Functional Activity Of P-glycoprotein Research Articles

A hydrogel model of the human blood-brain barrier using differentiated stem cells.

An in vitro model of the human blood-brain barrier was developed, based on a collagen hydrogel containing astrocytes, overlaid with a monolayer of endothelium, differentiated from human induced pluripotent stem cells (hiPSCs). The model was set up in transwell filters allowing sampling from apical and basal compartments. The endothelial monolayer had transendothelial electrical resistance (TEER) values >700Ω.cm2 and expressed tight-junction markers, including claudin-5. After differentiation of hiPSCs the endothelial-like cells expressed VE-cadherin (CDH5) and von-Willebrand factor (VWF) as determined by immunofluorescence. However, electron microscopy indicated that at set-up (day 8 of differentiation), the endothelial-like cells still retained some features of the stem cells, and appeared immature, in comparison with primary brain endothelium or brain endothelium in vivo. Monitoring showed that the TEER declined gradually over 10 days, and transport studies were best carried out in a time window 24-72hrs after establishment of the model. Transport studies indicated low permeability to paracellular tracers and functional activity of P-glycoprotein (ABCB1) and active transcytosis of polypeptides via the transferrin receptor (TFR1).

Rapid Increase of Gastrointestinal P-Glycoprotein Functional Activity in Response to Etoposide Stimulation.

We previously reported that exposure of human colon adenocarcinoma (Caco-2) cells to the bitter substance phenylthiocarbamide (PTC) rapidly enhanced the transport function of P-glycoprotein (P-gp). In this study, we investigated the short-term effect of etoposide, another bitter-tasting P-gp substrate, on P-gp transport function in the same cell line. We found that etoposide exposure significantly increased both the P-gp protein level in the plasma membrane fraction and the efflux rate of rhodamine123 (Rho123) in Caco-2 cells within 10 min. The efflux ratio (ratio of the apparent permeability coefficient in the basal-to-apical direction to that in the apical-to-basal direction) of Rho123 in etoposide-treated cells was also significantly increased compared with the control. These results indicated that etoposide rapidly enhances P-gp function in Caco-2 cells. In contrast, P-gp expression in whole cells at both the mRNA and protein level was unchanged by etoposide exposure, compared with the levels in non-treated cells. Furthermore, etoposide increased the level of phosphorylated ezrin, radixin and moesin (P-ERM) proteins in the plasma membrane fraction of Caco-2 cells within 10 min. P-gp functional changes were blocked by YM022, an inhibitor of cholecystokinin (CCK) receptor. These results suggest that etoposide induces release of CCK, causing activation of the CCK receptor followed by phosphorylation of ERM proteins, which recruit intracellular P-gp for trafficking to the gastrointestinal membrane, thereby increasing the functional activity of P-gp.

Functional activity of p-glycoprotein in blood-brain barrier during experimental par-kinson's syndrome

