Metabolism of a New Antiaggregant, Indolinone Derivative.

Cytochrome p450-mediated metabolism of GRS (indolinone antiaggregant) and its effects on activities of cytochrome p450 isoenzymes were studied. Inhibition of 6 isomers of cytochrome p450 in human liver microsomes was studied with the use of specific substrates. It was found that human liver cytochrome p450 enzymes could not induce degradation of GRS and that GRS was not an inductor or inhibitor of cytochrome p450 family members 1A2, 2C9, 2C19, 2D6, 2C8, and 3A4. Hence, clinical use of the prospective antiaggregant would not involve the risk of uncontrolled fluctuations in GRS concentrations in the organism because of interactions between the drugs.

Co-Transplantation of Pancreatic Islet Cells and Mesenchymal Bone Marrow Precursors on Titanium Nickelide Scaffolds in Alloxan-Induced Diabetes Mellitus.

We studied therapeutic activity of co-transplantation of allogeneic pancreatic islet cells and mesenchymal bone marrow progenitors on TiNi scaffolds in Wistar rats with experimental alloxan-induced diabetes mellitus. In preliminary experiments with co-culturing of cells in different proportions followed by their transplantation on tissue-engineered constructs, the optimum ratio of these cells was determined - 3:1. Regeneration was assessed by biochemical methods by the blood levels of glucose and glycosylated hemoglobin on days 15, 30, and 5. In the group with combined cell transplantation on TiNi scaffold, normalization of the studied biochemical parameters occurred earlier than after monotherapy with allogenic islet cells and was associated with an increase in animal lifespan. Normalization of the parameters of bone marrow hemopoiesis, in particular, the number of myelokaryocytes and erythroblasts was also noted.

[Oxidative stress in adipose tissue as a primary link in pathogenesis of insulin resistance].

Obesity is a leading risk factor of diabetes mellitus type 2, impairments of lipid metabolism and cardiovascular diseases. Dysfunctions of the accumulating weight of the visceral fat are primarily linked to pathogenesis of systemic insulin resistance. The review considers modern views about biochemical mechanisms underlying formation of oxidative stress in adipocytes at obesity, as one of key elements of impairments of their metabolism triggering formation of systemic insulin resistance.

[THE ROLE OF THE ENDOGENOUS CANNABINOID SYSTEM IN THE FORMATION OF THE GENERAL ADAPTATION SYNDROME].

Nonopioid stress-induced analgesia is the consequence of activation of CB1 receptors by the increased level of 2-arachidonoyl glycerol, anandamide in the periaqueductal gray matter in the midbrain. The activation of cannabinoid CB1 receptors inhibits stress-induced ulcerogenesis due to the strengthening of the antioxidant defense of the gastric mucosa. CB1 receptor antagonists promote an increase in ACTH and corticosterone concentrations in the blood of intact animals, the knockout of the gene encoding the CB1 receptor exhibits the same effect. Antagonists of CB1 receptors enhance the stressor elevation of ACTH and corticosterone levels in the blood of experimental animals. It was found an increase in stress-induced elevation of corticosterone and ACTH levels in the blood of mice with a knockout of the gene encoding the CB1 receptor. An increase in the endogenous anandamide level or disturbance of the reuptake of endogenous cannabi-noids after application of pharmacological agents promotes reducing corticosterone level in stressed animals. Consequently, endogenous cannabinoids inhibit basal and suppress stress-induced activity of the hypothalamic-pituitary-adrenal axis. The indicated regulation is carried out on the level of the hypothalamus, pituitary and adrenal cortex. Stimulation of central cannabinoid receptors leads to an activation of the sympathetic system. The activation of peripheral CB1 receptors leads to inhibition of norepinephrine release from sympathetic terminals and epinephrine release from the adrenal glands. The endogenous CB1 receptor agonists play an anxiolytic role and prevent the occurrence of pathological anxiety.

Evidence for factor IX-independent roles for factor XIa in blood coagulation.

