Apolipoprotein E-ε4 deficiency and cognitive function in hepatitis C virus-infected patients.

Hepatitis C virus (HCV) causes not only liver damage in certain patients but can also lead to neuropsychiatric symptoms. Previous studies have shown that the type 4 allele of the gene for apolipoprotein E (APOE) is strongly protective against HCV-induced damage in liver. In this study, we have investigated the possibility that APOE genotype is involved in the action of HCV in brain. One hundred HCV-infected patients with mild liver disease underwent a neurological examination and a comprehensive psychometric testing of attention and memory function. In addition, patients completed questionnaires for the assessment of fatigue, health-related quality of life and mood disturbances. Apolipoprotein E gene genotyping was carried out on saliva using buccal swabs. The APOE-ε4 allele frequency was significantly lower in patients with an impairment of working memory, compared to those with a normal working memory test result (P = 0.003). A lower APOE-ε4 allele frequency was also observed in patients with definitely altered attention ability (P = 0.008), but here, the P-value missed the level of significance after application of the Bonferroni correction. Our data suggest that the APOE-ε4 allele is protective against attention deficit and especially against poor working memory in HCV-infected subjects with mild liver disease. Considering the role of apolipoprotein E in the life cycle of the virus, the findings shed interesting new light upon possible pathomechanisms behind the development of neuropsychiatric symptoms in hepatitis C infection.

The helicase-primase inhibitor BAY 57-1293 reduces the Alzheimer's disease-related molecules induced by herpes simplex virus type 1.

Herpes simplex virus type 1 (HSV1) infection of cultured cells causes the formation of ß-amyloid (Aß) and abnormal tau (P-tau). These molecules comprise the main components of the abnormal protein deposits, amyloid plaques and neurofibrillary tangles, respectively, in Alzheimer's disease (AD) brains, and they have been implicated in disease development. The formation of P-tau, but not of Aß, depends on viral DNA replication, but nonetheless, three antiviral agents that inhibit HSV1 DNA replication, including acyclovir (ACV), were found to reduce greatly the level of Aß as well as P-tau, the former probably through prevention of viral spread. Previous studies showed that HSV1 DNA is present and is active in the brain of many elderly people, including AD patients, and that in combination with the type 4 allele of the apolipoprotein E gene, it is likely to play a role in the disease, perhaps via Aß and P-tau production. With the aim of finding the most suitable antiviral for inhibiting Aß and P-tau formation as well as HSV1 DNA replication, for future use in a clinical trial for treating AD, we compared the efficacy of ACV with that of another antiviral, BAY 57-1293, which acts by a different mechanism from ACV. We found that BAY 57-1293 is more efficient than ACV not only in inhibiting HSV1 replication, confirming previous studies, but also in decreasing Aß and P-tau formation. Also, the cell clusters that are formed during infection are reduced in size much more efficiently by BAY 57-1293 than by ACV. These data suggest that BAY 57-1293 would be a more effective agent than ACV for treating AD.

Herpes simplex virus type 1 DNA is located within Alzheimer's disease amyloid plaques.

The brains of Alzheimer's disease sufferers are characterized by amyloid plaques and neurofibrillary tangles. However, the cause(s) of these features and those of the disease are unknown, in sporadic cases. We previously showed that herpes simplex virus type 1 is a strong risk factor for Alzheimer's disease when in the brains of possessors of the type 4 allele of the apolipoprotein E gene (APOE-epsilon4), and that beta-amyloid, the main component of plaques, accumulates in herpes simplex virus type 1-infected cell cultures and mouse brain. The present study aimed to elucidate the relationship of the virus to plaques by determining their proximity in human brain sections. We used in situ polymerase chain reaction to detect herpes simplex virus type 1 DNA, and immunohistochemistry or thioflavin S staining to detect amyloid plaques. We discovered a striking localization of herpes simplex virus type 1 DNA within plaques: in Alzheimer's disease brains, 90% of the plaques contained the viral DNA and 72% of the DNA was associated with plaques; in aged normal brains, which contain amyloid plaques at a lower frequency, 80% of plaques contained herpes simplex virus type 1 DNA but only 24% of the viral DNA was plaque-associated (p < 0.001). We suggest that this is because in aged normal individuals, there is a lesser production and/or greater removal of beta-amyloid (Abeta), so that less of the viral DNA is seen to be associated with Abeta in the brain. Our present data, together with our finding of Abeta accumulation in herpes simplex virus type 1-infected cells and mouse brain, suggest that this virus is a major cause of amyloid plaques and hence probably a significant aetiological factor in Alzheimer's disease. They point to the usage of antiviral agents to treat the disease and possibly of vaccination to prevent it.

Apolipoprotein E-epsilon 4 and recurrent genital herpes in individuals co-infected with herpes simplex virus type 2 and HIV.

