There is a compelling body of evidence that autoimmune reactions in MS are initiated by a virus. A potential mechanism entails sporadic replication of a virus in glial cells. Although the virus does not cause extensive cell loss, the shedding of viral components elicits proinflammatory responses in neighboring cells. Double stranded RNA (dsRNA), a viral replication intermediate, is a potent proinflammatory activator. The binding of dsRNA to its receptor, Toll-like receptor 3 (TLR3) triggers the secretion of proinflammatory cytokines and other mediators. The proinflammatory milieu, ultimately activates the adaptive immune system that results in autoimmune attack. Although TLR3 is expressed by astrocytes, the chief mediators of innate immunity in the CNS, its function in astrocytes has not been characterized. The central hypothesis to be tested in the proposed study is that dsRNA elicits proinflammatory response in astrocytes through TLR3, and that this response can be attenuated by statins. Three interrelated specific aims are proposed to test this hypothesis.
In Aim 1 the ability of dsRNA to trigger proinflammatory response will be assessed. The production of cytokines and nitric oxide will be determined by RNase protection assay and by sandwich ELISA in cultured astrocytes challenged with dsRNA.
In Aim 2 the involvement of TLR3 in dsRNA-induced proinflammatory response will be determined. This will be achieved by determining the response of astrocytes prepared from TLR3 knockout mice, and by using specific inhibitors.
In Aim 3 the ability of statins to suppress dsRNA-induced proinflammatory response in astrocytes will be assessed. The suppressive potency of six different statins will be determined. Moreover, signaling pathways through which statins suppress dsRNA-induced proinflammatory response will be identified. The proposed study is expected to demonstrate that viral replication intermediate, dsRNA, can activate innate immune system in the CNS, and that this activation can be attenuated by statins through their interference with specific signaling pathways. These pilot experiments will provide a steppingstone for future studies to fully elucidate the mechanisms of proinflammatory responses of glial cells to viral components, and to develop rational and efficient therapeutic interventions to prevent inflammatory CNS tissue damage through pharmacological manipulation of sensitive signaling pathways in MS, and in other virus-triggered neurodegenerative conditions. Moreover, because TLR3 can also be activated by RNA released from damaged cells, the outcomes of this study will also be applicable to neurodegenerative conditions of non-viral etiology. ? ?
|Konat, Gregory W; Borysiewicz, Elizabeth; Fil, Daniel et al. (2009) Peripheral challenge with double-stranded RNA elicits global up-regulation of cytokine gene expression in the brain. J Neurosci Res 87:1381-8|
|Borysiewicz, Elizabeth; Fil, Daniel; Dlaboga, Daniel et al. (2009) Phosphodiesterase 4B2 gene is an effector of Toll-like receptor signaling in astrocytes. Metab Brain Dis 24:481-91|
|Borysiewicz, Elizabeth; Fil, Daniel; Konat, Gregory W (2009) Rho proteins are negative regulators of TLR2, TLR3, and TLR4 signaling in astrocytes. J Neurosci Res 87:1565-72|
|Krasowska-Zoladek, Alicja; Banaszewska, Monika; Kraszpulski, Michal et al. (2007) Kinetics of inflammatory response of astrocytes induced by TLR 3 and TLR4 ligation. J Neurosci Res 85:205-12|
|Konat, Gregory W; Kielian, Tammy; Marriott, Ian (2006) The role of Toll-like receptors in CNS response to microbial challenge. J Neurochem 99:1-12|