Pursuing an occult carcinoma in a patient with subacute cerebellar degeneration and anticerebellar antibodies. Need for vigorous follow-up.
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Antibodies, Anti-Idiotypic↗
Biomedical subjects
J E Greenlee
Publications and source records attributed to J E Greenlee.
The major site of murine K papovavirus persistence and reactivation is the renal tubular epithelium.
K virus, a murine papovavirus, produces a lethal pneumonia in newborn mice. Animals surviving acute illness develop a persistent infection which reactivates under conditions of immunosuppression. The present study was conducted to identify the cell populations which support persistent K virus infection and to determine the cell populations in which this persistent infection is reactivated during immunosuppression. Mice inoculated by the oral route with 100 50% newborn mouse lethal doses (LD50) of K virus at 14 days of age were followed over a period of 7 months. The distribution of infection was studied by virus assay, immunohistochemistry, and in situ nucleic acid hybridization methods. Viral replication during the acute phase of infection was confined to pulmonary and systemic vascular endothelial cells, as well as to scattered, apparently lymphoid cells within spleens. Beginning 2 months after inoculation, however, specific hybridization for K virus nucleic acids was detected in rare renal tubular epithelial cells, and by 6 months after inoculation renal tubular epithelial cells represented the major site of viral persistence. Positive cells were frequently present in groups of two or more, and a minority of positive cells also expressed viral capsid (V) antigen. Immunosuppression with cyclophosphamide resulted in reactivation of infection, with highest titers of virus being detected in kidneys and with increased numbers of renal tubular epithelial cells expressing viral capsid antigen. Capsid antigen was also detected in rare endothelial cells in kidneys, livers and lungs of these immunosuppressed mice. Although K virus behaves as an endotheliotrope during acute infection, the major site of K virus persistence and reactivation, the renal tubular epithelial cell, is similar to that involved during persistent infection by polyoma virus in mice, SV40 virus in monkeys, and BK and JC viruses in man. The observation that persistently infected renal tubular epithelial cells occur in groups of two or more and occasionally express capsid antigen suggests that virus may persist as a productive infection which is confined by antiviral antibody but maintains itself by cell-to-cell-spread. The present study represents the first instance in which the cell populations which support infection by a member of the polyomavirus subgroup in its natural host have been defined during acute, persistent, and reactivated infection.
Animals↗
Approach to diagnosis of meningitis. Cerebrospinal fluid evaluation.
CSF evaluation is the single most important aspect of the laboratory diagnosis of meningitis. Analysis of the CSF abnormalities produced by bacterial, mycobacterial, and fungal infections may greatly facilitate diagnosis and direct initial therapy. Basic studies of CSF that should be performed in all patients with meningitis include measurement of pressure, cell count and white cell differential; determination of glucose and protein levels; Gram's stain; and culture. In bacterial meningitis, Limulus lysate assay and tests to identify bacterial antigens may allow rapid diagnosis. Where there is strong suspicion of tuberculous or fungal meningitis, CSF should also be submitted for acid-fast stain, India ink preparation, and cryptococcal antigen; unless contraindicated by increased intracranial pressure, large volumes (up to 40-50 mL) should be obtained for culture. If a history of residence in the Southwest is elicited, complement-fixing antibodies to Coccidioides immitis should also be ordered. Newer tests based on immunologic methods or gene amplification techniques hold great promise for diagnosis of infections caused by organisms that are difficult to culture or present in small numbers. Despite the great value of lumbar puncture in the diagnosis of meningitis, injudicious use of the procedure may result in death from brain herniation. Lumbar puncture should be avoided if focal neurologic findings suggest concomitant mass lesion, as in brain abscess, and lumbar puncture should be approached with great caution if meningitis is accompanied by evidence of significant intracranial hypertension. Institution of antibiotic therapy for suspected meningitis should not be delayed while neuroradiologic studies are obtained to exclude abscess or while measures are instituted to reduce intracranial pressure.
Cerebrospinal Fluid↗
Anticerebellar antibodies in neurologically normal patients with ovarian neoplasms.