Apolipoprotein E (APOE) alleles have been associated with the severity of, or susceptibility to, infection by various microbes. We investigated the potential association between the APOE-epsilon 4 allele and the rate of recurrence of genital herpes in patients who were HIV positive and herpes simplex virus type 2 (HSV-2) seropositive. The APOE-epsilon 4 allele was significantly associated with recurrent genital ulceration independent of ethnicity, antiretroviral therapy and CD4 count (OR 8.3; 95% CI 2.4 to 28.5). To our knowledge, this is the first published study to demonstrate this association and suggests that APOE-epsilon 4 may represent a future prognostic marker for symptomatic recurrence of genital herpes in individuals with HIV.

The role of viruses and of APOE in dementia.

The virus, herpes simplex virus type 1 (HSV1), when present in brain, acts together with the type 4 allele of the APOE gene, a known susceptibility factor in Alzheimer disease (AD), to confer a strong risk of AD; in carriers of the other two main alleles of the gene, the virus does not confer a risk. It also has been shown that the outcome of infection in the case of five diseases known to be caused by viruses is determined by APOE. It is hoped that the discovery of the involvement of HSV1 in AD will lead to future antiviral therapy and possibly to immunization against the virus in infancy.

Infiltration of the brain by pathogens causes Alzheimer's disease.

Despite very numerous studies on Alzheimer's disease (AD), especially on amyloid plaques and neurofibrillary tangles, little information has been obtained thus on the causes of the disease. Evidence is described here that implicates firstly herpes simplex virus type 1 (HSV1) as a strong risk factor when it is present in brain of carriers of the type 4 allele of the gene for apolipoprotein E (APOE-4). Indirect support comes from studies indicating the role of APOE in several diverse diseases of known pathogen cause. A second putative risk factor is the bacterium, Chlamydia pneumoniae. This pathogen has been identified and localized in AD brain. Current studies aimed at "proof of principle" address the entry of the organism into the CNS, the neuroinflammatory response to the organism, and the role that the organism plays in triggering AD pathology. An infection-based animal model demonstrates that following intranasal inoculation of BALB/c mice with C. pneumoniae, amyloid plaques/deposits consistent with those observed in the AD brain develop, thus implicating this infection in the etiology of AD.

Does apolipoprotein E polymorphism influence susceptibility to malaria?

Outcome of infection varies greatly among people, and in the case of three very different viruses, it is determined by apolipoprotein E (APOE) genotype. APOE might affect outcome of malaria infection also, since apoE protein and the protozoon (like the viruses) share cell entry mediators (heparan sulphate proteoglycans and/or specific apoE receptors). APOE polymorphisms give rise to protein variants that differ in binding strength to these mediators; thus, the extent of competition between apoE and protozoon for cell entry, and hence magnitude of protozoan damage, might depend on apoE isoform. Genotypes of infants infected with malaria were examined. It was found that APOE epsilon 2 homozygotes became infected at an earlier age than those carrying the other genotypes, the difference being statistically significant. Parasite densities, all of which were low, did not differ significantly. This effect, although based on small numbers, suggests that APOE epsilon 2 may be a risk factor for early infection.

Vaccination prevents latent HSV1 infection of mouse brain.

Herpes simplex encephalitis (HSE) is a rare but very serious disorder caused by herpes simplex type 1 virus (HSV-1). Treatment with acyclovir decreases mortality but many patients still suffer cognitive impairment subsequently. A vaccine against HSV1 would therefore be of great value. HSV-1 has been implicated also in Alzheimer's disease (AD): we established that HSV1 resides in the brain of about two thirds of AD patients and aged normal people, and that in carriers of the type 4 allele of the apolipoprotein E gene, it is a strong risk factor for AD. Thus a vaccine against HSV-1 might prevent development of AD in some cases. To find whether a vaccine of mixed HSV-1 glycoproteins (ISCOMs), which protects mice from latent HSV-1 infection of sensory ganglia, prevents HSV1 latency in the CNS, ISCOM-vaccinated or unvaccinated animals were infected with HSV-1. Using polymerase chain reaction (PCR) we detected HSV-1 in brain from 16 of 39 unvaccinated mice (41%), but only 3 of 41 vaccinated mice (7%) (P < 0.001). Thus, ISCOMs protect the CNS also, suggesting their possible future usage in humans.

Herpes simplex encephalitis: involvement of apolipoprotein E genotype.

