Attenuation and immunogenicity of recombinant yellow fever 17D-dengue type 2 virus for rhesus monkeys.

A chimeric yellow fever (YF)-dengue serotype 2 (dengue 2) virus was constructed by replacing the premembrane and envelope genes of the YF 17D virus with those from dengue 2 virus strains of Southeast Asian genotype. The virus grew to high titers in Vero cells and, after passage 2, was used for immunogenicity and attenuation studies in rhesus monkeys. Subcutaneous immunization of naive rhesus monkeys with the 17D-D2 chimeric virus induced a neutralizing antibody response associated with the protection of 6 of 7 monkeys against viremia by wild-type dengue 2 virus. Neutralizing antibody titers to dengue 2 were significantly lower in YF-immune animals than in YF-naive monkeys and protection against challenge with wild-type dengue 2 virus was observed in only 2 of 11 YF-immune monkeys. An anamnestic response to dengue 2, indicated by a sharp increase of neutralizing antibody titers, was observed in the majority of the monkeys after challenge with wild-type virus. Virus attenuation was demonstrated using the standard monkey neurovirulence test. The 17D-D2 chimera caused significantly fewer histological lesions than the YF 17DD virus. The attenuated phenotype could also be inferred from the limited viremias compared to the YF 17DD vaccine. Overall, these results provide further support for the use of chimeric viruses for the development of a new live tetravalent dengue vaccine.

Construction, characterization and immunogenicity of recombinant yellow fever 17D-dengue type 2 viruses.

Chimeric yellow fever (YF)-dengue type 2 (Den 2) viruses were constructed by replacing the premembrane (prM) and envelope (E) genes of YF 17D virus with those from Den 2 virus strains of south-east Asian genotype. Whereas viable chimeric viruses were successfully recovered when the YF 17D C gene and the Den 2 prM gene were fused at the signalase cleavage site, no virus could be rescued from the constructions fused at the viral protease cleavage site. Unlike YF virus that replicated in all the cell lines tested and similar to the Den 2 virus, the recombinant viruses did not replicate in vaccine-production certified CEF and MRC5 cells. Besides, chimeric 17D/Den 2 viruses and their parental viruses reached similar growth titers in Vero and C6/36 cell cultures. Analysis of mouse neurovirulence, performed by intracerebral inoculation, demonstrated that the 17D/Den 2 chimera is more attenuated in this system than the YF 17DD virus. Immunization of mice with this chimera induced a neutralizing antibody response associated with a partial protection against an otherwise lethal dose of mouse neurovirulent Den 2 NGC virus. Overall, these results provide further support for the use of chimeric viruses as an attractive methodology for the development of new live flavivirus vaccines.

Purification and characterization of cold-active L-glutamate dehydrogenase independent of NAD(P) and oxygen.

L-Glutamate dehydrogenase (GLDH) independent of NAD(P) and oxygen was first obtained from the psychrotrophic bacterium Aeromonas sp. L101, originally isolated from the organs of salmon (Oncorhynchus keta). GLDH was purified by a series of chromatography steps on DEAE-Sepharose, Superdex 200pg, Q-Sepharose, CM-Sepharose, and Phenyl-Sepharose. The purified protein was determined to have a molecular mass of 110 kDa and a pI of 5.7. Maximum activity was obtained at 55 degrees C and pH 8.5. The activity of GLDH at 4 and 20 degrees C was 38 and 50%, respectively, of that at 50 degrees C. GLDH was coupled to cytochrome c and several redox dyes including 1-methoxy-5-methylphenazinium methylsulfate (1-Methoxy PMS), 2, 6-dichlorophenylindophenol (DCIP), 9-dimethylaminobenzo[alpha]phenoxazin-7-ium chloride (meldola's blue), 3,3'-[3,3'-dimethoxy-(1,1'-biphenyl)-4, 4'-diyl]-bis[2-(4-nitrophenyl)-5-phenyl-2H tetrazolium chloride] (nitroblue tetrazolium; NBT), and 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyl-2H tetrazolium (INT). The presence of NAD(P) and oxygen gave no oxidation activity to GLDH. Spectroscopic profile and ICP data indicated a b-type cytochrome containing iron.