Several groups of investigators have confirmed the occurrence of antibodies to Purkinje and other cerebellar neuronal populations in the serum and spinal fluid of patients with paraneoplastic cerebellar degeneration. Although this antibody response suggests that paraneoplastic cerebellar degeneration may have an autoimmune basis, it is not known what role anticerebellar antibodies play in the pathogenesis of this disorder or whether the presence of antibodies invariably results in cerebellar injury. We identified 3 patients with ovarian malignancies in whom high titers of circulating anticerebellar antibodies were present without clinical evidence of cerebellar disease. We followed these patients clinically and serologically until their deaths from their neoplasms. All 3 patients remained neurologically normal. In 2 of the patients, anticerebellar antibodies persisted at high titer. CSF obtained from 1 of these patients postmortem did not contain detectable levels of anticerebellar antibody, but histopathologic examination of her cerebellum revealed patchy loss of Purkinje cells. In the 3rd patient, antibody titers fell with removal of the primary tumor and chemotherapy but did not rise with tumor recurrence. Indirect immunofluorescence did not reveal anticerebellar antibodies in the serum or CSF of other patients with neoplasms, patients with other cerebellar disease, or normal controls. The present study demonstrates that patients with ovarian malignancies may occasionally develop antibodies that react with cerebellar neuronal antigens and can maintain this antibody response for protracted periods of time without clinically evident cerebellar injury. Tumor recurrence may not be accompanied by rise in titers of anticerebellar antibodies.
Aged↗
Progressive multifocal leukoencephalopathy.
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Animals↗
Immunoperoxidase labelling of rat brain sections with sera from patients with paraneoplastic cerebellar degeneration and systemic neoplasia.
Sera from patients with systemic cancer found by immunofluorescence staining to have antibodies to human cerebellar cell populations were reacted with vibratome sections of rat cerebellum and examined by peroxidase-antiperoxidase (PAP) methods. Seven patients with clinically or pathologically confirmed paraneoplastic cerebellar degeneration and two neurologically normal patients with high titers of anticerebellar antibodies were studied. Sera from all antibody-positive patients, but not from controls, produced intense staining of brain sections. Sera from patients with ovarian adenocarcinoma reacted predominantly with Purkinje cells and neurons within brainstem nuclei. Sera from patients with oat cell carcinoma and one patient with ductal carcinoma of the breast produced nuclear and cytoplasmic staining of neurons throughout the central nervous system. Serum from a patient with Hodgkin's disease labeled the peripheries of Purkinje cells and Golgi II cells. Serum from a patient with mixed mesodermal sarcoma of the ovary labeled Purkinje cells, basket cells, and scattered astrocytes. Staining of extraneural tissues was not observed. This study confirms the presence of antineural antibodies in patients with systemic neoplasia with and without paraneoplastic cerebellar degeneration and suggests that the antigens recognized by this antibody response may vary with the associated neoplasm.
Adenocarcinoma↗
Serial passage of murine K-papovavirus in primary cultures of mouse embryo cells. Brief report.
Murine K-papovavirus was serially passaged 10 times in primary cultures of mouse embryo cells, using cell suspensions and media from infected cultures to inoculate fresh flasks at each passage level. Titers of K virus infectivity in cell suspensions and media rose during serial passage, but viral hemagglutinating activity was not detected in cells or media before the 10th passage. Although the infectivity of K virus for mouse embryo cultures was enhanced by serial passage, lethality of the virus for suckling mice was lost. The present study represents the first successful serial transmission of K virus infection in vitro.
Animals↗
Anticerebellar antibodies in serum and cerebrospinal fluid of a patient with oat cell carcinoma of the lung and paraneoplastic cerebellar degeneration.
A 56-year-old man was seen with subacute cerebellar degeneration and was found to have oat cell carcinoma of the lung. Antibodies to cerebellar Purkinje cells and granule cells were detected in both serum and cerebrospinal fluid (CSF), and intrathecal antibody synthesis was suggested by serum CSF antibody ratios, CSF IgG index, and CSF IgG synthesis rate. The patient's condition improved slightly with plasmapheresis, corticosteroids, and therapy of his underlying tumor, but he continued to exhibit a severe cerebellar deficit until his death more than one year later. Paraneoplastic cerebellar degeneration may be accompanied by synthesis of anticerebellar antibodies, both systemically and within the central nervous system.