Background. P-glycoprotein (Pgp, ABCB1-protein) is a membrane transporter with broad substrate specificity that is localized in hepatocytes, enterocytes, epithelial renal tubules, and also in tissue barriers, including blood-brain barrier (BBB). Increased Pgp activity in BBB is one of the reasons for the pharmacoresistance of a number of CNS diseases.
 Aim. Analysis of Pgp functional activity in BBB during experimental Parkinson's syndrome.
 Materials and Methods. The work was performed on 90 Wistar rats, divided into 3 series (n=30 in each). The 1 series (control) was subcutaneously injected sunflower oil once a day for 7 days, and Pgp activity in BBB was assessed on the 8th day. The 2 and 3 series (pathology control) - were administered rotenone at a dose of 2.5 mg/kg once a day for 7 and 28 days respectively to simulate parkin-sonism. At the end of the experiment Pgp activity was estimated. To confirm Parkinson's syndrome, in addition to the clinical picture, level of dopamine in midbrain and striatum was determined using enzyme-linked immunosorbent assay. Pgp functional activity in BBB was assessed by the degree of penetration of its marker substrate fexofenadine into the brain after its intravenous administration at a dose of 10 mg/kg. The content of fexofenadine in the blood plasma and in brain tissue was estimated by the area under pharmacokinetic curve of the substance (in the blood or brain tissues) - AUC0-t(plasma) or AUC0-t(brain) respectively. To assess the BBB permeability the ratio AUC0-t(brain) / AUC0-t(plasma) was calculated.
 Results. Rotenone administration led to the development of parkinsonism typical picture: muscle stiffness, hypokinesia, gait instability. There was a decrease in dopamine level in the striatum after 7 days by 69.6% (p=0.095), after 28 days - by 93.9% (p=0.008), in midbrain - by 72.7% (p=0.095) and 68.7% (p=0.032) respectively. Fexofenadine AUC0-t(plasma) and AUC0-t(brain) after its intravenous administration to control rats were 266.2 (246.4; 285.6) μg/ml*min and 5.9 (5.8;6.6) µg/g*min respectively, AUC0-t(brain) /AUC0-t(plasma) - 0.020 (0.019; 0.022). When rotenone was for 7 days administered - fexofenadine AUC0-t(brain) increased 2.02 times (p=0.0163), AUC0-t(brain) / AUC0-t(plasma) - 2.4 times (p=0.0283). 28 days administration of rotenone led to augmentation of AUC0-t(brain) of fexofenadine by 1.75 times (p=0.0283), AUC0-t(brain) / AUC0-t(plasma) - by 2.27 times (p=0.0163).
 Conclusions. The development of Parkinson’s syndrome, caused by the administration of rotenone, inhibits Pgp functional activity in BBB, which is confirmed by the accumulation in the brain marker substrate of the transporter - fexofenadine.

P-glycoprotein functional activity in «soviet chinchilla» rabbits during pregnancy

P-glycoprotein (Pgp, ABCB1-protein) is a membrane transporter protein that plays the key role in pharmacokinetics of drugs with a broad spectrum of action. Substrates of this transporter are some medical drugs (antibacterial, antiretroviral, hypotensive) that are prescribed to pregnant women for long-term intake, sometimes throughout the whole gestation period.
 Aim to study the activity of Pgp on the organism level in rabbits of Soviet Chinchilla breed in pregnancy.
 Materials and Methods. The study was performed on 21 Soviet Chinchilla female rabbits (3000-3500 g). The animals were divided into 3 series. The first series (n=6) included rabbits with 7 days of pregnancy; the second series (n=5) - animals with 14 days of pregnancy; the third series (n=10) - rabbits with 21 days of pregnancy. 7 Days before the study and in the indicated gestation periods, functional activity of Pgp was assessed by the pharmacokinetics of marker transporter substrate – fexofenadine, after its single oral introduction (67.5 mg/kg). Besides, serum concentrations of progesterone, estradiol, testosterone and prolactin were determined by radio immune method
 Results. In all the studied gestational periods, serum concentrations of estradiol, testosterone and prolactin did not significantly differ from those before pregnancy, but the level of progesterone in blood serum was significantly elevated above the norm. On the 7th day of pregnancy pharmacokinetic parameters of fexofenadine did not show any reliable changes as compared to the initial va-lues. On the 14th day of pregnancy a reliable increase in Cmax, AUC0-t, T1/2 of fexofenadine was noted as compared to the parameters before pregnancy, which indicates a decrease in Pgp functional activity on the organism level. On the 21st day of pregnancy Cmax of fexofenadine remained elevated. Other pharmacokinetic parameters of fexofenadine did not show reliable changes. Conclusion. Reduction in Pgp functional activity, determined by the pharmacokinetics of its marker substrate (fexofenadine), was noted in rabbits of Soviet Chinchilla breed on the 14th and 21st days of pregnancy with the underlying significant increase in progesterone level.

