Inflammation is an important pathogenic mechanism in many neurodegenerative disorders. Activated microglia play a pivotal role in releasing proinflammatory factors including interleukin-1 (IL-1), tumor necrosis factor-α (TNF-α), and cyclooxygenase-2 (COX-2) for inducing inflammation. While microglia mediated inflammation is essential in maintaining CNS homeostasis, chronic inflammation results in activation of proteases for cell death. Here, we examined the effect of PPT (estrogen receptor α agonist), DPN (estrogen receptor β agonist), and estrogen on rat primary microglia following exposure to lipopolysaccharide (LPS). Exposure of microglia to LPS (200 ng/ml) for 24 h induced cell death. After LPS toxicity for 15 min, microglia were treated with 25 nM PPT, 25 nM DPN, or 100 nM estrogen that prevented cell death by attenuating the release of IL-1α, IL-1β, TNF-α, and COX-2. Treatment of cells with 100 nM fulvestrant (estrogen receptor antagonist) prior to addition of PPT, DPN, or estrogen significantly decreased their ability to prevent cell death, indicating involvement of estrogen receptor (ER) in providing PPT, DPN, or estrogen mediated cytoprotection. Reverse transcriptase polymerase chain reaction (RT–PCR) analyses showed alterations in mRNA expression of Bax, Bcl-2, calpain, and calpastatin during apoptosis. We also examined mRNA expression of ERβ and ERα following exposure of microglia to LPS and subsequent treatment with PPT, DPN, or estrogen. We found that estrogen or estrogen receptor agonists upregulated expression of ERs. Overall, results indicate that estrogen receptor agonist or estrogen uses a receptor mediated pathway to protect microglia from LPS toxicity.

Experimental autoimmune encephalomyelitis (EAE) is an animal model for studying multiple sclerosis (MS). Calpain has been implicated in many inflammatory and neurodegenerative events that lead to disability in EAE and MS. Thus, treating EAE animals with calpain inhibitors may block these events and ameliorate disability. To test this hypothesis, acute EAE Lewis rats were treated dose-dependently with the calpain inhibitor calpeptin (50 – 250 µg/kg). Calpain activity, gliosis, loss of myelin, and axonal damage were attenuated by calpeptin therapy, leading to improved clinical scores. Neuronal and oligodendrocyte death were also decreased with down regulation of pro-apoptotic proteins, suggesting that decreases in cell death were due to decreases in the expression or activity of pro-apoptotic proteins. These results indicate that calpain inhibition may offer a novel therapeutic avenue for treating EAE and MS.

Experimental autoimmune encephalomyelitis (EAE), a widely recognized animal model of multiple sclerosis (MS), is highly useful for studying inflammation, demyelination, and neurodegeneration in the central nervous system (CNS). EAE exhibits many similarities with MS, which is a chronic inflammatory disease affecting CNS white matter in humans. Various studies have indicated that EAE is a particularly useful animal model for understanding both the mechanisms of immune-mediated CNS pathology and also the progressive clinical course of MS. Demyelination and axonal dysfunction have previously been shown in MS and EAE but current evidences indicate that axonal damage and neuron death also occur, demonstrating that these diseases harbor a neurodegenerative component. Recent studies also have shown that the activation of calpain and caspase pathways contribute to the apoptotic death of oligodendrocytes and neurons, promoting the pathological events leading to neurological deficits. Apoptosis is involved in the disease-regulating as well as in the disease-promoting processes in EAE. This review discusses the major involvement of calpain and caspase pathways in causing demyelination and neurodegeneration in EAE animals.

