Richard T. Johnson

Measles encephalomyelitis. Clinical and immunologic studies

Abstract We studied 19 patients with postinfectious encephalomyelitis complicating natural measles-virus infections, and our results support the hypothesis that this demyelinating disease has a pathogenesis similar to that of experimental allergic encephalomyelitis. Early myelin destruction was demonstrated by the presence of myelin basic protein in cerebrospinal fluid, and lymphocyte proliferative responses to myelin basic protein were found in 8 of 17 patients tested. A lack of intrathecal synthesis of antibody against measles virus suggests that measles encephalomyelitis may not be dependent on virus replication within the central nervous system. Similar lymphoproliferative responses to myelin basic protein of lymphocytes from single patients with encephalomyelitis after rabies vaccine or after varicella or rubella virus infections suggest a common immune-mediated pathogenesis for the perivenular demyelinating disease that can follow the injection of neural tissues or infection by a variety of viruses....

Calcium-permeable AMPA receptor plasticity is mediated by subunit-specific interactions with PICK1 and NSF

A recently described form of synaptic plasticity results in dynamic changes in the calcium permeability of synaptic AMPA receptors. Since the AMPA receptor GluR2 subunit confers calcium permeability, this plasticity is thought to occur through the dynamic exchange of synaptic GluR2-lacking and GluR2-containing receptors. To investigate the molecular mechanisms underlying this calcium-permeable AMPA receptor plasticity (CARP), we examined whether AMPA receptor exchange was mediated by subunit-specific protein-protein interactions. We found that two GluR2-interacting proteins, the PDZ domain-containing Protein interacting with C kinase (PICK1) and N-ethylmaleimide sensitive fusion protein (NSF), are specifically required for CARP. Furthermore, PICK1, but not NSF, regulates the formation of extrasynaptic plasma membrane pools of GluR2-containing receptors that may be laterally mobilized into synapses during CARP. These results demonstrate that PICK1 and NSF dynamically regulate the synaptic delivery of GluR2-containing receptors during CARP and thus regulate the calcium permeability of AMPA receptors at excitatory synapses.

PKM-ζ is not required for hippocampal synaptic plasticity, learning and memory

Long-term potentiation (LTP), a well-characterized form of synaptic plasticity, has long been postulated as a cellular correlate of learning and memory. Although LTP can persist for long periods of time, the mechanisms underlying LTP maintenance, in the midst of ongoing protein turnover and synaptic activity, remain elusive. Sustained activation of the brain-specific protein kinase C (PKC) isoform protein kinase M-ζ (PKM-ζ) has been reported to be necessary for both LTP maintenance and long-term memory. Inhibiting PKM-ζ activity using a synthetic zeta inhibitory peptide (ZIP) based on the PKC-ζ pseudosubstrate sequence reverses established LTP in vitro and in vivo. More notably, infusion of ZIP eliminates memories for a growing list of experience-dependent behaviours, including active place avoidance, conditioned taste aversion, fear conditioning and spatial learning. However, most of the evidence supporting a role for PKM-ζ in LTP and memory relies heavily on pharmacological inhibition of PKM-ζ by ZIP. To further investigate the involvement of PKM-ζ in the maintenance of LTP and memory, we generated transgenic mice lacking PKC-ζ and PKM-ζ. We find that both conventional and conditional PKC-ζ/PKM-ζ knockout mice show normal synaptic transmission and LTP at Schaffer collateral–CA1 synapses, and have no deficits in several hippocampal-dependent learning and memory tasks. Notably, ZIP still reverses LTP in PKC-ζ/PKM-ζ knockout mice, indicating that the effects of ZIP are independent of PKM-ζ.

The virology of demyelinating diseases

Infectious agents have been postulated as causes of multiple sclerosis for over a century. The possible role of a virus or viruses is supported by data that (1) a childhood exposure is involved and “viral” infections may precipitate exacerbations of disease, (2) experimental infections in animals and natural infections in humans can cause diseases with long incubation periods, remitting and relapsing courses, and demyelination, and (3) patients with multiple sclerosis have abnormal immune responses to viruses. The pathogenesis of three human demyelinating diseases of known viral etiology is discussed. In progressive multifocal leukoencephalopathy, a papovavirus selectively infects oligodendrocytes and causes focal areas of demyelination. In postmeasles encephalomyelitis, the virus is lymphotrophic and disrupts immune regulation that can result in an autoimmune perivenular demyelinating illness without evidence of infection of the central nervous system. In human immunodeficiency virus‐encephalopathy and myelopathy virus is present in macrophages and microglia and the myelin abnormalities apparently are caused by soluble factors such as viral proteins, cytokines, or neurotoxins. These findings may have implications on how, when, and where to seek viruses in multiple sclerosis.

