Maternal and Fetal Expression of ATP-Binding Cassette and Solute Carrier Transporters Involved in the Brain Disposition of Drugs.

Evidence from preclinical and clinical studies demonstrate that pregnancy is a physiological state capable of modifying drug disposition. Factors including increased hepatic metabolism and renal excretion are responsible for impacting disposition, and the role of membrane transporters expressed in biological barriers, including the placental- and blood-brain barriers, has received considerable attention. In this regard, the brain disposition of drugs in the mother and fetus has been the subject of studies attempting to characterize the mechanisms by which pregnancy could alter the expression of ATP-binding cassette (ABC) and solute carrier (SLC) transporters. This chapter will summarize findings of the influence of pregnancy on the maternal and fetal expression of ABC and SLC transporters in the brain and the consequences of such changes on the disposition of therapeutic drugs.

[Pharmacokinetic interactions of fruit juices and herbal preparations]. / Jugos de fruta y productos herbáceos como perpetradores de interacciones farmacocinéticas mediadas por enzimas metabolizadoras y transportadores de membrana. Relevancia en clínica.

Drug disposition in the human body is strongly influenced by transporters and metabolizing enzymes expressed in key organs including intestine, liver and kidney. Since drugs and chemicals present in foods such as fruit juices and herb-based products are substrates of the above-mentioned proteins, there is a high probability of pharmacokinetic interactions. Findings from preclinical and clinical studies helped to characterize the mechanisms by which the components of fruit juices and herbs act as perpetrators of pharmacokinetic interactions. The aim of this review is to provide an overview of pharmacokinetic fruit juice- and herb-drug interactions that could be relevant in the clinical setting.

Deletion of the adenosine A2A receptor increases the survival rate in a mice model of polymicrobial sepsis.

We aim to investigate the role of A2A receptor in peritonitis-related sepsis by injection of a fecal solution (FS) as a model of polymicrobial infection. C57/black J6 wild-type (WT) and A2A-deficient mice (A2AKO) were exposed to sepsis induced by intraperitoneal injection of a FS (FS-induced peritonitis) or instead was injected with saline buffer (Sham). Survival rate and sepsis score were measured up to 48 h. The presence of bacteria in tissue homogenates was analyzed. Telemetry and speckle laser Doppler were used for systemic blood pressure and peripheral blood perfusion analysis, respectively. Histological analysis and identification of active caspase 3 were performed in selected organs, including the liver. The survival rate of A2AKO mice exposed to FS-induced peritonitis was significantly higher, and the sepsis score was lower than their respective WT counterpart. Injection of FS increases (50 to 150 folds) the number of colonies forming units in the liver, kidney, blood, and lung in WT mice, while these effects were significantly attenuated in A2AKO mice exposed to FS-induced peritonitis. A significant reduction in both systolic and diastolic blood pressure, as well as in the peripheral perfusion was observed in WT and A2AKO mice exposed to FS-induced peritonitis. Although, these last effects were significantly attenuated in A2AKO mice. Histological analysis showed a large perivascular infiltration of polymorphonuclear in the liver of WT and A2AKO mice exposed to FS-induced peritonitis, but again, this effect was attenuated in A2AKO mice. Finally, high expression of active caspase 3 was found only in the liver of WT mice exposed to FS-induced peritonitis. The absence of the A2A receptor increases the survival rate in mice exposed to polymicrobial sepsis. This outcome was associated with both hemodynamic compensation and enhanced anti-bacterial response.

Investigating Maternal Brain Alterations in Preeclampsia: the Need for a Multidisciplinary Effort.