Tau accumulation in a patient with pallidonigroluysian atrophy.

We studied the brain of a patient with pallidonigroluysian atrophy (PNLA) in whom argyrophilic and abnormally phosphorylated tau positive neurons and glia were identified in the brain on Gallyas-Braak silver staining and immunohistochemical analysis although neurofibrillary tangles were not seen by Bodian silver stain. Immunohistochemical studies using six anti-tau antibodies that recognize the different phosphorylated epitopes of tau protein revealed that these epitopes in neurons and glial cells share common characteristics with neurofibrillary tangles in Alzheimer's disease. Immunoblot analysis of phosphorylated tau protein showed major bands of 64 and 68 kDa and after dephosphorylation, tau consisted mainly of 4 repeat tau. No mutations were detected in the coding exons and their flanking intronic regions of the tau gene. This study suggests that PNLA is one of tauopathy and the biochemical characteristics of phosphorylated tau are similar to those found in progressive supranuclear palsy and corticobasal degeneration.

Family history of subarachnoid hemorrhage and the incidence of asymptomatic, unruptured cerebral aneurysms.

OBJECT: Previously the authors reported a significant correlation between a family history of subarachnoid hemorrhage (SAH) and the discovery of an unruptured aneurysm in a group of healthy volunteers. This study corroborates and extends previous findings regarding the relationship between genetic and acquired factors in the formation of cerebral aneurysms. METHODS: The incidence of asymptomatic, unruptured cerebral aneurysms was studied among patients with a family history of SAH within the second degree of consanguinity. Forty-one unruptured cerebral aneurysms were found in 34 (13.9%) of 244 patients. This incidence was significantly higher than that found in a control group of healthy volunteers (6%). Furthermore, patients who had a family history of SAH combined with multiple systemic risk factors were found to have the highest incidence of unruptured aneurysms (32%; odds ratio 3.49, 95% confidence interval 1.37-8.9). CONCLUSIONS: These findings suggest that patients with a family history of SAH with or without the presence of more than one systemic risk factor are at significantly higher risk of harboring cerebral aneurysms. This high-risk group should be periodically screened and treated with appropriate surgical or other forms of therapy when necessary.

Flow-through chemiluminescence sensor using immobilized histamine oxidase from Arthrobacter crystallopoietes KAIT-B-007 and peroxidase for selective determination of histamine.

A flow sensor with immobilized oxidases is proposed for the determination of histamine in fish meat. Chemiluminometric measurement of histamine was based on the luminol reaction with hydrogen peroxide produced by immobilized histamine oxidase (EC 1.4.3.-.) and peroxidase (EC 1.11.1.7.) within a flow cell. Histamine oxidase was found in cells of Arthrobacter crystallopoietes KAIT-B-007 isolated from soil. The oxidase and peroxidase were coimmobilized covalently on tresylated hydrophilic vinyl polymer beads and packed into transparent PTFE; the tubing was used as the flow cell. One assay for histamine was done at intervals of 2 min without carryover. The calibration curve for histamine was linear from 0.1 microM to 50 microM. The response was reproducible within 1.25% of the relative standard deviation for 115-replicate injections of 50 microM histamine. The sensor system was applied to the determination of histamine in fish meat extracts.

Cerebral venous sinus thrombosis associated with systemic multiple hemangiomas manifesting as chronic subdural hematoma--case report.

