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We demonstrated the presence of an in vitro transmissible cytotoxic agent (TCA) in the cerebrospinal fluid (CSF) of patients with different acute neurological diseases. The nature of this agent is still a matter of study since repeated attempts have failed to identify it as a conventional infectious agent. Here, we describe the mechanisms through which TCA affects human astrocytes, demonstrating: a late apoptotic process, mediated by caspases 9 and 3 activation, involving the Bcl2-Bak-axis; an early and late p38 MAPK activation; an interference with the IL-8 and MCP-1 secretory response. These in vitro data provide initial evidence of TCA involvement as a pro-apoptotic and pro-inflammatory signal, directly affecting astrocytic behavior. The implications of these findings in certain neurological diseases will be discussed.

A case of primary infection by human herpesvirus 6 (HHV-6) variant A in a 54-year-old woman, which occurred at the same time as the onset of encephalomyelitis, is reported. The correlation between the two events is discussed. It is speculated that, during the early phase of the infection, the HHV-6 spread to the central nervous system and triggered a pathogenic process that initially developed without symptoms. When the neurological disorders appeared, HHV-6 had already established a latent state: only the virus carried by infected blood cells was detected in the cerebrospinal fluid.

An infection by HHV-7 with presence of virus DNA in the spinal fluid was ascertained in a twenty five month old boy with an epileptic syndrome shortly after birth. The child was frequently admitted to hospital due to his basal disease and frequent bacterial infections. In the occasion of one of these hospital admissions for bacterial infections in conjunctiva, spleen and a lung, virological investigations were also carried out disclosing the presence of HHV-7 DNA in a sample of spinal fluid and of serum and the absence of DNAs from the other herpesviruses. The patient's monitoring for HHV-7 showed the presence of HHV-7 DNA in a sample of serum and in various samples of peripheral blood lymphocytes and saliva, but not in the cerebrospinal fluid sample successive to that positive for HHV-7 DNA. Forty seven days before the diagnosis of HHV-7 infection, the patient received a human gamma-globulin therapy which obscured the humoral response mounted against the virus by the host, so, whether the HHV-7 presence in the central nervous system was associated with a primary or a reactivated infection remained uncertain. The absence of detectable HHV-7 serum IgM antibody, however, suggests the HHV-7 DNA finding on the occasion of a virus reactivation rather than a primary infection. The virological data suggest that HHV-7 may have possibly reached the central nervous system in the course of a viremia. The absence of HHV-6 and HHV-7 antibodies, present in the patient's serum at a high level, support the integrity of the blood-brain barrier. A possible pathogenetic role of HHV-7 in the child's disease seems unlikely, since the epileptic syndrome was pre-existing.

Twenty-seven cervical condylomata were studied by morphological and immunohistochemical methods (peroxidase-antiperoxidase technique according to Sternberger, with some modifications). The antiserum was obtained from rabbits immunized by human papillomavirus virions; 37% of condylomata stained positively and the koilocytotic cells showed a dark brown nuclear stain. This technique, not particularly useful for diagnostic purposes, could be employed to obtain better understanding of the natural history of these cervical neoplasias.

Positive results were attained when human peripheral blood lymphocytes (PBLs) were investigated for their ability to propagate a transmissible cytotoxic activity (TCA) isolated on VERO cell cultures from a sample of cerebrospinal fluid (CSF) drawn from a woman with ischemic brain injury. In consideration of this finding it can be assumed that "in vivo" blood lymphocytes contributed to give rise to the TCA detected "in vitro" in the CSF inoculum.

Twenty five cases of meningitis occurred in urban areas surrounding a city (Modena) in Northern Italy, in the period May-July 1999. When the patients were admitted to the Infectious Diseases Division of the University of Modena and Reggio Emilia Hospital and studied by virological and serological methods, the meningitis proved to have an enteroviral origin and enterovirus ECHO 4 type was responsible for all cases of illness. An epidemiological characteristic of the enteroviral meninigitis outbreak was the adult age in 23 out of the 25 patients (mean age 24.50 +/- 7.84 years). The monthly distribution of the aseptic meningitis cases was the following: five cases occurred in May, 13 in June and seven in July. The origin of the spread of the virus infection and the reason for its sudden end remained unknown. The unusual drop in temperature which occurred in the geographic area involved in the aseptic meningitis outbreak at the beginning of August could have interfered with the slowdown in virus circulation.

