Cloning and expression analysis of two distinct HIF-alpha isoforms – gcHIF-1alpha and gcHIF-4alpha – from the hypoxia-tolerant grass carp,Ctenopharyngodon idellus

Background  

Hypoxia-inducible factors (HIFs) are involved in adaptive and survival responses to hypoxic stress in mammals. In fish, very little is known about the functions of HIFs.     

Strain-dependent Recovery Behavior of Single Chondrocytes

One of the challenges facing researchers studying chondrocyte mechanobiology is determining the range of mechanical forces pertinent to the problems they study. One possible way to deal with this problem is to quantify how the biomechanical behavior of cells varies in response to changing mechanical forces. In this study, the compressibility and recovery behaviors of single chondrocytes were determined as a function of compressive strains from 6 to 63%. Bovine articular chondrocytes from the middle and deep zones were subjected to this range of strains, and digital videocapture was used to track changes in cell dimensions during and after compression. The normalized volume change, apparent Poisson’s ratio, residual strain after recovery, cell volume fraction after recovery, and characteristic recovery time constant were analyzed with respect to axial strain. Normalized volume change varied as a function of strain, demonstrating that chondrocytes exhibited compressibility. The mean Poisson’s ratio of chondrocytes was found to be 0.29 ± 0.14, and did not vary with axial strain. In contrast, residual strain, recovered volume fraction, and recovery time constant all depended on axial strain. The dependence of residual strain and recovered volume fraction on axial strain showed a change in behavior around 25–30% strain, opening up the possibility that this range of strains represents a critical value for chondrocytes. Quantifying the mechanical behavior of cells as a function of stress and strain is a potentially useful approach for identifying levels of mechanical stimulation that may be germane to normal cartilage physiology, functional tissue engineering of cartilage, and the etiopathogenesis of osteoarthritis.     

Effect of sucrose and methyl jasmonate on biomass and anthocyanin production in cell suspension culture of Melastoma malabathricum (Melastomaceae)

Melastoma malabathricum, belongs to the Melastomaceae family, is an important medicinal plant widely distributed from Madagascar to Australia, that is used in traditional remedies for the treatment of various ailments. Besides its medicinal properties, it has been identified as a potential source of anthocyanin production. The present study was carried out to investigate the effect of sucrose and methyl jasmonate and feeding time on cell biomass yield and anthocyanin production in cell suspension culture of M. malabathricum. Addition of different concentrations of sucrose into the cell culture of M. malabathricum influenced cell biomass and pigment accumulation. The addition of methyl jasmonate was found to have no effect on cell biomass but the presence of higher amount (12.5-50 mg/L) had caused a reduction in anthocyanin production and accumulation. MS medium supplemented with 30 g/L sucrose and 3.5 mg/L of MeJA added on cero day and 3rd day produced high fresh cell mass at the end of nine days of culture but did not support the production of anthocyanins. However, cells cultured in the medium supplemented with 45 g/L sucrose without MeJA showed the highest pigment content (0.69 +/- 0.22 CV/g-FCM). The cells cultured in MS medium supplemented with 30 g/L sucrose with 3.5 mg/L MeJA added on the 3rd and 6th day of culture, showed the lowest pigment content (0.37-0.40 CV/g-FCM). This study indicated that MeJA was not necessary but sucrose was needed for the enhancement of cell growth and anthocyanin production in M. malabathricum cell cultures.     

Lack of association between Gly82Ser, 1704G/T and 2184A/G of RAGE gene polymorphisms and retinopathy susceptibility in Malaysian diabetic patients

Diabetic retinopathy is the most common diabetic eye disease, occurring in about 60% of type 2 diabetic patients. Other than known clinical risk factors, the influence of genes has been suggested as part of the development of diabetic retinopathy. We investigated the association of Gly82Ser, 1704G/T and 2184A/G polymorphisms in the RAGE gene with retinopathy in type 2 diabetic patients in Malaysia. Ninety-eight unrelated retinopathy patients and 185 unrelated healthy controls from all over Malaysia were recruited in this study. The allele and genotype frequencies of the three gene polymorphisms were investigated using PCR-RFLP. The allele frequency of the three polymorphisms did not differ significantly between the control and the retinopathy group (P > 0.05). Analysis of the frequency of GA+AA, GT+TT and AG+GG in the retinopathy group did not reveal significant differences (P > 0.05) compared to the control group. We conclude that RAGE gene Gly82Ser, 1704G/T and 2184A/G polymorphisms are not associated with retinopathy development in the Malaysian population.     

Oligosaccharide recognition and binding to the carbohydrate binding module of AMP-activated protein kinase

The AMP-activated protein kinase (AMPK) contains a carbohydrate-binding module (beta1-CBM) that is conserved from yeast to mammals. Beta1-CBM has been shown to localize AMPK to glycogen in intact cells and in vitro. Here we use Nuclear Magnetic Resonance spectroscopy to investigate oligosaccharide binding to 15N labelled beta1-CBM. We find that beta1-CBM shows greatest affinity to carbohydrates of greater than five glucose units joined via alpha,1-->4 glycosidic linkages with a single, but not multiple, glucose units in an alpha,1-->6 branch. The near identical chemical shift profile for all oligosaccharides whether cyclic or linear suggest a similar binding conformation and confirms the presence of a single carbohydrate-binding site.     

