VAPB and C9orf72 mutations in 1 familial amyotrophic lateral sclerosis patient

Previously, we have reported amyotrophic lateral sclerosis (ALS) families with multiple mutations in major ALS-associated genes. These findings provided evidence for an oligogenic basis of ALS. In our present study, we screened a cohort of 755 sporadic ALS patients, 111 familial ALS patients (97 families), and 765 control subjects of Dutch descent for mutations in vesicle-associated membrane protein B (VAPB). We have identified 1 novel VAPB mutation (p.V234I) in a familial ALS patient known to have a chromosome 9 open reading frame 72 (C9orf72) repeat expansion. This p.V234I mutation was absent in control subjects, located in a region with high evolutionary conservation, and predicted to have damaging effects. Taken together, these findings provide additional evidence for an oligogenic basis of ALS.     

Coronary thrombi neovascularization in patients with ST-elevation myocardial infarction - clinical and angiographic implications

Introduction

Coronary artery thrombosis in ST-elevation myocardial infarction (STEMI) is a dynamic process often preceded by episodes of silent plaque rupture and subocclusive thrombosis. Thrombus organization is achieved by ingrowth of endothelial and smooth muscle cells. Clinical significance and impact of thrombus neovascularization on primary percutaneous coronary intervention (pPCI) outcome remain unclear. Therefore we investigated composition and neovascularization of thrombi aspirated during pPCI and their association with clinical and angiographic parameters of STEMI patients.

Methods

Aspirated thrombi retrieved from 84 STEMI patients were classified as fresh (< 1 day), lytic (1-5 days) or organized (> 5 days). Thrombus neovascularization was evaluated immunohistochemically using CD34, CD31 and VEGF antibodies. CD34 and CD31 immunopositive (CD34/CD31 +) cells were organized as single, clusters and microvessels. VEGF positivity was graded as low or high, based on thrombus surface immunopositive area.

Results

CD34/CD31 + cells were present in 67% of all aspirated thrombi. Thrombus CD34/CD31 positivity was associated with previous history of angina pectoris (χ² = 6.142, p = 0.013) and lower myocardial blush grade (MBG < 3, χ² = 12.602, p < 0.001). Organization of CD34/CD31 + cells showed inverse association with the extent of VEGF positivity (χ² = 10.607, p = 0.005). Fresh thrombi were associated with shorter ischemic time (U = 237.5, p = 0.002) and MBG 3 (χ² = 6.379, p = 0.012).

Conclusions

Older thrombus age and neovascularization are associated with suboptimal myocardial perfusion in STEMI patients. Thrombus VEGF expression is inversely associated with degree of CD34 + cell organization. Therefore, neovascularization of aspirated thrombi may indicate the duration of thrombosis, coronary microcirculation status and outcome in STEMI patients.     

17β-Estradiol-Induced Synaptic Rearrangements Are Accompanied by Altered Ectonucleotidase Activities in Male Rat Hippocampal Synaptosomes

17β-Estradiol (E2) rapidly, by binding to membrane estrogen receptors, activates cell signaling cascades which induce formation of new dendritic spines in the hippocampus of males as in females, but the interaction with other metabolic processes, such as extracellular adenine nucleotides metabolism, are currently unknown. Extracellular adenine nucleotides play significant roles, controlling excitatory glutamatergic synapses and development of neural circuits and synaptic plasticity. Their precise regulation in the synaptic cleft is tightly controlled by ecto-nucleoside triphosphate diphosphohydrolase (NTPDase)/ecto-5′-nucleotidase (eN) enzyme chain. Therefore, we sought to clarify whether a single systemic injection of E2 in male rats is accompanied by changes in the expression of the pre- and postsynaptic proteins and downstream kinases linked to E2-induced synaptic rearrangement as well as alterations in NTPDase/eN pathway in the hippocampal synaptosomes. Obtained data showed activation of mammalian target of rapamycin and upregulation of key synaptic proteins necessary for spine formation, 24 h after systemic E2 administration. In E2-mediated conditions, we found downregulation of NTPDase1 and NTPDase2 and attenuation of adenine nucleotide hydrolysis by NTPDase/eN enzyme chain, without changes in NTPDase3 properties and augmentation of synaptic tissue-nonspecific alkaline phosphatase (TNAP) activity. Despite reduced NTPDase activities, increased TNAP activity probably prevents toxic accumulation of ATP in the extracellular milieu and also hydrolyzes accumulated ADP due to unchanged NTPDase3 activity. Thus, our initial evaluation supports idea of specific roles of different ectonucleotidases and their coordinated actions in E2-mediated spine remodeling and maintenance.     

