the LCX. Case 2 was a 68-year-old man. His ECG showed ST elevation in inferior and anterior leads. Emergent CAG demonstrated a 99% stenosis of RCA-segment 1 and total obstruction in LAD-segment 6. The distal LAD was not visualized by antegrade and retrograde flow from collaterals. We considered the state of hyper-coagulability and reducing of coronary blood flow as the mechanism of simultaneous occlusions in our cases.