Eva Neuen-Jacob

Alterations in Expression of Glutamatergic Transporters and Receptors in Sporadic Alzheimer's Disease

Excitatory neurotransmitter dysfunction has been discussed to be involved in the pathophysiology of Alzheimers disease (AD). In the current study we investigated gene and protein expression patterns of glutamatergic receptors and transporters in brains of AD patients in various stages of disease using gene chip arrays, real time PCR and immunohistochemistry. We found marked impairment in the expression of excitatory amino acid transporters (EAAT1 and EAAT 2) at both gene and protein levels in hippocampus and gyrus frontalis medialis of AD patients, already in early clinical stages of disease. The loss of EAAT immunoreactivity was particularly obvious in the vicinity of amyloid plaques. In contrast, EAAT expression was up-regulated in the cerebellum of these patients. Furthermore, a significant up-regulation of the glutamatergic kainate (GRIK4) receptor observed by gene arrays was confirmed by quantitative RT-PCR in late stages in the hippocampus of AD patients. Moreover, there were down-regulations of other glutamatergic receptors such as NMDA (GRINL1A) and AMPA (GRIA4) receptors. Our data show marked changes in the functional elements of the glutamatergic synapses such as glutamatergic receptors and transporters and indicate impaired glutamate clearing rendering neurons susceptible to excess extracellular glutamate and support further the involvement of excitotoxic mechanisms in the pathogenesis of AD.

Memantine upregulates BDNF and prevents dopamine deficits in SIV-infected macaques: a novel pharmacological action of memantine.

N-methyl-D-aspartate (NMDA) receptor activation is involved in the pathogenetic cascades of neurodegenerative disorders including human immunodeficiency virus (HIV) dementia. Memantine, an uncompetitive NMDA receptor antagonist, which has been recently approved for the treatment of Alzheimers disease, is being discussed as a potential adjunctive therapeutic substance for HIV dementia. We used simian immunodeficiency virus-infected rhesus macaques to assess the effects of memantine on brain dysfunction and brain pathology within 3–5 months after initial infection during early asymptomatic stage of disease. We had shown previously that within this time frame, marked changes were evident in the dopaminergic systems. Memantine was administered two weeks post infection, at peak viremia, in order to prevent early NMDA receptor activation due to immune mediators. We found that memantine prevented onset of dopamine deficits in the brains of SIV-infected macaques, without affecting early brain pathology or peripheral course of infection. Memantine specifically upregulated mRNA and protein expression of the neurotrophic factor brain-derived neurotrophic factor (BDNF), suggesting that the protective effect of memantine on dopamine function may be mechanistically remote from NMDA receptor antagonism. This novel pharmacological action of memantine may also be relevant for other neurodegenerative disorders and supports the involvement of neurotrophic factors in adult brain neuroprotection.

ADAM-10 and ADAM-17 in the inflamed human CNS.

Inflammatory demyelinating disorders of the CNS, such as multiple sclerosis (MS), are mediated, at least in part, by various cytokines and proteases. In the present study, we investigated the expression of A disintegrin and metalloproteinase (ADAM)‐17, an important sheddase for various proteins, including tumor necrosis factor‐α (TNF‐α), and the p75‐ and p55‐TNF receptors, as well as ADAM‐10, a protease implicated in myelin degradation, in post mortem CNS tissue samples from patients with MS, and normal brain tissue (as control) by immunohistochemistry. ADAM‐10 was found to be expressed by astrocytes in all MS and control sections studied; however, in some MS sections, perivascular macrophages were determined as an additional cellular source as well. ADAM‐17 could be observed exclusively in acute and chronic active MS plaques and localized to invading T lymphocytes. The staining pattern of ADAM‐17 in MS plaques was mirrored in distribution and extent by the pattern obtained with an antibody against the p75–TNF‐receptor (TNFR‐2), whereas TNF‐α was found to be expressed primarily by perivascular macrophages. In studying cerebrospinal fluid (CSF) samples from MS patients, we were able to detect increased protein levels of ADAM‐17 as compared with noninflammatory controls. In addition, increased levels of soluble TNFR‐2 could be measured, suggestive of an active shedding process mediated by ADAM‐17. The stimulation of peripheral blood mononuclear cells (PBMC) obtained from MS patients and healthy individuals corroborated these findings by revealing expression of ADAM‐17 by T lymphocytes and ADAM‐10 by macrophages in vitro. Our results indicate that ADAM‐10 is expressed constitutively by astrocytes in the normal and inflamed human CNS. In contrast, under inflammatory conditions, ADAM‐10, expressed by perivascular macrophages, and ADAM‐17, expressed by invading T cells, may actively contribute to the pathogenesis of inflammatory disorders of the CNS. GLIA 42:398–405, 2003.

