Infectivity of influenza B virus in cultured human muscle.

Severe muscle symptoms, particularly in children, are frequently associated with influenza B virus infection. In this study we examined the effects of influenza B virus (Lee Strain) on cultured human muscle by light and electron microscopy (EM), immunofluorescence, hemadsorption and plaque assays. Muscle injury was also evaluated by the appearance of muscle-specific creatine kinase (CK) in the culture medium. By fluorescence immunocytochemistry viral antigen was demonstrated in muscle cell nuclei within 3 h postinoculation (p.i.) and in the cytoplasm at 6 h p.i. Membrane-associated viral antigen was seen at 16 h p.i., at which time budding influenza virus-like particles could be demonstrated by EM, both in myoblasts and multinucleated myotubes. At 16 h all cells were hemadsorption positive. Plaque assays showed peak virus production at 48 h (p.i.), at which time cytopathic effects (cell retraction, pycnosis and cytoplasmic vacuolization) were prominent and some cells detached from the substratum. Leakage of muscle-specific CK isozyme into the culture medium could be demonstrated as early as 6 h p.i. with peak enzyme activity around 40-48 h p.i. Cytopathic changes and virus production were observed both in myoblasts and myotubes indicating that both cell types are susceptible.

In vivo immunization against autologous glioblastoma-associated antigens.

A glioblastoma patient was immunized in vivo with a mixture of autologous and homologous glioblastoma cells coupled to adjuvant peptide and cord-factor analog. Immune activity of peripheral blood lymphocytes was measured in a short-term 51chromium-release assay against autologous tumor target cells. The patient developed direct cell-mediated cytotoxicity against tumor-associated antigens, which appeared to be T-cell mediated.

Delayed onset of post-rabies vaccination encephalitis.

A 31-year-old veterinarian developed seizures, left hemiparesis, loss of memory, and behavioral disorders 5 months after intensive antirabies vaccination, the longest incubation period yet recorded. Computed tomographic scan revealed a right frontal contrastenhanced mass that extended to the left frontal lobe through the corpus callosum. Brain biopsy showed foci of primary demyelination largely confined to the white matter. The lesions were characteristic of the demyelinating encephalomyelitis that follows treatment with certain vaccines against rabies. The hemiplegia improved, but seizures, memory impairment, and abnormal behavior persisted.

Hyperreactivity to myelin basic protein in multiple sclerosis.

Multiple sclerosis patients, control patients with other neurological diseases, and normal volunteers were assayed in a short-term 51chromium release assay for cell-mediated cytotoxicity against lymphocyte targets coated with myelin basic protein. Multiple sclerosis patients, compared to the other two groups, were hyperreactive to myelin basic protein, both before and after in vitro boost with additional myelin basic protein. The boost served to augment the difference between multiple sclerosis patients and controls.

Symmetric sarcoid polyneuropathy: analysis of a sural nerve biopsy.

A 20-year-old woman with sarcoidosis had uveitis, subacute symmetric sensorimotor neuropathy, and noncaseous granulomas in biopsies of gastrocnemius muscle and sural nerve. Morphologic studies of the nerve confirmed the electrodiagnostic impression of an acute axonal and demyelinating neuropathy. Of 100 teased myelinated nerve fibers, 15% contained myelin ovoids and 24% demonstrated segmental demyelination. Quantitative analysis showed a reduction in the numerical density of myelinated fibers. By electronmicroscopy, unmyelinated fibers were largely spared. The exact mechanism of nerve fiber damage was not determined, but a local effect of the granuloma seemed likely, because most lesions were found in the endoneurium.

Recurrent herpetic angular blepharitis in an adult.

A 36-year-old man had several episodes of left angular blepharoconjunctivitis associated with upper respiratory infections, herpes labialis, or emotional stress. The clinical diagnosis had always been a blepharitis of bacterial origin. During the last attack, cytologic examination of a scraping obtained from an eyelid pustule vesicle revealed multinucleated keratinocytes, diagnostic of a herpetic infection. Virologic studies proved that herpes simplex virus type 1 was the causal agent.

Isolation of influenza virus from muscle in myoglobinuric polymyositis.

We report the first isolation of influenza virus from muscle in a man with myoglobinuria and acute polymyositis. Influenza virus was isolated from cultures of Madin Darby bovine kidney and primary rhesus monkey kidney cells inoculated with muscle homogenates in the presence of trypsin; the virus was identified by neutralization and hemagglutination inhibition studies using influenza B/Lee antiserum. Viral plaque assay was performed with Madin Darby canine cells. Viral antigen was also detected by specific immunofluorescence in muscle, and myxovirus-like particles were seen in subsarcolemmal vacuoles by electronmicroscopy. The pathologic findings were similar to those of childhood dermatomyositis, except for a large proportion of necrotic muscle fibers. The evidence suggests that the pathogenesis of influenzal polymyositis in this patient involved direct viral infection of muscle.