A 35-year-old male was admitted with headache, nausea, and vomiting persisting for 2 days. Computed tomography (CT) revealed a left chronic subdural hematoma. Cerebral angiography demonstrated cerebral venous sinus thrombosis (CVST). He had presented with a subcutaneous mass involving the neck at age 2 years, which was shown to be a cavernous angioma, and thereafter shown signs of consumptive coagulopathy with systemic multiple hemangiomas. Burr hole aspiration of the hematoma was performed. Seventy-two-hours later, he developed clouding of consciousness and right hemiparesis. CT revealed a fresh hematoma in the operated subdural cavity and hemorrhagic diathesis manifested. A frontotemporoparietal large craniotomy was performed to remove the hematoma. Extensive electrocauterization was required. He had a satisfactory postoperative course. Collateral venous pathways, resulting from the CVST due to systemic multiple hemangiomas, may have caused hemodynamic stress in the bridging veins which subsequently induced chronic subdural hematoma.

[Seroconversion study of Biken CAM-70 measles vaccine formulations]. / Estudo de soroconversão com formulações da vacina Biken CAM-70 contra sarampo.

OBJECTIVES: To compare seroconversion rates induced by Biken CAM-70 measles vaccines at different viral concentrations. METHODS: Healthy children aged 9 to 18 months from a primary health care unit in Rio de Janeiro, Brazil, and whose guardians agreed with their participation, were randomly assigned to receive one of the following vaccine formulations: 5,000, 1,000 or 200 CCID50 (50% Tissue Culture Infective Dose). The research team, participants, and data analysts were blinded to the type of vaccine administered. Pre- and post-vaccination antibody levels were assessed through Plaque Reduction Neutralization Test. Two interim data analyses were planned to assess unequivocal evidence of the superiority of one of the vaccine types. RESULTS: From 223 recruited children, 84% completed the whole course. Of them, 79% were less than 10 months of age, and 93% did not show detectable measles antibodies in pre-vaccination serum. Seroconversion (four-fold increase in antibody levels) in groups vaccinated with 5,000, 1,000 or 200 CCID50, were 82%, 55%, and 37% (p<0.0000), respectively. Differences in the mean concentration of post-vaccination antibodies were also substantial and statistically significant (p<0.000). Seroconversion rates (pooling data from all vaccine formulations) were 73% to children aged 10 months or more, and 53% in those below 10 months. CONCLUSIONS: Vaccines with concentrations below 5,000 CCID50 did not produce satisfactory seroconversion rates. The vaccine performance by age was consistent with that seen in other studies using Biken CAM-70 strain in which a sizable proportion of 9-month-old children failed to achieve full immunological response.

["Dynamic" MR imaging of the cervical cord in patients with cervical spondylosis and ossification of the posterior longitudinal ligament--significance of dynamic cord compression].

This investigation was designed to assess the influence of dynamic cord compression on severity and course of myelopathy. Sixty-seven patients studied comprised 54 cases of cervical spondylosis and 13 cases of ossification of the posterior longitudinal ligament. These patients underwent "dynamic" MR imaging of the cervical spine. MR images in the sagittal view were obtained in three different neck positions: flexion, neutral, and extension. MR imaging was performed with a 0.15 T resistive unit. For technical reasons, the body coil was used. The pulse sequence was 500/30 (Tr msec/echo time msec) for T1 images. The spinal cord compression was accelerated in 32 cases when extended, in 2 cases when flexed, and in 4 cases when both extended and flexed. In 21 cases, we compared myelograms with MR images in a same neck position. Findings of myelograms well corresponded with those of MR images on 83 percent of intervertebral levels. The patients with dynamic cord compression were proved to have severer long tract signs, and their disability was regressive or progressive case by case for an average of 21-month follow-up. The "dynamic" MR imaging can provide dynamic nature of spinal cord compression, and prognostic clues.

[Cerebellar arteriovenous malformation presenting as trigeminal neuralgia].

A 44-year-old female had been suffering from left facial pain for six years. On admission, intermittent facial pain, hypesthesia and paresthesia were recognized in the 2nd division of the left trigeminal nerve. CT scan revealed an enhanced curvilinear mass on the left cerebellar hemisphere. The left vertebral angiogram demonstrated cerebellar arteriovenous malformation fed by the duplicated superior cerebellar arteries and drained into the inferior petrosal vein. The feeding arteries and the draining vein compressed the trigeminal nerve at its nerve entry zone. Removal of the nidus combined with neurovascular decompression of the trigeminal nerve resulted in complete relief of any facial pain.

