Validated HPLC method for the determination of gabapentin in human plasma using pre-column derivatization with 1-fluoro-2,4-dinitrobenzene and its application to a pharmacokinetic study

A rapid, sensitive and accurate high-performance liquid chromatographic method with UV detection was developed and validated for the quantification of gabapentin in human plasma. Gabapentin was quantified using pre-column derivatization with 1-fluoro-2,4-dinitrobenzene following protein precipitation of plasma with acetonitrile. Amlodipine was used as internal standard. The chromatographic separation was carried out on a Nova-Pak C(18) column using a mixture of 50 mM NaH(2)PO(4) (pH=2.5)-acetonitrile (30:70, v/v) as mobile phase with UV detection at 360 nm. The flow rate was set at 1.5 ml/min. The method was linear over the range of 0.05-5 microg/ml of gabapentin in plasma (r(2)>0.999). The within-day and between-day precision values were in the range of 2-5%. The limit of quantification of the method was 0.05 microg/ml. The method was successfully used to study the pharmacokinetics of gabapentin in healthy volunteers.     

Activation of Wnt/β-Catenin Pathway in Monocytes Derived from Chronic Kidney Disease Patients

Patients with chronic kidney disease (CKD) have significantly increased morbidity and mortality resulting from infections and cardiovascular diseases. Since monocytes play an essential role in host immunity, this study was directed to explore the gene expression profile in order to identify differences in activated pathways in monocytes relevant to the pathophysiology of atherosclerosis and increased susceptibility to infections. Monocytes from CKD patients (stages 4 and 5, estimated GFR <20 ml/min/1.73 m²) and healthy donors were collected from peripheral blood. Microarray gene expression profile was performed and data were interpreted by GeneSpring software and by PANTHER tool. Western blot was done to validate the pathway members. The results demonstrated that 600 and 272 genes were differentially up- and down regulated respectively in the patient group. Pathways involved in the inflammatory response were highly expressed and the Wnt/β-catenin signaling pathway was the most significant pathway expressed in the patient group. Since this pathway has been attributed to a variety of inflammatory manifestations, the current findings may contribute to dysfunctional monocytes in CKD patients. Strategies to interfere with this pathway may improve host immunity and prevent cardiovascular complications in CKD patients.     

Change in membrane fatty acid compositions and cold-induced responses in chickpea

Plant cells often increase cold tolerance by reprogramming their genes expression which results in adjusted metabolic alternations, a process enhanced under cold acclimation (CA) phase. In present study, we assessed the changes of membrane fatty acid compositions and defense machine (like antioxidative enzymes) along with damage indexes like electrolyte leakage index (ELI) and malondialdehyde (MDA) during CA, cold stress (CS) and recovery (R) phases in chickpea (Cicer arietinum L.). Results showed an increase in unsaturated fatty acids ratio compare to saturated ones which is a sign of cold tolerance especially after CA phase. Antioxidant enzymes had an important role during CA and R phases while CS affected their activity which can be a sign for associating other metabolites or enzymes activities to create cold tolerance in plants. To investigation of enzymes assay under experimental treatments, the expression pattern of some enzymes including superoxide dismutase (sod), catalase (cat) and lipoxygenase (lox) was studied using quantitative real time PCR. LOX activity has shown a bilateral behavior: a positive relation with membrane damage index in CA and an interesting link with double bond index (DBI) in CS indicating probably its role in secondary metabolites like jasmonic acid signaling pathway. It was suggested that increased DBI and low LOX activity under CS could be a reason for plant cold tolerance.     

Association of a GPI-anchored protein with detergent-resistant membranes facilitates its trafficking through the early secretory pathway

Membrane microdomains are implicated in the trafficking and sorting of several membrane proteins. In particular GPI-anchored proteins cluster into Triton X-100 resistant, cholesterol- and sphingolipid-rich membrane microdomains and are sorted to the apical membrane. A growing body of evidence has pointed to the existence of other types of microdomains that are insoluble in detergents, such as Lubrol WX and Tween-20. Here, we report on the role of detergent-resistant membranes formed at early stages in the biosynthesis of membrane dipeptidase (MDP), a GPI-anchored protein, on its trafficking and sorting. Pulse-chase experiments revealed a retarded maturation rate of the GPI-anchor deficient mutant (MDPΔGPI) as compared to the wild type protein (wtMDP). However, Golgi to cell surface delivery rate did not show a significant difference between the two variants. On the other hand, early biosynthetic forms of wtMDP were partially insoluble in Tween-20, while MDPΔGPI was completely soluble. The lack of association of MDPΔGPI with detergent-resistant membranes prior to maturation in the Golgi and the reduction in its trafficking rate strongly suggest the existence of an early trafficking control mechanisms for membrane proteins operating at a level between the endoplasmic reticulum and the cis-Golgi.     

MiR-101 and miR-144 Regulate the Expression of the CFTR Chloride Channel in the Lung

The Cystic Fibrosis Transmembrane conductance Regulator (CFTR) is a chloride channel that plays a critical role in the lung by maintaining fluid homeostasis. Absence or malfunction of CFTR leads to Cystic Fibrosis, a disease characterized by chronic infection and inflammation. We recently reported that air pollutants such as cigarette smoke and cadmium negatively regulate the expression of CFTR by affecting several steps in the biogenesis of CFTR protein. MicroRNAs (miRNAs) have recently received a great deal of attention as both biomarkers and therapeutics due to their ability to regulate multiple genes. Here, we show that cigarette smoke and cadmium up-regulate the expression of two miRNAs (miR-101 and miR-144) that are predicted to target CFTR in human bronchial epithelial cells. When premature miR-101 and miR-144 were transfected in human airway epithelial cells, they directly targeted the CFTR 3′UTR and suppressed the expression of the CFTR protein. Since miR-101 was highly up-regulated by cigarette smoke in vitro, we investigated whether such increase also occurred in vivo. Mice exposed to cigarette smoke for 4 weeks demonstrated an up-regulation of miR-101 and suppression of CFTR protein in their lungs. Finally, we show that miR-101 is highly expressed in lung samples from patients with severe chronic obstructive pulmonary disease (COPD) when compared to control patients. Taken together, these results suggest that chronic cigarette smoking up-regulates miR-101 and that this miRNA could contribute to suppression of CFTR in the lungs of COPD patients.     