In vivo observation of intracellular oximetry in perfluorocarbon‐labeled glioma cells and chemotherapeutic response in the CNS using fluorine‐19 MRI

Preclinical development of therapeutic agents against cancer could greatly benefit from noninvasive markers of tumor killing. Potentially, the intracellular partial pressure of oxygen (pO₂) can be used as an early marker of antitumor efficacy. Here, the feasibility of measuring intracellular pO₂ of central nervous system glioma cells in vivo using ¹⁹F magnetic resonance techniques is examined. Rat 9L glioma cells were labeled with perfluoro‐15‐crown‐5‐ether ex vivo and then implanted into the rat striatum. ¹⁹F MRI was used to visualize tumor location in vivo. The mean ¹⁹F T₁ of the implanted cells was measured using localized, single‐voxel spectroscopy. The intracellular pO₂ in tumor cells was determined from an in vitro calibration curve. The basal pO₂ of 9L cells (day 3) was determined to be 45.3 ± 5 mmHg (n = 6). Rats were then treated with a 1× LD10 dose of bischloroethylnitrosourea intravenously and changes in intracellular pO₂ were monitored. The pO₂ increased significantly (P = 0.042, paired T‐test) to 141.8 ± 3 mmHg within 18 h after bischloroethylnitrosourea treatment (day 4) and remained elevated (165 ± 24 mmHg) for at least 72 h (day 6). Intracellular localization of the perfluoro‐15‐crown‐5‐ether emulsion in 9L cells before and after bischloroethylnitrosourea treatment was confirmed by histological examination and fluorescence microscopy. Overall, noninvasive ¹⁹F magnetic resonance techniques may provide a valuable preclinical tool for monitoring therapeutic response against central nervous system or other deep‐seated tumors. Magn Reson Med, 2010. © 2010 Wiley‐Liss, Inc.     

Height and prostate cancer risk: a large nested case-control study (ProtecT) and meta-analysis

BACKGROUND: Height, a marker of childhood environmental exposures, is positively associated with prostate cancer risk, perhaps through the insulin-like growth factor system. We investigated the relationship of prostate cancer with height and its components (leg and trunk length) in a nested case-control study and with height in a dose-response meta-analysis. METHODS: We nested a case-control study within a population-based randomized controlled trial evaluating treatments for localized prostate cancer in British men ages 50 to 69 years, including 1,357 cases detected through prostate-specific antigen testing and 7,990 controls (matched on age, general practice, assessment date). Nine bibliographic databases were searched systematically for studies on the height-prostate cancer association that were pooled in a meta-analysis. RESULTS: Based on the nested case-control, the odds ratio (OR) of prostate-specific antigen-detected prostate cancer per 10 cm increase in height was 1.06 [95% confidence interval (95% CI): 0.97-1.16; p(trend) = 0.2]. There was stronger evidence of an association of height with high-grade prostate cancer (OR: 1.23; 95% CI: 1.06-1.43), mainly due to the leg component, but not with low-grade disease (OR: 0.99; 95% CI: 0.90-1.10). In general, associations with leg or trunk length were similar. A meta-analysis of 58 studies found evidence that height is positively associated with prostate cancer (random-effects OR per 10 cm: 1.06; 95% CI: 1.03-1.09), with a stronger effect for prospective studies of more advanced/aggressive cancers (random-effects OR: 1.12; 95% CI: 1.05-1.19). CONCLUSION: These data indicate a limited role for childhood environmental exposures-as indexed by adult height-on prostate cancer incidence, while suggesting a greater role for progression, through mechanisms requiring further investigation.     

Regulatory mechanisms of fear extinction and depression-like behavior.

Human anxiety is frequently accompanied by depression, and when they co-occur both conditions exhibit greater severity and resistance to treatment. Little is known, however, about the molecular processes linking these emotional and mood disorders. Based on previously reported phosphorylation patterns of extracellular signal-regulated kinase (ERK) in the brain, we hypothesized that ERK's upstream activators intertwine fear and mood regulation through their hippocampal actions. We tested this hypothesis by studying the upstream regulation of ERK signaling in behavioral models of fear and depression. Wild-type and ERK1-deficient mice were used to study the dorsohippocampal actions of the putative ERK activators: mitogen-activated and extracellular signal-regulated kinase (MEK), protein kinase C (PKC), and cAMP-dependent protein kinase (PKA). Mice lacking ERK1 exhibited enhanced fear extinction and reduced depression caused by overactivation of ERK2. Both behaviors were reversed by inhibition of MEK, however the extinction phenotype depended on hippocampal, whereas the depression phenotype predominantly involved extrahippocampal MEK. Unexpectedly, inhibition of PKC accelerated extinction and decreased depression by ERK-independent mechanisms, whereas inhibition of PKA did not produce detectable molecular or behavioral effects in the employed paradigm. These results indicate that, contrary to fear conditioning but similar to mood stabilization, extinction of fear required upregulation of MEK/ERK and downregulation of ERK-independent PKC signaling. The dissociation of these pathways may thus represent a common mechanism for fear and mood regulation, and a potential therapeutic option for comorbid anxiety and depression.     

