Reduced Treg frequency in LFA-1-deficient mice allows enhanced T effector differentiation and pathology in EAE.

The αLß2-integrin LFA-1 (CD11a/CD18) is known as an important molecule for leukocyte migration. However, the precise role of LFA-1 in the pathogenesis of EAE has so far remained unclear. We describe here the disease development in LFA-1(-/-) mice compared with WT controls. Ablation of LFA-1 resulted in more severe EAE with increased demyelination and increased numbers of myelin oligodendrocyte glycoprotein-reactive CD4(+) T cells in the CNS. However, the production of the pro-inflammatory cytokines IL-17 and IFN-γ was unchanged on the level of antigen-specific T cells. Interestingly, LFA-1-deficient mice showed a clearly reduced frequency of Treg in the inflamed CNS. Moreover, Treg counts in spleens and thymi of unimmunized LFA-1(-/-) mice were lower in comparison to the WT controls, indicating an impairment of Treg generation. In combination, these results suggest a substantial role of LFA-1 in Treg generation and subsequent expansion of effector T cells and highlight the importance of Treg in limiting EAE.

Neuroinflammation in prion diseases: concepts and targets for therapeutic intervention.

Prion infections of the central nervous system (CNS) are characterized by a reactive gliosis and the subsequent degeneration of neuronal tissue. The activation of glial cells, which precedes neuronal death, is likely to be initially caused by the deposition of misfolded, in part proteinase K-resistant, isoforms (termed PrP(TSE)) of the normal cellular prion protein (PrP(c)) in the brain. Proinflammatory cytokines and chemokines released by PrP(TSE)-activated glial cells and stressed neurons may contribute directly or indirectly to the disease development by enhancement and generalization of the gliosis and via cytotoxicity for neurons. Recent studies have illustrated that interfering with inflammatory responses may represent a therapeutic approach to slow down the course of disease development. Hence, a better understanding of driving factors in neuroinflammation may well contribute to the development of improved strategies for treatment of prion diseases.

Prion disease development in slow Wallerian degeneration (Wld(S)) mice.

Axon destruction represents one aspect of prion disease-associated neurodegeneration. We characterized here the scrapie infection of Wld(S)-mice in comparison to wild-type C57Bl/6 controls to determine whether mechanisms involved in Wallerian degeneration contribute to disease development in this murine model system. The Wld(S) mutation had neither an effect on survival times, nor on typical hallmarks of a prion infection like deposition of misfolded PrP(Sc) and glia activation. At the ultrastructural level, axonal damage like loss of axoplasms and disintegration of myelin sheaths was evident. Moreover, lysosomes accumulated in neuronal cell bodies. These alterations occured however similarly in Wld(S)- and wild-type mice. In conclusion, it appears unlikely that axonal damage of the kind, which is slowed down in Wld(S)-mice, contributes significantly to disease progression. These findings distinguish the neurodegeneration occuring in this prion model from chronic neurodegenerative diseases, in which the Wld(S)-mutation provides axon protection and greatly improves the clinical outcome.

Accelerated prion replication in, but prolonged survival times of, prion-infected CXCR3-/- mice.

Prion diseases have a significant inflammatory component. Glia activation, which is associated with increased production of cytokines and chemokines, may play an important role in disease development. Among the chemokines upregulated highly and early upregulated during scrapie infections are ligands of CXCR3. To gain more insight into the role of CXCR3 in a prion model, CXCR3-deficient (CXCR3(-/-)) mice were infected intracerebrally with scrapie strain 139A and characterized in comparison to similarly infected wild-type controls. CXCR3(-/-) mice showed significantly prolonged survival times of up to 30 days on average. Surprisingly, however, they displayed accelerated accumulation of misfolded proteinase K-resistant prion protein PrP(Sc) and 20 times higher infectious prion titers than wild-type mice at the asymptomatic stage of the disease, indicating that these PrP isoforms may not be critical determinants of survival times. As demonstrated by immunohistochemistry, Western blotting, and gene expression analysis, CXCR3-deficient animals develop an excessive astrocytosis. However, microglia activation is reduced. Quantitative analysis of gliosis-associated gene expression alterations demonstrated reduced mRNA levels for a number of proinflammatory factors in CXCR3(-/-) compared to wild-type mice, indicating a weaker inflammatory response in the knockout mice. Taken together, this murine prion model identifies CXCR3 as disease-modifying host factor and indicates that inflammatory glial responses may act in concert with PrP(Sc) in disease development. Moreover, the results indicate that targeting CXCR3 for treatment of prion infections could prolong survival times, but the results also raise the concern that impairment of microglial migration by ablation or inhibition of CXCR3 could result in increased accumulation of misfolded PrP(Sc).