BACKGROUND: Factor XIa is traditionally assigned a role in FIX activation during coagulation. However, recent evidence suggests this protease may have additional plasma substrates. OBJECTIVE: To determine whether FXIa promotes thrombin generation and coagulation in plasma in the absence of FIX, and to determine whether FXI-deficiency produces an antithrombotic effect in mice independently of FIX. METHODS: FXIa, FXIa variants and anti-FXIa antibodies were tested for their effects on plasma coagulation and thrombin generation in the absence of FIX, and for their effects on the activation of purified coagulation factors. Mice with combined FIX and FXI deficiency were compared with mice lacking either FIX or FXI in an arterial thrombosis model. RESULTS: In FIX-deficient plasma, FXIa induced thrombin generation, and anti-FXIa antibodies prolonged clotting times. This process involved FXIa-mediated conversion of FX and FV to their active forms. Activation of FV by FXIa required the A3 domain on the FXIa heavy chain, whereas activation of FX did not. FX activation by FXIa, unlike FIX activation, was not a calcium-dependent process. Mice lacking both FIX and FXI were more resistant to ferric chloride-induced carotid artery occlusion than FXI-deficient or FIX-deficient mice. CONCLUSION: In addition to its predominant role as an activator of FIX, FXIa may contribute to coagulation by activating FX and FV. As the latter reactions do not require calcium, they may make important contributions to in vitro clotting triggered by contact activation. The reactions may be relevant to FXIa's roles in hemostasis and in promoting thrombosis.

[Effect of the energy metabolism regulator Yantar-Antitox on the system of energy production in rat liver during experimental pathology of beta-oxidation].

Effects of the Yantar-Aantitox (succinic acid preparation) preparation on bioenergetic processes in mitochondria of rat liver during the experimental disorders of beta oxidation process evoked by 4-pentenoic acid have been studied. It is established that the course administration of Yantar-Antitox leads to normalization of disturbed bioenergetic processes in rat liver, which is due to stimulation of the rapid metabolic cluster of mitochondria.

[Functional state of a sphingomyeline cycle and free radical lipid oxidation activity of a rat's liver during different phases of starvation].

The functional state of a sphingomyeline cycle and character of its mutual relations with the processes of free radical lipid oxidation during starvation of animals without any restriction of access to drinking water at 1, 2, 3 day (I phase) and 6 day (II phase of starvation) were studied at the liver of rats. The maximal values of the ceramide/sphingomyeline ratio and activity neutral sphingomyelinase and executive caspase-3 were reached in a liver of animals at the 3rd day of starvation. From the 3rd day of starvation the concentration of the tumour necrosis factor-alpha which is one of activators neutral sphingomyelinase was increase in rats blood serum. During the extent of large part of the I phase of starvation the intensity of free radical lipid peroxidation in a liver had almost the same level as in control group--that was a result of the high-grade functioning of antioxidant defense system. After transition the I phase of starvation into the II phase (6 day of experiment) the oxidative stress was developed as result of an exhaustion of system antioxidant defense potential in a liver. The results of this data can testify that during I phase of starvation in a liver the conditions was raised for display of the ceramide-mediated proapoptotic signalling. We assume that ceramide-mediated apoptosis is one of mechanisms of optimization of liver cellular population at the frames of metabolic adaptation. The I phase of starvation in a liver proves by the ceramide-mediated proapoptotic signaling developing. During the II phase of starvation the oxidative stress process were prevailed.

[The parameters of collagen proteolytic and metabolic systems in chronic liver diseases of viral and toxic etiologies].

AIM: To study a relationship of the plasma activity of elastase-like and collagenase-like proteinases and their inhibitors to hepatic collagen metabolism and to detect the serum markers of fibrosis severity. SUBJECTS AND METHODS: Three hundred and fifty-nine patients with chronic liver diseases (CLD), including 118 patients with chronic viral hepatitis (CVH), 113 with CVH concurrent with alcoholic liver disease (ALD), 109 with ALD, and 19 with CLD in the presence of opiomania were examined. The activities of alpha1-proteinase inhibitor and alpha2-macroglobulin (alpha2-MG) were determined by the unified spectrophometric assay from the inhibition of N-benzoyl-arginine ethyl ester hydrolysis. The activity of elastase-like proteinases was determined by enzymatic assay from the hydrolysis of the synthetic substrate N-butyloxycarbonyl-L-alanine-para-nitrophenyl ester. That of collagenase-like proteinases was determined, by using a collagen type 1 substrate and expressed in terms of micromoles of the resultant hydroxyproline. The content of hydroxyproline was determined by a color reaction with demethylbenzaldehyde, a free, peptide- and protein-bound hydroxyproline; their fraction was obtained under various conditions of plasma protein isolation and hydrolysis. Plasma fibronectin levels were measured by solid-phase immunoassay. Liver biopsy specimens were morphologically studied in the majority of patients to determine the histological hepatitis activity index and the stage of fibrosis. RESULTS: Fibrois formation in the liver in its chronic diseases was attended by a significant reduction in the activity of collagenase-like proteinases hydrolyzing collagen and by the lower activity of alpha2-MG, an inhibitor limiting collagen formation. CONCLUSION: The identified changes make themselves evident just in early fibrosis, which suggests the rapid onset of imbalance in the mechanisms responsible for regulation of connective tissue synthesis and promotes intensified fibrosis formation.