Early impairment in dopaminergic neurotransmission in brains of SIV‐infected rhesus monkeys due to microglia activation

Movement disorders are a common neurological complication of immunodeficiency virus infection and are thought to result from dopaminergic dysfunction in the basal ganglia. We measured levels of dopamine, and its metabolites homovanillic acid and 3,4‐dihydroxyphenylacetic acid, in the putamen of healthy and simian immunodeficiency virus (SIV)‐infected rhesus monkeys from infection until the development of AIDS. Changes in expression levels of cAMP response element binding protein (CREB), a transcription factor involved in the signalling pathway of dopamine, were also examined. Furthermore, we isolated microglia from the same animals and investigated their activation status in order to explore whether neurochemical findings are associated with immune activation. Plasma and CSF viral RNA load, T‐cell analysis and basal ganglia histopathology provided information about disease progression in the animals. Putamen dopamine content was significantly reduced within 3 months of SIV infection, due to decreased dopamine synthesis initially, followed by loss of tyrosine hydroxylase‐positive cells in substantia nigra, and accompanied by a decrease in total CREB expression. Pharmacological manipulation of dopaminergic tone with l‐DOPA and selegiline showed that the reduction in CREB expression was due to reduced levels of dopamine. These neurochemical changes were significantly correlated with microglia activation in the absence of gross histopathological lesions. Our data demonstrate that putamen dopaminergic function is impaired during SIV infection and indicate that microglia may trigger endogenous mechanisms involved in the dysfunction of dopaminergic systems.

Local inflammation and devascularization — in vivo mechanisms of the “bystander effect” in VPC-mediated HSV-Tk/GCV gene therapy for human malignant glioma

Somatic gene therapy with the herpes simplex virus type I thymidine kinase gene/ganciclovir (HSV-Tk/GCV) system and murine retroviral vector producer cells (VPCs) was introduced as a new adjuvant treatment modality to treat tumor bulk and to prevent tumor recurrence in patients harboring malignant glioma. The single-center experience after treatment of 27 patients undergoing tumor resection followed by intracerebral VPC injection for HSV-Tk suicide gene therapy will be presented focused on findings of systematic and close MRI follow-up and a few histological specimens. The data indicate that hemorrhagic necrosis due to endothelial cell transfection mediated vessel necrosis and that local inflammatory immune response occurs frequently after gene therapy. These phenomena seem to be specific because none of the patients of a control group showed any similar features. The prognosis (time to progression, survival) of the patients with “bystander effects” after gene therapy was better, but compared to those patients without bystander effects, they were also privileged by a favorable constellation of prognostic factors. Therefore, the appearance of these neuroradiologic features cannot serve as an indicator for treatment effectiveness and outcome. Cancer Gene Therapy (2001) 8, 843–851

Osteopontin: A novel axon-regulated Schwann cell gene

Osteopontin (OPN) is a RGD‐containing glycoprotein with cytokine‐like, chemotactic, and pro‐adhesive properties. During wound healing, OPN is abundantly expressed by infiltrating macrophages and has been implicated in posttraumatic tissue repair. To delineate a role in the regenerative response to axotomy we examined the expression of OPN in Wallerian degeneration of the sciatic nerve in rats. Unexpectedly, we found high constitutive expression of OPN by myelinating Schwann cells (SCs) in uninjured control nerves. OPN mRNA expression was confirmed in primary cultures of rat SCs. Upon axotomy, SC‐expressed OPN in the degenerating distal nerve stump transiently increased during the first days after injury, but was continuously downregulated thereafter, reaching its minimum at Day 14. Macrophages invading axotomized nerves were OPN‐negative. During late stages after axotomy, SC‐OPN was reexpressed in regenerating but not permanently transected nerves. We also found OPN expression by myelinating SCs in human sural nerves with a dramatic reduction in severe axonal polyneuropathies. Taken together, our study identifies OPN as a novel Schwann cell gene regulated by axon‐derived signals. The lack of OPN induction in infiltrating macrophages indicates fundamental differences in tissue repair between axonal injury in the peripheral nervous system and structural lesions in other organ systems.

Successful treatment of hepatitis B virus associated polyarteritis nodosa with a combination of prednisolone, α-interferon and lamivudine

Therapy of hepatitis B virus (HBV)-associated poly-arteritis nodosa is still evolving. Here we report a successful treatment with a short-term steroid administration in combination with a-interferon and lamivudine and a complete sequence analysis of the HBV genome. A 58-year-old man presented with the symptoms of mononeuritis multiplex associated in time with the onset of highly replicative hepatitis B. Polyarteritis nodosa was confirmed by biopsy. During an initial course with alpha-interferon and prednisolone no clinical improvement or hepatitis B virus seroconversion was observed. After addition of lamivudine to the protocol with fast tapering of prednisolone, HBV DNA fell to undetectable levels within 1 month and liver transaminases normalized. After 6 months of treatment HBeAg seroconversion took place, followed by HBsAg seroconversion 2 months later. Clinical symptoms of polyarteritis improved. No relapse of polyarteritis or hepatitis B was seen during the follow up of 9 months. Complete sequence analysis of the HBV genome revealed 6 nucleotide mutations but none in a relevant antigenic epitope. The present protocol of short-term prednisolone administration combined with alpha-interferon and lamivudine was effective for the treatment of HBV-related polyarteritis nodosa and may be a promising new therapeutic approach.