Experimental Studies of the Effects of Progesterone on the Functional Activity of P-Glycoprotein

Experiments on female Chinchilla rabbits addressed the effects of progesterone on the functional activity of P-glycoprotein (Pgp). Animals were divided into four groups: animals of group 1 underwent sham operations; group 2 underwent ovariectomy; groups 3 and 4 underwent ovariectomy followed 15 days later by p.o. treatment with progesterone (2 and 15 mg/rabbit respectively). The functional activity of Pgp was determined seven days before the experiment started and on post-operative days 14, 28, and 42 by analysis of the pharmacokinetics of its marker substrate fexofenadine and serum sex hormone concentrations. At the end of the study, animals of each group were harvested by air embolism and specimens of large intestine were collected for immunohistochemical analysis. Ovariectomy led to a decrease in the functional activity of Pgp and a decrease in the staining intensity of the apical and lateral membranes of enterocytes, providing indirect evidence of a decrease in the expression of the transporter protein. Administration of progesterone at a dose of 2 mg/rabbit on the background of ovariectomy increased the functional activity of Pgp from the level seen in the ovariectomy group, though its activity remained decreased compared with the baseline level. This group showed no changes in the staining intensity of enterocyte membranes as compared with the ovariectomy group. Use of progesterone at a dose of 15 mg/rabbit after ovariectomy increased the functional activity of Pgp and the staining intensity of enterocyte apical and lateral membranes from the level seen in the ovariectomy group, restoring the level of transporter protein activity to that of intact animals, probably because of increased expression.

Влияние эстрадиола на функциональную активность гликопротеина-Р в эксперименте

Цель - изучение функциональной активности белка-транспортера гликопротеина-Р (Pgp) при овариоэктомии и последующем введении эстрадиола в эксперименте in vivo . Методика. Исследование выполнено на 23 кроликах-самках Шиншилла массой 4300-4700 г. 1-я группа - ложнооперированные животные, 2-я - овариоэктомия, кроликам 3-й и 4-й групп с 15-х сут. после овариоэктомии перорально вводили эстрадиол в дозах 0,5 мг и 2 мг, соответственно. За 7 сут. до начала исследования, на 14-е, 28-е и 42-е сут. после оперативного вмешательства у всех животных определяли функциональную активность гликопротеина-Р по фармакокинетике его маркерного субстрата фексофенадина методом ВЭЖХ и сывороточные концентрации половых гормонов (эстрадиола, прогестерона, тестостерона) радиоиммунным методом. Результаты. Овариоэктомия приводила к снижению функциональной активности гликопротеина-Р, что проявлялось изменением фармакокинетики его маркерного субстрата фексофенадина: повышением С, AUC и снижением Cl. Введение эстрадиола в дозе 0,5 мг в течение 28 сут. на фоне овариоэктомии повышало функциональную активность гликопротеина-Р по сравнению с показателями группы «овариоэктомия», однако активность белка-транспортера оставалась сниженной по сравнению с исходными данными. Применение эстрадиола в дозе 2 мг на фоне овариоэктомии повышало функциональную активность гликопротеина-Р по сравнению с показателями группы «овариоэктомия», и восстанавливало активность белка-транспортера до показателей интактных животных. Корреляционных зависимостей между содержанием эстрадиола и функциональной активностью гликопротеина-Р выявлено не было. Заключение. Введение эстрадиола в низкой дозе стимулирует Pgp на уровне организма, но недостаточно для восстановления его исходной активности. Возможно, для полного восстановления функциональной активности Pgp до исходного уровня необходимо дополнительное введение прогестерона. При введении эстрадиола в высокой дозе, активность Pgp повышается до исходного, дооперационного уровня даже без нормализации содержания прогестерона. The aim of the study was to evaluate the P-glycoprotein functional activity after ovariectomy followed by estradiol treatment. Methods. The study was conducted on 23 Chinchilla rabbits weighing 4300-4700 g. Group 1 underwent a sham surgery; Group 2 was ovariectomized; Groups 3 and 4 underwent ovariectomy followed by oral estradiol treatment (0.5 mg and 2 mg, respectively). On day 7 prior to the experiment and on days 14, 28, and 42 after the surgery, P-glycoprotein functional activity was determined in rabbits of all groups by measuring fexofenadine pharmacokinetics using HPLC and serum concentrations of sex hormones (estradiol, progesterone, testosterone) using the radio immune method. Results. Ovariectomy decreased the P-glycoprotein functional activity as evident from changes in the fexofenadine pharmacokinetics - increased C and AUC and decreased Cl. The estradiol administration at a dose of 0.5 mg for 28 days after ovariectomy increased the P-glycoprotein functional activity compared to the ovariectomy group; however, the P-glycoprotein functional activity remained lower than at baseline. The estradiol treatment at a dose of 2 mg after ovariectomy increased the P-glycoprotein functional activity compared to the ovariectomy group and restored the P-glycoprotein functional activity to the value for intact animals. No correlations were found between the maintenance of estradiol content and the P-glycoprotein functional activity. Conclusion. The treatment with low-dose estradiol stimulated Pgp at the level of a whole body but was not sufficient to restore its original activity. Perhaps, for a complete recovery of the Pgp functional activity to the baseline level, additional administration of progesterone is required. When estradiol was administered at a high dose the Pgp activity increased to the baseline, preoperative level even without normalization of the progesterone content.