[Activity of the sphingomyelin cycle enzymes and concentration of products of sphingomyelin degradation in the rat liver in the course of acute toxic hepatitis].

Activity of key enzymes of a sphingomyelin cycle and the maintenance of its components (sphingomyelin, ceramide and sphingosine-1-phosphate) have been studied in livers of rats in dynamics of the acute toxic hepatitis caused by hypodermic introduction of an oil solution of CCl4. Sphingomyelinase activity significally increased already on early terms and remained increased over the whole period of observation. Activity of ceramidase insignificantly differed from the control level. The levels of sphingomyelin and sphingosine-1-phosphate did not undergo marked changes while ceramide content significally increased. Thus, balance between liver content of ceramide (proapoptotic) and the sphingosine-1-phosphate, being the antiapoptotic factor, was shifted towards ceramide. In sphingomyelin molecules there was a significant decrease in the content of fatty acids C18: and C22:2, while in ceramide molecules and sphingosine-1-phosphate only fatty acid C22:2 changed. In spite of significant decrease in content of some unsaturated fatty acids, calculated unsaturation coefficients of the fatty acid component of the sphingomyelin cycle metabolites. Thus, our results together with literature data suggests involvement of ceramide-mediated apoptosis in the pathogenesis of acute toxic hepatitis. Elimination of damaged hepatocytes facilitates realization of repair processes and optimization of cellular community of a liver.

Activity of sphingomyelinase in rat liver in acute and chronic toxic hepatitis: proportion between peroxidative and phospholipase pathways of lipid bilayer modification.

We showed that sphingomyelinase activity in the liver increased only during the acute phase of toxic hepatitis. Peroxidative modification of hepatocyte membrane bilayer prevailed during the acute phase, while after transformation of the process to the chronic phase phospholipase pathway predominated.

Functional activity of sphingomyelin cycle in rat liver in chronic toxic hepatitis.

Activities of sphingomyelinase and ceramidase decreased in the liver in chronic toxic hepatitis and the balance between the levels of proapoptotic ceramide and antiapoptotic sphyngosine-1-phosphate shifts towards the latter substance. Pronounced changes in the qualitative and quantitative composition of fatty acids in the sphingomyelin cycle effector molecules were revealed.

[Experimental evaluation of the biological risks of introduction of the genetically modified microorganism (GMM) B. subtilis VKPM B-7092 into the environment].

Experimental evaluation of the biological risks of introducing the genetically modified microorganism (GMM) B. subtilis VKPM B-7092, an active ingredient of the probiotic VETOM 1.1, into an open system was performed. The following features of the GMM were studied: the survival rate of the GMM in bovine gastroenteric tract; its influence on the microbiocenosis; the species composition of microflora of the gastroenteric tract of the animal species; the possibility of transfer of the DNA fragment cloned in the B. subtilis bacterium and containing the gene of human leukocyte alpha2 interferon to the representatives of intestinal microflora of animals fed on the probiotic VETOM 1.1, as well as the GMM transfer to other microorganism species spread in the areas of potential getting of the GMM into the environment (soil). The study revealed no negative effects of the GMM on the animal organism and the environment, including remote aftereffects.

[The effect of entomopathogenic fungi on the activity of detoxicating enzymes of larvae of bee pyralid Galleria mellonella L. (Lepidoptera, Pyralidee) and the role of detoxicating enzymes in the formation of the insect' resistance to entomopathogenic fungi].

In the work it was shown that fungal infection of insects increased the total esterase and glutathione-S-transferase activities of hemolymph. Acid and alkaline phosphatase activity in infected larvae remained at the same level as in intact larvae. Fungi-infected bee pyralid larvae turned out to be 1.46 times more resistant to phosphoorganic insecticide malation as compared to intact larvae. On the other hand, treating insects with the inhibitors of detoxicating enzymes caused a sharp decrease in the insects' resistance to entomophathogenic fungi. The work discusses possible participation of detoxicating enzymes in the formation of the insects' resistance to entomopathogenic fungi as well as new strategies of the development and creation of complex biopreparations based on entomopathogenic microorganisms and inhibitors of detoxicating enzymes.