Antibodies↗
Immunoenzymatic labelling of JC papovavirus T antigen in brains of patients with progressive multifocal leukoencephalopathy.
Formalin-fixed, paraffin-embedded autopsy sections of brains from two patients with progressive multifocal leukoencephalopathy (PML) were stained by peroxidase-antiperoxidase methods for human papovavirus T antigen, a nonstructural protein expressed in cells lytically infected or transformed by JC, BK, and SV40 viruses. Adjacent sections were stained for papovavirus common structural antigen, a component of JC, BK, and SV40 virions which is synthesized in productively infected but not transformed cells. Intense immunoperoxidase labelling specific for T antigen was detected in large numbers of oligodendrocytes at the edges of demyelinated areas and in occasional oligodendrocytes within otherwise normal brain. Occasional morphologically normal astrocytic cells exhibited similar specific staining, but only rate atypical astrocytic cells contained detectable amounts of T antigen. Examination of adjacent sections stained with antisera to common structural antigen revealed an identical pattern of immunoenzymatic labelling, indicating that most of the cells expressing T antigen were also expressing viral structural proteins. The present study demonstrates that T antigen can be identified by immunoperoxidase methods in routinely processed autopsy material from cases of PML, but that detectable amounts of antigen are found almost exclusively in cells undergoing lytic infection.
Adult↗
Chronic infection of nude mice by murine K papovavirus.
Nude (nu/nu) mice were inoculated intracranially with 10(6.5) newborn mouse 50% lethal doses of K virus and were studied over a period of 28 weeks using serological methods, virus assay and immunohistological staining for viral antigens. K virus infection of nude mice, although clinically asymptomatic, was slowly progressive despite prompt IgM and IgG antibody response. The highest titres of K virus infectivity were reached in spleens, kidneys and intestines. Vascular endothelial cells represented the major site of viral replication, as has been shown to be the case in immunologically normal mice, with extensive involvement of intestinal capillaries. In addition, however, unlike immunologically normal mice, nude mice inoculated with K virus developed multifocal infection of renal tubular epithelial cells. Nude mice did not develop histologically detectable evidence of central nervous system involvement by K virus, and K virus infection did not result in neoplasia. Infected vascular endothelial cells and renal tubular epithelial cells in animals studied at 16 and 27 weeks after inoculation were grouped in scattered clusters, suggesting local spread of infection. The present study indicates that nude mice with preserved B cell function but impaired T cell-mediated immunity are able to limit systemic dissemination of K virus but are unable to prevent local progression of infection by cell-to-cell spread. K virus is capable of altering its cellular tropism during chronic infection.
Animals↗
Acute febrile cerebrovasculitis: a syndrome of unknown, perhaps rickettsial, cause.
In late 1983, five patients living near Charlottesville, Virginia, were treated for an unusual syndrome of fever, headache, altered mentation, multifocal neurologic signs, and cerebrospinal fluid pleocytosis. Clinical signs of brainstem disease developed in four patients. All five had had recent exposure to forests or wood and contact with flea-infested dogs. Two patients died; one survivor has had recurrent seizures. Brain biopsy samples in two patients and autopsy findings in another showed cerebral vasculitis and perivasculitis involving mostly venules and capillaries. In the autopsy, the severest vascular lesions involved the brainstem and thalami, where they were accompanied by acute fibrinoid necrosis, but discrete vascular lesions of lesser intensity were randomly distributed in the white matter and cortex. Serologic studies on paired specimens in four patients showed significant cross-reacting antibody responses to rickettsial (typhus-group) antigens in the indirect hemagglutination, latex agglutination, and IgM microimmunofluorescence tests, but no agent was visualized or isolated. The cause of this serious inflammatory disorder is unknown.
Acute Disease↗
Immunofluorescent labeling of nonhuman cerebellar tissue with sera from patients with systemic cancer and paraneoplastic cerebellar degeneration.