Increased dopaminergic neurotransmission in therapy-naïve asymptomatic HIV patients is not associated with adaptive changes at the dopaminergic synapses.

Central dopaminergic (DA) systems are affected during human immunodeficiency virus (HIV) infection. So far, it is believed that they degenerate with progression of HIV disease because deterioration of DA systems is evident in advanced stages of infection. In this manuscript we found that (a) DA levels are increased and DA turnover is decreased in CSF of therapy-naïve HIV patients in asymptomatic infection, (b) DA increase does not modulate the availability of DA transporters and D2-receptors, (c) DA correlates inversely with CD4+ numbers in blood. These findings show activation of central DA systems without development of adaptive responses at DA synapses in asymptomatic HIV infection. It is probable that DA deterioration in advanced stages of HIV infection may derive from increased DA availability in early infection, resulting in DA neurotoxicity. Our findings provide a clue to the synergism between DA medication or drugs of abuse and HIV infection to exacerbate and accelerate HIV neuropsychiatric disease, a central issue in the neurobiology of HIV.

Changes in canine skeletal muscles during experimental tibial lengthening.

In 24 beagles, lengthening of the right tibia was performed by callus distraction after osteotomy and application of a ring fixator. Distraction was started at the fifth postoperative day, with a distraction rate of 0.5 mm twice per day, and ended after 25 days. A control group of six additional dogs underwent tibial osteotomy and external fixation without distraction. Twelve animals with and three animals without leg lengthening were euthanized immediately after the distraction period of 25 days (Group A); the remaining 15 dogs were euthanized after an additional consolidation phase of another 25 days (Group B). From the distracted right leg and from the left control leg the tibialis anterior muscle, extensor digitorum longus muscle, peroneus longus muscle, and gastrocnemius muscle were removed and studied by means of routine histologic, histochemical, and immunohistochemical analyses, and electron microscopic examination. The muscles of the control group showed no differences between the right and left sides. However, in the other 24 dogs of Groups A and B, the authors saw marked alterations affecting only the lengthened muscles but not the muscles of the control limbs. These changes were highly significant and included muscle fiber degeneration and regeneration, target fibers, central cores, minicores, marked endomysial and perimysial fibrosis, and atrophy of Type 1 and Type 2 fibers. In the consolidation period (Group B) fiber type grouping indicated that reinnervation had occurred. In addition, an increase in satellite cells and myoblasts and proliferation of nuclei were observed. The findings of the current study indicate that leg lengthening results not only in muscle fiber degeneration followed by regeneration and reinnervation but also in formation of new muscle tissue.

Diagnostic criteria and clinical procedures in HIV-1 associated progressive multifocal leukoencephalopathy

The diagnosis of definite progressive multifocal leukoencephalopathy (PML) has been a neuropathological domain. We reviewed all Human Immunodeficiency Virus Type 1 (HIV-1) seropositive patients in our institution between 01.01.1989 and 31.12.1994 and identified 20/823 cases with PML by clinical and imaging criteria. Diagnosis was neuropathologically confirmed in 5 cases. Diagnostic criteria included rapid onset (< 2 weeks) of multifocal neurological signs and symptoms, advanced immunosuppression and asymmetric uni- or multifocal white matter lesions without mass effect, contrast enhancement or cortical atrophy in magnetic resonance imaging (MRI). The overall incidence of PML was stable over the observation period (approximately equal to 2.5%). The mean age at onset (41.7 years) was significantly lower compared to HIV-1 seronegative PML patients (peak in the sixth decade of life), male patients prevailed (100%). Mean survival (3.9 months) was extremely short. Human polyoma virus JC (JCV) polymerase chain reaction (PCR) in the cerebrospinal fluid (CSF) demonstrated a considerable rate of possible cerebral co-infection with HIV-1 and JCV as well as subclinical infection with JCV. Therefore demonstration of JCV deoxyribonucleic acid by PCR in the CSF alone is not sufficient for clinical PML diagnosis. We present diagnostic criteria on the basis of epidemiological, neuroradiological and CSF parameters that allow us to make the clinical diagnosis of PML. Although quick and safe, routine stereotactic brain biopsy is not necessary to confirm the diagnosis.