[Tethered cord syndrome of adult onset: report of a case and a review of the literature].

A 47-year-old female was admitted with complaints of spontaneous pain at the medial surface of the right thigh, and worsening of urinary incontinence and weakness of the legs. These phenomena had been noticed previously 20 years ago, and again 10 years ago, respectively. There was a scar like skin abnormality of 4 X 6 cm over the lumbosacral region. Compression on this region induced neuralgic pain around the anus radiating down to both legs. Neurologically, weakness of the legs, sensory impairment on the L2-S5 dermatomes and urinary incontinence were recognized. Plain X-P revealed spina bifida on the lower lumber and sacral vertebrae (L4-S1), where a low density mass was found by spinal CT. Metrizamide CT myelography and MRI disclosed medullaris to be low and the spinal cord to be elongated. There was a meningocele surrounded by subcutaneous lipoma at the caudal end of the spinal cord at L5 level. Upon operation, a meningocele containing neural elements and adherent to the intra and extra-dural lipoma was found protruding through the laminar defect. Lipoma was removed partially. The conus medullaris and cauda equina were freed from the surrounding tissue. Dural plasty using the Lyodura was made. Postoperatively, the pain disappeared and her gait disturbance and sensory impairment were significantly improved, but urinary incontinence remained unchanged. Fifty-six cases of tethered cord syndrome of adult onset were reviewed from the literatures. Clinical symptoms, etiologies of the tethering, surgical outcomes and factors causing symptoms were discussed.(ABSTRACT TRUNCATED AT 250 WORDS)

[Percutaneous transluminal angioplasty for cervical internal carotid artery stenosis: case report].

We encountered one case of right cervical internal carotid artery (ICA) stenosis with ulcerative plaque. The patient was treated using percutaneous transluminal angioplasty (PTA). A 70 year-old man complained of left motor weakness and left hemidysesthesia. CT scan on the day of admission showed no abnormal findings. Angiographical findings revealed a severe right ICA stenosis with ulcerative plaque. We first treated the patient conservatively because of high-positioned carotid bifurcation, bronchial asthma and hypertension. 20 hours after the stroke, left motor weakness gradually worsened and CT scan revealed multiple low-density areas in the right fronto-parieto-occipital watershed region. Because of this, we treated the stenosed ICA by PTCA balloon dilatation catheter, and successfully obtained adequate dilatation with no complications. We propose that PTA of ICA stenosis, as well as PTA of the vertebral region, is a useful method for patients who have conditions which would make surgical operations risky.

[Percutaneous transluminal angioplasty for progressing stroke caused by severe basilar artery stenosis: case report].

We encountered one case of progressing stroke caused by severe basilar artery stenosis. The patient was treated with emergent percutaneous transluminal angioplasty (PTA) with satisfactory results. A 54 year-old woman was admitted 15 minutes after the sudden onset of rt. hemiparesis, rt. hemidysesthesia, dysarthria and consciousness disturbance. CT scan on admission showed no abnormal findings. The consciousness level of the patient deteriorated rapidly from JCS 1 to JCS 20 within 30 minutes from the ictus. An emergent angiogram revealed severe basilar artery stenosis at its middle portion and poorly developed collateral circulation. One hour after the stroke occurred, PTA was performed. Using a 3mm diameter balloon catheter, we introduced the balloon into the stenotic lesion and inflated it 6 times from 4 atm to 8 atm pressure. The patient recovered immediately on the operating table improving from JCS 20 to JCS 1. Rt. hemiparesis also improved. Follow-up MRI showed a small area of ischemic change in the brain stem, but no large infarction appeared. The patient was discharged with no neurosurgical deficits after 30 days of PTA. PTA for basilar artery stenosis is still a controversial subject. One reason is the risk of basilar artery perforating branches occlusion by balloon catheter and the other is the difficulty of introducing a flexible balloon catheter into the basilar artery beyond the acute angulation of the vertebral artery. If those two problems were solved, PTA would be the first-choice therapy for basilar artery stenosis in both acute and chronic stages.