Prion infection of mice transgenic for human APPSwe: increased accumulation of cortical formic acid extractable Abeta(1-42) and rapid scrapie disease development.

Neuropathological, epidemiological and experimental data indicate a potential interrelationship between Alzheimer's disease and prion diseases. Proteolytic processing of amyloid precursor protein (APP) by beta-secretase was recently suggested to be controlled by prion protein expression. Here, we characterized the prion infection of Tg2576 mice, which overexpress the human APP(Swe) protein. Prion infection of Tg2576-mice led to an early death of the animals, which was preceded by a relatively short symptomatic stage. However, disease-associated gliosis and deposition of misfolded prion protein PrP(Sc) were identical in infected Tg2576-mice and non-transgenic littermate controls. To analyze the effect of prion infection on APP processing and generation of beta-amyloid we determined cortical levels of SDS- and formic acid (FA)-extractable forms of beta-amyloid (1-40) and (1-42) by ELISA. Formic acid-extractable Abeta (1-42) levels were 10-fold higher in infected versus uninfected Tg2576 mice whereas other forms of Abeta were essentially unaffected by the prion infection. Hence, the experimental model demonstrates that a prion infection of the CNS promotes selectively formation of FA-extractable Abeta(1-42) in Tg2576 mice.

Evaluation of drugs for treatment of prion infections of the central nervous system.

Prion diseases are fatal and at present there are neither cures nor therapies available to delay disease onset or progression in humans. Inspired in part by therapeutic approaches in the fields of Alzheimer's disease and amyotrophic lateral sclerosis, we tested five different drugs, which are known to efficiently pass through the blood-brain barrier, in a murine prion model. Groups of intracerebrally prion-challenged mice were treated with the drugs curcumin, dapsone, ibuprofen, memantine and minocycline. Treatment with antibiotics dapsone and minocycline had no therapeutic benefit. Ibuprofen-treated mice showed severe adverse effects, which prevented assessment of therapeutic efficacy. Mice treated with low- but not high-dose curcumin and mice treated with memantine survived infections significantly longer than untreated controls (P<0.01). These results encourage further research efforts to improve the therapeutic effect of these drugs.

Role of galectin-3 in prion infections of the CNS.

Galectin-3 is a multi-functional protein and participates in mediating inflammatory reactions. The pronounced overexpression of galectin-3 in prion-infected brain tissue prompted us to study the role of this protein in a murine prion model. Immunofluorescence double-labelling identified microglia as the major cell type expressing galectin-3. Ablation of galectin-3 did not affect PrP(Sc)-deposition and development of gliosis. However, galectin-3(-/-)-mice showed prolonged survival times upon intracerebral and peripheral scrapie infections. Moreover, protein levels of the lysosomal activation marker LAMP-2 were markedly reduced in prion-infected galectin-3(-/-)-mice suggesting a role of galectin-3 in regulation of lysosomal functions. Lower mRNA levels of Beclin-1 and Atg5 in prion-infected wild-type and galectin-3(-/-)-mice indicated an impairment of autophagy although autophagosome formation was unchanged. The results point towards a detrimental role of galectin-3 in prion infections of the CNS and suggest that endo-/lysosomal dysfunction in combination with reduced autophagy may contribute to disease development.

Simvastatin prolongs survival times in prion infections of the central nervous system.

Prion diseases are fatal and at present there are neither cures nor palliative therapies known/available, which delay disease onset or progression. Cholesterol-lowering drugs have been reported to inhibit prion replication in infected cell cultures and to modulate inflammatory reactions. We aimed to determine whether simvastatin-treatment could delay disease onset in a murine prion model. Groups of mice were intracerebrally infected with two doses of scrapie strain 139A. Simvastatin-treatment commenced 100 days postinfection. The treatment did not affect deposition of misfolded prion protein PrP(res). However, expression of marker proteins for glia activation like major histocompatibility class II and galectin-3 was found to be affected. Analysis of brain cholesterol synthesis and metabolism revealed a mild reduction in cholesterol precursor levels, whereas levels of cholesterol and cholesterol metabolites were unchanged. Simvastatin-treatment significantly delayed disease progression and prolonged survival times in established prion infection of the CNS (p < or = 0.0003). The results suggest that modulation of glial responses and the therapeutic benefit observed in our murine prion model of simvastatin is not due to the cholesterol-lowering effect of this drug.