Sera from patients with paraneoplastic cerebellar degeneration have been shown to contain high titers of antibody to human Purkinje cells. It is not known, however, whether these sera react with cerebellar material from species other than man. The present study was conducted to determine whether cerebellar tissue of nonhuman species could be used to screen human sera for anticerebellar antibodies and whether similarities in cerebellar antigens between nonhuman and human material might permit attempts to transmit cerebellar degeneration to experimental animals by passive transfer of patient sera. Sections of human, monkey, pig, sheep, cat, rabbit, and rat cerebellar tissue were overlaid with serial dilutions of positive sera from patients with cancer and with paraneoplastic cerebellar degeneration and were stained using indirect immunofluorescence methods. All sera stained specific Purkinje cells when reacted with monkey, pig, and rabbit cerebellar sections, but not all sera stained sheep, cat, or rat tissue. Immunofluorescent labeling of animal cerebellar tissue was less bright than that obtained with human cerebellar sections, and anticerebellar antibody titers were invariably lower when assayed with nonhuman than with human material. Although human anticerebellar antibodies react with cerebellar tissue from other animal species, patient-to-patient variation in staining is sufficiently great that not all patient sera might be suitable for passive transfer experiments, and that attempts to identify anticerebellar antibodies by reacting patient sera with nonhuman cerebellar tissue could be negative where these antibodies are in fact present and could be demonstrated using human material.
Animals↗
Interaction of K papovavirus with hamster cells: transformation of glial cells in vitro but failure of the virus to produce central nervous system tumors in vivo.
Primary cultures of lungs, kidneys, and glial cells derived from midgestation Syrian hamsters were inoculated with 10(5) hemagglutinating units of murine K-papovavirus and were serially subcultivated to allow appearance of lines of persistently infected or transformed cells. K virus did not replicate in renal cell cultures and produced only transient productive infection of lung cells. Evidence of K virus-induced cell transformation was not detected in either of these cultures. Inoculation of glial cultures with K virus, however, resulted initially in a protracted infection in which 80--100 percent of cells expressed K virus V antigen for 18 subcultivations and in which cloning experiments suggested that all cells in the culture contained the viral genome. After 18 subcultivations numbers of positive cells rapidly diminished, and cells appeared which exhibited altered morphology and density dependence. These altered cells (KVHG3 cells) grew well in serum-free media, could be cloned in soft agar, and were negative for infectious virus or K virus V antigen. Although KVHG3 cells did not exhibit staining when reacted with antisera to K virus T antigen, Southern blot analysis of these cultures demonstrated the presence of K virus DNA integrated into the host chromosomal DNA and indicated that some rearrangement of the viral genome had occurred. Attempts to produce tumors in hamsters with these cells were unsuccessful, as were attempts to induce tumors in newborn hamsters by intracranial inoculation of K virus. The present study demonstrates that K virus is capable of causing productive infection and cell transformation in primary cultures of fetal hamster glial cells but that other hamster cell types are relatively resistant to the virus and that both K virus and K virus-transformed hamster cells are poorly oncogenic for hamsters in vivo.
Animals↗
Persistent infection and transformation of mouse glial cultures by K virus, a murine papovavirus.
Foetal mouse glial cultures were inoculated with murine K papovavirus and subjected to serial subcultivation. Two cell lines were developed. The first of these, KVBCG2A, remained positive for viral infectivity and K virus capsid (V) antigen for over 30 subcultivations. Productive infection was not abolished by serial subcultivation in the presence of antiviral antibody. The second cell line, KVBCG1B, became negative for infectious virus and K virus V antigen, could be cloned from single cells and produced tumours in mice. Sera from tumour-bearing animals produced nuclear fluorescence of KVBCG1B cells and K virus-infected mouse embryo cells but did not react with uninfected mouse embryo cells or with cells infected by polyoma virus. DNA hybridization studies confirmed the presence of K virus DNA in KVBCG1B cells and suggested integration of the viral genome into host chromosomal DNA. K virus produces both persistent infection and cell transformation in glial cultures derived from its natural host.
Animals↗
Reactivation of persistent papovavirus K infection in immunosuppressed mice.
