Caffeine Consumption Prevents Diabetes-Induced Memory Impairment and Synaptotoxicity in the Hippocampus of NONcZNO10/LTJ Mice

Diabetic conditions are associated with modified brain function, namely with cognitive deficits, through largely undetermined processes. More than understanding the underlying mechanism, it is important to devise novel strategies to alleviate diabetes-induced cognitive deficits. Caffeine (a mixed antagonist of adenosine A1 and A2A receptors) emerges as a promising candidate since caffeine consumption reduces the risk of diabetes and effectively prevents memory deficits caused by different noxious stimuli. Thus, we took advantage of a novel animal model of type 2 diabetes to investigate the behavioural, neurochemical and morphological modifications present in the hippocampus and tested if caffeine consumption might prevent these changes. We used a model closely mimicking the human type 2 diabetes condition, NONcNZO10/LtJ mice, which become diabetic at 7–11 months when kept under an 11% fat diet. Caffeine (1 g/l) was applied in the drinking water from 7 months onwards. Diabetic mice displayed a decreased spontaneous alternation in the Y-maze accompanied by a decreased density of nerve terminal markers (synaptophysin, SNAP25), mainly glutamatergic (vesicular glutamate transporters), and increased astrogliosis (GFAP immunoreactivity) compared to their wild type littermates kept under the same diet. Furthermore, diabetic mice displayed up-regulated A2A receptors and down-regulated A1 receptors in the hippocampus. Caffeine consumption restored memory performance and abrogated the diabetes-induced loss of nerve terminals and astrogliosis. These results provide the first evidence that type 2 diabetic mice display a loss of nerve terminal markers and astrogliosis, which is associated with memory impairment; furthermore, caffeine consumption prevents synaptic dysfunction and astrogliosis as well as memory impairment in type 2 diabetes

NatF Contributes to an Evolutionary Shift in Protein N-Terminal Acetylation and Is Important for Normal Chromosome Segregation

N-terminal acetylation (N-Ac) is a highly abundant eukaryotic protein modification. Proteomics revealed a significant increase in the occurrence of N-Ac from lower to higher eukaryotes, but evidence explaining the underlying molecular mechanism(s) is currently lacking. We first analysed protein N-termini and their acetylation degrees, suggesting that evolution of substrates is not a major cause for the evolutionary shift in N-Ac. Further, we investigated the presence of putative N-terminal acetyltransferases (NATs) in higher eukaryotes. The purified recombinant human and Drosophila homologues of a novel NAT candidate was subjected to in vitro peptide library acetylation assays. This provided evidence for its NAT activity targeting Met-Lys- and other Met-starting protein N-termini, and the enzyme was termed Naa60p and its activity NatF. Its in vivo activity was investigated by ectopically expressing human Naa60p in yeast followed by N-terminal COFRADIC analyses. hNaa60p acetylated distinct Met-starting yeast protein N-termini and increased general acetylation levels, thereby altering yeast in vivo acetylation patterns towards those of higher eukaryotes. Further, its activity in human cells was verified by overexpression and knockdown of hNAA60 followed by N-terminal COFRADIC. NatF's cellular impact was demonstrated in Drosophila cells where NAA60 knockdown induced chromosomal segregation defects. In summary, our study revealed a novel major protein modifier contributing to the evolution of N-Ac, redundancy among NATs, and an essential regulator of normal chromosome segregation. With the characterization of NatF, the co-translational N-Ac machinery appears complete since all the major substrate groups in eukaryotes are accounted for

Hypoxia induced amoeboid microglial cell activation in postnatal rat brain is mediated by ATP receptor P2X4