Neonatal polycystic encephalomalacia: four new cases and review of the literature.

Four patients with pathologically documented polycystic cavitation of the brain had an acute illness characterised by stupor, seizures, CSF erythrocytic and monocytic pleocytosis, increased CSF protein, and diminished CSF glucose. The acute phase was followed by chronic decerebation, disappearance of the CSF abnormalities, and radiological evidence of polycystic cavitation of the brain. In one patient Herpes simplex was isolated from a cutaneous vesicle. The CSF abnormalities in the disorder have received scant attention, and have not previously been correlated with the acute and chronic stages. Clearly some cases are associated with Herpes simplex virus. The clinical profile should now be sufficiently distinctive to permit future identification of the aetiology in more neonates.

Susceptibility of human skeletal muscle culture to influenza virus infection. Part 2. Ultrastructural cytopathology.

Cultured post-fused human skeletal muscle monolayers exposed to WSN influenza A virus were analyzed by scanning and transmission electron microscopy. At 12-14h post-inoculation (p.i.), affected mononuclear cells retracted from the cell surface, but remained anchored to the substrate by taut filar processes. Retraction was accompanied by shortening of microvilli, appearance of hemispherical cytoplasmic protrusions and corrugation of the surface proper. These changes were more pronounced at 24 and 48h p.i. The rounded, moribund mononuclear cells eventually detached from the substratum. Surface alterations were accompanied by the intracellular appearance of electron-dense nuclear inclusions (often associated with the nucleolus) and paracrystalline ribosomestudded cytoplasmic bodies, which increased in size and number with time. In myotubes, distinct surface alterations appeared later (24h p.i.). Early myotube retraction was accompanied by accentuation of the longitudinally oriented surface pleats and appearance of "blebs" followed by cell-rounding. At 48-72 h, many myotubes detached from the substratum. The surfaces of those still adhering appeared corrugated. Intranuclear and cytoplasmic inclusions accumulated, and budding virions, often filamentous, could be demonstrated at the plasmalemma of mononuclear cells and myotubes. Late (end-stage) cytopathic effects included clumping of chromatin, breakdown of the nuclear envelope, disappearance of cortical and endoplasmic cytofilaments, mitochondrial swelling, and vesiculation of surface membranes. The lesions leading to cell injury and cell death appeared to be due to massive accumulation of virus-induced products that altered cellular metabolism, with physical and functional abnormalities of surface membranes.

Susceptibility of human skeletal muscle culture to influenza virus infection. I. Cytopathology and immunofluorescence.

We found that fused human muscle in culture supports neurotropic influenza A viral infection, as demonstrated by viral growth experiments, hemadsorption, observation of cytopathic changes and detection of intracellular viral antigen. The time of peak virion production and the appearance of cytopathic effects in these experiments were similar to previously described characteristics in influenza A-susceptible organotypic cultures of other tissues. Cytopathological changes occurred earlier in mononucleated cells than in myotubes and included cell-rounding; cytoplasmic retraction, granularity and vacuolization; ribonucleic acid-containing cytoplasmic inclusions; nucleolar enlargement; and clumping of chromatin. Immunofluorescent staining demonstrated early nuclear fluorescence, followed by spread of viral antigen into the cytoplasm. A subpopulation of mononucleated cells, shown by cloning studies to be myoblasts, showed no cytopathic effects or evidence of intracellular viral antigen and was presumably resistant to neurotropic influenza A infection. The adverse effects of influenza A on fused human muscle cells in tissue culture contrast with the inability of the virus to replicate in skeletal muscle after animal inoculation in acute experiments. Therefore, host factors may affect attempts to produce an animal model of human muscle disease with this virus.

Murine influenza virus encephalomyelitis. III. Effect of defective interfering virus particles.

Defective interfering influenza virus particles lessened the yield of infective virus recovered from brain after intracerebral challange of 3-week old mice, but failed to affect the appearance of disease or lethality. In 7-week old mice, the presence of defective interfering influenza virus particles reduced both lethality and virus yield. Viral inocula containing significant numbers of defective particles appeared to diminish later inflammatory changes, nerve cell degeneration and the appearance of viral antigen in deeper cerebral regions. The protective effect exerted by defective virus particles does not appear related to interferon production in brain.