Characteristics of immunomodulating effect of histamine in inbred strain mice]

The activity of 5'-nucleotidase of peritoneal exudate in subcutaneous injection of histamine in a dose of 1.0-100 microliters was studied in mice of different lines (CBA, C57, B1/6, Balb/c, NFS/n, NFR/n). There were interline differences in the influence of histamine on this metabolic index.     

Purification and properties of leucyl-tRNA synthetase from the cow mammary gland

Three forms (E1, E2 and E3) of leucyl-tRNA synthetase (LeuRS) were separated by DEAE-cellulose chromatography of total aminoacyl-tRNA synthetases from cow lactating mammary gland. The method of purification of all three components is described. E1 is a dimeric molecule (alpha 2) of molecular weight 182 000. Two other forms of molecular weight 67 000 and 64,000 consist of a single polypeptide chain as determined by polyacrylamide gel electrophoresis. Optimum conditions and kinetic parameters of leucyl-tRNA formation were studied for every enzyme form. The low values of Vmax and thermostability are characteristic of E3. All forms of LeuRS interact with 6 isoaccepting tRNA(Leu) from lactating mammary gland and can activate leucine in the absence of tRNA. E2 and E3 are supposed to derive from the native enzyme by endogenous proteolysis. The physico-chemical properties of native LeuRS from lactating mammary gland are compared with those of LeuRS's from other sources.     

Plant virus infection development as affected by heavy metal stress

The work was focused on the investigation of possible dependencies between the development of viral infection in plants and the presence of high heavy metal concentrations in soil. Field experiments have been conducted in order to study the development of systemic tobacco mosaic virus (TMV) infection in Lycopersicon esculentum L. cv. Miliana plants under effect of separate salts of heavy metals Cu, Zn and Pb deposited in soil. As it is shown, simultaneous effect of viral infection and heavy metals in tenfold maximum permissible concentration leads to decrease of total chlorophyll content in experiment plants mainly due to the degradation of chlorophyll a. The reduction of chlorophyll concentration under the combined influence of both stress factors was more serious comparing to the separate effect of every single factor. Plants' treatment with toxic concentrations of lead and zinc leaded to slight delay in the development of systemic TMV infection together with more than twofold increase of virus content in plants that may be an evidence of synergism between these heavy metal's and virus' effects. Contrary, copper although decreased total chlorophyll content but showed protective properties and significantly reduced amount of virus in plants.     

The determination of lipopolysaccharide admixtures in protein preparations

A new method for the determination of lipopolysaccharide (LPS) admixtures in protein solutions has been developed. The method includes the periodate oxidation of LPS, biotinylation with biotin hydraside, immobilization on a nitrocellulose membrane and the development of biotinylated LPS in the streptavidin--alkaline phosphatase system. Proteins are previously removed from the solution by treatment with hot phenol. Development with the use of 5-bromoinodyl phosphate and nitrotetrazolium blue makes it possible to detect about 30 pg of LPS immobilized on the nitrocellulose membrane.     

Effect of dehydration and dDAVP on water permeability of basolateral membranes of epithelial cells in the kidney collecting tubules

Water permeability of the outer medullary collecting duct's (OMCD) basolateral membrane was determined in vitro in the tubules isolated from hyperhydrated or dehydrated Wistar rats. Oil was injected into the lumen to block apical membrane water permeability. OMCD fragments underwent a hypoosmic shock (600/300 mOsm) and epithelial cells volume increased ad recorded with a digital camera. The latter's rate was used to calculate apparent water permeability of the membrane (Pf). Treatment of the tubules with Hg2Cl2 suppressed the water permeability. Water deprivation and dDAVP induced an increase in the basolateral water permeability. The data obtained suggest that the water permeability of the OMCD basolateral membrane may be stimulated by vasopressin and water deprivation.     

Oligonucleotide conjugates with minor groove ligands as probes for hybridization microarray chips

A possibility of using oligonucleotide conjugates with minor groove ligands as probes for hybridization microarray chips was studied. The oligonucleotide conjugates contain a hairpin ligand (MGB) composed of two tripyrrolcarboxamide residues with an aminocaproic acid residue as a linker and bound to the oligonucleotide duplex AT tract in a site-specific manner. We used as (5'-3') probes GACAAGAp, GACAAAAp, GACAAGA-MGB, and GACAAAA-MGB. The oligonucleotides labeled with Cy3 cyanine dye, Cy3-ACTAATTTTGTC and Cy3-ACTAATCTTGTC, were used as targets. The maximal MGB effect on the fluorescence level of microarray chip spots, which caused its fourfold increase as compared with the initial unmodified duplex, was observed for the duplex containing only AT pairs in the ligand binding site. The presence of A-C and G-T mutations in the binding site (imperfect duplexes) or a C-G pair (perfect duplex) affects the change in fluorescence level to a considerably lesser degree.     