<p>Abstract</p> <p>Background</p> <p>Activation of amoeboid microglial cells (AMC) and its related inflammatory response have been linked to the periventricular white matter damage after hypoxia in neonatal brain. Hypoxia increases free ATP in the brain and then induces various effects through ATP receptors. The present study explored the possible mechanism in ATP induced AMC activation in hypoxia.</p> <p>Results</p> <p>We first examined the immunoexpression of P2X4, P2X7 and P2Y12 in the corpus callosum (CC) and subependyma associated with the lateral ventricles where both areas are rich in AMC. Among the three purinergic receptors, P2X4 was most intensely expressed. By double immunofluorescence, P2X4 was specifically localized in AMC (from P0 to P7) but the immunofluorescence in AMC was progressively diminished with advancing age (P14). It was further shown that P2X4 expression was noticeably enhanced in P0 day rats subjected to hypoxia and killed at 4, 24, 72 h and 7 d versus their matching controls by double labeling and western blotting analysis. P2X4 expression was most intense at 7 d whence the inflammatory response was drastic after hypoxia. We then studied the association of P2X4 with cytokine release in AMC after hypoxic exposure. In primary microglial cells exposed to hypoxia, IL-1β and TNF-α protein levels were up-regulated. Blockade of P2X4 receptor with 2', 3'-0-(2, 4, 6-Trinitrophenyl) adenosine 5'-triphosphate, a selective P2X1-7 blocker resulted in partial suppression of IL-1β (24% <it>vs </it>hypoxic group) and TNF-α expression (40% <it>vs </it>hypoxic group). However, pyridoxal phosphate-6-azo (benzene-2, 4-disulfonic acid) tetrasodium salt hydrate, a selective P2X1-3, 5-7 blocker did not exert any significant effect on the cytokine expression.</p> <p>Conclusions</p> <p>It is concluded that P2X4 which is constitutively expressed by AMC in postnatal rats was enhanced in hypoxia. Hypoxia induced increase in IL-1β and TNF-α expression was reversed by 2', 3'-0-(2, 4, 6-Trinitrophenyl) adenosine 5'-triphosphate suggesting that P2X4 mediates ATP induced AMC activation and its production of proinflammatory cytokines.</p

Intravenous magnesium prevents atrial fibrillation after coronary artery bypass grafting: a meta-analysis of 7 double-blind, placebo-controlled, randomized clinical trials

<p>Abstract</p> <p>Background</p> <p>Postoperative atrial fibrillation (POAF) is the most common complication after coronary artery bypass grafting (CABG). The preventive effect of magnesium on POAF is not well known. This meta-analysis was undertaken to assess the efficacy of intravenous magnesium on the prevention of POAF after CABG.</p> <p>Methods</p> <p>Eligible studies were identified from electronic databases (Medline, Embase, and the Cochrane Library). The primary outcome measure was the incidence of POAF. The meta-analysis was performed with the fixed-effect model or random-effect model according to heterogeneity.</p> <p>Results</p> <p>Seven double-blind, placebo-controlled, randomized clinical trials met the inclusion criteria including 1,028 participants. The pooled results showed that intravenous magnesium reduced the incidence of POAF by 36% (RR 0.64; 95% confidence interval (CI) 0.50-0.83; <it>P </it>= 0.001; with no heterogeneity between trials (heterogeneity <it>P </it>= 0.8, <it>I</it>²= 0%)).</p> <p>Conclusions</p> <p>This meta-analysis indicates that intravenous magnesium significantly reduces the incidence of POAF after CABG. This finding encourages the use of intravenous magnesium as an alternative to prevent POAF after CABG. But more high quality randomized clinical trials are still need to confirm the safety.</p