Dispersal and field progeny production of Trichogramma species released in an olive orchard in Egypt

Dispersal ability and field progeny production of augmentative released biological control agents depend on ecological adaptations of the particular species or strains used. Four species of the egg parasitoid genus Trichogramma were compared aiming to select suitable candidates for control of lepidopteran olive pests. Three of them (T. bourarachae Pintureau and Babault, T. cordubensis Vargas and Cabello, T. euproctidis Girault) had been previously collected from olive groves, whereas the commercially available strain used (T. evanescens Westwood) was originally isolated from sugarcane fields. During five consecutive field releases in an olive orchard near Cairo, dispersal and/or progeny production of these species was monitored using sentinel eggs placed at different heights in the release tree canopies as well as in neighboring trees (“distance effect”). The cardinal direction of dispersal was random for T. euproctidis and T. evanescens. Significant higher parasitism occurred on sentinel eggs placed on the middle part of tree canopy and highest parasitism was observed in trees where wasps had been released. Field progeny production was highest for T. bourarachae, followed by T. euproctidis and T. cordubensis. T. evanescens propagated less under field conditions. Inter-tree dispersal of all species except T. bourarachae was limited and, for biological control, releasing material should therefore be distributed on each olive tree, preferably also at different levels of the canopy.     

Piecing together the transport pathway of aliphatic glucosinolates

Glucosinolates are sulfur-rich secondary metabolites characteristic of the Brassicales order. Transport of glucosinolates was suggested more than 30 years ago through a number of studies which indicated that glucosinolates are produced in maternal tissue and subsequently transported to the seed. These observations laid the foundation for numerous studies on glucosinolate transport which have provided a wealth of information on biochemical properties of glucosinolate transport, source–sink relationships between organs and on the transport routes of glucosinolates. However, most of the conclusions and hypotheses proposed in these studies have not been discussed in context of each other to provide a complete overview of the current state of knowledge on glucosinolate transport. In this review, we are thus piecing together the glucosinolate pathway by presenting and critically analyzing all data on glucosinolate research. Furthermore, the data on glucosinolate transport is considered in the light of the newest findings on glucosinolate synthesis and distribution. The aim is to provide a comprehensive and updated set of hypotheses which may prove useful in directing future research on glucosinolate transport.     

Head and Neck Mycetoma: The Mycetoma Research Centre Experience

Mycetoma is a unique neglected tropical disease which is endemic in what is known as the “mycetoma belt”. The disease has many devastating impacts on patients and communities in endemic area and is characterised by massive deformity, destruction and disability. Mycetoma is commonly seen in the foot and hand and less frequent in other parts of the body. Mycetoma of the head and neck is a rarity and is associated with high morbidity and even mortality if not treated early. In this communication we report on 49 patients with head and neck mycetoma followed up at the Mycetoma Research Centre in Khartoum. Most of the reported patients had actinomycetoma and the majority were young adult males from mycetoma endemic areas in the Sudan. Most of them were students, farmers and workers. Prior to presentation the majority had long disease duration and the cause was multifactorial. Advanced disease with massive lesion, deformity and disability was the common presentation. There was no obvious history of local trauma, familial tendency or other predisposing factor identified in this group of patients. MRI and CT scan were the most accurate diagnostic tools to determine the disease extent. The treatment outcome was rather poor and characterised by low cure rate, poor outcome and high follows-up dropout. Such a gloomy outcome calls for structured and objective health education programs.     

The role of chick Ebf genes in the mediolateral patterning of the somites

This study was conducted to check whether the three chick Early B‐cell Factor (Ebf) genes, particularly cEbf1, would be targets for Shh and Bmp signals during somites mediolateral (ML) patterning. Tissue manipulations and gain and loss of function experiments for Shh and Bmp4 were performed and the results revealed that cEbf1 expression was initiated in the cranial presomitic mesoderm by low dose of Bmp4 from the lateral mesoderm and maintained in the ventromedial part of the epithelial somite and the medial sclerotome by Shh from the notochord; while cEbf2/3 expression was induced and maintained by Bmp4 and inhibited by high dose of Shh. To determine whether Ebf1 plays a role in somite patterning, transfection of a dominant‐negative construct was carried out; this showed suppression of cPax1 expression in the medial sclerotome and upregulation and medial expansion of cEbf3 and cPax3 expression in sclerotome and dermomyotome, respectively, suggesting that Ebf1 is important for ML patterning. Thus, it is possible that low doses of Bmp4 set up Ebf1 expression which, together with Shh from the notochord, leads to establishment of the medial sclerotome and suppression of lateral identities. These data also conclude that Bmp4 is required in both the medial and lateral domain of the somitic mesoderm to keep the ML identity of the sclerotome through maintenance of cEbf gene expression. These striking findings are novel and give a new insight on the role of Bmp4 on mediolateral patterning of somites.