Increased insulin-like growth factor-I gene expression precedes left ventricular cardiomyocyte hypertrophy in a rapidly-hypertrophying rat model system

Chronic pressure overload leads to an increase in the size, i.e. hypertrophy, of cardiomyocytes in the heart. However, the molecular mechanisms underlying this hypertrophy are not understood. Insulin-like growth factor-I (IGF-I) synthesized locally in the heart is known to be associated with the hypertrophic process. So far, however, cardiac IGF-I gene expression in the widely used rat model system has only been shown to be increased when the hypertrophy induced by pressure-overload was already established. Therefore, the question of whether IGF-I serves as an initiating or early-enhancing factor for the cardiac hypertrophy remains unanswered. Here, cardiac hypertension and hypertrophy were rapidly induced in the rat by complete constriction of the abdominal aorta between the origins of the renal arteries. Carotid arterial systolic blood pressure remained unchanged in sham rats but increased rapidly in the pressure-overloaded constricted rats with a sustained hypertension established by 3 days. Hypertrophy of left ventricular (LV) cardiomyocytes in constricted rats also occurred by 3 days. However, this hypertrophy was preceded by increases in LV IGF-I mRNA and protein which occurred within 1 day. These results support the hypothesis that cardiac-synthesized IGF-I is an initiating or early-enhancing factor for hypertrophy of LV cardiomyocytes.     

Characterisation of the gene for Drosophila amphiphysin

A sequence similarity search of the Drosophila nucleotide database using vertebrate amphiphysin as a query identified a cDNA that encodes a Drosophila amphiphysin. The predicted protein has conserved sequence domains that should enable it to dimerise and bind to dynamin. Structural modelling suggests that the Src-homology-3 (SH3) domains of vertebrate and Drosophila amphiphysins are highly similar, supporting the putative ability of the latter to bind dynamin. However, the fly amphiphysin shows less conservation to sequences in the vertebrate amphiphysins that bind other endocytic components such as clathrin, AP-2 and endophilin. Amphiphysin is a single-copy gene that maps to position 49B on polytene chromosomes. Messenger RNA of this amphiphysin is expressed widely during embryogenesis and has elevated expression in a number of sites including the foregut, hindgut and epidermis, but not in the central nervous system. Taken together, these data are consistent with a role for Drosophila amphiphysin in endocytosis, but the details of this role may differ from that of vertebrate amphiphysins.     

The Use of Fluorogenic Substrates To Measure Fungal Presence and Activity in Soil