Papovavirus K (K virus) is a murine papovavirus that produces a fatal interstitial pneumonia in newborn mice and a clinically inapparent infection in older animals. The present study was conducted to determine whether the virus produces latent infection in animals surviving acute infection and whether the infection can be reactivated by immunosuppression. Mice were inoculated by the oral route with 100 newborn mouse 50% lethal doses at 12 days of age and followed for 8 months by using immunofluorescence staining. Cells positive for K virus capsid antigen were found in lungs, livers, kidneys, intestines, and brains for 6 months, but not thereafter. Organ examined at 8 months were negative for virus by tissue culture assay, mouse inoculation, explantation, and cocultivation. Immunosuppression of the remaining animals with 8 weekly injections of cyclophosphamide (150 mg/kg) resulted in the reappearance of viral antigen and infectious virus in multiple organs including brains. The highest titers of virus were present in kidneys. One animal sacrificed after 42 days of immunosuppression was found to have a small pulmonary adenoma or alveologenic carcinoma, but efforts to explant this tumor into tissue culture were unsuccessful. The present study demonstrates that K virus produces a latent infection that is reactivated by immunosuppression, and our results raise questions as to whether reactivated infection may occasionally be associated with the development of neoplasia.
Animals↗
Antibodies to cerebellar Purkinje cells in patients with paraneoplastic cerebellar degeneration and ovarian carcinoma.
Sera from 2 patients with ovarian carcinoma and paraneoplastic cerebellar degeneration confirmed postmortem were reacted with frozen sections of human cerebellum and stained using indirect immunofluorescence methods. Both sera produced bright cytoplasmic staining of Purkinje cells and of neurons within deep cerebellar nuclei. Titration of these sera to end point revealed staining at final dilutions of 1:640 and 1:2,560, respectively. Neither of these sera reacted with sections of human cerebrum, basal ganglia, spinal cord, peripheral nerve, lung, liver, kidney, or ovary. Staining of Purkinje cells was not obtained with sera from 34 normal, healthy controls, 5 patients with oat cell carcinoma of the lung, 6 patients with inflammatory central nervous system disorders, or 12 of 14 neurologically normal patients with ovarian carcinoma. Sera from 2 neurologically normal patients with ovarian carcinoma, however, produced staining of Purkinje cells and deep nuclei similar to that obtained with sera from patients with paraneoplastic cerebellar degeneration. The present study documents the presence of antibodies to Purkinje cells in patients with ovarian carcinoma and cerebellar degeneration and demonstrates that development of these antibodies may antedate the onset of clinically evident cerebellar degeneration.
Antibodies↗
Pathogenesis of K virus infection in mice.
The interaction of a murine papovavirus, K virus, with its natural host, the mouse, has been studied as a possible means of gaining insight into papovavirus infections of man. These investigations have delineated the pathogenesis of K virus infection in suckling, weanling, and adult animals, have demonstrated that the virus does not produce transplacental infection, and have shown that the virus has a strong affinity for vascular endothelium but does not appear to replicate in epithelial cells in kidneys, ureters, or bladder. Preliminary work suggests that, within brains, the virus replicates not only in capillary endothelial cells but also in astrocytes and perineuronal satellite cells. In animals surviving acute infection, K virus produces protracted infection of scattered cells within multiple organs including brains and is capable of persisting despite high titers of circulating antibody. K virus infection of mice provides a useful model of acute and persistent infection by a member of the papovavirus group.
Animals↗
An in vitro assay for K-papovavirus of mice.
Primary cultures of mouse embryo cells were inoculated with K virus, a murine papovavirus, and were examined for cytopathic effect (CPE) of for the development of fluorescent antibody staining specific for K virus V antigen. CPE was not observed. However, numerous cells in infected cultures exhibited positive nuclear fluorescence, and the presence of papovavirus virions was demonstrated by electron microscopy. Extracts from infected cultures produced typical K virus pneumonia in newborn mice. Inoculation of cultures with serial dilutions of virus demonstrated that these cells provide a fluorescent antibody assay for K virus equal in sensitivity to animal inoculation methods. Although specific K virus fluorescence was also detected in cultures of fetal mouse endocardial cells, livers, placentas, and brains, positive cells were much less abundant in these cultures than in cultures of mouse embryo cells. The mouse embryo culture assay described in the present paper represents the first method of measuring K virus infectivity in vitro.
Animals↗