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J. Neurochem. (2011) 118, 205–214. Abstract Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis. Dark Agouti rats immunized with spinal cord homogenate (SCH) and carbonyl iron (CI), as an adjuvant, develop severe hyperacute form of EAE. They succumb to EAE earlier and have higher clinical scores and lethality rate in comparison to counterparts immunized with SCH + complete Freund’s adjuvant. There is no difference in the number of cells or in histological presentation of the CNS infiltrates of rats immunized with the two adjuvants. However, there are more granulocytes, NK and NKT cells, and less CD4+ T cells in the spinal cord infiltrates of SCH + CI‐immunized animals. Nitric oxide (NO)‐generating enzyme inducible NO synthase have higher expression in spinal cord of SCH + CI‐immunized rats, and this corresponds to more intensive nitrotyrosine formation in the CNS tissue of these rats. Abundant infiltration of granulocytes and NK cells into the CNS and excessive generation of peroxynitrite within the CNS of SCH + CI‐immunized rats might account for the severe neurological deficits induced by immunization with CI. These factors should be closely examined in the fulminant forms of multiple sclerosis and acute disseminated encephalomyelitis, as they could represent a promising targets for therapy.

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We examined the status and role of autophagy, a process of lysosomal recycling of cellular material, in clear cell renal cell carcinoma (ccRCC). Paired samples of tumor and adjacent non-malignant tissue were collected from 20 patients with ccRCC after radical nephrectomy. The mRNA levels of apoptosis (BAD, BAX, BCL2, BCLXL, BIM) and autophagy (ATG4, BECN1, GABARAP, p62, UVRAG) regulators were measured by RT-qPCR. The protein levels of autophagosome-associated LC3-II, autophagy receptor p62, apoptotic marker PARP, as well as phosphorylation of autophagy initiator Unc 51-like kinase 1 (ULK1), its activator AMP-activated protein kinase (AMPK) and 4EBP1, the substrate of ULK1 inhibitor mechanistic target of rapamycin (mTOR), were analyzed by immunoblotting. The mRNA levels of pro-apoptotic BAX, anti-apoptotic BCLXL and pro-autophagic ATG4, p62 and UVRAG were higher in ccRCC tumors. Autophagy induction was confirmed by an increase in phospho-ULK1 and degradation of the autophagic target p62, while apoptotic PARP cleavage was unaltered. AMPK phosphorylation was reduced and 4EBP1 phosphorylation was increased in ccRCC tissue. The expression of apoptosis regulators did not correlate with clinicopathological features of ccRCC. Conversely, high mRNA levels of ATG4, GABARAP and p62 were associated with lower tumor stage, as well as with smaller tumor size and better disease-specific 5-year survival (ATG4 and p62). Accordingly, low p62 protein levels, corresponding to increased autophagic flux, were associated with lower tumor stage, reduced metastasis and improved 5-year survival. These data demonstrate that transcriptional induction of autophagy in ccRCC is accompanied by AMPK/mTOR-independent increase in ULK1 activation and autophagic flux, which might slow tumor progression and metastasis independently of apoptosis.

Arylpiperazine-based dopaminergic/serotonergic ligands exert neuroprotective activity. We examined the effect of arylpiperazine D 2 /5-HT 1A ligands, N-{4-[2-(4-phenyl-piperazin-1-yl)-ethyl}-phenyl]-picolinamide (6a) and N-{3-[2-(4-phenyl-piperazin-1-yl)-ethyl]-phenyl}-picolinamide (6b), in experimental autoimmune encephalomyelitis (EAE), a model of neuroinflammation. Both compounds (10 mg/kg i.p.) reduced EAE clinical signs in spinal cord homogenate-immunized Dark Agouti rats. Compound 6b was more efficient in delaying the disease onset and reducing the maximal clinical score, which correlated with its higher affinity for D 2 and 5-HT 1A receptors. The protection was retained if treatment was limited to the effector (from day 8 onwards), but not the induction phase (day 0-7) of EAE. Compound 6b reduced CNS immune infiltration and expression of mRNA encoding the proinflammatory cytokines tumor necrosis factor, IL-6, IL-1, and GM-CSF, T H 1 cytokine IFN-c, T H 17 cytokine IL-17, as well as the signature transcription factors of T H 1 (T-bet) and T H 17 (RORct) cells. Arylpiperazine treatment reduced apoptosis and increased the activation of anti-apoptotic mediators Akt and p70S6 kinase in the CNS of EAE animals. The in vitro treatment with 6b protected oligodendrocyte cell line OLN-93 and neuronal cell line PC12 from mitogen-activated normal T cells or myelin basic proteinactivated encephalitogenic T cells. In conclusion, arylpiperazine dopaminergic/serotonergic ligands suppress EAE through a direct neuroprotective action and decrease in CNS inflammation.