Thymoquinone Prevents β-Amyloid Neurotoxicity in Primary Cultured Cerebellar Granule Neurons

Thymoquinone (TQ), a bioactive constituent of Nigella sativa Linn (N. sativa) has demonstrated several neuropharmacological attributes. In the present study, the neuroprotective properties of TQ were investigated by studying its anti-apoptotic potential to diminish β-amyloid peptide 1–40 sequence (Aβ_1–40)-induced neuronal cell death in primary cultured cerebellar granule neurons (CGNs). The effects of TQ against Aβ_1–40-induced neurotoxicity, morphological damages, DNA condensation, the generation of reactive oxygen species, and caspase-3, -8, and -9 activation were investigated. Pretreatment of CGNs with TQ (0.1 and 1 μM) and subsequent exposure to 10 μM Aβ_1–40 protected the CGNs against the neurotoxic effects of the latter. In addition, the CGNs were better preserved with intact cell bodies, extensive neurite networks, a loss of condensed chromatin and less free radical generation than those exposed to Aβ_1–40 alone. TQ pretreatment inhibited Aβ_1–40-induced apoptosis of CGNs via both extrinsic and intrinsic caspase pathways. Thus, the findings of this study suggest that TQ may prevent neurotoxicity and Aβ_1–40-induced apoptosis. TQ is, therefore, worth studying further for its potential to reduce the risks of developing Alzheimer’s disease.     

Scan-Rescan Reproducibility of High Resolution Magnetic Resonance Imaging of Atherosclerotic Plaque in the Middle Cerebral Artery

Purpose

To evaluate the scan-rescan reproducibility of high-resolution magnetic resonance imaging (MRI) of middle cerebral artery (MCA) plaque, and calculate the number of subjects needed for future longitudinal clinical studies.

Material and Methods

Twenty two patients with MCA plaque were scanned twice by a T₂-weighted fast-spin-echo sequence at 3T. Areas and volumes of MCA lumen, total vessel and plaque were quantified and compared between two repeated scans. Agreement and measurement error was quantified by intraclass correlation coefficient (ICC) and coefficient of variance (CV) as defined by standard deviation (SD) of pair wise difference / mean. Sample size needed to detect 5% to 20% changes in area/volume was calculated using 80% power and 5% significance level.

Results

There was no significant different between the area and volume measurements of two repeated scans (p>0.05) with good agreement (ICC range 0.97–0.98 for area and 0.99 for volume). Relatively small measurement errors were observed with CVs range 6.1%-11.8% for area quantification and 4.9%-8.0% for volume quantification. Volume measurements tended to have 19.7% to 32.2% smaller CVs compared with area measurements. Sample size calculation showed a group of 47 patients was sufficient to detect 5% to 10% changes in MCA area/volume.

Conclusion

High resolution MRI is feasible for quantifying intracranial plaque area and volume in longitudinal clinical studies with low scan-rescan variability. Volume measurement tends to be more reproducible compared with area measurements.     

Effect of feeding frequency as a rearing system on biological performance,survival, body chemical composition and economic efficiency of Asian seabass Lates calcarifer (Bloch, 1790) reared under controlled environmental conditions

A 70-day rearing trial was done to determine the optimal frequency of feeding on growth performance (GP), feed conversion rate (FCR), cannibalism, survival rate (SR), body chemical composition and economic efficiency of the Asian sea bass. This study tested four different treatments of feeding frequencies (FF), once (T1), twice (T2), three times (T3), and four times (T4) per day. An average initial weight of Asian sea bass fry was 0.2 g (SD = ±0.12) were stocked 10 individuals per m³ (9.14 m × 1.82 m × 1.22 m, L × W × H; water depth 0.61 m) with two replicates per treatment (4 × 2 = 8). Fry were fed a mixture of larval commercial feed and shrimp with a pellet diet containing (46% CP). Initially, the feeding rate of 8% biomass per day was further adjusted according to fish biomass on a weekly basis. Results showed that, the FF significantly affected (p < 0.05) on growth indictors and survival rate (SR). Specifically fry fed three times a day (T3) had the best FBW, FL, SGR, ADWG and FCR followed by T4 and T2 while fry fed one time a day was the lowest in these parameters. Also, VSI, HSI and CF (k) significantly differed among the treatments. The fish whole body content of protein, moisture and ash did not significantly (p < 0.05) be affected by feeding frequency, but lipid content differed and both T3, T4 were the highest. It could be concluded that, increasing FF up to three times a day had a positive effect on weight gain, survival rate and feed utilization of Lates calcarifer. The second degree polynomial regression indicates that fed three times a day is optimum for best growth performance and survival for Asian sea bass.     

Increased adipose protein carbonylation in human obesity

Insulin resistance is associated with obesity but mechanisms controlling this relationship in humans are not fully understood. Studies in animal models suggest a linkage between adipose reactive oxygen species (ROS) and insulin resistance. ROS oxidize cellular lipids to produce a variety of lipid hydroperoxides that in turn generate reactive lipid aldehydes that covalently modify cellular proteins in a process termed carbonylation. Mammalian cells defend against reactive lipid aldehydes and protein carbonylation by glutathionylation using glutathione-S-transferase A4 (GSTA4) or carbonyl reduction/oxidation via reductases and/or dehydrogenases. Insulin resistance in mice is linked to ROS production and increased level of protein carbonylation, mitochondrial dysfunction, decreased insulin-stimulated glucose transport, and altered adipokine secretion. To assess protein carbonylation and insulin resistance in humans, eight healthy participants underwent subcutaneous fat biopsy from the periumbilical region for protein analysis and frequently sampled intravenous glucose tolerance testing to measure insulin sensitivity. Soluble proteins from adipose tissue were analyzed using two-dimensional gel electrophoresis and the major carbonylated proteins identified as the adipocyte and epithelial fatty acid-binding proteins. The level of protein carbonylation was directly correlated with adiposity and serum free fatty acids (FFAs). These results suggest that in human obesity oxidative stress is linked to protein carbonylation and such events may contribute to the development of insulin resistance.     