Влияние прогестерона на функциональную активность гликопротеина-P в эксперименте

В исследовании на кроликах-самках породы шиншилла изучено влияние прогестерона на функциональную активность гликопротеина-P (Pgp). Животные были разделены на 4 группы: первой группе животных была проведена «ложная операция»; второй группе – овариоэктомия; третьей и четвертой группе проводили овариоэктомию и с 15 сут после операции per os вводили прогестерон (2 и 15 мг/кролик соответственно). За 7 сут до начала опыта, на 14, 28 и 42 сут после операции определяли функциональную активность Pgp по анализу фармакокинетики его маркерного субстрата фексофенадина и сывороточные концентрации половых гормонов. В конце исследования животных каждой группы выводили из эксперимента методом воздушной эмболии и забирали образец тощей кишки для иммуногистохимического анализа. Овариоэктомия приводила к снижению функциональной активности Pgp и уменьшению интенсивности окрашивания апикальной и латеральных мембран энтероцитов, косвенно свидетельствующей о снижении экспрессии белка-транспортера. Введение прогестерона в дозе 2 мг/кролик на фоне овариоэктомии повышало функциональную активность Pgp по сравнению с показателями группы овариоэктомии, однако его активность оставалась сниженной по сравнению с исходными данными. Также в данной группе не выявлено изменения интенсивности окрашивания мембран энтероцитов в сравнении с серией овариоэктомии. Применение прогестерона в дозе 15 мг/кролик при овариоэктомии повышало функциональную активность Pgp и интенсивность окрашивания апикальной и латеральных мембран энтероцитов по сравнению с показателями группы овариоэктомии, восстанавливая активность белка-транспортера до показателей интактных животных, вероятно, за счет повышения его экспресии.

Effects of Testosterone on the Functional Activity of P-Glycoprotein

The effects of testosterone undecanoate on the functional activity of P-glycoprotein (ABCB1 protein) were studied in male Chinchilla rabbits. Testosterone was given as single intramuscular doses of 12 or 24 mg/kg. The functional activity of P-glycoprotein was determined in terms of the pharmacokinetics of its marker substrate - fexofenadine - on days 7, 14, and 21 after hormone administration. Administration of testosterone at a dose of 12 mg/kg (n = 7) increased the C max of fexofenadine on days 7 and 14 and AUC 0-t on day 14 and decreased total clearance on day 14. Use of testosterone at a dose of 24 mg/kg (n = 7) led to increases in the C max of fexofenadine on day 7 and AUC 0-t on days 7, 14, and 21 and decreased total clearance on days 7 and 14. These changes in the pharmacokinetics of fexofenadine provide evidence of decreases in the functional activity of Pgp on the background of testosterone administration. Correlational relationships were found between measures of the pharmacokinetics of fexofenadine and serum testosterone levels.