A transmissible cytotoxic agent thought to be associated with one or more misfolded protein(s) was found in several cerebrospinal fluid (CSF) samples from neurological patients. Since some experiments carried out to identify this unusual infectious factor showed the block of its propagation by rabbit gammaglobulins (IgGs), the search for such an activity by human IgGs was programmed. Neutralizing assays carried out using human sera as IgGs source showed a blocking property displayed by: twenty serum samples from as many patients with a diagnosis of acute infection, two of ten sera from healthy subjects and four serum samples from patients with lupus erythematous (SLE). When neutralizing sera were tested on cell cultures in immunofluorescence assays for the serum ability to label specific protein( s), similar fluorescent pictures resulted in treated and control cells. On the other hand, the SLE serum samples disclosed a granulosity of the nuclear material of cytotoxic cells in accordance with the DNA apoptotic laddering reported in previous papers. Oxidative disorders, as suggested by the immunoblotting analysis of the antioxidant enzymes Mn-superoxide dismutase (SOD2) and heme-oxygenase 1 (HO-1), point to an alteration of the oxidative pathway among the causes of the DNA damage induced by the cytotoxic transmissible agent under study.

Protein misfolding and aggregation are the common mechanisms in a variety of aggregation-dependent diseases. The compromised proteins often assemble into toxic, accumulating amyloid-like structures of various lengths and their toxicity can also be transferred both in vivo and in vitro a prion-like behavior. The characterization of protein interactions, degradation and conformational dynamics in biological systems still represents an analytical challenge in the prion-like protein comprehension. In our work, we investigated the nature of a transferable cytotoxic agent, presumably a misfolded protein, through the coupling of a multi-detector, non-destructive separation platform based on hollow-fiber flow field-flow fractionation with imaging and downstream in vitro tests. After purification with ion exchange chromatography, the transferable cytotoxic agentwas analyzed with Atomic Force Microscopy and statistical analysis, showing that the concentration of protein dimers and low n-oligomer forms was higher in the cytotoxic sample than in the control preparation. To assess whether the presence of these species was the actual toxic and/or self-propagating factor, we employed HF5 fractionation, with UV and Multi-Angle Light Scattering detection, to define proteins molar mass distribution and abundance, and fractionate the sample into size-homogeneous fractions. These fractions were then tested individually in vitro to investigate the direct correlation with cytotoxicity. Only the later-eluted fraction, which contains high-molar mass aggregates, proved to be toxic onto cell cultures. Moreover, it was observed that the selective transfer of toxicity also occurs for one lower-mass fraction, suggesting that two different mechanisms, acute and later induced toxicity, are in place. These results strongly encourage the efficacy of this platform to enable the identification of protein toxicants.

Previous infection, both of bacterial and viral origin, is reported to represent an independent risk factor for ischemic stroke in children and young adults. The authors describe the case of an immunocompetent young woman who developed a middle cerebral artery thrombosis and stroke in course of a recurrence of human parvovirus B19 (PVB19) infection. A previously healthy 25-year-old woman developed right ataxic hemiparesis, 5 days after the onset of a flulike syndrome. Magnetic resonance imaging of the brain revealed acute multiple left frontal-parietal ischemic lesions. Conventional and magnetic resonance angiograms revealed a stenosis in the left middle cerebral artery. Nested polymerase chain reaction detected PVB19-specific DNA sequences in the cerebrospinal fluid and blood, and serology showed high titers of high avidity immunoglobulin G against PVB19. After 10 days, the patient's recovery was nearly complete. One month later, PVB19 disappeared from the serum, whereas it persisted in the peripheral blood mononuclear cells. This case report suggests that PVB19 infection may play a trigger role in the development of ischemic stroke, and that it should be considered in the screening of infectious risk factors for cerebrovascular diseases in young adults.