It was previously found that herpes simplex type 1 virus (HSV1) when present in the brain, is a risk factor for Alzheimer's disease in carriers of the type 4 allele of the gene for apolipoprotein E (apoE epsilon4), and apoE epsilon4 is a risk factor for herpes labialis. Whether a specific allele of the gene is involved in susceptibility to another disorder caused by HSV1-herpes simplex encephalitis (HSE)-has now been investigated. DNA was prepared from formalin-fixed, paraffin-embedded blocks of specimens from the brain or spleen of 14 United Kingdom patients with HSE, confirmed by necropsy, and from the CSF of seven United Kingdom clinical patients with HSV1 in their CSF detected by polymerase chain reaction (PCR). ApoE genotype of the DNA from blocks was determined by seminested PCR, and of the DNA from CSF by one step PCR, followed by restriction endonuclease digestion. The apoE allele frequencies were compared with values previously obtained for 238 normal people from the United Kingdom. The apoE epsilon2 allele frequency of the patients with HSE was 26%, significantly higher than the value of 7% for the normal subjects (OR=4.6, 95% confidence interval (95% CI) 2. 0-10.8). The apoE epsilon3 and epsilon4 allele frequencies did not differ significantly between the two groups. Thus, it seems that apoE epsilon2 is a risk factor for HSE.

Herpes simplex virus type 1 and Alzheimer's disease.

Until recently, the only risk factors implicated in noninherited cases of Alzheimer's disease were increasing age, Down's syndrome, and probably, head injury. Having found that herpes simplex type 1 virus (HSV1) is present in the brain of many elderly people, we discovered that it is a risk factor for Alzheimer's disease when in the central nervous system of APOE-epsilon4 allele carriers. On the basis of this result and our finding that apoE-epsilon4 is a risk factor for herpes labialis, we suggested that the combination of virus and genetic factor is particularly damaging in the nervous system. The present review describes 1) the search for HSV1 in human brain; 2) HSV1 infection of the peripheral nervous system; 3) HSV1 infection of the central nervous system; 4) how APOE genotype might influence HSV1 infection; 5) possible APOE genotype effect on viral latency and its reactivation; 6) interactions of viruses with lipoproteins, their components, and lipoprotein receptors; 7) the role of APOE in repair; 8) pathological processes in AD and their relationship to prior damage; and 9) implications for the prevention or treatment of Alzheimer's disease.

Apolipoprotein E and herpes virus diseases: herpes simplex keratitis.

Our previous studies showed that herpes simplex type 1 virus (HSV1) is present in a high proportion of the brain of elderly normal people and Alzheimer's disease (AD) patients. We subsequently discovered that the combination of HSV1 in brain and carriage of the type 4 allele of the gene for apolipoprotein E (apoE-epsilon 4) is a strong risk factor for AD, and also that apoE-epsilon 4 is a strong risk factor for herpes labialis. In this study we have examined apoE genotypes of sufferers from another disorder caused by HSV1, namely, herpes simplex keratitis (HSK), to find if an apoE allele is involved in the disorder. In 46 HSK patients the apoE-epsilon 4 allele frequency was 15%-the same as that found in 238 unaffected controls. The apoE-epsilon 2 allele frequency was 13%-higher than the value of 7% for unaffected people, but the difference is not statistically significant.

Mechanisms of uptake of gallium by human neuroblastoma cells and effects of gallium and aluminum on cell growth, lysosomal protease, and choline acetyl transferase activity.

We have studied the uptake and removal of gallium, used as an analogue of aluminum, and the effects of aluminum itself on cultured human neuroblastoma cells treated with soluble metal complexes. The prohibitively high cost of measurement of the only available radioisotope of aluminum (26Al) precluded its usage, and so we considered that gallium, which is chemically extremely similar, would be the most suitable model. Gallium has been used thus in a number of previous biological studies and has been found to behave like aluminum in many respects. We have previously shown that Al-EDTA treatment results in uptake of aluminum and expression of hyperphosphorylated tau, a key component of Alzheimer's disease paired helical filaments. Here we demonstrate that gallium uptake can occur by two separate methods, both leading to physiologically relevant intracellular metal concentrations. Uptake from medium containing bovine transferrin occurred mainly by pinocytosis, but in the presence of human transferrin (hTf), uptake by transferrin-mediated endocytosis occurred also, despite a very low level of hTf saturation, indicating that Tf-mediated uptake is a very effective method of Ga internalization. The intracellular gallium is relatively stable, though partially removable by (1 mM) EDTA, desferrioxamine, or 1,2-dimethyl-3-hydroxypyrid-4-one. Aluminum and gallium treatment were found to increase the overall activity of lysosomal proteases, enzymes implicated in amyloid precursor protein cleavage. No effects were detected on choline acetyl transferase activity, cell growth, or tritiated thymidine incorporation or on the structure of the cells, as judged by light or electron microscopy.

Location of aluminium and gallium in human neuroblastoma cells treated with metal-chelating agent complexes.

