Differential Regulation of Cysteinyl Leukotriene Receptor Signaling by Protein Kinase C in Human Mast Cells

Cysteinyl leukotrienes (cys-LTs) are a group of lipid mediators that are potent bronchoconstrictors, powerful inducers of vascular leakage and potentiators of airway hyperresponsiveness. Cys-LTs play an essential role in asthma and are synthesized as well as activated in mast cells (MCs). Cys-LTs relay their effects mainly through two known GPCRs, CysLT₁R and CysLT₂R. Although protein kinase C (PKC) isoforms are implicated in the regulation of CysLT₁R function, neither the role of PKCs in cys-LT-dependent MC inflammatory signaling nor the involvement of specific isoforms in MC function are known. Here, we show that PKC inhibition augmented LTD₄ and LTE₄-induced calcium influx through CysLT₁R in MCs. In contrast, inhibition of PKCs suppressed c-fos expression as well MIP1β generation by cys-LTs. Interestingly, cys-LTs activated both PKCα and PKCε isoforms in MC. However, knockdown of PKCα augmented cys-LT mediated calcium flux, while knockdown of PKCε attenuated cys-LT induced c-fos expression and MIP1β generation. Taken together, these results demonstrate for the first time that cys-LT signaling downstream of CysLT₁R in MCs is differentially regulated by two distinct PKCs which modulate inflammatory signals that have significant pathobiologic implications in allergic reactions and asthma pathology.     

On the relationship between the activity and structure of PEG-alpha-chymotrypsin conjugates in organic solvents

Enzymes are attractive catalysts for the production of optically active compounds in organic solvents. However, their often low catalytic activity in such applications hampers their practical use. To overcome this, we investigated the effectiveness of the covalent modification of alpha-chymotrypsin with methoxy poly(ethylene glycol) (PEG) with a Mw of 5,000 to enhance its activity. The model transesterification reaction between sec-phenethyl alcohol and vinyl butyrate in various neat dry organic solvents and at a controlled water activity of 0.008 in two solvents was employed to measure the effect of PEGylation on activity and enantioselectivity. Synthesis conditions were varied to obtain various conjugates with average molar ratios of PEG-to-chymotrypsin ranging from ca. 1 to 7. While the enantioselectivity increased only modestly from ca. 4.4 to 6.1 when averaging results in all solvents, PEG was very efficient in increasing the activity of alpha-chymotrypsin up to more than 400-fold compared to that of the powder lyophilized from buffer alone. The activity increase was more pronounced in apolar than in polar organic solvents and also depended on the amount of PEG bound to the enzyme. For example, the activity of the modified enzyme towards the most reactive "S" enantiomer in octane increased 440-fold but increasing the molar ratio of PEG-to-enzyme from 1.1 to 7.1 resulted in a more than twofold decrease in enzyme activity. Controlling the water activity did not prevent the drop in activity. To investigate the possible origin of the activity changes, Fourier transform infrared (FTIR) spectroscopy experiments were conducted. It was found that PEGylation reduced lyophilization-induced structural perturbations, but exposure to the organic solvents caused structural perturbations. These perturbations were more pronounced in polar than in apolar solvents. The pronounced activity drop in polar solvents at increasing PEG-modification levels correlated with an increasing level of solvent-induced structural perturbations. This correlation was less pronounced in apolar solvents where both, activity drop and structural perturbations, were less pronounced at increasing PEGylation levels. In summary, PEG-modified alpha-chymotrypsin might be an interesting system to catalyze reactions, particularly in apolar organic solvents.     

Ciprofloxacin-Induced Antibacterial Activity is Reversed by Vitamin E and Vitamin C

In the present study, we investigated the possible involvement of oxidative stress in ciprofloxacin-induced cytotoxicity against several reference bacteria including Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213, and clinical isolate of methicillin-resistant Staphylococcus aureus (MRSA). Oxidative stress was assessed by measurement of hydrogen peroxide generation using a FACScan flow cytometer. The antibacterial activity of ciprofloxacin was assessed using the disk diffusion method and by measuring the minimum inhibitory concentration (MIC). Ciprofloxacin induced a dose-dependent antibacterial activity against all bacteria where the highest tested concentration was 100 ug/ml. Results revealed that E. coli cells were highly sensitive to ciprofloxacin (MIC = 0.21 μg/mL ± 0.087), P. aeruginosa and S. aureus cells were intermediately sensitive (MIC = 5.40 μg/mL ± 0.14; MIC = 3.42 μg/mL ± 0.377, respectively), and MRSA cells were highly resistant (MIC = 16.76 μg/mL ± 2.1). Pretreatment of E. coli cells with either vitamin E or vitamin C has significantly protected cells against ciprofloxacin-induced cytotoxicity. These results indicate the possible antagonistic properties for vitamins C or E when they are used concurrently with ciprofloxacin.     