[De novo aneurysm associated with a persistent primitive trigeminal artery: case report].

This paper presents a case of a de novo unruptured anterior communicating aneurysm associated with a persistent primitive trigeminal artery. The patient was a 65-year-old female with tinitis, for whom computed tomography with contrast medium indicated right dural enhancement. Dural arteriovernous malformation was suspected and angiography was thus conducted. A right persistent primitive trigeminal artery was demonstrated but no aneurysm could be found. Six years later, magnetic resonance angiography indicated an aneurysm on the anterior communicating artery. It was subsequently removed by surgery. A persistent primitive artery is frequently found to accompany aneurysm but de novo aneurysm associated with such an artery does not appear in the literature to date. The authors emphasize the importance of follow-up angiography in cases of persistent primitive artery not accompanied by an aneurysm.

[A case of distal posterior cerebral artery aneurysm associated with occlusion of the internal carotid artery].

Hemodynamic stress is considered one of the most important factors in the growth of cerebral aneurysms. The authors report a rare case of cerebral aneurysm located at the distal posterior cerebral artery (PCA) in which collateral circulation developed due to occlusion of the internal carotid artery (ICA). A 73-year-old male was admitted to our hospital with a sudden headache and nausea. Computerized tomography (CT) revealed an intracerebral hematoma in the right parieto-occipital lobe and the acute subdural hematoma in both the cerebral interhemispheric fissure and the convexity. Moreover, angiography revealed a saccular aneurysm at the P4 portion of the PCA. The right ICA was occluded at the cervical portion and collateral circulation which had developed in the PCA was extended to the region of the right middle cerebral artery. The aneurysm was clipped 15 days after admission without new neurological complications. This case demonstrates that increased hemodynamic stress plays a role in the growth and rupture of cerebral aneurysm.

[Cerebellar infarcts that require differentiation from tumors: diagnosis with MRI].

It is often difficult to differentiate cerebellar infarct with cerebellar swelling from neoplastic disorders, because former can be shown as cerebellar mass with marked contrast enhancement on CT scan. We analyzed radiologically three cases with cerebellar infarction by using MRI, conventional CT scan and angiography. Two cases in the acute stage could be diagnosed as cerebellar infarction by MRI alone based on the following findings: 1) the lesion was distributed in the territory of cerebellar arteries; 2) the normal pattern of cerebellar folia and fissures was preserved in Gd enhancement MRI image; 3) characteristics of MRI intensity were compatible with hemorrhagic infarction. The other case in the chronic stage showed peculiar enhancement, which was unusual for infarction. It was diagnosed as cellular infarction with reference to the angiographic findings. MRI is generally useful to obtain early diagnosis of tumor-like cerebellar infarcts, and proper treatment should be started as early as possible.

[Percutaneous transluminal angioplasty for cervical carotid artery stenosis].