Влияние афобазола на функциональную активность и экспрессию гликопротеина-P в эксперименте

На 17 кроликах-самцах породы Шиншилла изучено влияние афобазола (5-этокси-2-[2-(морфолино)этилтио]бензилимидазола дигидрохлорида) в дозе 3,8 мг/кг на функциональную активность эффлюксного белка-транспортера гликопротеина-P и его экспрессию в печени и тонком кишечнике. Активность белка-транспортера оценивали по фармакокинетике его маркерного субстрата — фексофенадина после однократного внутрижелудочного введения. Уровень экспрессии гликопротеина-P определяли иммуногистохимически. Установлено, что введение афобазола (5-этокси-2-[2-(морфолино)этилтио]бензилимидазола дигидрохлорида) в дозе 3,8 мг/кг внутрь 3 раза в день в течение 14 дней приводило к повышению площади под фармакокинетической кривой концентрация — время фексофенадина и снижению общего клиренса, что свидетельствует об ингибировании функциональной активности белка-транспортера на уровне целостного организма. Иммуногистохимическое исследование показало уменьшение экспрессии гликопротеина-P в печени кролика на фоне курсового введения афобазола.

The role of testosterone in the regulation of P-glycoprotein functioning

The functional activity of P-glycoprotein (Pgp) was studied in Chinchilla rabbits after orchiectomy and administration of testosterone undecanoate after surgery. The functional activity of Pgp was determined by the pharmacokinetics of fexofenadine, its marker substrate. In addition, the localization of the transport protein was assessed in the liver, kidney, and jejunum by the immunohistochemistry. It was found that a decrease in the serum testosterone concentration after orchiectomy leads to an increase in Pgp activity on the 14th and 21st days after the surgery, which is accompanied by an increase in the staining intensity of Pgp-positive epithelium membranes of the proximal and distal renal tubules. It was found that the administration of testosterone at a dose of 6 mg/kg body weight in the period after the surgery neutralizes these changes.

INFLUENCE OF VILDAGLIPTIN AND GLIQUIDONE COMBINATION ON P-GLYCOPROTEIN FUNCTIONAL ACTIVITY AND EXPRESSION ON THE BACKGROUND OF NORM AND IN EXPERIMENTAL ALLOXAN-INDUCED DIABETES MELLITUS TYPE 2

In the in vivo study on rabbits investigated the effect of the vildagliptin and gliqui-done combination on the background of norm and experimental alloxan-induced diabetes mellitus (DM) type 2 on the P-glycoprotein (P-gp, ABCB1 protein) functional activity and its expression in the tissues of the jejunum, liver, kidney and the blood-brain barrier. Introduction of combination of vildagliptin at a dose of 5 mg/kg and gliquidone 10 mg/kg body weight on the background of norm for 14 days did not result in P-glycoprotein changes. On the 5-day of the cancellation of this combination observed inhibition of transporter protein functional activity without altering its expression. In the combination therapy of vildagliptin and gliquidone alloxan-induced diabetes type 2 showed complete recovery of P-glycoprotein functional activity and expression reduced on the pathology background. On the 5-day of treatment withdrawal again occurred inhibition of functional activity of the transporter protein and a decrease in its expression.

The transport activity of P-glycoprotein upon a change of the redox balance in lymphocytes of patients with chronic B-lymphocytic leukemia

The effect of drugs used in the treatment of chronic B-lymphocytic leukemia on the functional activity of P-glycoprotein, which is a membrane transporter associated with the phenomenon of multiple drug resistance, in B-lymphocytes of leukemia patients has been investigated. The level of reactive oxygen species and the content of low molecular weight antioxidants in the leukemic cells during the metabolic processing of chemotherapy drugs has been assessed. The degree of involvement of low molecular weight antioxidants in the maintenance of redox balance in the lymphocytes of drug-treated leukemia patients has been characterized. Variation of the levels of reactive oxygen species in leukemic B-lymphocytes within a certain range, as well as the subsequent return of these values to the levels of intact cells, was shown to enhance the P-glycoprotein transport activity. The viability of leukemic B-cells decreased upon exposure to antitumor compounds and depended on the redox balance. No statistically significant correlation between the P-glycoprotein transport activity and the viability of lymphocytes during the detoxification from drugs in B-cell chronic lymphocytic leukemia patients has been detected; however, the transport activity of the protein was directly correlated to the viability of leukemia cells within 15 h after exposure of the cells to H2O2.

