Region-specific changes in gene expression are associated with cognitive deficits in the alpha-synuclein-induced model of Parkinson's disease: A transcriptomic profiling study.

Mild cognitive impairment (MCI) is a common trait of Parkinson's disease (PD), often associated with early motor deficits, eventually evolving to PD with dementia in later disease stages. The neuropathological substrate of MCI is poorly understood, which weakens the development and administration of proper therapies. In an α-synuclein (αSyn)-based model of PD featuring early motor and cognitive impairments, we investigated the transcriptome profile of brain regions involved in PD with cognitive deficits, via a transcriptomic analysis based on RNA sequencing (RNA-seq) technology. Rats infused in the substantia nigra with human α-synuclein oligomers (H-SynOs) developed mild cognitive deficits after three months, as measured by the two-trial recognition test in a Y-maze and the novel object recognition test. RNA-seq analysis showed that 17,436 genes were expressed in the anterior cingulate cortex (ACC) and 17,216 genes in the hippocampus (HC). In the ACC, 51 genes were differentially expressed between vehicle and H-αSynOs treated samples, which showed N= 21 upregulated and N = 30 downregulated genes. In the HC, 104 genes were differentially expressed, the majority of them not overlapping with DEGs in the ACC, with N = 41 upregulated and N = 63 downregulated in H-αSynOs-treated samples. The Gene Ontology (GO) and the Kyoto Encyclopedia of Gene and Genomes (KEGG) analysis, followed by the protein-protein interaction (PPI) network inspection of DEGs, revealed that in the ACC most enriched terms were related with immune functions, specifically with antigen processing/presentation via the major histocompatibility complex (MHC) class II and phagocytosis via CD68, supporting a role for dysregulated immune responses in early PD cognitive dysfunction. Immunofluorescence analysis confirmed the decreased expression of CD68 within microglial cells. In contrast, the most significantly enriched terms in the HC were mainly involved in mitochondrial homeostasis, potassium voltage-gated channel, cytoskeleton and fiber organisation, suggesting that the gene expression in the neuronal population was mostly affected in this region in early disease stages. Altogether results show that H-αSynOs trigger a region-specific dysregulation of gene expression in ACC and HC, providing a pathological substrate for MCI associated with early PD.

Somatosensory evoked potentials and transcranial color Doppler monitoring in subarachnoid hemorrhage.

OBJECTIVES: The outcome of patients with subarachnoid hemorrhage (SAH) is broadly influenced by the complications that may result from the hemorrhage. We describe a series of subjects, in which neurophysiological monitoring executed by simultaneous recording of somatosensory evoked potentials (SEPs) and transcranial color Doppler (TCD) was performed to reveal possible, early complications following acute SAH. MATERIALS AND METHODS: We described the absolute and interhemispheric values of SEPs from the upper limb and TCD examinations of the cerebral arteries in 13 subjects with acute SAH. RESULTS: In cases with middle cerebral artery (MCA) vasospasm, N20 SEP amplitude absolute values for the hemisphere involved in the vasospasm were much lower than the contralateral ones. The N20 amplitude ratio reduction correlated with reciprocal of MCA mean flow velocity values detected within each patient. In the subjects with early ischemic damage following SAH, the affected hemisphere showed N20 amplitude drop; in addition, the relationship between SEPs and TCD findings was missing. CONCLUSION: Our findings emphasize the utility of simultaneous evaluation of SEPs and TCD in SAH follow-up, since the two methods reflect different pathomechanisms of possible secondary brain damage in aneurysmal SAH.

Validation of I'screen AFLA M1 Milk for Detection of Aflatoxin M1 in Raw Bovine Milk and Powdered Milk: AOAC Performance Tested MethodSM 072002.