Our objective was to determine if 4-methylumbelliferyl-labelled enzyme substrates could be used to detect and quantify specific components of chitinase and cellulase activities as specific indicators of the presence and activity of fungal biomass. The fluorogenic substrates 4-methylumbelliferyl (MUF) N-acetyl-β-d-glucosaminide and MUF β-d-lactoside were used for the detection and quantification of β-N-acetylglucosaminidase (EC 3.2.1.30) (NAGase) and endo 1,4-β-glucanase (EC 3.2.1.4)/cellobiohydrolase (EC 3.2.1.91) (CELase), respectively. Culture screenings on solid media showed a widespread ability to produce NAGase among a taxonomically diverse selection of fungi on media with and without added chitin. NAGase activity was expressed only in a limited number of bacteria and on media supplemented with chitin. The CELase activity was observed only in a limited number of fungi and bacteria. Bacterial CELase activity was expressed on agar media containing a cellulose-derived substrate. In soil samples, NAGase activity was significantly correlated with estimates of fungal biomass, based on the content of two fungus-specific indicator molecules, 18:2ω6 phospholipid fatty acid (PLFA) and ergosterol. CELase activity was significantly correlated with the PLFA-based estimate of fungal biomass in the soil, but no correlation was found with ergosterol-based estimates of fungal biomass.The determination of enzyme activities is a simple approach to the study of microbially mediated processes within the soil environment. Thus, soil enzyme activities have been interpreted as indirect measures of microbial biomass, rhizosphere effects, soil productivity, and mineralization potential of naturally occurring substrates or xenobiotics (4). However, few studies have attempted to correlate soil enzyme activities with the presence and activities of specific components of the microbial community. The ability of soil-inhabiting fungi to produce a range of enzymes capable of degrading complex litter substances could make the use of an enzymatic approach to study soil fungal populations possible. These enzymes must be specific for fungal presence and activity. In one study of chitinase in soil (24), chitinase activity and the number of fungal propagules in chitin-amended soils were strongly correlated. The same correlation was not found for actinomycetes or bacteria. Thus, chitinase activity appears to be a suitable indicator of actively growing fungi in the soil. The hydrolysis of cellulose requires the interaction of a number of hydrolases produced by cellulolytic microorganisms. A major role is played by the cellulase system, which consists of several distinct enzymes that are produced by a large number of microorganisms, including fungi, actinomycetes, and bacteria. However, fungi have been suggested as the predominant source of β-d-glucosidase (EC 3.2.1.21) (16, 17) and endo 1,4-β-glucanase (EC 3.2.1.4) (23) activity in soils.Fluorogenic 4-methylumbelliferyl (MUF)-labelled enzyme substrates have been introduced for process-oriented studies in aquatic systems (3, 18) and, more recently, in peatlands (11). MUF substrates have been used to assay cell-bound activities in pure cultures of fungi, as the soluble substrate can enter the cell wall, making periplasmic enzyme activity detectable (15). These substrates have been used to detect fungal chitinolytic activities (17a) and cellulases (6) in vitro. The substrates may be added to environmental samples and, when hydrolyzed, release 4-methyl-umbelliferone (4-MU), which fluoresces and can be quantified in nanomolar concentrations (3).A variety of methods to quantify fungi in soil have been described. The techniques include direct microscopic observation and extraction of fungus-specific indicator molecules such as glucosamine or ergosterol (9). More recently, the phospholipid fatty acid (PLFA) 18:2ω6 has been proposed as an indicator of fungal biomass (7, 12). Our objectives in the present study were to determine if (i) components of chitinase and cellulase activities could be used as indicators of the presence and activity of fungal biomass and (ii) enzyme activities detected with specific MUF substrates in soil samples were correlated with the content of the fungus-specific indicator molecules 18:2ω6 PLFA and ergosterol.     

Modelling the Health Impact of an English Sugary Drinks Duty at National and Local Levels

Background

Increasing evidence associates excess refined sugar intakes with obesity, Type 2 diabetes and heart disease. Worryingly, the estimated volume of sugary drinks purchased in the UK has more than doubled between 1975 and 2007, from 510ml to 1140ml per person per week. We aimed to estimate the potential impact of a duty on sugar sweetened beverages (SSBs) at a local level in England, hypothesising that a duty could reduce obesity and related diseases.

Methods and Findings

We modelled the potential impact of a 20% sugary drinks duty on local authorities in England between 2010 and 2030. We synthesised data obtained from the British National Diet and Nutrition Survey (NDNS), drinks manufacturers, Office for National Statistics, and from previous studies. This produced a modelled population of 41 million adults in 326 lower tier local authorities in England. This analysis suggests that a 20% SSB duty could result in approximately 2,400 fewer diabetes cases, 1,700 fewer stroke and coronary heart disease cases, 400 fewer cancer cases, and gain some 41,000 Quality Adjusted Life Years (QALYs) per year across England. The duty might have the biggest impact in urban areas with young populations.

Conclusions

This study adds to the growing body of evidence suggesting health benefits for a duty on sugary drinks. It might also usefully provide results at an area level to inform local price interventions in England.     

