N-Methyl-D-Aspartate Receptor Function Observed by Rate of Ligand Dialysis From Proteoliposome Solution

Abstract

This study reports rate-of-dialysis of an iodinated N-methyl-D-aspartate antagonist drug, [125-1] MK-801, from solutions of lipid vesicles and from proteoliposomes containing purified membrane proteins. A 170 kd protein precipitated from proteoliposomes cross reacts with monoclonal antibodies against cloned NMDA-NR2(A) and NR2(B) subunits. Drug binding in proteoliposomes includes contributions from lipid and from protein, in addition to lipid. A significant change in drug binding was observed in proteoliposomes in response to 10 uM agonist, NMDA. Rate-of-dialysis from agonist-stimulated proteoliposomes was sensitive to perturbation by decreased aqueous ligand concentration in a manner consistent with a lipid-mediated receptor/antagonist equilibrium.     

Electrotonic and action potentials in the Venus flytrap

The electrical phenomena and morphing structures in the Venus flytrap have attracted researchers since the nineteenth century. We have observed that mechanical stimulation of trigger hairs on the lobes of the Venus flytrap induces electrotonic potentials in the lower leaf. Electrostimulation of electrical circuits in the Venus flytrap can induce electrotonic potentials propagating along the upper and lower leaves. The instantaneous increase or decrease in voltage of stimulating potential generates a nonlinear electrical response in plant tissues. Any electrostimulation that is not instantaneous, such as sinusoidal or triangular functions, results in linear responses in the form of small electrotonic potentials. The amplitude and sign of electrotonic potentials depend on the polarity and the amplitude of the applied voltage. Electrical stimulation of the lower leaf induces electrical signals, which resemble action potentials, in the trap between the lobes and the midrib. The trap closes if the stimulating voltage is above the threshold level of 4.4 V. Electrical responses in the Venus flytrap were analyzed and reproduced in the discrete electrical circuit. The information gained from this study can be used to elucidate the coupling of intracellular and intercellular communications in the form of electrical signals within plants.     

Sensitivity of antioxidant status of plant cells to furostanol glycosides

The present paper reports that in vitro plant objects (test tube plants and cell cultures), when subjected to furostanol glycosides (FG), underwent nonspecific reactions related to antioxidant status—decrease in peroxidation of lipids (POL) and increase in guaiacol-dependent peroxidase activity. The level of superoxide increased as early as after 5 min from contact with yam (Dioscorea deltoidea Wall) cells with FG. In this case, changes in POL processes and in activities of peroxidase and aldehyde-disposing emzymes were also observed. Upon a short-term cell exposure to FG, the levels of the primary POL products (conjugated dienes) increased, and that of the secondary POL products decreased compared to the control. These events were preceded by a rise in SOD activity and in an antioxidant activity of peroxidase along with a concurrent decrease in its oxidase (prooxidant) activity. The elevated activities of aldehyde-disposing enzymes aldehyde dehydrogenase and aldehyde reductase favored the reduction in the content of the secondary products of POL. Upon a long contact of FG with cells, the effect of FG was seen only at the initial and final phases of the culture growth cycle. Namely, FG diminished the POL level at the exponential growth phase and at the end of the cell degradation phase but had no effect at the stationary phase and the onset of the degradation phase. Therefore, the treatment with FG retarded the cell culture degradation and made the fall in cell viability not so dramatic by the end of the growth cycle. Actually, by the end of the degradation phase, the viability diminished down to 40% in the control but remained at 70% in the FG-treated counterpart.     

Matrix metalloproteinases as target genes for gene regulatory networks driving molecular and cellular pathways related to a multistep pathogenesis of cerebrovascular disease

The present study investigated a joint contribution of matrix metalloproteinases (MMPs) genes to ischemic stroke (IS) development and analyzed interactions between MMP genes and genome-wide associated loci for IS. A total of 1288 unrelated Russians (600 IS patients and 688 healthy individuals) from Central Russia were recruited for the study. Genotyping of seven single nucleotide polymorphisms (SNPs) of MMP genes (rs1799750, rs243865, rs3025058, rs11225395, rs17576, rs486055, and rs2276109) and eight genome-wide associated loci for IS were done using Taq-Man–based assays and MALDI-TOF mass spectrometry iPLEX platform, respectively. Allele − 799T at rs11225395 of the MMP8 gene was significantly associated with a decreased risk of IS after adjustment for sex and age (OR = 0.82; 95%CI, 0.70-0.96; P = 0.016). The model-based multifactor dimensionality reduction method has revealed 21 two-order, 124 three-order, and 474 four-order gene-gene (G×G) interactions models meaningfully (P_perm < 0.05) associated with the IS risk. The bioinformatic analysis enabled establishing the studied MMP gene polymorphisms possess a clear regulatory potential and may be targeted by gene regulatory networks driving molecular and cellular pathways related to the pathogenesis of IS. In conclusion, the present study was the first to identify an association between polymorphism rs11225395 of the MMP8 gene and IS risk. The study findings also indicate that MMPs deserve special attention as a potential class of genes influencing the multistep mechanisms of cerebrovascular disease including atherosclerosis in cerebral arteries, acute cerebral artery occlusion as well as the ischemic injury of the brain and its recovery.     

