Background. Glycoprotein-P (Pgp, АВСВ1) is a transporter protein participating in pharmacokinetics of medical drugs, and also in development of resistance of tumor cells to chemotherapy. Aim. To study the influence of progesterone on the activity of Pgp in vitro on a cell model of human small intestinal epithelium. Materials and Methods. The work was conducted on Caco-2 cells. The activity of Pgp was evaluated by transport of fexofenadine in a special transwell-system. Concentration of fexofenadine was analyzed by HPLC method. The amount of Pgp was determined by EIA method. Four series of experiments were conducted: control cells preincubated with clean transport medium without addition of any substances; influence of rifampicin on the activity and synthesis of Pgp in the concentration 10 mol/l in preincubation for 3 days (induction control); influence of progesterone on the activity of Pgp in concentrations 1, 10 and 100 mol/l in preincubation for 30 min; influence of progesterone on the activity and synthesis of Pgp in concentrations 1, 10 and 100 mol/l in preincubation for 3 days. Results. Progesterone in the concentrations 1 and 10 M in incubation with cells within 30 minutes did not show any reliable influence on the activity of Pgp, however, in concentration 100 M it reduced the activity of the transporter protein. In incubation of Caco-2 cells with progesterone in concentrations 1, 10 and 100 M within 3 days the activity of Pgp remained unchanged. Progesterone in concentration 100 M in incubation within 3 days significantly increased synthesis of Pgp in enterocytes by 114.3% as compared to control, and in other used concentrations (1 and 10 M) it produced no reliable effect. Conclusion. In in vitro experiments on Caco-2 cells progesterone in concentration 100 M produces a direct inhibiting effect on the activity of Pgp; however, in incubation within 3 days it increases synthesis of the transporter protein, which cancels out its inhibitory activity.

Objective. To study the effect of Mexidol on the level of factors regulating neuroregenesis. Material and methods. The study was performed on male Wistar rats. Focal cerebral ischemia was reproduced by endovascular occlusion-reperfusion of the right medial cerebral artery. The duration of occlusion was 60 min. During the beginning of reperfusion, the animals were given a single intravenous injection of saline solution or the drug Mexidol at a dose of 50 mg/kg. 4, 8 and 24 hours after the beginning of reperfusion in the ischemic hemisphere, the relative amount of molecules regulating neurogenesis was estimated by Western blot method. Additionally, 24 hours after the start of reperfusion, the size of the brain infarction was analyzed after staining with a 1% solution of 2.3,5,-triphenyltetrazolium. Results. When modeling occlusion-reperfusion of the middle cerebral artery, the volume of necrosis in the affected hemisphere of animals injected with saline was 37.75±7.46%, administration of Mexidol at a dose of 50 mg /kg led to a decrease in the volume of necrosis to 20.48±2.33% (p=0.0006). The simulation of occlusion-reperfusion of the middle cerebral artery was accompanied by activation of neurotrophic factors IGF-1, NGF and vascular factor VEGF. A single intravenous administration of Mexidol at a dose of 50 mg/kg during reperfusion increased the level of neurotrophic factors IGF-1, NGF, BDNF and VEGF in the ischemic area of the brain compared with the administration of saline, which leads to increased neuroregeneration at all follow-up periods (4, 8 and 24 hours after reperfusion), the marker of which is tubulin-3. Conclusion. The results suggest that Mexidol not only has a protective effect on neurons, but can also stimulate neuroregeneration by increasing the level of the main regulatory molecules.

The article describes modern approaches for testing of drugs belonging to substrates and inhibitors of P-glycoprotein (Pgp, ABCB1-protein, MDR1-protein) according to the recommendations of Food and Drug Administration (United States) and European Medicines Agency. In vitro methods on cell lines with hyperexpression of the transporter are presented. The same analysis was done on human colon adenocarcinoma cell line (Caco-2).

The article presents an original method for determining belonging of drugs to the substrates of the transport protein P-glycoprotein (Pgp). This methodics is based on a comparison of the pharmacokinetic parameters of the test substrate before and after a course of administration of the inductor (rifampicin) and inhibitor (verapamil) of the transport protein. If the content of test substance in organism is reduced after Pgp inductor administration, and increased after the inhibitor administration, we conclude that this substance is Pgp substrate. For approbation of developed technique we examined by blind method the pharmacokinetics of fexofenadine - known Pgp substrate at a dose of 67,5 mg/kg, before and after the 14-days administration of rifampicin (40 mg/kg twice daily) and verapamil (a daily dose of 20 mg/kg three times a day). It was revealed that after a course administration of verapamil maximum concentration of fexofenadine, its area under the concentration-time curve and half-life were significantly increased, as well as the time of maximum concentration was decreased. In contrast, rifampicin course administration resulted in a decrease in the area under the curve concentration-time and an increase in total clearance of marker substrate of Pgp, wich proves the adequacy of methods.

The investigation of pharmacokinetics of marker substrates of carrier protein P-glycoprotein (Pgp, ABCB1-protein) including fexofenadine, is one of the methods of its functional activity evaluation.The aim of the study was to work out the HPLC methods of the quantitative determination of fexofenadine in rabbits’ liver.Materials and methods. The quantitative determination of fexofenadine was performed using Stayer chromatographic system (Akvilon, Russia) with UVV 104 ultraviolet detector. Reverse-phased chromatographic column Luna C18 100Å (250*4.6) was used with 5 µm granulation at 45°С. The сoncentration of fexofenadine was determined by methods of absolute peak area calibration.Results. The work was conducted in the isocratic mode. The composition of the mobile phase consisted of deionized water, acetonitrile and glacial acetic acid at the ratio of 267.4:120:4.33 brought to pH=6.7 with triethylamine. The sample processing was in the form of homogenization of 500 mg of ground liver in 500 µl of purified water with the subsequent centrifugation (1750 g) and selection of the supernatant. The proteins were precipitated by acetonitrile (2.5 ml) acidified with 375 µl of hydrochloric acid by shaking at 500 rev/min. The supernatant was transported into a separate test tube, where methylene chloride, diethyl ether and ethyl acetate were added (2 ml each). Then the solution was again shaken for 10 minutes (500 rev/min). After that, the solution was centrifuged (1750 g) and the supernatant was evaporated on a rotor-vacuum evaporator at 50°С. 300 µl of the mobile phase was added to the dry residue, and 100 µl was injected into the chromatograph. The method was validated in the linear range from 3 to 60 µg/g of fexofenadine with the acceptable intra- and intercycle accuracy, precision and stability. The method was tested on rabbits after the intravenous administration of fexofenadine at the dose of 11 mg/kg.Conclusion. The HPLC methods of fexofenadine quantitative determination in the hepatic tissue of rabbits has been worked out. It can be used for the evaluation of the functional activity of Pgp in preclinical studies.Abbreviations: Pgp – P-glycoprotein, HPLC – high performance liquid chromatography, rev/min – revolutions per minute