[Paticipation of cannabinoid receptors in the regulation of cardiac rhythm and cardiac contractility].

It has been found that i. v. administration of cannabinoid receptor (CB) agonists (HU-210, ACPA, anandamide, methanandamide) induced a decrease in the heart rate (HR) in anesthetized rats. Pretreatment with CB1 receptor antagonist SR141716A completely abolished a negative chronotropic effect of CB receptor agonist HU-210. The CB2 receptor antagonist SRI 44528 did not prevent a HU-210-induced decrease in the HR. Pretreatment with the ganglion blocker hexamethonium had no effect on the negative chronotropic action of HU-210. Addition of HU-210 (100 nM) to perfusion solution induced a decrease in the HR, left ventricular development pressure, rate of contractility and relaxation of isolated perfused rate heart without change in end diastolic pressure. These data suggest that cardiac CBI receptor activation induces a decrease in the HR both in vivo and in vitro. An occupancy of the same receptors mediates a negative inotropic effects of cannabinoids.

Negative chronotropic effect of cannabinoids and their water-soluble emulsion is related to activation of cardiac CB1 receptors.

Intravenous injection of cannabinoids dissolved in cremophore EL:ethanol:NaCl mixture and water-soluble emulsion of the same cannabinoids caused identical negative chronotropic effects in chloralose-narcotized rats. Selective CB1 and CB2 receptor antagonist HU-210 also induced a negative chronotropic effect in rats, while pre-injection of CB1 receptor antagonist SR 141716A completely abolished this effect of HU-210. Selective CB2 receptor antagonist SR 144528 had no effect on HU-210-induced bradycardia. Preinjection of ganglioblocker hexamethonium also did not abolish the negative chronotropic effect of HU-210 and ACPA. Perfusion of isolated rat heart with Krebs-Henseleit solution containing HU-210 in a final concentration of 100 nM reduced heart rate. It was shown that the negative chronotropic effect of cannabinoids is mediated through activation of cardiac CB1 receptors.

[The effect of K(ATP)-channel activation on the electrical stability of myocardium in rats with postinfarction cardiosclerosis]. / Vliianie aktivatsii K(ATP)-kanalov na élektricheskuiu stabil'nost' miokarda pri postinfarktnom kardioskleroze.

Opening of the ATP-dependent K-channels (K(ATP) channels) upon intravenous administration of the cardioselective activator BMS 180448 (3 mg/kg) decreased the ventricular fibrillation threshold (VFT) in rats with postinfarction cardiosclerosis (PIC). Preliminary injection of the selective K(ATP) channel blocker glibenclamide (0.3 mg/kg, i.v.) completely abolished the profibrillatory effect of BMS 180448. At the same time, the mitochondrial K(ATP) channel blocker 5-hydroxydecanoic acid (5 mg/kg) did not influence the proarrhythmogen activity of BMS 180448. Simultaneous administration of the sarcoK(ATP) channel inhibitor HMR 1098 (3 mg/kg) and BMS 180448 increased the VFT up to a level in intact animals. Administration of the mitoK(ATP) channel activator diazoxide (5 mg/kg) after preliminary treatment with guanethidine (50 mg/kg) increased the VFT in rats with PIC. It is concluded that opening of the mitoK(ATP) channels increases the cardiac electrical stability in rats with PIC.

[Lipid spectrum of lymphocyte plasma membranes in abstinent syndrome in patients with opioid dependence]. / Spektr lipidov plazmaticheskikh membran pri abstinentnom sindrome u bol'nykh opioidnoi narkomaniei.

The lipid composition of the lymphocyte plasmatic membranes was studied in the withdrawal syndrome as observed in patients with opioid addiction. The method of thin-layer chromatography was used to identify the lipid fractions. Higher cholesterol content, a bigger trans-membrane asymmetry index and a higher ratio between cholesterol and total phospholipids were detected. The contents of the lysophospholipid fraction and of the total fraction of phosphatidylcholine and serine were increasing. The contents of cholesterol ethers as well as of the fractions of sphingomyelin and phosphatidyl ethanolamine were found to be lower.