A family of highly diverse human and mouse genes structurally links leukocyte FcR,gp42 and PECAM-1

A group of genes encoding proteins structurally related to the leukocyte Fc receptors (FcRs) and termed the IFGP family was identified in human and mouse. Sequences of four human and two mouse cDNAs predict proteins differing by domain composition. One of the mouse cDNAs encodes a secreted protein, which, in addition to four immunoglobulin (Ig)-like domains, contains a scavenger receptor superfamily-related domain at the C-terminus. The other cDNAs code for the type I transmembrane proteins with the extracellular parts comprised of one to six Ig-like domains. Five homologous types of the Ig-like domains were defined and each protein was found to have a unique combination of the domain types. The cytoplasmic tails of the transmembrane proteins show different patterns of the tyrosine-based signal motifs. While the human IFGP members appear to be B-cell antigens, the mouse genes have a broader tissue distribution with predominant expression in brain. Sequence comparisons revealed that the IFGP family may be regarded as a phylogenetic link joining the leukocyte FcRs with the rat NK cell-specific gp42 antigen and platelet endothelial cell adhesion molecule-1 (PECAM-1), two mammalian leukocyte receptors whose close relatives were not found previously. It is suggested that FcRs, the IFGP proteins and gp42 have arisen by a series of duplications from a common ancestor receptor comprised of five Ig-like domains. The organization of the human genes shows that the IFGP family evolved through differential gain and loss of exons due to recombination and/or mutation accumulation in the duplicated copies.     

Structure prediction for peptides capable of inducing antibody formation in mice

A simple method for the sequence prediction of peptides capable of the in vivo stimulation of antibody production in mice without conjugation with protein carriers was proposed on the basis of literature data on the structure of T-helper epitopes active in vivo. According to this approach, a potentially active peptide should contain a nine-membered sequence with a hydrophobic amino acid residue in the first position and a positively charged residue in the ninth position. The efficiency of this approach was confirmed by the presence of such sequences in the previously described synthetic peptides with immune activities, by the application of this approach to the choice of immunogenic fragments within the sequences of various proteins that exhibited further the specific activity, and by the construction of immunogenic peptides on the basis of inactive natural sequences.     

Phorbol12-Myristate13-Acetate Relieves the Inhibitory Effect of A23187 on Erythroid Differentiation of K562 Cells Induced by Dimethylsulfoxide

The differentiating effect of DMSO on K562 cells was studied against the background of pretreatment of the cells by the modulators of activities of protein kinase C and Ca signaling, phorbol 12-myristate 13-acetate, and ionophore A23187. The 2-hour pretreatment of K562 cells with A23187 (1 M), rather than phorbol 12-myristate 13-acetate (0.1 M), led to inhibition of the differentiating effect of DMSO (0.1%) during the subsequent four days of incubation. When the cells were pretreated jointly with A23187 and phorbol 12-myristate 13-acetate, the DMSO-induced erythroid differentiation was restored. Analysis of the DNA-degrading effect of the reagents used on K562 cells by fluorescent dyes under the same conditions suggests that this activity is induced in the presence of DMSO in the cells pretreated with a combination of phorbol 12-myristate 13-acetate and A23187 or without pretreatment.     

Ecdysterone Modulates Antitumor Activity of Cytostatics and Biosynthesis of Macromolecules in Tumor-Bearing Mice

The influence of therapeutic and half doses of cisplatin and adriamycin combination with the anabolic drug ecdysterone (20-hydroecdison) on development of subcutaneously and intraperitoneally transplanted P388 and L1210 leukemia and metastasizing B16 melanoma was studied. Ecdysterone significantly stimulated the chemotherapeutic effect of low doses of the cytostatics: inhibition of tumor growth, mice survival rate, their lifespan, and the antimetastatic activity index were comparable or better than after therapy with high doses of the antitumor drugs. The influence of high and low doses of cisplatin and its low dose in combination with ecdysterone on the dynamics of protein and DNA biosynthesis in the liver, pancreas, thymus, spleen, and adrenals of tumor-bearing mice were also studied. Although the therapeutic effect of 4 mg/kg cisplatin by activated protein biosynthesis and DNA repair is comparable or better than that of its low dose (2 mg/kg) in combination with ecdysterone, in terms of chemotherapy the combination looks preferable since the therapeutic dose of cisplatin is toxic for the intact tissues.     

Inhibition of LDL oxidation by flavonoids in relation to their structure and calculated enthalpy

Twenty flavonoid compounds of five different subclasses were selected, and the relationship of their structure to the inhibition of low-density lipoprotein (LDL) oxidation in vitro was investigated. The most effective inhibitors, by either copper ion or 2,2'-azobis (2-amidino-propane) dihydrochloride (AAPH) induction, were flavonols and/or flavonoids with two adjacent hydroxyl groups at ring B. In the presence of the later catechol group, the contribution of the double bond and the carbonyl group at ring C was negligible. Isoflavonoids were more effective inhibitors than other flavonoid subclasses with similar structure. Substituting ring B with hydroxyl group(s) at 2' position resulted in a significantly higher inhibitory effect than by substituting ring A or ring B at other positions. The type of LDL inducer had no effect in flavonoids with catechol structure. Calculated heat of formation data (deltadeltaH(f)) revealed that the donation of a hydrogen atom from position 3 was the most likely result, followed by that of a hydroxyl from ring B. Position 3 was favored only in the presence of conjugated double bonds between ring A to ring B. This study makes it possible to assign the contribution of different functional groups among the flavonoid subclasses to in vitro inhibition of LDL oxidation.