Inhibition of beta-amylase activity by calcium, magnesium and zinc ions determined by spectrophotometry and isothermal titration calorimetry

The inhibition effect of metal ions on beta amylase activity was studied. The inhibitor-binding constant (Ki) was determined by spectrophotometric and isothermal titration calorimetric (ITC) methods. The binding of calcium, magnesium and zinc ion as inhibitors at the active site of barley beta amylase was studied at pH = 4.8 (sodium acetate 16 mM) and T = 300K. The Ki and enthalpy of binding for calcium (13.4, 13.1 mM and -14.3 kJ/mol), magnesium (18.6, 17.8mM and -17.7 kJ/mol) and zinc (17.5, 17.7 mM and -20.0 kJ/mol) were found by spectrophotometric and ITC methods respectively.     

Effects of sediment deposition on the seaweed Sargassum on a fringing coral reef

 Coral reef degradation may involve shifts from coral to algal dominance and may be caused in part by increased sediment loads. Inshore fringing reef flats in the central Great Barrier Reef region are often subjected to periods of high sedimentation and are often dominated by macroalgae such as Sargassum. Experiments reported here examine the impacts of sediments on the recruitment, growth, survival, degeneration and vegetative regeneration of Sargassum microphyllum on a fringing coral reef flat in the central Great Barrier Reef. Comparison of three levels of sediment deposition (experimental addition, control (ambient condition) and experimental removal) showed that increased amounts of sediment significantly decreased rates of recruitment, growth, survival and vegetative regeneration, but not degeneration of S. microphyllum. In addition, the regenerative ability of S. microphyllum thalli with short, persistent erect branches (untreated) was compared with that of thalli experimentally cut back to the holdfast. This experimental damage significantly reduced regeneration. Accepted: 6 October 1997     

A case of amniotic membrane transplantation in non-healing Nocardia asteroides keratitis

A patient with severe non-healing microscopically proven Nocardia keratitis was treated with maximal topical amikacin followed by amniotic membrane transplantation (AMT) 2 months later (single-layer epithelial side-down). Epithelial healing was achieved, but neovascularization continued. This case report indicates that AMT combined with topical antibiotic provides pain relief and allows epithelial healing in severe Nocardia keratitis.     

Oxidative Stress Inhibits Nuclear Protein Export by Multiple Mechanisms That Target FG Nucleoporins and Crm1

Nuclear transport of macromolecules is regulated by the physiological state of the cell and thus sensitive to stress. To define the molecular mechanisms that control nuclear export upon stress, cells were exposed to nonlethal concentrations of the oxidant diethyl maleate (DEM). These stress conditions inhibited chromosome region maintenance-1 (Crm1)-dependent nuclear export and increased the association between Crm1 and Ran. In addition, we identified several repeat-containing nucleoporins implicated in nuclear export as targets of oxidative stress. As such, DEM treatment reduced Nup358 levels at the nuclear envelope and redistributed Nup98. Furthermore, oxidative stress led to an increase in the apparent molecular masses of Nup98, Nup214, and Nup62. Incubation with phosphatase or β-N-acetyl-hexosaminidase showed that oxidative stress caused the phosphorylation of Nup98, Nup62, and Nup214 as well as O-linked N-acetylglucosamine modification of Nup62 and Nup214. These oxidant-induced changes in nucleoporin modification correlated first with the increased binding of Nup62 to the exporter Crm1 and second with the reduced interaction of Nup62 with other FxFG-containing nucleoporins. Together, oxidative stress up-regulated the binding of Crm1 to Ran and affected multiple repeat-containing nucleoporins by changing their localization, phosphorylation, O-glycosylation, or interaction with other transport components. We propose that the combination of these events contributes to the stress-dependent regulation of Crm1-mediated protein export.     

Structure-activity relationships of diastereomeric lysine ring size analogs of the antimicrobial peptide gramicidin S: mechanism of action and discrimination between bacterial and animal cell membranes

Structure-activity relationships were examined in seven gramicidin S analogs in which the ring-expanded analog GS14 [cyclo-(VKLKVdYPLKVKLdYP)] is modified by enantiomeric inversions of its lysine residues. The conformation, amphiphilicity, and self-association propensity of these peptides were investigated by circular dichroism spectroscopy and reversed phase high performance liquid chromatography. (31)P nuclear magnetic resonance spectroscopic and dye leakage experiments were performed to evaluate the capacity of these peptides to induce inverse nonlamellar phases in, and to permeabilize phospholipid bilayers; their growth inhibitory activity against the cell wall-less mollicute Acholeplasma laidlawii B was also examined. The amount and stability of beta-sheet structure, effective hydrophobicity, propensity for self-association in water, ability to disrupt the organization of phospholipid bilayers, and ability to inhibit A. laidlawii B growth are strongly correlated with the facial amphiphilicity of these GS14 analogs. Also, the magnitude of the parameters segregate these peptides into three groups, consisting of GS14, the four single inversion analogs, and the two multiple inversion analogs. The capacity of these peptides to differentiate between bacterial and animal cell membranes exhibits a biphasic relationship with peptide amphiphilicity, suggesting that there may only be a narrow range of peptide amphiphilicity within which it is possible to achieve the dual therapeutic requirements of high antibiotic effectiveness and low hemolytic activity. These results were rationalized by considering how the physiochemical properties of these GS14 analogs are likely to be reflected in their partitioning into lipid bilayer membranes.