PURPOSE OF REVIEW: To provide insight into the mechanisms underlying cerebral pathophysiology and to highlight possible methods for evaluation, screening, and surveillance of cerebral complications in preeclampsia. RECENT FINDINGS: The pathophysiology of eclampsia remains enigmatic. Animal studies show that the cerebral circulation in pregnancy and preeclampsia might be affected with increased permeability over the blood-brain barrier and altered cerebral blood flow due to impaired cerebral autoregulation. The increased blood pressure cannot be the only underlying cause of eclampsia and cerebral edema, since some cases of eclampsia arise without simultaneous hypertension. Findings from animal studies need to be confirmed in human tissues. Evaluation of brain alterations in preeclampsia and eclampsia is challenging and demands a multidisciplinary collaboration, since no single method can accurately and fully describe how preeclampsia affects the brain. Cerebral complications of preeclampsia are significant factors in maternal morbidity and mortality worldwide. No single method can accurately describe the full picture of how preeclampsia affects the brain vasculature and parenchyma. We recommend an international and multidisciplinary effort not only to overcome the issue of limited sample availability but also to optimize the quality of research.

Regulation of the ATP-binding cassette transporters ABCB1, ABCG2 and ABCC5 by nuclear receptors in porcine blood-brain barrier endothelial cells.

BACKGROUND AND PURPOSE: Blood-brain barrier (BBB) ABCB1, ABCG2 and ABCC5 transporters influence central therapeutic drug distribution. Transporter expression is regulated by the NR3C1, NR1I3 and NR1I2 nuclear receptors, but their precise roles in brain are poorly understood. We investigated the effects of selective ligand-based activation of NR3C1, NR1I3, NR1I2 and NR2B1 in porcine brain endothelial cells (PBECs). EXPERIMENTAL APPROACH: Primary cultures of PBECs were exposed to NR3C1, NR1I3 and NR1I2 ligands and ABCB1, ABCG2 and ABCC5 transporter activities determined by measuring intracellular accumulation of fluorescent probes. Western blotting was used to determine the effects of receptor ligands on expression of ABCB1, ABCG2, ABCC5, NR1I2, NR1I3, NR3C1 and NR2B1. Fluorescent immunocytochemistry was employed to assess the effects of receptor ligands on the cellular localisation of NR1I2 and NR1I3. KEY RESULTS: The NR1I2 agonist rifampicin significantly up-regulated ABCG2 activity, which is counteracted by co-treatment with NR1I2 antagonist l-sulforaphane. The NR1I3 agonist 6-(4-chlorophenyl)-imidazo[2,1-b]thiazole-5-carbaldehyde and inverse agonist meclizine significantly down-regulated ABCB1, ABCG2 and ABCC5 activity. NR3C1 agonist dexamethasone significantly increased ABCB1, ABCG2 and ABCC5 activity and ABCG2 and ABCC5 protein expression, which was counteracted by co-treatment with the NR3C1 antagonist mifepristone. This first study demonstrates that NR1I3 and NR3C1 regulate ABCC5 activity and protein expression in BBB endothelial cells. CONCLUSIONS AND IMPLICATIONS: In PBECs, expression of key ATP-binding cassette (ABC) transporters and nuclear receptors is differentially regulated by NR1I3, NR1I2, NR3C1 and NR2B1. This will help to better understand the response of the BBB to physiological and pharmacological activation of nuclear receptors.

Brain Vascular Dysfunction in Mothers and Their Children Exposed to Preeclampsia.

Preeclampsia is a maternal syndrome characterized by the new onset of hypertension and proteinuria after 20 weeks of gestation associated with multisystemic complications, including brain alterations. Indeed, brain complications associated with preeclampsia are the leading direct causes of fetal and maternal morbidity and mortality, especially in low- and middle-income countries. In addition to the well-recognized long-term adverse cardiovascular effects of preeclampsia, women who have had preeclampsia have higher risk of stroke, dementia, intracerebral white matter lesions, epilepsy, and perhaps also cognitive decline postpartum. Furthermore, increasing evidence has also associated preeclampsia with similar cognitive and cerebral disorders in the offspring. However, the mechanistic links between these associations remain unresolved. This article summarizes the current knowledge about the cerebrovascular complications elicited by preeclampsia and the potential pathophysiological mechanisms involved, emphasizing the impaired brain vascular function in the mother and their offspring.