The Arabian race camel normal parameters--I. Haemogram, enzymes and minerals

1. Racing camels' (Camelus dromedarius) normal blood parameters were determined in Al-Ain, U.A.E. The parameters were: packed cell volume, haemoglobin, total red and white blood cells, the activities of glutamate oxaloacetate (GOT), creatinine kinase (CK), gamma-glutamyl transferase (gamma-GT) and CK muscle-brain isoenzyme and the concentrations of creatinine, urea, total protein, albumin, calcium, magnesium, phosphorus, sodium, potassium, zinc, copper and iron. 2. Most blood parameters were found to differ from those of other domestic animals and from non-race camels. 3. The data are discussed in relation to the management system practised.     

The effect of oxytetracycline on growth and lipid metabolism in poultry

The effect of oxytetracycline at doses of 0.291, 0.461, 0.922, 1.383 and 1.844 g/l in drinking water on the growth rate, lipid metabolism, GOT, GPT, calcium and magnesium was studied on one-day-old chicks and laying hens (Gallus domesticus). Oxytetracycline at a dose of 0.461 g/l increased body weight gain in one-day-old chicks. Oxytetracycline had no effect on hepatic triglyceride and phospholipid levels while cholesterol levels were decreased in one-day-old chicks and increased in laying hens. Oxytetracycline tended to decrease serum cholesterol and to increase serum triglyceride concentrations while its effect on serum phospholipids were inconsistent. Oxytetracycline, although inconsistent, tended to increase GPT and GOT activities in both young chicks and laying hens. Higher doses of oxytetracycline resulted in fatty changes in the hepatocytes and cells of the kidney tubules and lungs in both young chicks and laying hens. With the exception of hepatic phospholipids, all other parameters were higher in laying hens than in young chicks.     

Identification of a flunixin metabolite in camel by gas chromatography–mass spectrometry

A flunixin metabolite, a hydroxylated product, has been identified in camel urine and plasma samples using gas chromatography–mass spectrometry (GC–MS) and GC–MS–MS in the electron impact and chemical ionization modes. Its major fragmentation pattern has been verified by GC–MS–MS in daughter ion and parent ion scan modes. The method could detect flunixin and its metabolite in camel urine after a single intravenous dose of 2.2 mg of flunixin/kg body weight for 96 and 48 h, respectively, which increases the reliability of antidoping control analysis.     

The effect of oxytetracycline on the fat content and fatty acid composition of the egg yolk

The effects of oxytetracycline (OXT) at doses of 0.291, 0.461, 0.922, 1.383 and 1.844 g/l in drinking water on the egg quality and on the fat content and fatty acid composition of egg yolk were studied. OXT had no effect on egg weight, yolk weight or shell thickness. Increasing availability of OXT in water reduced cholesterol and triglyceride in the egg yolk while it had no effect on the phospholipid content. OXT at higher doses favoured unsaturated and polyunsaturated fatty acids relative to saturated fatty acids in the yolk.     

The contribution of platelet glycoproteins (GPIa C807T and GPIba C-5T) and cyclooxygenase 2 (COX-2G-765C) polymorphisms to platelet response in patients treated with aspirin

Objective

Aspirin is an antiplatelet agent commonly used in treatment of patients with high risk to develop stroke and myocardial infarction. However, inter-individual variability regarding the inhibition of platelet function by aspirin is well documented. In this study, the correlation between platelet glycoproteins (GPIa C807T and GPIba C-5T) and cyclooxygenase 2 (COX-2G-765C) polymorphisms and antiplatelet response in patients treated with aspirin was investigated.

Methods

Jordanian adult patients (n = 584) who are taking aspirin as an antiplatelet agent participated in the study. Platelet aggregation response was measured using Multiplate Analyzer® system. Polymerase chain reaction–restriction fragment length polymorphism assay (PCR–RFLP) was used for genotyping of the examined polymorphisms.

Results

Aspirin resistance was found in 15.8% of patients. Response to aspirin was significantly associated with GPIba C-5T polymorphism (P < 0.05). However, the GPIa C807T and COX-2G-765C polymorphisms were not related to aspirin resistance (P > 0.05).

Conclusion

A considerable fraction of the Jordanian population is resistant to the antiplatelet effect of aspirin, which might be related to GPIba C-5T polymorphism.     

Genetic association of adiponectin with type 2 diabetes in Jordanian Arab population

Adiponectin, a protein exclusively secreted by adipose tissue and present at low levels in obese individuals, is now widely recognized as a key determinant of insulin sensitivity and protection against obesity-associated metabolic syndrome. In Jordan, prevalence of diabetes (17.1%) is twice that of the United States (7.8%). In this study, we examined the contribution of the promoter variant rs266729 (− 11377C>G) of the ADIPOQ gene as a risk factor for diabetic patients in Jordan. DNA was extracted from blood samples for patients and controls .Polymerase chain reaction and restriction fragment length polymorphism were used to genotype this variant. A total of 420 type 2 diabetic patients and 230 controls were successfully genotyped. The results showed a significant genotypic (p = 0.00001) and allelic (p = 0.01) association with variant in the diabetic patients as compared to controls. This suggests that the ADIPOQ gene plays a major role in increasing the risk of diabetes, at least in the Jordanian Arab population.