SCFAs Induce Mouse Neutrophil Chemotaxis through the GPR43 Receptor

Short chain fatty acids (SCFAs) have recently attracted attention as potential mediators of the effects of gut microbiota on intestinal inflammation. Some of these effects have been suggested to occur through the direct actions of SCFAs on the GPR43 receptor in neutrophils, though the precise role of this receptor in neutrophil activation is still unclear. We show that mouse bone marrow derived neutrophils (BMNs) can chemotax effectively through polycarbonate filters towards a source of acetate, propionate or butyrate. Moreover, we show that BMNs move with good speed and directionality towards a source of propionate in an EZ-Taxiscan chamber coated with fibrinogen. These effects of SCFAs were mimicked by low concentrations of the synthetic GPR43 agonist phenylacetamide-1 and were abolished in GPR43−/− BMNs. SCFAs and phenylacetamide-1 also elicited GPR43-dependent activation of PKB, p38 and ERK and these responses were sensitive to pertussis toxin, indicating a role for Gi proteins. Phenylacetamide-1 also elicited rapid and transient activation of Rac1/2 GTPases and phosphorylation of ribosomal protein S6. Genetic and pharmacological intervention identified important roles for PI3Kγ, Rac2, p38 and ERK, but not mTOR, in GPR43-dependent chemotaxis. These results identify GPR43 as a bona fide chemotactic receptor for neutrophils in vitro and start to define important elements in its signal transduction pathways

MicroRNA Expression Analysis: Clinical Advantage of Propranolol Reveals Key MicroRNAs in Myocardial Infarction

BACKGROUND: As playing important roles in gene regulation, microRNAs (miRNAs) are believed as indispensable involvers in the pathogenesis of myocardial infarction (MI) that causes significant morbidity and mortality. Working on a hypothesis that modulation of only some key members in the miRNA superfamily could benefit ischemic heart, we proposed a microarray based network biology approach to identify them with the recognized clinical effect of propranolol as a prompt. METHODS: A long-term MI model of rat was established in this study. The microarray technology was applied to determine the global miRNA expression change intervened by propranolol. Multiple network analyses were sequentially applied to evaluate the regulatory capacity, efficiency and emphasis of the miRNAs which dysexpression in MI were significantly reversed by propranolol. RESULTS: Microarray data analysis indicated that long-term propranolol administration caused 18 of the 31 dysregulated miRNAs in MI undergoing reversed expression, implying that intentional modulation of miRNA expression might show favorable effects for ischemic heart. Our network analysis identified that, among these miRNAs, the prime players in MI were miR-1, miR-29b and miR-98. Further finding revealed that miR-1 focused on regulation of myocyte growth, yet miR-29b and miR-98 stressed on fibrosis and inflammation, respectively. CONCLUSION: Our study illustrates how a combination of microarray technology and functional protein network analysis can be used to identify disease-related key miRNAs

A Rapid and Simple Procedure for the Establishment of Human Normal and Cancer Renal Primary Cell Cultures from Surgical Specimens

The kidney is a target organ for the toxicity of several xenobiotics and is also highly susceptible to the development of malignant tumors. In both cases, in vitro studies provide insight to cellular damage, and represent adequate models to study either the mechanisms underlying the toxic effects of several nephrotoxicants or therapeutic approaches in renal cancer. The development of efficient methods for the establishment of human normal and tumor renal cell models is hence crucial. In this study, a technically simple and rapid protocol for the isolation and culture of human proximal tubular epithelial cells and human renal tumor cells from surgical specimens is presented. Tumor and normal tissues were processed by using the same methodology, based on mechanical disaggregation of tissue followed by enzymatic digestion and cell purification by sequential sieving. The overall procedure takes roughly one hour. The resulting cell preparations have excellent viabilities and yield. Establishment of primary cultures from all specimens was achieved successfully. The origin of primary cultured cells was established through morphological evaluation. Normal cells purity was confirmed by immunofluorescent staining and reverse transcription-polymerase chain reaction analysis for expression of specific markers

Development of mandibular, hyoid and hypobranchial muscles in the zebrafish: homologies and evolution of these muscles within bony fishes and tetrapods