Influence of different hypoxia types on p-glycoprotein functional activity and expression

The review of literature is devoted to researches in vitro and in vivo in which functioning changes P-glycoprotein (Pgp) under the influence of hypoxia of different types are shown. In the majority of scientific works activation of expression and functional activity of Pgp is shown that authors connect with activation of HIF-1 and Sp1 which are described in more detail. However a number of researches disproves the conclusion about induction of activity of Pgp in the conditions of hypoxia that testifies about difficult mechanisms involved in regulation of Pgp activity, their dependence on a type and duration of hypoxia, redox status and other parameters.

Оценка принадлежности мексидола к субстратам, ингибиторам или индукторам гликопротеина-P

На 18 половозрелых кроликах-самцах породы шиншилла изучена принадлежность препарата мексидол к числу субстратов, ингибиторов или индукторов гликопротеина-P. Для оценки принадлежности мексидола к субстратам гликопротеина-P изучали его концентрацию в плазме крови животных методом ВЭЖХ с УФ-детектированием до и после курсового введения рифампицина и верапамила в качестве соответственно индуктора и ингибитора транспортера. Влияние мексидола на функциональную активность гликопротеина-P в плазме крови кроликов оценивали по фармакокинетике его маркерного субстрата — фексофенадина, концентрацию которого определяли методом ВЭЖХ с УФ-детектированием. Установлено, что мексидол не является субстратом гликопротеина-P, но оказывает ингибирующее влияние на его функциональную активность.

FUNCTIONAL ACTIVITY OF P-GLYCOPROTEIN DURING EXPERIMENTAL MANIPULATIONS

In a study on rabbits the effect of experimental manipulations (false surgery and distilled water administration) on functional P-glycoprotein activity was studied. P-glycoprotein activity was assessed by pharmacokinetic analysis of its marker substrate - fexofenadine. Found that manipulations applied do not lead to modulation of Pgp activity at all stages of the research period.

Development, validation, and application of a novel 7-day Caco-2 cell culture system

IntroductionMonolayers of Caco-2 cells have been widely accepted as one of the well-established in vitro models to predict intestinal drug permeability and absorption in humans. However, the procedure for culturing the traditional 21-day Caco-2 model is labor intensive and time consuming, which limits its wide application in drug development. The objective of the present study was to develop a rapid Caco-2 model with a 7-day cell culture process. MethodsA few modifications of the BIOCOAT® HTS Caco-2 Assay System were introduced including changing the cell seeding density, the composition of cell culture media, and the interval to change media. The monolayer structure was visualized through confocal microscopy. The transepithelial electrical resistance (TEER), apparent permeability coefficients (Papp), and functional activity of P-glycoprotein (P-gp) were determined and compared with the 21-day model. Moreover, pharmacokinetic studies were performed in animal models to evaluate the absolute bioavailability of oral doses (Foral) of different compounds. ResultsOur newly developed 7-day Caco-2 model displayed comparable cellular morphology and integrity with the traditional 21-day model. No significant difference in paracellular and transcellular permeability was observed between the two systems. The efflux ratios of transporting digoxin, the prototypical substrate of P-gp in 21- and 7-day systems were 16.04 and 24.92, respectively. When the P-gp inhibitor verapamil was present, the efflux ratios of 21- and 7-day systems were 1.37 and 0.86, respectively, suggesting the comparability of the P-gp functional activity in both systems. Furthermore, pharmacokinetic studies of several compounds performed in animal models revealed that the absolute bioavailability of oral doses in vivo was well correlated with the Caco-2 permeability in vitro. DiscussionThe novel system provides a rapid and economical option for assessing the drug permeability, and is applicable to the studies of intestinal drug absorption.

Flavonoid compounds as reversal agents of the P-glycoprotein-mediated multidrug resistance: biology, chemistry and pharmacology

