NMDA excitotoxicity and free radical generation in rat brain homogenates: Application of a chemiluminescence assay

NMDA, the specific agonist of glutamate gated ion channels permeable to calcium, is implicated as a causal factor in the pathogenesis of several neurobiological disorders such as stroke, seizures, ischemia, and chronic neurodegenerative disease. On the other hand, evidence on the roles of oxidative mechanisms involved in NMDA-induced neurotoxicity is accumulating. In this study, we have used chemiluminescence measurements as an easy, rapid and sensitive assay to investigate the effects of NMDA and oxidative stress on brain cell vulnerability. Rat brain homogenates were incubated with increasing concentrations of glutamate and NMDA. Production of reactive oxygen species was followed by single photon emission measurements using the specific enhancers luminol and lucigenin. Increases in emission were observed at excitotoxic concentrations of glutamate and NMDA. Other parameters of oxidative stress such as diene conjugates, TBARS and carbonyl groups were also investigated. Our results indicated that chemiluminescence measurements may be used to study involvement of oxidative stress in neurotoxicity.     

Synthesis of some 4-arylidenamino-4H-1,2,4-triazole-3-thiols and their antituberculosis activity

The increasing clinical importance of drug-resistant mycobacterial pathogens has lent additional urgency to microbiological research and new antimycobacterial compound development. For this purpose, new triazoles were synthesized and evaluated for antituberculosis activity. A series of 4-arylidenamino-4H-1,2,4-triazole-3-thiol derivatives (2a–n) were synthesized from the treatment of 4-amino-4H-1,2,4-triazoles-3-thiol (1) with the respective aldehydes and were evaluated for antituberculosis activity against Mycobacterium tuberculosis H₃₇Rv (ATCC 27294), using the BACTEC 460 radiometric system and BACTEC 12B medium. Compound 2k showed an intereting activity at 6.25 μg/mL with a 87 percentage inhibition.     

Synthesis and evaluation of naphthalene-based thiosemicarbazone derivatives as new anticancer agents against LNCaP prostate cancer cells

Fourteen new naphthalene-based thiosemicarbazone derivatives were designed as anticancer agents against LNCaP human prostate cancer cells and synthesized. MTT assay indicated that compounds 6, 8 and 11 exhibited inhibitory effect on LNCaP cells. Among these compounds, 4-(naphthalen-1-yl)-1-[1-(4-hydroxyphenyl)ethylidene)thiosemicarbazide (6), which caused more than 50% death on LNCaP cells, was chosen for flow cytometric analysis of apoptosis. Flow cytometric analysis pointed out that compound 6 also showed apoptotic effect on LNCaP cells. Compound 6 can be considered as a promising anticancer agent against LNCaP cells owing to its potent cytotoxic activity and apoptotic effect.     

Contractile responses of the human umbilical artery to KCl and serotonin in Ca-free medium and the effects of levcromakalim

It is known that K(ATP) channel openers inhibit the release and refilling of Ca(2+) from intracellular stores. The present study was designed to test the effects of levcromakalim in human umbilical artery (HUA) rings stimulated by serotonin (5-HT) and KCl in Ca-free medium. Umbilical cords were obtained at vaginal or cesarean deliveries from healthy, term pregnancies. After the isolation, HUA rings were placed in organ baths in solution with indomethacin (10(-5) M) and N(G)-nitro-L-arginine methyl ester (L-NAME) (10(-3) M) at 37 degrees C and aerated with 95% O(2) and 5% CO(2) for the measurement of isometric force. In Ca-free solution with Ethylene glycol-bis (ss-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) (2 mM) the contractions produced by 5-HT (10(-6) M) and KCl (40 mM) decreased significantly. Afterwards, HUA rings were treated with 5-HT and KCl in repeated manner in Ca-free medium. In contrast to KCl, 5-HT induced contractions reduced in each application, progressively. Levcromakalim (10(-4) M) abolished the contractions elicited by 5-HT. On the other hand, levcromakalim had a little but significant inhibitory effect on KCl induced contraction in Ca-free medium. These results suggest that Ca(2+) is not the only transduction pathway in KCl produced contractions of HUA smooth muscle cells.     

Electron microscopic immunolocalization of basic fibroblast growth factor in peripheral nerves

In spite of ample information about the distribution and the effects of basic fibroblast growth factor (bFGF) in the central nervous system, few data are available concerning the localization of this protein in the peripheral nervous system. In view of the role of bFGF in the regulation of trophic and non-trophic functions, we focused on the presence and precise localization of this growth factor in normal peripheral nerves at the electron microscopic level. The study shows that bFGF is mainly located in the Schwann cells, especially in the nuclei. There is slight labeling in the myelin sheath and in the axon cytoplasm. The study provides morphologic evidence for an association between bFGF expression and Schwann cells. Such as association argues for a role of this peptide in the maintenance or regeneration of peripheral nerves.     