Myelin Basic Protein as an Encephalitogen in Encephalomyelitis and Polyneuritis Following Rabies Vaccination

Encephalitis and polyneuritis occurring after rabies vaccination are believed to be immunologically mediated. We studied antibody responses to neural antigens in 36 patients with major neurologic complications, 25 with minor complications, and 39 with no complications after immunization with a brain-derived, Semple rabies vaccine. Patients with major complications had significantly elevated levels of antibody to brain white matter as compared with the other groups (P less than 0.001). Assays for antibody to selected central nervous system antigens showed that high levels of serum and cerebrospinal fluid antibody to myelin basic protein correlated with the presence of major neurologic complications (both central and peripheral nervous systems). The level of antibody to cerebroside correlated best with the number of injections of vaccine, but like antibody to myelin basic protein, the antibody to cerebroside was present in the cerebrospinal fluid of patients with major complications. Some patients with major complications also had antibodies directed to the gangliosides GD1b and GT1b. No antibodies to myelin-associated glycoprotein were detected in any of the samples. These data implicate myelin basic protein as an encephalitogen in these autoimmune diseases of the human nervous system, but suggest that immune responses to cerebroside and certain gangliosides may have an augmentative role in severe disease.

Cellular immune responses during complicated and uncomplicated measles virus infections of man

Lymphocytes from patients with measles showed profound and prolonged suppression of proliferative responses to mitogens. The degree of suppression was similar in patients with uncomplicated measles virus infection and in those with pneumonia or postinfectious encephalitis. Despite this suppression, lymphocyte responses to measles antigen and PPD were demonstrated in patients with encephalitis and uncomplicated disease, even early in infection. Most patients with pneumonia did not have demonstrable antigen-specific responses. The proportions of T helper (OKT 4) and T suppressor (OKT 8) cells and functional tests of Con A suppressor cell activity showed no significant difference between control and measles patients but, in contrast to controls, cells from measles patients cultured in the absence of any stimulant significantly suppressed the proliferation of allogeneic responder cells. Nine of 20 supernatant fluids from these cultures possessed a soluble suppressor factor. These studies indicate varied disruptions of immune reactivity during measles.

The neurobiology of human immunodeficiency virus infections.

A variety of diseases of the central and peripheral nervous systems evolves during the course of human immunodeficiency virus (HIV) infections. Most are not related to documented opportunistic infections and may be the direct result of HIV infections, as large proportions of healthy and ill HIV‐infected persons show evidence of nervous system infection. These diseases occur at different times during the infection and have diverse inflammatory, demyelinating, or degenerative pathological features that suggest different pathogenetic mechanisms. The route and determinants of HIV invasion of the nervous system are unknown. Within the brain, viral antigen and RNA are found predominantly in macrophages, but the reason why profound dementia and cortical atrophy result from this infection remains a mystery. By analogy to other lentivirus infections, particularly visna virus in sheep, neuropathological changes may be mediated by cytokines. Other possible pathogenetic mechanisms include toxicity of viral polypeptides, transactivation of viral or cellular genes, autoimmunity, or other opportunistic infections. Clarification of the pathogenesis of HIV‐related diseases is critical to the design of rational therapies.— Johnson, R. T.; McArthur, J. C.; Narayan, O. The neurobiology of human immunodeficiency virus infection. FASEB J. 2: 2970‐2981; 1988.

Etiology of progressive multifocal leukoencephalopathy. Identification of papovavirus.

Abstract Papovaviruses were identified in brains of 13 patients with progressive multifocal leukoencephalopathy, asubacute human demyelinating disease. Fluorescent-antibody staining and electron microscopical agglutination technics demonstrated the JC type of papovavirus in 11 patients. Simian-virus 40 and the virus of progressive multifocal leukoencephalopathy were isolated from the first two patients studied, but all subsequent isolates were of the JC-virus type, and the BK type was not implicated in the disease. The use of monospecific rabbit serums to identify viral antigens in the brain tissue of patients and to serotype by electron microscopy virions extracted directly from brain demonstrates that rapid diagnostic methods can be employed in this disease. (N Engl J Med 289:1278–1282, 1973)

Viral infections and demyelinating diseases.