Study of the effects of oral zinc supplementation on peroxynitrite levels, arginase activity and NO synthase activity in seminal plasma of Iraqi asthenospermic patients

Background

Low concentrations of nitric oxide (NO) are necessary for the biology and physiology of spermatozoa, but high levels of NO are toxic and have negative effects on sperm functions. Although several studies have considered the relationship between infertility and semen NO concentrations, no study on the effects of asthenospermia treatments such as oral zinc supplementation on concentrations of NO, which are important in fertility, has been reported. Studies have shown that oral zinc supplementation develops sperm count, motility and the physical characteristics of sperm in animals and in some groups of infertile men. The present study was conducted to study the effect of zinc supplementation on the quantitative and qualitative characteristics of semen, along with enzymes of the NO pathway in the seminal plasma of asthenospermic patients.

Methods

Semen samples were obtained from 60 fertile and 60 asthenozoospermic infertile men of matched age. The subfertile group was treated with zinc sulfate; each participant took two capsules (220 mg per capsule) per day for 3 months. Semen samples were obtained (before and after zinc sulfate supplementation). After liquefaction of the seminal fluid at room temperature, routine semen analyses were performed. The stable metabolites of NO (nitrite) in seminal plasma were measured by nitrophenol assay. Arginase activity and NO synthase activity were measured spectrophotometrically.

Results

Peroxynitrite levels, arginase activity, NO synthase activity and various sperm parameters were compared among fertile controls and infertile patients (before and after treatment with zinc sulfate). Peroxynitrite levels and NO synthase activity were significantly higher in the infertile patients compared to the fertile group. Conversely, arginase activity was significantly higher in the fertile group than the infertile patients. Peroxynitrite levels, arginase activity and NO synthase activity of the infertile patient were restored to normal values after treatment with zinc sulfate. Volume of semen, progressive sperm motility percentage and total normal sperm count were increased after zinc supplementation.

Conclusions

Treatment of asthenospermic patients with zinc supplementation leads to restored peroxynitrite levels, arginase activity and NO synthase activity to normal values and gives a statistically significant improvement of semen parameters compared with controls.

Trial registration

ClinicalTrials.gov identifier: NCT01684059     

Chronic inhibition of the mitochondrial ATP synthase in skeletal muscle triggers sarcoplasmic reticulum distress and tubular aggregates

Tubular aggregates (TA) are honeycomb-like arrays of sarcoplasmic-reticulum (SR) tubules affecting aged glycolytic fibers of male individuals and inducing severe sarcomere disorganization and muscular pain. TA develop in skeletal muscle from Tubular Aggregate Myopathy (TAM) patients as well as in other disorders including endocrine syndromes, diabetes, and ageing, being their primary cause unknown. Nowadays, there is no cure for TA. Intriguingly, both hypoxia and calcium dyshomeostasis prompt TA formation, pointing to a possible role for mitochondria in their setting. However, a functional link between mitochondrial dysfunctions and TA remains unknown. Herein, we investigate the alteration in muscle-proteome of TAM patients, the molecular mechanism of TA onset and a potential therapy in a preclinical mouse model of the disease. We show that in vivo chronic inhibition of the mitochondrial ATP synthase in muscle causes TA. Upon long-term restrained oxidative phosphorylation (OXPHOS), oxidative soleus experiments a metabolic and structural switch towards glycolytic fibers, increases mitochondrial fission, and activates mitophagy to recycle damaged mitochondria. TA result from the overresponse of the fission controller DRP1, that upregulates the Store-Operate-Calcium-Entry and increases the mitochondria-SR interaction in a futile attempt to buffer calcium overloads upon prolonged OXPHOS inhibition. Accordingly, hypoxic muscles cultured ex vivo show an increase in mitochondria/SR contact sites and autophagic/mitophagic zones, where TA clusters grow around defective mitochondria. Moreover, hypoxia triggered a stronger TA formation upon ATP synthase inhibition, and this effect was reduced by the DRP1 inhibitor mDIVI. Remarkably, the muscle proteome of TAM patients displays similar alterations in mitochondrial dynamics and in ATP synthase contents. In vivo edaravone treatment in mice with restrained OXPHOS restored a healthy phenotype by prompting mitogenesis and mitochondrial fusion. Altogether, our data provide a functional link between the ATP synthase/DRP1 axis and the setting of TA, and repurpose edaravone as a possible treatment for TA-associated disorders.Subject terms: Musculoskeletal abnormalities, Energy metabolism     