The subcellular location of aluminium is unknown, probably because of difficulties in investigating aluminium biochemistry and the use of varied experimental approaches of uncertain sensitivity. We have studied levels of uptake and the localization of gallium and of aluminium in cultured human neuroblastoma cells treated with soluble metal complexes (mainly Al- or Ga-EDTA), radiolabeled with 26Al or 67Ga, respectively. Crude nuclei and cytoplasm were obtained by two separate methods, and DNA, RNA, and proteins were prepared from the nuclei by centrifugation in high salt; also, cytosol and noncytosol were separated using a nondissociating method. Levels of uptake were of similar order for the two metals-on average about 50 pmol/10(6) cells for aluminium and 120 pmol/10(6) cells for gallium, after 4 to 8 days treatment at 250 microM, and approximately 50 to 70% of the metal was found in the cytosol. About 20% of the aluminium and 10 to 25% of the gallium was associated with nuclear protein. A lower proportion was bound to DNA and to nuclear RNA. In cells treated with gallium-citrate/transferrin mixtures, 30 to 35% of the gallium in the cytosol was bound to protein, at least 35 being loosely bound; the main gallium-associated protein was probably intracellular transferrin. The remaining 65 to 70% of the metal in the cytosol was in low-molecular-weight form, and we suggest that the latter metal could affect structures such as the cytoskeleton and also metabolic processes in the cytoplasm. The similarity in distribution of the two metals supports the use of gallium as a "surrogate" for aluminium, at least in cell culture studies.

Neurotropic viruses and Alzheimer's disease: a search for varicella zoster virus DNA by the polymerase chain reaction.

BACKGROUND: In studies on the possible role of viruses in the aetiopathogenesis of Alzheimer's disease, herpes simplex virus type 1 (HSV1) DNA was detected by the polymerase chain reaction (PCR) in a high proportion of normal elderly people and of patients with Alzheimer's disease. The combination of HSV1 and a host factor, the type 4 allele of the gene for apolipoprotein E, is a strong risk factor for the disease. METHODS: Brain specimens were examined for another herpes virus, varicella zoster (VZV), which, like HSV1, is neurotropic, has a predilection for residing latently in the peripheral nervous system, and can reactivate. RESULTS: Using primers for sequences in the VZV origin of replication gene or thymidine kinase gene, VZV DNA was not found in any of 24 samples (18 HSV1 positive), from 17 patients with Alzheimer's disease, nor in 20 samples (12 HSV1 positive from 12 aged normal people. Hybridisation of the PCR products with a radiolabelled oligonucleotide probe capable of detecting less than 10 copies of the target sequence, confirmed the absence of VZV DNA. CONCLUSION: The presence of one neurotropic virus--HSV1--and the absence of another--VZV--in aged human brains is consistent with a role for HSV1 in the aetiology of Alzheimer's disease.

Herpes simplex virus type 1 in brain and risk of Alzheimer's disease.

BACKGROUND: The apolipoprotein E epsilon 4 (APOE-epsilon 4) allele is a risk factor for Alzheimer's disease (AD), but it is neither essential nor sufficient for development of the disease. Other factors-genetic or environmental-must therefore have a role. By means of a PCR we have detected herpes simplex virus type 1 (HSV1) in latent form in brains of elderly people with and without AD. We have postulated that limited reactivation of the virus causes more damage in AD patients than in elderly people without AD because of a difference in the hosts. We now report the APOE genotypes of AD patients and non-AD sufferers with and without HSV1 in brain. METHODS: DNA was extracted from 84 samples of brain from 46 AD patients (39 temporal lobe, 39 frontal lobe, three hippocampus) and from 75 samples of brain from 44 non-AD elderly people (33 temporal lobe, 36 frontal lobe, six hippocampus). PCR amplification was used to detect HSV1 thymidine kinase gene and the host APOE gene. FINDINGS: By multiple logistic regression, the APOE-epsilon 4 allele frequency was significantly higher in the patients positive for HSV1 in brain than in the HSV1-negative AD group, the HSV1-positive non-AD group, or the HSV1-negative non-AD group (52.8% vs 10.0%, 3.6%, and 6.3%, respectively). The odds ratio for APOE-epsilon 4 in the HSV1-positive AD group compared with HSV1-negative non-AD group was 16.8 (95% CI 3.61-77.8) and in the HSV1-negative AD group, 1.67 (0.21-13.4). We also compared APOE genotypes of 40 people who had recurrent cold sores and 33 non-sufferers; the APOE-epsilon 4 allele frequencies were 36% and 9%, respectively (p < 0.0001). INTERPRETATION: These findings suggest that the combination of HSV1 in brain and carriage of an APOE-epsilon 4 allele is a strong risk factor for AD, whereas either of these features alone does not increase the risk of AD. The findings in people with cold sores support our hypothesis that APOE-epsilon 4 and HSV1 together are damaging in the nervous system.