VIRAL infections can cause demyelinating disease in human beings and laboratory animals. Recent studies of these infections along with epidemiologic data have led to growing speculation that a viru...

West Nile Virus: Pathogenesis and Therapeutic Options

Dr. Juan Gea-Banacloche (Infectious Diseases, Experimental Transplantation, and Immunology Branch, National Cancer Institute, National Institutes of Health [NIH], Bethesda, Maryland): A 55-year-old man with refractory chronic lymphocytic leukemia presented for evaluation to the Clinical Center, NIH, on 30 July 2002, 19 days after his third cycle of EPOCH-F (etoposide, prednisone, vincristine, cyclophosphamide, and fludarabine) chemotherapy. His temperature was 39.3 C. The rest of his vital signs were normal. His history was unremarkable except for numerous mosquito bites during the previous weeks. Results of laboratory tests and imaging studies were unrevealing. The patient remained febrile (temperature, 39 C to 40 C) but clinically stable until day 5, when he reported leg weakness and diplopia. A magnetic resonance imaging (MRI) scan of the brain was normal. A lumbar puncture was performed (Table 1). Broad-spectrum antibiotics and acyclovir therapy were started. On day 6, the patient was drowsy, could not ambulate, and developed a coarse tremor. On day 7, he developed dysphagia and dysarthria and was transferred to the intensive care unit. Electromyographic studies showed mild axonal polyneuropathy. Findings on repeated lumbar puncture on day 7 were unchanged. Intubation was required for airway protection on day 8. On day 10, Maryland State Laboratory reported a positive result for West Nile virus on polymerase chain reaction (PCR) of the cerebrospinal fluid. Generalized flaccid weakness persisted. On day 14, the patient developed status epilepticus, which presented with flickering of the eyelids and tachycardia. Between days 15 and 21, intravenous immunoglobulin with a high titer against West Nile virus (Omr-IgG-am) was administered, without improvement. On day 23, a negative PCR result for West Nile virus in the cerebrospinal fluid was reported for the first time, but the patient never regained consciousness. Ventilatory support was discontinued on day 42. Table 1. Diagnostic Test Results for the Patient West Nile virus belongs to the family Flaviviridae, a large family of positive-strand RNA viruses with 3 main genera (flavivirus, hepacivirus, and pestivirus). Among the more than 70 viruses in the genus flavivirus, several neurotropic and hepatotropic viruses that are important in human disease are transmitted by arthropods (dengue, Japanese encephalitis, yellow fever, and tick-borne encephalitis). West Nile virus belongs to the Japanese encephalitis serocomplex, which also includes Japanese encephalitis and St. Louis encephalitis, among others. West Nile virus was associated with West Nile fever, a nonspecific febrile illness that was found in several countries in Africa and the Middle East, either in epidemics or as an endemic mild febrile illness (1). The association with high rates of encephalitis and death is relatively new (2-5) and suggests the presence of a new strain of virus. The first U.S. cases of West Nile virus natural infection occurred in 1999 in New York, New York (6). The number of human cases did not increase during 2000 and 2001, but the virus spread in animal reservoirs. In 2002, there were 4156 cases with 284 deaths (7). In 2003, there were more than 9000 cases and 220 deaths, and the disease has been identified in almost all parts of the United States (8) (Table 2). Table 2. West Nile Virus Disease in the United States, 19992003 Birds are the main reservoir of West Nile virus in nature; more than 200 species in the United States have been found to be infected. Several species of mosquitoes can acquire the virus after biting a bird with high-level viremia and may then transmit it to the next animals they bite. Transmission between birds in the absence of mosquitoes has been documented in the laboratory, but whether this occurs under natural conditions remains unknown (9). Many species of vertebrates can be infected by virus-carrying mosquitoes. Horses have a high mortality rate (30%); in contrast, cats and dogs are infected frequently, but the case-fatality rate is low. Humans typically do not develop high-level viremia, so they are considered to be a dead end for the virus under normal circumstances. However, transmission through organ transplantation or blood transfusion has been documented (10, 11). One case of transmission through breastfeeding was reported, but the infant remained asymptomatic (12). The proportion of patients who develop disease after acquiring the virus is unknown. The commonly reported estimates (1 in 5 infected patients develops fever and 1 in 150 infected patients develops severe neurologic disease) come from serologic surveillance data from the New York epidemic (13). Of the cases reported in 2003, approximately 70% were reported as West Nile fever (milder disease) and 30% were reported as West Nile meningitis or encephalitis (8). Age older than 70 years seems to be the main risk factor for severe meningoencephalitis and death. Whether immunosuppression is a risk factor remains unclear (3). Dr. Richard T. Johnson (The Johns Hopkins University School of Medicine and Bloomberg School of Public Health, Baltimore, Maryland): West Nile virus was first isolated from the blood of a febrile woman in the West Nile province of Uganda in 1937 (14). Subsequent investigations established that the virus in Africa cycled between culicine species of mosquitoes and various species of birds (15, 16). A mild dengue-like illness with fever, malaise, lymphadenopathy, and rash was thought to be the only manifestation of the infection. During outbreaks in the 1950s, patients occasionally had nuchal rigidity and pleocytosis, which first associated benign aseptic meningitis with West Nile virus. Encephalitis was not recorded (17). The first documented cases of human encephalitis due to West Nile virus occurred in New York City in 1952, when 95 debilitated patients with advanced cancer were inoculated with an Egyptian strain on the premise that the virus might have an oncolytic effect. Encephalitis developed in 9 patients, virus was recovered from spinal fluid of 3 patients, and classic encephalitis was seen in 1 autopsy (18). During the following decades, only few encephalitis cases were reported in Israel, India, and the Mediterranean region (19). A major change in neurovirulence was evident with the epidemics of West Nile virus encephalitis in 1996 in Bucharest, Romania (2), and in 1999 in Volgograd, Russia (4). The 2000 outbreak in Israel showed unprecedented high rates of encephalitis, a higher mortality rate, and more involvement of the elderly population (5), similar to the recent experience in the United States. Greater neurotropism and neurovirulence seem to have evolved in strains of West Nile virus circulating in the Middle East, Europe, and the United States. The virus reported in New York in 1999 closely resembles a recent Israeli isolate. In Central Africa, West Nile virus infections have been associated with fulminant hepatitis resembling yellow fever, suggesting the appearance of hepatotropic strains in that region (20). After a female mosquito takes an infected blood meal, West Nile virus penetrates the gut; replicates in tissues, often including the nervous system; and then invades and replicates in the mosquitos salivary glands. This noncytopathic infection of mosquitoes is lifelong. During subsequent feedings, the mosquito injects virus-laden saliva into the warm-blooded host. Virus may initially infect local fibroblasts, vascular endothelial cells, or cells of the reticuloendothelial system (21). This extraneural infection leads to viremia, which is the probable route for invasion of the central nervous system. Viremia is also essential to provide an infected blood meal to the next mosquito. Clinical disease is not observed in most avian infections; the massive deaths of crows during the past 3 years in the United States also indicate a change in the neurovirulence of the virus. The human incubation period of West Nile virus is 2 to 14 days. Approximately 20% of infected patients develop a mild febrile illness with malaise, myalgias, headache, and lymphadenopathy; few patients develop a maculopapular rash. About 1 in 150 patients develops meningitis or encephalitis. The mean age of patients with encephalitis in 2002 was 56 years, and the mean age of decedents was 79 years. Stiff neck and altered mental status are the most common neurologic findings. Seizures are relatively rare (<5%), and flaccid paralysis is common (about 10%). Immunocytochemical staining of brains of patients with the related Japanese encephalitis showed infection that was limited to neurons scattered throughout the brain but with greater intensity of infection in the basal ganglia, thalamus, and brainstem (22). Older studies have shown inflammation and neuronophagia in anterior horns of the cord, explaining the flaccid paralysis (23, 24). The intense weakness and flaccid paralysis in West Nile virus infections were initially thought to represent an axonal form of the GuillainBarr syndrome (3). The observations of asymmetry of paralysis, preserved tendon reflexes, and electrophysiologic studies support the localization to the anterior horn cells (25). The finding of a paralytic poliomyelitis syndrome has seldom been seen with West Nile virus but is well described with other flavivirus infections, such as Japanese encephalitis and St. Louis encephalitis (26-28). A neurovirulent strain of West Nile virus has spread across the United States and established natural cycles in a wide variety of mosquitoes and birds. This virus will probably be with us for future generations; rates of human disease, however, cannot be predicted. As with St. Louis encephalitis in North America, rates of disease may alternately increase and decrease in the future; in contrast, as with Japanese encephalitis in parts of southeast Asia, high rates of seasonal disease may recur yearly. Dr. Anto Bagic (Clinical Epilepsy Se