We examined neurodegeneration in spinal cord (SC) and role of such extra‐nigral degeneration in MPTP‐induced experimental parkinsonism in C57BL/6N mice. HPLC‐photodiode array analysis confirmed presence of the active neurotoxin MPP+ in SC after single injection of MPTP (25 mg/kg, i.p.). Mitochondrial enzyme monoamine oxidase‐B (MAO‐B) responsible for in vivo conversion of MPTP to MPP+ was inhibited in SC by pre‐treatment with l‐deprenyl, a specific inhibitor of MAO‐B. Besides in vitro conversion of MPTP to MPP+ occurred by SC mitochondrial preparation, which was inhibited by l‐deprenyl implicating SC as a specific target of MPTP‐neurotoxicity. Double immunofluorescent labeling and spectrofluorimetric assay via kynuramine oxidation showed MAO‐B expression and activity in SC neurons. Localization of dopamine transporter immunoreactivity in SC along with specific uptake of 3H‐MPP+ by SC synaptosomal preparation further confirmed SC as target of MPTP‐neurotoxicity. Compared with control, increased neuronal death on the seventh day in SC of mice injected with MPTP (2 × 25 mg/kg, at 6 h interval) strongly suggested SC degeneration in pre‐symptomatic phase of MPTP‐induced experimental parkinsonism. Such extra‐nigral neurodegeneration in Parkinson’s disease indicated novel molecular mechanism preceding nigrostriatal degeneration and suggested designing broad therapeutic intervention for this complex movement disorder.

Massive T cell infiltration into the central nervous system is a hallmark of multiple sclerosis (MS) and its rodent model experimental autoimmune encephalomyelitis (EAE), resulting in the induction of many of the pathophysiological events that lead to neuroinflammation and neurodegeneration. Thus, blocking T cell migration into the central nervous system may reduce disease severity in MS and EAE. One potential target for reducing T cell migration is inhibition of the Ca 2+ -activated neutral protease calpain. Previous studies in other cell types have demonstrated that migration is reduced by incubation of cells with calpain inhibitors. Thus, we hypothesize that calpain inhibition will reduce migration of T cells in response to and toward the chemokine CCL2. To test this hypothesis, the intracellular free Ca 2+ levels in Jurkat E6-1 T cells was first measured by the fura-2 assay to assess whether the intracellular ion environment would support calpain activation. The intracellular free Ca 2+ levels were found to increase in response to CCL2. The cells were next treated with the calpain inhibitor calpeptin in a multiwelled Boyden chamber with CCL2 used as the chemoattractant. These studies demonstrate that inhibition of calpain with its inhibitor calpeptin produces a dose-dependent inhibition of chemotaxis. Calpain activity, as measured by live cell imaging, was also increased in response to CCL2, providing further evidence of its involvement in the process of chemotaxis and migration. These studies provide evidence for the involvement of calpain in the mechanisms of chemotaxis and warrants further exploration in MS patient and EAE animal samples. Keywords calpain; chemotaxis; EAE/MS; T cellsT cell involvement in the disease process of multiple sclerosis (MS) (Hickey, 1991;McFarland and Martin, 2007) and its animal model experimental autoimmune encephalomyelitis (EAE) are well established (Waksman and Adams, 1962;Swanborg, 2001). T cell infiltration into the central nervous system (CNS) is greatly increased in the plaques of MS patients (Hickey, 1991) as well as of EAE animals immediately before the onset of pathological symptoms and during clinical course of the disease (Waksman and Adams, 1962). Increased activity and expression of the Ca 2+ -activated protease calpain has been demonstrated in inflammatory cells (Shields et al., 1998(Shields et al., , 1999 during the disease state in EAE animals. Moreover, inhibition of *Correspondence to: Naren L. Banik, PhD, Department of Neurosciences, Medical University of South Carolina, 96 Jonathan Lucas Street, Suite 309 CSB, PO Box 250606, Charleston, SC 29425. E-mail: E-mail: baniknl@musc.edu. calpain has been demonstrated to decrease the T cell infiltration during acute EAE (Hassen et al., 2006). Calpain has signaling functions and is involved in a multitude of cellular processes including apoptosis , T cell activation (Penna et al., 1999;Schaecher et al., 2004), cell cycle (Janossy et al., 2004), and cell migration (Huttenlocher et al., 1997;Franco and Huttenl...