<p>Abstract</p> <p>Background</p> <p>During vertebrate head evolution, muscle changes accompanied radical modification of the skeleton. Recent studies have suggested that muscles and their innervation evolve less rapidly than cartilage. The freshwater teleostean zebrafish (<it>Danio rerio</it>) is the most studied actinopterygian model organism, and is sometimes taken to represent osteichthyans as a whole, which include bony fishes and tetrapods. Most work concerning zebrafish cranial muscles has focused on larval stages. We set out to describe the later development of zebrafish head muscles and compare muscle homologies across the Osteichthyes.</p> <p>Results</p> <p>We describe one new muscle and show that the number of mandibular, hyoid and hypobranchial muscles found in four day-old zebrafish larvae is similar to that found in the adult. However, the overall configuration and/or the number of divisions of these muscles change during development. For example, the undivided adductor mandibulae of early larvae gives rise to the adductor mandibulae sections A0, A1-OST, A2 and Aω, and the protractor hyoideus becomes divided into dorsal and ventral portions in adults. There is not always a correspondence between the ontogeny of these muscles in the zebrafish and their evolution within the Osteichthyes. All of the 13 mandibular, hyoid and hypobranchial muscles present in the adult zebrafish are found in at least some other living teleosts, and all except the protractor hyoideus are found in at least some extant non-teleost actinopterygians. Of these muscles, about a quarter (intermandibularis anterior, adductor mandibulae, sternohyoideus) are found in at least some living tetrapods, and a further quarter (levator arcus palatini, adductor arcus palatini, adductor operculi) in at least some extant sarcopterygian fish.</p> <p>Conclusion</p> <p>Although the zebrafish occupies a rather derived phylogenetic position within actinopterygians and even within teleosts, with respect to the mandibular, hyoid and hypobranchial muscles it seems justified to consider it an appropriate representative of these two groups. Among these muscles, the three with clear homologues in tetrapods and the further three identified in sarcopterygian fish are particularly appropriate for comparisons of results between the actinopterygian zebrafish and the sarcopterygians.</p

Water-Borne Cues of a Non-Indigenous Seaweed Mediate Grazer-Deterrent Responses in Native Seaweeds, but Not Vice Versa

Plants optimise their resistance to herbivores by regulating deterrent responses on demand. Induction of anti-herbivory defences can occur directly in grazed plants or from emission of risk cues to the environment, which modifies interactions of adjacent plants with, for instance, their consumers. This study confirmed the induction of anti-herbivory responses by water-borne risk cues between adjoining con-specific seaweeds and firstly examined whether plant-plant signalling also exists among adjacent hetero-specific seaweeds. Furthermore, differential abilities and geographic variation in plant-plant signalling by a non-indigenous seaweed as well as native seaweeds were assessed. Twelve-day induction experiments using the non-indigenous seaweed Sargassum muticum were conducted in the laboratory in Portugal and Germany with one local con-familiar (Portugal: Cystoseira humilis, Germany: Halidrys siliquosa) and hetero-familiar native species (Portugal: Fucus spiralis, Germany: F. vesiculosus). All seaweeds were grazed by a local isopod species (Portugal: Stenosoma nadejda, Germany: Idotea baltica) and were positioned upstream of con- and hetero-specific seaweeds. Grazing-induced modification in seaweed traits were tested in three-day feeding assays between cue-exposed and cue-free ( = control) pieces of both fresh and reconstituted seaweeds. Both Fucus species reduced their palatability when positioned downstream of isopod-grazed con-specifics. Yet, the palatability of non-indigenous S. muticum remained constant in the presence of upstream grazed con-specifics and native hetero-specifics. In contrast, both con-familiar (but neither hetero-familiar) native species reduced palatability when located downstream of grazed S. muticum. Similar patterns of grazer-deterrent responses to water-borne cues were observed on both European shores, and were almost identical between assays using fresh and reconstituted seaweeds. Hence, seaweeds may use plant-plant signalling to optimise chemical resistance to consumers, though this ability appeared to be species-specific. Furthermore, this study suggests that native species may benefit more than a non-indigenous species from water-borne cue mediated reduction in consumption as only natives responded to signals emitted by hetero-specifics