Diverse Brain Myeloid Expression Profiles Reveal Distinct Microglial Activation States and Aspects of Alzheimer’s Disease Not Evident in Mouse Models

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Plasmid-mediated degradation of o- and p-phthalate by Pseudomonas fluorescens

A Pseudomonas fluorescens strain SKP3 capable of utilizing both phthalic acid and terephthalic acid as sole source of carbon and energy was isolated by enrichment technique. Phthalic acid, terephthalic acid and protocatechuic acid were easily oxidized by both phthalate-grown and glucose-grown cells without a lag period. Phthalic acid is metabolized through the ortho cleavage pathway and terephthalic acid through the meta cleavage pathway and the enzymes of the two pathways are constitutive in nature. A large plasmid of approximately 140kb in size was found to be involved in the degradation of phthalates. The catabolic plasmid pSKL was transferable to different hosts.     

Cardioprotection Resulting from Glucagon-Like Peptide-1 Administration Involves Shifting Metabolic Substrate Utilization to Increase Energy Efficiency in the Rat Heart

Previous studies have shown that glucagon-like peptide-1 (GLP-1) provides cardiovascular benefits independent of its role on peripheral glycemic control. However, the precise mechanism(s) by which GLP-1 treatment renders cardioprotection during myocardial ischemia remain unresolved. Here we examined the role for GLP-1 treatment on glucose and fatty acid metabolism in normal and ischemic rat hearts following a 30 min ischemia and 24 h reperfusion injury, and in isolated cardiomyocytes (CM). Relative carbohydrate and fat oxidation levels were measured in both normal and ischemic hearts using a 1-¹³C glucose clamp coupled with NMR-based isotopomer analysis, as well as in adult rat CMs by monitoring pH and O₂ consumption in the presence of glucose or palmitate. In normal heart, GLP-1 increased glucose uptake (↑64%, p<0.05) without affecting glycogen levels. In ischemic hearts, GLP-1 induced metabolic substrate switching by increasing the ratio of carbohydrate versus fat oxidation (↑14%, p<0.01) in the LV area not at risk, without affecting cAMP levels. Interestingly, no substrate switching occurred in the LV area at risk, despite an increase in cAMP (↑106%, p<0.05) and lactate (↑121%, p<0.01) levels. Furthermore, in isolated CMs GLP-1 treatment increased glucose utilization (↑14%, p<0.05) and decreased fatty acid oxidation (↓15%, p<0.05) consistent with in vivo finding. Our results show that this benefit may derive from distinct and complementary roles of GLP-1 treatment on metabolism in myocardial sub-regions in response to this injury. In particular, a switch to anaerobic glycolysis in the ischemic area provides a compensatory substrate switch to overcome the energetic deficit in this region in the face of reduced tissue oxygenation, whereas a switch to more energetically favorable carbohydrate oxidation in more highly oxygenated remote regions supports maintaining cardiac contractility in a complementary manner.     

A System for Distinguishing Octoploid Strawberry Cultivars Using High-Throughput SNP Genotyping

Cultivated strawberry (Fragaria × ananassa) is an important commercial berry crop grown throughout the world. Improved strawberry cultivars are developed to meet the needs of consumers and breeders. Strawberries are usually propagated through runners, which sometimes lead to mislabeling or misinterpretation of cultivars. However, perfect identification of strawberry cultivars is essential for germplasm maintenance and for breeding programs. Molecular marker technology has been widely used to distinguish cultivars of other crops, but marker development in octoploid strawberries is complicated. Therefore, SNP marker with high-density and even distribution in the genome has been used currently as efficient DNA markers. In this report, previously published high-quality poly high resolution (PHR) SNPs from the 90 K Axiom® SNP array were utilized to develop a Fluidigm 24 SNPs genotyping system. Hundred nine (109) octoploid strawberry cultivars were screened using this 24 SNPs chip set. In addition, 24 SNPs were mapped to six chromosomes of diploid strawberry (Fragaria vesca). Our developed SNPs fluidigm genotyping is automatable, easy and reliable for processing and interpretation of data. Thus, this high-throughput SNP genotyping system will be a useful tool for distinguishing strawberry cultivars and find out parent-offspring relationship.     

Detection and measurement of alternative splicing using splicing-sensitive microarrays

Splicing and alternative splicing are major processes in the interpretation and expression of genetic information for metazoan organisms. The study of splicing is moving from focused attention on the regulatory mechanisms of a selected set of paradigmatic alternative splicing events to questions of global integration of splicing regulation with genome and cell function. For this reason, parallel methods for detecting and measuring alternative splicing are necessary. We have adapted the splicing-sensitive oligonucleotide microarrays used to estimate splicing efficiency in yeast to the study of alternative splicing in vertebrate cells and tissues. We use gene models incorporating knowledge about splicing to design oligonucleotides specific for discriminating alternatively spliced mRNAs from each other. Here we present the main strategies for design, application, and analysis of spotted oligonucleotide arrays for detection and measurement of alternative splicing. We demonstrate these strategies using a two-intron yeast gene that has been altered to produce different amounts of alternatively spliced RNAs, as well as by profiling alternative splicing in NCI 60 cancer cell lines.     