G(i) proteins in regulation of water permeability of the rat kidney collecting tube epithelium cell membrane

Principal mechanism of the transepithelial water permeability increase in the kidney collecting ducts in response to vasopressin involves insertion of aquaporin 2 (AQP2) into the apical membrane. Previously we have shown that water permeability of the basolateral membrane also may be increased with stimulation of V2-receptors. It is known that inhibition of G(i)-proteins with pertussis toxin blocks redistribution of AQP2 into the apical membrane following the application of vasopressin or forskolin. The aim of the present study was to investigate potential involvement of G(i)-proteins in regulation of basolateral membrane water permeability. Effect of pertussis toxin on the ability of desmopressin to increase the basolateral membrane osmotic water permeability was investigated, and the expression of Galpha(i)2 and Galpha(i)3 genes under normal conditions and after 2 days of water deprivation were evaluated. We demonstrated that dehydration leds to a 30% increase of Galpha(i)3 mRNA content while the Galpha(i)2 mRNA level remains unchanged. In control experiments, basolateral membrane water permeability increased in response to desmopressin from 59.2 +/- 6.61 to 70.6 +/- 9.2 microm/s (p < 0.05, paired t-test). Pertussis toxin completely blocked this reaction (53.5 +/- 5.18 vs 50.1 +/- 6.50 microm/s, respectively). We conclude that G(i)-proteins participate in the mechanism of the basolateral membrane water permeability increase in response to stimulation of V2-receptors. Clarification of the G(i)-proteins role in this process requires further investigation, but most likely they are involved in regulation of aquaporin transport and insertion into the cell membrane.     

Inductive characteristics of proteins secreted by retinal cells

The studies of the development of eye rudiments and formation of adult eye tissues have always been among priorities in developmental biology and then in developmental genetics, which is associated with the peculiarities of the development and structure of the eye. In the late 80s, it was established by the group of developmental factors of the Institute of Gene Biology of RAS that many differentiated tissues are able to produce proteins causing homologous differentiations in polypotent cells of early gastrula ectoderm. The aim of our present study was isolation of proteins secreted by mammalian and fish retinal cells and determination of their inductive properties in early gastrula ectoderm of Xenopus laevis. The sets of proteins secreted by retina induce tissues homologous to the inducer, that is, neural tissue, brain, retina, pigmented epithelium, and also lenses and ear vesicles. The retinal inductive proteins retain their homologous inductive capacity after lyophilization. Biological testing shows that a total mixture of proteins secreted by retinal cells induces in polypotent gastrula ectoderm of X. laevis a narrower spectrum of tissues than the fractions obtained from this mixture. The above-outlined results obtained in thecourse of investigations of inductive peculiarities of retina and its fractions help in the elucidation of questions concerning embryonic induction and factors determining it, as well as questions concerning the maintenance of tissue specifity and regenerative capacity of the tissue studied.     

Role of water channels in the regulation of the volume of principal cells of rat kidney collecting ducts in hypoosmotic medium

The effect of plasma membrane water permeability on the rate of changes in the volume of principal cells of collecting ducts of the outer substantia medullaris under conditions of hypoosmotic shock has been studied. Changes in cell volume were studied by the fluorescent method. It was shown that the hypotonic shock induced a rapid increase in the cell volume with the characteristic time that depended on plasma membrane water permeability. The decrease in volume occurred much more slowly, and the rate of volume decrease directly correlated with the rate of swelling. The inhibition of potassium transport by barium chloride decreased the rate of volume restoration, without affecting substantially the duration of the swelling phase. The inhibition of mercury-sensitive water channels by mercury caused a significant increase in the time of both cell swelling and volume restoration. It was concluded that the state of water channels largely determines the rate of the regulatory response of epithelial cells of collecting ducts to hypoosmotic shock and affects the exchange of cell osmolites.