POTENTIAL OF THE AQUATIC FERN AZOLLA FILICULOIDES IN BIODEGRADATION OF AN AZO DYE: MODELING OF EXPERIMENTAL RESULTS BY ARTIFICIAL NEURAL NETWORKS

The potential of an aquatic fern, Azolla filiculoides, in phytoremediation of a mono azo dye solution, C.I. Acid Blue 92 (AB92), was studied. The effects of operational parameters such as reaction time, initial dye concentration, fern fresh weight, pH, temperature and reusability of the fern on biodegradation efficiency were investigated. The intermediate compounds produced by biodegradation process were analyzed using GC–MS analysis. An artificial neural network (ANN) model was developed to predict the biodegradation efficiency. The findings indicated that ANN provides reasonable predictive performance (R² = 0.961). The effects of AB92 solutions (10 and 20 mg L^?1) on growth, chlorophylls and carotenoids content, activity of antioxidant enzymes such as superoxide dismutase, peroxidase and catalase and formation of malondialdehyde were analyzed. AB92 generally showed inhibitory effects on the growth. Moreover, photosynthetic pigments in the fronds significantly decreased in the treatments. An increase was detected for lipid peroxidation and antioxidant enzymes activity, suggesting that AB92 caused reactive oxygen species production in Azolla fronds, which were scavenged by induced activities of antioxidant enzymes.     

Structure,composition and regeneration of riparian forest along an altitudinal gradient in northern Iran

In order to protect and understand the regeneration of riparian forests, it is important to understand the environmental conditions that lead to their vegetation differentiation. We evaluated the structure, composition, density and regeneration of woody species in forests along the river Safaroud in Ramsar forest in northern Iran in relation to elevation, soil properties and distance from the river. Using 60 transects located perpendicularly to river and 300 quadrats, we examined forests 0, 50, 100, 150 and 200 m from the stream along an elevation gradient spanning from 350 to 2,400 m a.s.l. We found that total density, regeneration, diameter and basal area of trees were significantly higher in the interior of the forest whereas shrub density was higher close to the river. The uneven-aged forest structure showed no significant differences among gradient from the river to the forest interior. Hydro-geomorphic processes, flooding, the elevation gradient, distance from the river and soil properties were the most important factors that influenced plant community distribution in relation to the river.     

A fast inverse dynamics model of walking for use in optimisation studies

Computer simulation of human gait, based on measured motion data, is a well-established technique in biomechanics. However, optimisation studies requiring many iterative gait cycle simulations have not yet found widespread application because of their high computational cost. Therefore, a computationally efficient inverse dynamics model of 3D human gait has been designed and compared with an equivalent model, created using a commercial multi-body dynamics package. The fast inverse dynamics model described in this paper led to an eight fold increase in execution speed. Sufficient detail is provided to allow readers to implement the model themselves.     

Multiplex PCR and Next Generation Sequencing for the Non-Invasive Detection of Bladder Cancer

Background

Highly sensitive and specific urine-based tests to detect either primary or recurrent bladder cancer have proved elusive to date. Our ever increasing knowledge of the genomic aberrations in bladder cancer should enable the development of such tests based on urinary DNA.

Methods

DNA was extracted from urine cell pellets and PCR used to amplify the regions of the TERT promoter and coding regions of FGFR3, PIK3CA, TP53, HRAS, KDM6A and RXRA which are frequently mutated in bladder cancer. The PCR products were barcoded, pooled and paired-end 2 x 250 bp sequencing performed on an Illumina MiSeq. Urinary DNA was analysed from 20 non-cancer controls, 120 primary bladder cancer patients (41 pTa, 40 pT1, 39 pT2+) and 91 bladder cancer patients post-TURBT (89 cancer-free).

Results

Despite the small quantities of DNA extracted from some urine cell pellets, 96% of the samples yielded mean read depths >500. Analysing only previously reported point mutations, TERT mutations were found in 55% of patients with bladder cancer (independent of stage), FGFR3 mutations in 30% of patients with bladder cancer, PIK3CA in 14% and TP53 mutations in 12% of patients with bladder cancer. Overall, these previously reported bladder cancer mutations were detected in 86 out of 122 bladder cancer patients (70% sensitivity) and in only 3 out of 109 patients with no detectable bladder cancer (97% specificity).