Synthetic LXR agonists increase LDL in CETP species

Liver X receptor (LXR) nuclear receptors regulate the expression of genes involved in whole body cholesterol trafficking, including absorption, excretion, catabolism, and cellular efflux, and possess both anti-inflammatory and antidiabetic actions. Accordingly, LXR is considered an appealing drug target for multiple indications. Synthetic LXR agonists demonstrated inhibition of atherosclerosis progression in murine genetic models; however, these and other studies indicated that their major undesired side effect is an increase of plasma and hepatic triglycerides. A significant impediment to extrapolating results with LXR agonists from mouse to humans is the absence in mice of cholesteryl ester transfer protein, a known LXR target gene, and the upregulation in mice but not humans of cholesterol 7alpha-hydroxylase. To better predict the human response to LXR agonism, two synthetic LXR agonists were examined in hamsters and cynomolgus monkeys. In contrast to previously published results in mice, neither LXR agonist increased HDL-cholesterol in hamsters, and similar results were obtained in cynomolgus monkeys. Importantly, in both species, LXR agonists increased LDL-cholesterol, an unfavorable effect not apparent from earlier murine studies. These results reveal additional problems associated with current synthetic LXR agonists and emphasize the importance of profiling compounds in preclinical species with a more human-like LXR response and lipoprotein metabolism.     

Extreme chemical mutagen sensitivity of respiratory adaptation in yeast

The respiratory adaptation process (i.e essentially mitochondrial biogenesis) in the cells of both wild-type Saccharomyces cerevisiae and strains sensitive to ultraviolet radiation (UV) undergoing transition from the anaerobic to the aerobic state (1–2 h aeration) could be arrested by a prior incubation for 15–30 min with several chemical mutagens and other DNA-acting chemicals at very low concentrations (10^?7 to 10^?6 M added to cells suspended at the density of 10⁷ cells/ml). At the same concentrations, these chemicals also inhibited DNA and RNA biosynthesis in maturing mitochondria during respiratory adaptation. This provides suggestive evidence for the view that the inhibitory effect of the chemical mutagens on respiratory adaptation could be due to lesions introduced into the DNA of promitochondria in the anaerobic cells. The system of respiratory adaptation in S. cerevisiae cells could serve as a rapid test for ascertaining the potentiality of a chemical to affect DNA and probably, in turn, its potentiality to be mutagenic.     

Defect in oxidative phosphorylation in LV papillary muscle mitochondria of patients undergoing mitral valve replacement

Mitochondria play a pivotal role in cellular metabolism, especially in energy production. Myocardial function depends on adenosine triphosphate (ATP) supplied by oxidation of several substrates. In the adult heart, this energy is obtained primarily from fatty acid oxidation through oxidative phosphorylation (OXPHOS). With this in view, we studied OXPHOS, Total-ATPase and cytochrome content in the mitochondria of the left ventricular (LV) papillary muscles in excised mitral valves of patients who underwent mitral valve replacement (MVR). The mitochondrial OXPHOS, cytochrome content and ATPase activity were studied in 70 patients (ranging from 22 to 40 years) operated on for mitral valve disease. Control study includes 25 normal mitral valves removed at necropsy from patients who died of extracardiac causes. In the presence of glutamate and succinate as substrates, the rate of mitochondrial oxygen consumption was significantly lower in LV papillary muscles of pathological mitral valves (P<0.001) by using with and without addition of ADP. The ADP/O ratio indices for glutamate and succinate were not significantly affected. Using glutamate as substrate, respiratory control index was significantly raised (P<0.05) as compared with control. A significant reduction of total cytochrome content and ATPase activity (P<0.001) was noted in LV papillary muscles of patients operated for mitral valve disease. Our results showed that OXPHOS, cytochromes 'a', 'b', 'c+c(1)' and ATP activity are significantly impaired in LV papillary muscles in patients with pathological mitral valve. Cardiac mitochondrial oxygen consumption is a very valuable tool to investigate the regulation of cardiac mitochondrial energy metabolism. There is increasing evidence that mitochondrial diseases, such as mitochondrial cardiomyopathy, valvular disease and some myopathies, can be responsive to treatment with metabolic intermediates such as coenzyme Q(10), thiamine, prednisone, and vitamin therapy.     

Synergistic effect of calcium stearate and photo treatment on the rate of biodegradation of low density polyethylene spent saline vials

The biodegradation of spent saline bottles, a low density polyethylene product (LDPE) by two selected Arthrobacter sp. under in vitro conditions is reported. Chemical and UV pretreatment play a vital role in enhancing the rate of biodegradation. Treated LDPE film exhibits a higher weight loss and density when compared to untreated films. Arthrobacter oxydans and Arthrobacter globiformis grew better in medium containing pretreated film than in medium containing untreated film. The decrease in density and weight loss of LDPE was also more for pretreated film when compared to untreated film indicating the affect of abiotic treatment on mechanical properties of LDPE. The decrease in the absorbance corresponding to carbonyl groups and double bonds that were generated during pretreatment suggest that some of the double bonds were cut by Arthrobacter species. Since Arthrobacter sp. are capable of degrading urea, splitting of urea group were also seen in FTIR spectrum indicating the evidence of biodegradation after microbial incubation. The results indicated that biodegradation rate could be enhanced by exposing LDPE to calcium stearate (a pro-oxidant) which acts as an initiator for the oxidation of the polymers leading to a decrease of molecular weight and formation of hydrophilic group. Therefore, the initial step for biodegradation of many inert polymers depends on a photo-oxidation of those polymers. The application in sufficient details with improved procedures utilizing recombinant microorganism with polymer degradation capacity can lead to a better plastic waste management in biomedical field. The present plastic disposal trend of waste accumulation can be minimized with this promising eco-friendly technique.