Percutaneous transluminal angioplasty (PTA) was attempted in 16 patients (17 procedures) with cervical internal carotid artery (ICA) stenosis. Among the 16 patients, 14 were male and 2 were female aged from 44 to 76 years (average 63.4 years). One had cerebral infarction on the acute stage, and the other 15 were in the chronic stage. On CT scan and MRI, there were nine multiple lacunar infarctions and seven watershed infarctions. On angiographical findings, 13 had Rt.-ICA stenosis and 4 had Lt. ICA stenosis. Stenotic lesion existed beyond the level of the third cervical vertebral body in eleven cases, and so-called long segmental stenosis ranged from 3 to 5 cervical vertebral bodies in 3 cases. Before PTA, 14 patients underwent a balloon occlusion test for 3 to 20 minutes (average 9 minutes). Neurological symptoms of hemiparesis or sensory disturbance occurred in 3 patients during balloon inflation, but these disappeared completely after balloon deflation. It took from 1.5 to 2 hours (average 1.7 hours) to carry out PTA including the balloon occlusion test. All cases had satisfactory results with no morbidity or mortality. The mean stenosis ratio of pre-PTA, approximately 80% (55-93%), improved to that of 22% (0-50%) after PTA. Bradycardia and hypotension occurred transiently in 9 cases during and after PTA, but no symptoms remained by atropine sulfate and catecholamine infusion intravenously. In the following 1 to 26 months (mean 9.0 months) after PTA, 3 cases restenosed. The restenosis was recognized by MR angiography after 8 to 26 months (average 15.7 months) of PTA.(ABSTRACT TRUNCATED AT 250 WORDS)

[A 75-year-old man with parkinsonism and sudden death].

We report a 75-year-old man with parkinsonism who died suddenly. The patient was well until 64 years of the age when he had an onset of tremor in his left hand. He was treated with a medicine in another hospital, and his tremor subsided. Five years after the onset, he started to note difficulty in fine finger movements and gait disturbance. He tended to lean backward with frequent falls. He was treated with bromocriptine, trihexyphenydil, and L-dops without apparent improvement. He visited our out patient clinic on November 11, 1993 when he was 75 years of the age. Neurologic examination at that time revealed an alert and well oriented man in no acute distress. Higher cerebral functions were intact. In the cranial nerves, he showed restriction in the upward as well as down ward gaze (40% of normal). He showed masking of the face and spoke in small voice. He walked in a stooped posture with small steps; retropulsion was present. Muscle rigidity was moderately positive in the neck, however, no rigidity was noted in the limbs. No abnormal involuntary movements were seen. He showed moderate bradykinesia and difficulty in finger tapping. Muscle stretch reflexes were normally elicited and the plantar response was flexor bilaterally. Sensation was intact. The autonomic nervous system appeared intact. He was treated with 300 mg/day of Sinemet with marginal improvement in his balance. In February 4, 1994, he had a common cold. On the next day, his parkinsonism worsened and he became unable to walk by himself. He was found unconscious in the bathroom on the same day. He was brought to our hospital by an ambulance. Upon arrival, he was unresponsive and was not breathing. Blood pressure could not be measured. Pupils were dilated without reaction to light. Cardiac resuscitation was attempted, however, ventricular fibrillation appeared on an EEG monitor, and he was pronounced dead at eleven o'clock in the morning. The patient was discussed in a neurological CPC, and the chief discussant arrived at the conclusion that the patient had progressive supranuclear palsy because of vertical gaze palsy, axial rigidity, and poor response to levodopa. Regarding the cause of his sudden death, the chief discussant thought that he developed pulmonary embolism. Postmortem examination revealed non-bacterial thrombotic endocarditis in the heart, but this did not appeared to be related to his sudden death. Multiple disseminated small emboli were found occluding small arteries of the left lung; this was consistent with acute pulmonary embolism, and this was thought to be the cause of his sudden death. In the central nervous system, marked atrophy of the globus pallidus was noted; both internal as well as external segments showed marked atrophy; no myelinated fibers were seen in the globus pallidus. Neuronal cell loss was marked in the globus pallidus, the subthalamic nucleus, and the substantia nigra. No Lewy bodies or tangles were seen. The histologic diagnosis was consistent with pallido-nigro-luysian atrophy. Brownish pigments such as seen in Hallervorden-Spatz disease were seen in the globus pallidus. In addition, formy spheroids were seen in the substantia nigra. However, iron deposits were not so strong as to suggest Hallervorden-Spatz disease. Pallido-nigro-luysian atrophy is a rare neurodegenerative disorder. It is interesting to note that this condition may mimic progressive supranuclear palsy or pure akinesia clinically.