Conclusion

This simple, cost-effective approach could be used for the non-invasive surveillance of patients with non-muscle-invasive bladder cancers harbouring these mutations. The method has a low DNA input requirement and can detect low levels of mutant DNA in a large excess of normal DNA. These genes represent a minimal biomarker panel to which extra markers could be added to develop a highly sensitive diagnostic test for bladder cancer.     

Metformin Ameliorates Dysfunctional Traits of Glibenclamide- and Glucose-Induced Insulin Secretion by Suppression of Imposed Overactivity of the Islet Nitric Oxide Synthase-NO System

Metformin lowers diabetic blood glucose primarily by reducing hepatic gluconeogenesis and increasing peripheral glucose uptake. However, possible effects by metformin on beta-cell function are incompletely understood. We speculated that metformin might positively influence insulin secretion through impacting the beta-cell nitric oxide synthase (NOS)-NO system, a negative modulator of glucose-stimulated insulin release. In short-time incubations with isolated murine islets either glibenclamide or high glucose augmented insulin release associated with increased NO production from both neural and inducible NOS. Metformin addition suppressed the augmented NO generation coinciding with amplified insulin release. Islet culturing with glibenclamide or high glucose revealed pronounced fluorescence of inducible NOS in the beta-cells being abolished by metformin co-culturing. These findings were reflected in medium nitrite-nitrate levels. A glucose challenge following islet culturing with glibenclamide or high glucose revealed markedly impaired insulin response. Metformin co-culturing restored this response. Culturing murine islets and human islets from controls and type 2 diabetics with high glucose or high glucose + glibenclamide induced a pronounced decrease of cell viability being remarkably restored by metformin co-culturing. We show here, that imposed overactivity of the beta-cell NOS-NO system by glibenclamide or high glucose leads to insulin secretory dysfunction and reduced cell viability and also, importantly, that these effects are relieved by metformin inhibiting beta-cell NO overproduction from both neural and inducible NOS thus ameliorating a concealed negative influence by NO induced by sulfonylurea treatment and/or high glucose levels. This double-edged effect of glibenclamide on the beta-cellsuggests sulfonylurea monotherapy in type 2 diabetes being avoided.     

Long-term infusion of nutrients (total parenteral nutrition) suppresses circulating ghrelin in food-deprived rats

BACKGROUND: Ghrelin derives from endocrine cells (A-like cells) in the stomach (mainly the oxyntic mucosa). Its concentration in the circulation increases during fasting and decreases upon re-feeding. This has fostered the notion that the absence of food in the upper gastrointestinal (GI) tract stimulates the secretion of ghrelin. The purpose of the present study was to determine the concentration of ghrelin in serum and oxyntic mucosa after replacing food with intravenous (iv) infusion of nutrients for 8 days using the technique known as total parenteral nutrition (TPN) MATERIALS AND METHODS: Male Sprague-Dawley rats (200-250 g) were given nutrients (lipids, glucose, amino acids, minerals and vitamins) by iv infusion for 8 days during which time they were deprived of food and water; another group was deprived of food for 24-48 h (fasted controls), while fed controls had free access to food and water. Serum ghrelin, gastrin and pancreastatin concentrations were measured together with the ghrelin content of the oxyntic mucosa. Plasma insulin and glucose as well as serum lipid concentrations were also determined. RESULTS: Fasted rats had higher serum ghrelin than TPN rats and fed controls. The oxyntic mucosal ghrelin concentration (and content) was lower in TPN rats than in fasted rats or fed controls. The serum gastrin and pancreastatin concentrations were lower in TPN rats and fasted rats than in fed controls. The plasma insulin concentration was 87 pmol/l+/-8 (SEM) in TPN rats compared to 101+/-16 pmol/l in fed controls; it was 26+/-14 pmol/l in fasted rats. The basal plasma glucose level was 11+/-0.6 mmol/l in TPN rats and 12+/-0.8 mmol/l in fed controls; it was 7+/-0.3 mmol/l in fasted rats. In TPN rats, the serum concentrations of free fatty acids, triglycerides and cholesterol were increased by 100%, 50% and 25%, respectively, compared to fed controls. Fasted rats had higher circulating concentrations of free fatty acids (20%) and lower concentrations of triglycerides (-40%) than fed controls; fasted rats did not differ from fed controls with respect to serum cholesterol. CONCLUSION: The circulating ghrelin concentration is high in situations of nutritional deficiency (starvation) and low in situations of nutritional plenty (free access to food or TPN). The actual presence or absence of food in the GI tract seems irrelevant. Circulating insulin and glucose concentrations did not differ much between TPN rats and fed controls; serum lipids, however, were elevated in the TPN rats. We suggest that elevated blood lipid levels contribute to the suppression of circulating ghrelin in rats subjected to TPN for 8 days.     