Petri Net and Probabilistic Model Checking Based Approach for the Modelling,Simulation and Verification of Internet Worm Propagation

Internet worms are analogous to biological viruses since they can infect a host and have the ability to propagate through a chosen medium. To prevent the spread of a worm or to grasp how to regulate a prevailing worm, compartmental models are commonly used as a means to examine and understand the patterns and mechanisms of a worm spread. However, one of the greatest challenge is to produce methods to verify and validate the behavioural properties of a compartmental model. This is why in this study we suggest a framework based on Petri Nets and Model Checking through which we can meticulously examine and validate these models. We investigate Susceptible-Exposed-Infectious-Recovered (SEIR) model and propose a new model Susceptible-Exposed-Infectious-Recovered-Delayed-Quarantined (Susceptible/Recovered) (SEIDQR(S/I)) along with hybrid quarantine strategy, which is then constructed and analysed using Stochastic Petri Nets and Continuous Time Markov Chain. The analysis shows that the hybrid quarantine strategy is extremely effective in reducing the risk of propagating the worm. Through Model Checking, we gained insight into the functionality of compartmental models. Model Checking results validate simulation ones well, which fully support the proposed framework.     

Effect of Oral Zinc Supplementation on the Thiol Oxido-Reductive Index and Thiol-Related Enzymes in Seminal Plasma and Spermatozoa of Iraqi Asthenospermic Patients

Despite increasing evidence indicating the essential involvement of selenium (Se) on growth performance, antioxidant capacity, and meat quality of commercial broilers, the effects of different Se sources on local Chinese Subei chickens is unclear. A total of 360 50-day-old male chickens were individually weighed and randomly allocated to four treatment groups. Chickens in each of the four groups were fed diets supplemented with 0.3 mg Se/kg as sodium Se (SS), Se-enriched yeast (SY), selenomethionine (Met-Se), or nano red element Se (Nano-Se) for 40 days. At the end of the experiment, one bird of approximately average weight from each cage was selected and slaughtered, and blood and breast muscles samples were collected. The results showed that there was no significant difference in feed intake, body weight gain, or feed to gain ratio among treatments (P > 0.05). Dietary SY, Met-Se, and Nano-Se supplementation increased the activity of glutathione peroxidase in serum and breast muscles and decreased the concentration of malondialdehyde in serum and carbonyl in breast muscles compared with the SS group (P < 0.05). Moreover, SY, Met-Se, and Nano-Se supplementation increased pH_45min, total protein solubility, and myofibrillar protein solubility, as well as decreased the shear force value compared with the SS group (P < 0.05). In addition, birds in the SY and Met-Se groups exhibited lower cooking loss compared with the SS group (P < 0.05). In conclusion, organic Se and Nano-Se supplementation resulted in an improvement of antioxidant capacity and meat quality in local Chinese Subei chickens relative to inorganic Se.     

A Simple Procedure to Detect a dsRNA Associated with Groundnut Rosette

A low-molecular weight double-stranded (ds) RNA [900 base pairs (bp)] associated with groundnut rosette disease can be used as a diagnostic tool. A simple procedure has been developed that is rapid, reliable, and requires only standard electrophoresis equipment and ultraviolet light for detection of nucleic acid bands. Using this procedure, the dsRNA was detected only in groundnut plants with green rosette of chlorotic rosette symptoms. It was not found in uninoculated groundnut plants, in symptomless groundnut plants with groundnut rosette assistor virus, or in groundnut plants infected with several other known groundnut viruses. In studies with northern blots of extracts from rosette-diseased and healthy plants, 5′-endlabeled dsRNA only hybridized to a 900 bp dsRNA from diseased plants. The 900 bp dsRNA was not infectious and its origin remains obscure.