Multidrug resistance (MDR) represents one of the major problems in pharmacotherapy of important diseases (e.g., cancer, epilepsy). Although many factors may contribute to the development of MDR phenotype, the increased expression and/or functional activity of P-glycoprotein (P-gp; active drug efflux transporter) across the cell membrane has been recognized as the main one. Therefore, a great attention has been given to the search of P-gp inhibitors as therapeutic agents to reverse the MDR mediated by P-gp. Since the chemical entities identified over the last three decades as potential P-gp inhibitors did not show suitable pharmacological properties, more recently herbal components, such as flavonoid compounds, have gained a great interest as safe P-gp inhibitors. The interest in flavonoids as P-gp inhibitors is increasing due to their potential favourable characteristics, including selectivity and non-cytotoxic effects. Flavonoids integrate the third-generation non-pharmaceutical category of P-gp inhibitors, and some of them exhibited effects comparable to those of the classic P-gp inhibitors. In fact, some flavonoids found in foods and beverages of herbal origin appear to be quite promising to inhibit the P-gp–mediated drug efflux, indicating their potential value to enhance the systemic/cellular bioavailability of P-gp drug substrates when administrated in co-therapy. This review paper summarizes the current evidence of P-gp inhibitory effects produced by flavonoids, taking into account studies performed in cell-based in vitro models, in vivo animal models and clinical trials.

Effects of serum lipoproteins on cyclosporine A cellular uptake and renal toxicity in vitro

In-vitro studies were performed to shed light on previous findings that showed increased uptake of cyclosporine A in the kidneys and liver of hyperlipidemic rats, and increased signs of kidney toxicity. Hepatocytes were obtained from rats, cultured, and exposed to a diluted serum from hyperlipidemic rats. Some cells were also exposed to lipid-lowering drugs. After washing out the rat serum or lipid-lowering drugs, cells were exposed to cyclosporine A embedded in serum lipoproteins. Pretreatment with hyperlipidemic serum and lipid-lowering drugs was associated with an increased uptake of cyclosporine A. As expected, atorvastatin caused an increase in low density lipoprotein receptor and a decrease in MDR1A mRNA in the hepatocytes. A decrease in NRK-52E rat renal tubular cellular viability caused by cyclosporine A was noted when cells were preincubated with diluted hyperlipidemic serum. This was matched with evidence of hyperlipidemic-serum-associated increases in the NRK-52E cellular uptake of cyclosporine A and rhodamine-123. The findings of these experiments suggested that in hyperlipidemia the expression and (or) the functional activity of P-glycoprotein was diminished, leading to greater hepatic and renal uptake of cyclosporine A, and renal cellular toxicity.

Oral hormonal contraceptive "janine" influence on P-glycoprotein functional activity

In the research on rabbits influence of combined oral contraceptive, "Janine" on the functional activity of the protein efflux transporter P-glycoprotein (Pgp) was studied. Pgp activity was assessed by pharmacokinetic analysis of marker substrate fexofenadine. It was found that administration of the drug "Janine" for 14 and 21 days, resulting in inhibition of Pgp functional activity.

An immortalised astrocyte cell line maintains the in vivo phenotype of a primary porcine in vitro blood–brain barrier model

Whilst it is well documented that all components of the neurovascular unit contribute to the restrictive nature of the blood–brain barrier (BBB), astrocytes have been identified as the cellular component most likely to play an essential role in maintaining the barrier properties. The aim of this study was to examine the impact of the rat astrocyte cell line, CTX-TNA2, on the structural and functional characteristics of an in vitro BBB and determine the capacity of this astrocyte cell line to maintain the BBB phenotype. Co-culture of the CTX-TNA2 cells with primary porcine brain endothelial cells produced an in vitro BBB model which retains key features of the in vivo BBB. High transendothelial electrical resistances, comparable to those reported in vivo, were obtained. Ultrastructural analysis revealed distinct intercellular tight junction protein complexes and immunocytochemistry confirmed expression of the tight junction proteins ZO-1 and occludin. Western blotting and fluorescent tracer assays confirmed expression and functional activity of P-glycoprotein (ABCB1) and breast cancer resistance protein (ABCG2) efflux transporters. Studies employing Alexa-fluor 555-conjugated human transferrin revealed temperature-sensitive internalisation indicating the BBB model retains functional receptor-mediated transferrin uptake. The findings of this study indicate that a robust BBB model has been produced and this is the first report of the inductive capacity of the CTX-TNA2 cell line. Since this in vitro BBB model possesses many key characteristics of the BBB in vivo it has the potential to be a valuable tool for the study of biochemical and physiological processes associated with the BBB.