Palmitate-Induced β-Cell Dysfunction Is Associated with Excessive NO Production and Is Reversed by Thiazolidinedione-Mediated Inhibition of GPR40 Transduction Mechanisms

Background

Type 2 diabetes often displays hyperlipidemia. We examined palmitate effects on pancreatic islet function in relation to FFA receptor GPR40, NO generation, insulin release, and the PPARγ agonistic thiazolidinedione, rosiglitazone.

Principal Findings

Rosiglitazone suppressed acute palmitate-stimulated GPR40-transduced PI hydrolysis in HEK293 cells and insulin release from MIN6c cells and mouse islets. Culturing islets 24 h with palmitate at 5 mmol/l glucose induced β-cell iNOS expression as revealed by confocal microscopy and increased the activities of ncNOS and iNOS associated with suppression of glucose-stimulated insulin response. Rosiglitazone reversed these effects. The expression of iNOS after high-glucose culturing was unaffected by rosiglitazone. Downregulation of GPR40 by antisense treatment abrogated GPR40 expression and suppressed palmitate-induced iNOS activity and insulin release.

Conclusion

We conclude that, in addition to mediating acute FFA-stimulated insulin release, GPR40 is an important regulator of iNOS expression and dysfunctional insulin release during long-term exposure to FFA. The adverse effects of palmitate were counteracted by rosiglitazone at GPR40, suggesting that thiazolidinediones are beneficial for β-cell function in hyperlipidemic type 2 diabetes.     

A KATP Channel-Dependent Pathway within α Cells Regulates Glucagon Release from Both Rodent and Human Islets of Langerhans

Glucagon, secreted from pancreatic islet α cells, stimulates gluconeogenesis and liver glycogen breakdown. The mechanism regulating glucagon release is debated, and variously attributed to neuronal control, paracrine control by neighbouring β cells, or to an intrinsic glucose sensing by the α cells themselves. We examined hormone secretion and Ca²⁺ responses of α and β cells within intact rodent and human islets. Glucose-dependent suppression of glucagon release persisted when paracrine GABA or Zn²⁺ signalling was blocked, but was reversed by low concentrations (1–20 μM) of the ATP-sensitive K⁺ (K_ATP) channel opener diazoxide, which had no effect on insulin release or β cell responses. This effect was prevented by the K_ATP channel blocker tolbutamide (100 μM). Higher diazoxide concentrations (≥30 μM) decreased glucagon and insulin secretion, and α- and β-cell Ca²⁺ responses, in parallel. In the absence of glucose, tolbutamide at low concentrations (<1 μM) stimulated glucagon secretion, whereas high concentrations (>10 μM) were inhibitory. In the presence of a maximally inhibitory concentration of tolbutamide (0.5 mM), glucose had no additional suppressive effect. Downstream of the K_ATP channel, inhibition of voltage-gated Na⁺ (TTX) and N-type Ca²⁺ channels (ω-conotoxin), but not L-type Ca²⁺ channels (nifedipine), prevented glucagon secretion. Both the N-type Ca²⁺ channels and α-cell exocytosis were inactivated at depolarised membrane potentials. Rodent and human glucagon secretion is regulated by an α-cell K_ATP channel-dependent mechanism. We propose that elevated glucose reduces electrical activity and exocytosis via depolarisation-induced inactivation of ion channels involved in action potential firing and secretion.