Ultrastructural immunolocalization of basic fibroblast growth factor in endothelial cells: morphologic evidence for unconventional secretion of a novel protein

Basic fibroblast growth factor (bFGF) is one of the most potent angiogenic factors. Unlike many other growth factors, bFGF lacks a classic peptide sequence for its secretion. Recent studies suggest that there is an unconventional secretory pathway for this growth factor. The aim of this study was to identify the specific location of bFGF in endothelial cells and to find morphologic evidences concerning its synthesis, storage and release from endothelial cells. The capillaries in hippocampus, adrenal gland, kidney, peripheral nerves as well as the vessels in connective tissues were analysed by using immunogold labeling techniques at electron microscope level. Results show that endogenous bFGF is mainly located in the nuclei of endothelial cells. Slight immunoreactivity is found in the cytoplasm. Immunolabeling is notably absent in pinocytotic vesicles, Golgi complexes, endoplasmic reticulum, nuclear membrane and intercellular junctions. These results provide morphologic evidence suggesting that endothelial cells might export bFGF via unique cellular pathways that are clearly distinct from classical signal peptide mediated secretion and/or release of this protein could be directly through mechanically induced disruptions of these cells. The current study support the recent hypothesis related with unconventional secretory pathway for bFGF as some other “cargo” proteins.     

Frequencies of pyrethroid resistance-associated mutations of Vssc1 and CYP6D1 in field populations of Musca domestica L. in Turkey

House flies were collected from 16 different provinces in the Aegean and Mediterranean regions of Turkey, and the frequencies of pyrethroid resistance-associated mutations in Vssc1 and CYP6D1 in these field-collected populations were studied. Although there is no organized resistance management program for house fly control in Turkey, it is known that different groups of insecticides, including pyrethroids, are used. The frequencies of both Vssc1 and CYP6D1 alleles were weighted toward the susceptibles, with Vssc1-susceptible alleles having higher frequencies in both regions (0.75 in Aegean and 0.69 in Mediterranean populations) than CYP6D1-susceptible alleles (0.65 in Aegean and 0.56 in Mediterranean populations). The frequencies of kdr-his alleles were higher than the frequencies of kdr alleles in these populations. While the frequencies of kdr-his alleles were close to each other in the Aegean (0.23) and Mediterranean (0.17) populations, the frequencies of kdr alleles remarkably differed in these two regions, with values of 0.02 and 0.14, respectively. In contrast to Europe, Asia, and the U.S.A., no super-kdr allele was detected in the samples from both regions. We identified six and eight different Vssc1+CYP6D1 genotype classes in the Aegean and Mediterranean regions, respectively. The three most common genotype classes in the regions were susceptible Vssc1 with heterozygous CYP6D1v1 (29%), sus/kdr-his1 with heterozygous CYP6D1v1 (23%), and susceptible Vssc1 with CYP6D1 (22%). The total frequencies of these three most common genotype classes (approximately 75%) obtained in our study were very close to the value obtained in Florida in a previous study, which was related by the similarity of temperature patterns between Florida and the corresponding regions of Turkey. This may reflect the lack of overwintering fitness cost associated with resistance alleles in both climates.     

Kinases and glutathione transferases: selective and sensitive targeting

Kinases, representing almost 500 proteins in the human genome, are responsible for catalyzing the phosphorylation reaction of amino acid residues at their targets. As the largest family of kinases, the protein tyrosine kinases (PTKs) have roles in controlling the essential cellular activities, and their deregulation is generally related to pathologic conditions. The recent efforts on identifying their signal transducer or mediator role in cellular signaling revealed the interaction of PTKs with numerous enzymes of different classes, such as Ser/Thr kinases (STKs), glutathione transferases (GSTs), and receptor tyrosine kinases (RTKs). In either regulation or enhancing the signaling, PTKs are determined in close interaction with these enzymes, under specific cellular conditions, such as oxidative stress and inflammation. In this concept, intensive research on thiol metabolizing enzymes recently showed their involvement in the physiologic functions in cellular signaling besides their well known traditional role in antioxidant defense. The shared signaling components between PTK and GST family enzymes will be discussed in depth in this research review to evaluate the results of recent studies important in drug targeting for therapeutic intervention, such as cell viability, migration, differentiation and proliferation.     

Effects of simvastatin treatment on oxidant/antioxidant state and ultrastructure of streptozotocin-diabetic rat lung

In the present study, we investigated the effects of simvastatin, a 3-hydroxy-3-methyl-glutaryl coenzyme A reductase inhibitor, on lipid metabolism, lipid peroxidation, antioxidant enzyme activities and ultrastructure of diabetic rat lung. Diabetes was induced by a single injection of streptozotocin (45 mg kg(-1), i.p.). After 8 weeks induction of diabetes, some control and diabetic rats were treated with simvastatin (10 mg kg(-1) rat day(-1); orally) for 4 weeks. Diabetes resulted in significantly high levels of blood glucose and plasma lipids. Malondialdehyde levels were unchanged after 12-week-old diabetic rats, whereas catalase activity significantly decreased in the lung. Glutathione peroxidase activity and nitric oxide level were significantly elevated in the diabetic lung. Histological analysis of the diabetic lung revealed some deterioration in the structure. Simvastatin treatment reduced plasma lipid levels and partially decreased the severity of hyperglycaemia. Catalase, glutathione peroxidase activities and nitric oxide levels were partially restored and accompanied by improved structure in diabetic lung by the simvastatin treatment. These results suggest that structural disturbances and alteration of antioxidative enzyme activities occurred in diabetic lung. Simvastatin treatment may provide some benefits in the maintenance of antioxidant status and structural organization of diabetes-induced injury of lung.