Protective effect of basil (Ocimum basilicum L.) against oxidative DNA damage and mutagenesis.

Mutagenic and antimutagenic properties of essential oil (EO) of basil and its major constituent Linalool, reported to possess antioxidative properties, were examined in microbial tests. In Salmonella/microsome and Escherichia. coli WP2 reversion assays both derivatives (0.25–2.0 μl/plate) showed no mutagenic effect. Salmonella. typhimurium TA98, TA100 and TA102 strains displayed similar sensitivity to both basil derivatives as non-permeable E. coli WP2 strains IC185 and IC202 oxyR. Moreover, the toxicity of basil derivatives to WP2 strains did not depend on OxyR function. The reduction of t-BOOH-induced mutagenesis by EO and Linalool (30–60%) was obtained in repair proficient strains of the E. coli K12 assay (Nikoli?, B., Stanojevi?, J., Miti?, D., Vukovi?-Ga?i?, B., Kne?evi?-Vuk?evi?, J., Simi?, D., 2004. Comparative study of the antimutagenic potential of vitamin E in different E. coli strains. Mutat. Res. 564, 31–38), as well as in E. coli WP2 IC202 strain. EO and Linalool reduced spontaneous mutagenesis in mismatch repair deficient E. coli K12 strains (27–44%). In all tests, antimutagenic effect of basil derivatives was comparable with that obtained with model antioxidant vitamin E. Linalool and vitamin E induced DNA strand breaks in Comet assay on S. cerevisiae 3A cells, but at non-genotoxic concentrations (0.075 and 0.025 μg/ml, respectively) they reduced the number of H₂O₂-induced comets (45–70% Linalool and 80–93% vitamin E). Obtained results indicate that antigenotoxic potential of basil derivatives could be attributed to their antioxidative properties.     

Variability of hemagglutinin-neuraminidase and nucleocapsid protein of vaccine and wild-type mumps virus strains.

The mumps virus (MuV) molecular evolution is characterized by the co-circulation of numerous distinct strains. Standardized phylogenetic analyses based on the nucleotide sequences of the SH gene are important for mumps surveillance, but lack the information regarding antigenic properties. So far, the location of antigenic epitopes has been determined for two MuV proteins, the hemagglutinin-neuraminidase (HN) and the nucleocapsid (N) protein. We performed multiple sequence comparisons of putative HN and N protein sequences in order to describe their diversity and plasticity, and to determine the level of similarity between vaccine and wild-type strains. The results of full-length HN or N protein phylogeny showed that MuV strains form a number of differing clades which are in concordance with grouping obtained by standard MuV genotyping. When vaccine strains are compared to all wild-type strains, the highest mean percentage of amino acid differences in both HN and N protein analysis was found for Jeryl Lynn 5 and Jeryl Lynn 2 strains while the lowest value was obtained for Leningrad-3 and L-Zagreb strains. When only 3 antigenic regions of the HN protein, comprising 45 amino acids in total, were investigated, the diversity is considerably diminished: 51.5% of all putative HN proteins show identical sequences (including those of vaccine strains L-Zagreb, Leningrad-3, Hoshino and Urabe). Another 26.5% proteins (including Miyahara vaccine strain) differ in only one amino acid, while the others differ in two to five amino acids from the most common sequence. Jeryl Lynn 2 and Jeryl Lynn 5 strains differ in four amino acids each. N protein antigenic sites have been mapped within its hypervariable C-terminus. Our results indicate that there might be genotype-specific amino acids residing in this antigenic region. The results of our study present the background information for investigations of MuV heterogeneity and antigenic diversity.     

Correlation between age-related changes in open field behavior and plaque forming cell response in DA female rats

We investigated the relationship between immunological and behavioral changes during ageing in Dark Agouti female rats. Results showed that ageing was associated with decreased exploratory behavior and increased emotionality (open field test) and decreased pain perception (writhing assay), but not with altered depression-like behavior (forced swim test). The observed behavioral changes were paralleled with decreased innate immunity in middle-aged and old rats, as revealed by reduced peroxide production of peritoneal macrophages; and decreased specific immunity, measured by the plaque-forming cell response, in old rats in comparison with young rats. Correlation analyses between behavioral and immune parameters demonstrated a significant correlation between the lines crossed in the open field test and the plaque-forming cell response. Taken together, the demonstrated age-dependent association between exploratory behavior and specific immune response suggests a senescent decline of a common neuroimmune regulatory mechanism.