BACKGROUND: Aflatoxin M1 (AFM1) is the major metabolite of Aflatoxin B1 (AFB1) and can be found in the milk of animals fed with feed containing AFB1. The frequency of occurrence of AFM1 in milk has led to the development of specific quantitative methods of analysis to mitigate the risk of adversely affecting human health. OBJECTIVE: To demonstrate that the I'screen AFLA M1 Milk ELISA kit can quantify AFM1 in raw bovine milk and powdered milk. METHOD: Assay performance was evaluated studying lot-to-lot consistency, assay stability, robustness, and possible interferences of related molecules. Raw bovine milk samples spiked at 0, 5.0, 20, 50, 100, and 200 ng/L of AFM1 and powdered milk reference materials and spiked samples at 100 and 200 ng/L were tested to determine recovery, repeatability, and bias. LOD and LOQ were also determined for both matrices. RESULTS: High selectivity for AFM1 was demonstrated and performances were consistent, robust, and stable. The LOQ was validated at 5 ng/L for raw milk and 50 ng/L for powdered milk. Recoveries for spiked raw and powdered milk were 97-122%, with RSDr < 10%, and 106-111% for reference materials, with RSDr < 5%. CONCLUSIONS: The data collected validate the method as a selective, specific, sensitive, accurate, and precise tool for the analysis of AFM1 in raw bovine milk and powdered milk. HIGHLIGHTS: We demonstrated that I'screen AFLA M1 is a reliable kit and a proper screening tool suitable for high analytical throughputs.

Validation of two enzyme immunoassays for aminoglycoside residues according to European Decision 657/2002.

Aminoglycoside antibiotics are commonly used to treat bacterial infections in human and veterinary practice. Owing to their toxicity, the European Community has established maximum residue limits (MRL) in foodstuffs of animal origin (EEC No 2377/90). In the present study, the performance of two new enzyme immunoassays (EIA), I'screen Gentamicin and I'screen Neomycin, for the quantitative detection of the aminoglycosides, gentamicin and neomycin, in milk and tissue are described. Validation of these EIAs has been performed in accordance to the criteria of European Decision 657/2002. Assays sensitivity at the MRLs was 95% for milk samples and 100% for tissue samples, while specificity was 100% at 33 and 25% of the MRLs for milk and tissues, respectively. The performance of these EIAs indicates that they can be used as easy screening methods for the analysis of aminoglycosides in milk and tissue samples.

Differential HMGA expression and post-translational modifications in prostatic tumor cells.

The HMGA architectural nuclear factors are involved in chromatin dynamics and their overexpression has been strongly linked to the neoplastic transformation process. Here we investigate the expression and post-translational modifications (PTMs) of HMGA proteins (HMGA1a, HMGA1b and HMGA2) in the rat prostatic cancer Dunning model (G, AT-1, and MAT-Ly-Lu cell lines). We demonstrate the expression of HMGA2, in addition to HMGA1a and HMGA1b, in both the anaplastic cell lines AT-1 and MAT-Ly-Lu and an extremely specific HMGA1a mono-methylation only in the most metastatic cell line MAT-Ly-Lu. The HMGA ectopic expression in HMGA-negative Dunning G cells does not significantly alter their growth ability, suggesting that, although HMGA expression is necessary for the progression of neoplastic transformation in several cellular models, in these cells it is not sufficient. These data suggest exploring HMGA2 as a potential marker in human prostate tumor and moreover indicate PTMs as an additional tool in the staging of tumor progression.

During apoptosis of tumor cells HMGA1a protein undergoes methylation: identification of the modification site by mass spectrometry.

Programmed cell death is characterized by posttranslational modifications of a limited and specific set of nuclear proteins. We demonstrate that during apoptosis of different types of tumor cells there is a monomethylation of the nuclear protein HMGA1a that is associated to its previously described hyperphosphorylation/dephosphorylation process. HMGA1a methylation is strictly related to the execution of programmed cell death and is a massive event that involves large amounts of the protein. In some tumor cells, HMGA1a protein is already methylated to an extent that depends on cell type. The degree of methylation in any case definitely increases during apoptosis. In the studied cell systems (human leukaemia, human prostate tumor, and rat thyroid transformed cells) among the low-molecular-mass HMG proteins, only HMGA1a was found to be methylated. A tryptic digestion map of HPLC-purified HMGA1a protein showed that methylation occurs at arginine 25 in the consensus G(24)R(25)G(26) that belongs to one of the DNA-binding AT-hooks of the protein. An increase of HMGA1a methylation could be related to heterochromatin and chromatin remodeling of apoptotic cells.