Increasement of Heterologous Expression of Recombinant Vit v 1 in Pichia pastoris KM71 by Nonionic Detergents as a Cost-effective Approach

Applied Biochemistry and Microbiology - Vit v 1 as a lipid-transfer protein is a major allergen of grapes (Vitis vinifera) that elicits food allergy in many patients in Iran. Todays, recombinant...     

Molecular Surveillance of Antiviral Drug Resistance of Influenza A/H3N2 Virus in Singapore, 2009-2013

Adamantanes and neuraminidase inhibitors (NAIs) are two classes of antiviral drugs available for the chemoprophylaxis and treatment of influenza infections. To determine the frequency of drug resistance in influenza A/H3N2 viruses in Singapore, large-scale sequencing of neuraminidase (NA) and matrix protein (MP) genes was performed directly without initial culture amplification. 241 laboratory-confirmed influenza A/H3N2 clinical samples, collected between May 2009 and November 2013 were included. In total, 229 NA (95%) and 241 MP (100%) complete sequences were obtained. Drug resistance mutations in the NA and MP genes were interpreted according to published studies. For the NAIs, a visual inspection of the aligned NA sequences revealed no known drug resistant genotypes (DRGs). For the adamantanes, the well-recognised S31N DRG was identified in all 241 MP genes. In addition, there was an increasing number of viruses carrying the combination of D93G+Y155F+D251V (since May 2013) or D93G (since March 2011) mutations in the NA gene. However, in-vitro NAI testing indicated that neither D93G+Y155F+D251V nor D93G alone conferred any changes in NAI susceptibility. Lastly, an I222T mutation in the NA gene that has previously been reported to cause oseltamivir-resistance in influenza A/H1N1/2009, B, and A/H5N1, was detected from a treatment-naïve patient. Further in-vitro NAI testing is required to confirm the effect of this mutation in A/H3N2 virus.     

Mechanosensitive axon outgrowth mediated by L1-laminin clutch interface

Mechanical properties of the extracellular environment modulate axon outgrowth. Growth cones at the tip of extending axons generate traction force for axon outgrowth by transmitting the force of actin filament retrograde flow, produced by actomyosin contraction and F-actin polymerization, to adhesive substrates through clutch and cell adhesion molecules. A molecular clutch between the actin filament flow and substrate is proposed to contribute to cellular mechanosensing. However, the molecular identity of the clutch interface responsible for mechanosensitive growth cone advance is unknown. We previously reported that mechanical coupling between actin filament retrograde flow and adhesive substrates through the clutch molecule shootin1a and the cell adhesion molecule L1 generates traction force for axon outgrowth and guidance. Here, we show that cultured mouse hippocampal neurons extend longer axons on stiffer substrates under elastic conditions that correspond to the soft brain environments. We demonstrate that this stiffness-dependent axon outgrowth requires actin-adhesion coupling mediated by shootin1a, L1, and laminin on the substrate. Speckle imaging analyses showed that L1 at the growth cone membrane switches between two adhesive states: L1 that is immobilized and that undergoes retrograde movement on the substrate. The duration of the immobilized phase was longer on stiffer substrates; this was accompanied by increases in actin-adhesion coupling and in the traction force exerted on the substrate. These data suggest that the interaction between L1 and laminin is enhanced on stiffer substrates, thereby promoting force generation for axon outgrowth.     

A Pilot Study to Examine the Correlation between Cognition and Blood Biomarkers in a Singapore Chinese Male Cohort with Type 2 Diabetes Mellitus

Background

Diabetes is reported to be linked to poorer cognitive function. The purpose of this study is to examine (a) clinical correlation between cognitive function and the biochemical perturbations in T2DM, and (b) the impact of statin treatment on cognitive function in diabetic subjects.

Methods

Forty Singaporean Chinese males with diabetes and twenty Singaporean Chinese males without diabetes were recruited for this study. Twenty-two of the diabetic subjects were on statin therapy and all subjects were non-demented. This was a 2-period non-interventional case-control study in which subjects were assessed for cognitive function in period 1 and blood samples taken over 2 periods, approximately 1 week apart. Blood was collected to determine the level of total cholesterol, high-density lipoprotein (HDL), low-density lipoprotein (LDL), triglycerides, glucose and insulin. Cognitive performance was measured by a neuropsychological battery covering domains of attention, language, verbal and visual memory, visuomotor speed and executive function. Z-scores were derived for each cognitive domain using the mean and standard deviations (SDs), and they were used to compare between (a) diabetic and non-diabetic groups, and (b) diabetic subjects with and without statin treatment. ANCOVAs with age, education, BMI, and the duration of diabetes as covariates were employed to examine differences in mean score of cognitive domains and subtests between the two groups.

Results

Overall cognitive function was similar among diabetics and age matched non-diabetic controls. Among diabetic statin users, HDL, LDL and total cholesterol were negatively correlated with executive function, whereas peripheral insulin levels and insulin resistance were negatively associated with attention.

Conclusion

Diabetic statin users were likely to have poorer performance in attention and executive function. Increasing levels of the peripheral biomarkers are likely to contribute to poorer cognitive performance.