No abstract

Neuronal ceroid lipofuscinoses (NCLs) are a genetically heterogeneous group of neurodegenerative diseases characterized by cognitive and motor decline, epilepsy, visual loss and by lysosomal autofluorescent inclusions. Two distinct clinical phenotypes, the progressive epilepsy with mental retardation (EPMR) and a late-infantile variant of NCLs (CLN8-vLINCL) are associated with mutations in the CLN8 gene that encodes a transmembrane protein predominantly located to the endoplasmic reticulum (ER). To gain insight into the function of CLN8 protein, we employed the split-ubiquitin membrane-based yeast two-hybrid (MYTH) system, which detects protein-protein interactions in a membrane environment, using the full-length human CLN8 as bait and a human brain cDNA library as prey. We identified several potential protein partners of CLN8 and especially referred to VAPA, c14orf1/hERG28, STX8, GATE16, BNIP3 and BNIP3L proteins that are associated with biologically relevant processes such as synthesis and transport of lipids, vesicular/membrane trafficking, autophagy/mitophagy and apoptosis. Interactions of CLN8 with VAPA and GATE16 were further validated by co-immunoprecipitation and co-localization assays in mammalian cells. Using a new C-terminal-oriented CLN8 antibody, CLN8-VAPA interaction was also confirmed by co-staining in close spatial proximity within different CNS tissues. The results of this study shed light on potential interactome networks of CLN8 and provide a powerful starting point for understanding protein function(s) and molecular aspects of diseases associated with CLN8 deficiency.

Obesity and metabolic disorders can be risk factors for the onset and development of neurodegenerative diseases. The aim of the present study was to investigate the protective effects of a natural dietary supplement (NDS), containing Curcuma longa, silymarin, guggul, chlorogenic acid and inulin, on dysmetabolism and neurodegeneration in the brains of high fat diet (HFD)-fed mice. Decrease in the expression of FACL-4, CerS-1, CerS-4, cholesterol concentration and increase in the insulin receptor expression and insulin signaling activation, were found in brains of NDS-treated HFD brains in comparison with HFD untreated-mice, suggesting that NDS is able to prevent brain lipid accumulation and central insulin resistance. In the brains of NDS-treated HFD mice, the levels of RNS, ROS and lipid peroxidation, the expression of p-ERK, H-Oxy, i-NOS, HSP60, NF-kB, GFAP, IL-1β, IL-6 and CD4 positive cell infiltration were lower than in untreated HFD mice, suggesting antioxidant and anti-inflammatory effects of NDS. The decreased expression of p-ERK and GFAP in NDS-treated HFD mice was confirmed by immunofluorescence. Lastly, a lower number of apoptotic nuclei was found in cortical sections of NDS-treated HFD mice. The present data indicate that NDS exerts neuroprotective effects in HFD mice by reducing brain fat accumulation, oxidative stress and inflammation and improving brain insulin resistance.

Cyanobacteria have been recognized as a source of bioactive molecules to be employed in nutraceuticals, pharmaceuticals, and functional foods. An extract of Aphanizomenon flos-aquae (AFA), commercialized as Klamin®, was subjected to chemical analysis to determine its compounds. The AFA extract Klamin® resulted to be nontoxic, also at high doses, when administered onto LAN5 neuronal cells. Its scavenging properties against ROS generation were evaluated by using DCFH-DA assay, and its mitochondrial protective role was determined by JC-1 and MitoSOX assays. Klamin® exerts a protective role against beta amyloid- (Aβ-) induced toxicity and against oxidative stress. Anti-inflammatory properties were demonstrated by NFβB nuclear localization and activation of IL-6 and IL-1β inflammatory cytokines through ELISA. Finally, by using thioflavin T (ThT) and fluorimetric measures, we found that Klamin® interferes with Aβ aggregation kinetics, supporting the formation of smaller and nontoxic structures compared to toxic Aβ aggregates alone. Altogether, these data indicate that the AFA extract may play a protective role against mechanisms leading to neurodegeneration.