Dysregulation of vascular endothelial growth factor receptor 2 phosphorylation is associated with disruption of the blood-brain barrier and brain endothelial cell apoptosis induced by plasma from women with preeclampsia.

Disruption of the blood-brain barrier (BBB) is central in the pathophysiology of acute cerebral complications in women who have preeclampsia. Underling mechanisms are unclear. Using female human brain endothelial cells as an in vitro model of BBB, we show that plasma of women with preeclampsia increases cell apoptosis and permeability via activation of the vascular endothelial growth factor receptor 2 (VEGFR2). Since plasma of women with preeclampsia also enhanced VEGFR2 phosphorylation in the tyrosine 951 but decreased phosphorylation at the tyrosine 1175, we propose the former would be the more likely active form of VEGFR2 responsible for BBB alterations.

Cerebral Biomarkers and Blood-Brain Barrier Integrity in Preeclampsia.

Cerebral complications in preeclampsia contribute substantially to maternal mortality and morbidity. There is a lack of reliable and accessible predictors for preeclampsia-related cerebral complications. In this study, plasma from women with preeclampsia (n = 28), women with normal pregnancies (n = 28) and non-pregnant women (n = 16) was analyzed for concentrations of the cerebral biomarkers neurofilament light (NfL), tau, neuron-specific enolase (NSE) and S100B. Then, an in vitro blood−brain barrier (BBB) model, based on the human cerebral microvascular endothelial cell line (hCMEC/D3), was employed to assess the effect of plasma from the three study groups. Transendothelial electrical resistance (TEER) was used as an estimation of BBB integrity. NfL and tau are proteins expressed in axons, NSE in neurons and S100B in glial cells and are used as biomarkers for neurological injury in other diseases such as dementia, traumatic brain injury and hypoxic brain injury. Plasma concentrations of NfL, tau, NSE and S100B were all higher in women with preeclampsia compared with women with normal pregnancies (8.85 vs. 5.25 ng/L, p < 0.001; 2.90 vs. 2.40 ng/L, p < 0.05; 3.50 vs. 2.37 µg/L, p < 0.001 and 0.08 vs. 0.05 µg/L, p < 0.01, respectively). Plasma concentrations of NfL were also higher in women with preeclampsia compared with non-pregnant women (p < 0.001). Higher plasma concentrations of the cerebral biomarker NfL were associated with decreased TEER (p = 0.002) in an in vitro model of the BBB, a finding which indicates that NfL could be a promising biomarker for BBB alterations in preeclampsia.

How Soluble Fms-Like Tyrosine Kinase 1 Could Contribute to Blood-Brain Barrier Dysfunction in Preeclampsia?

Preeclampsia is a pregnancy-related syndrome that courses with severe cerebrovascular complications if not properly managed. Findings from pre-clinical and clinical studies have proposed that the imbalance between pro- and anti-angiogenic factors exhibited in preeclampsia is a major component of its pathophysiology. In this regard, measurement of circulating levels of soluble tyrosine kinase-1 similar to fms (sFlt-1), a decoy receptor for vascular endothelial growth factor (VEGF), is a moderately reliable biomarker for the diagnosis of preeclampsia. However, few studies have established a mechanistic approach to determine how the high levels of sFlt-1 are responsible for the endothelial dysfunction, and even less is known about its effects at the blood-brain barrier (BBB). Since the expression pattern of VEGF receptors type 1 and 2 in brain endothelial cells differs from the observed in peripheral endothelial cells, and components of the neurovascular unit of the BBB provide paracrine secretion of VEGF, this compartmentalization of VEGF signaling could help to see in a different viewpoint the role of sFlt-1 in the development of endothelial dysfunction. In this article, we provide a hypothesis of how sFlt-1 could eventually be a protective factor for brain endothelial cells of the BBB under preeclampsia.

Disruption of the Blood-Brain Barrier by Extracellular Vesicles From Preeclampsia Plasma and Hypoxic Placentae: Attenuation by Magnesium Sulfate.

[Figure: see text].

Preeclampsia and Increased Permeability Over the Blood-Brain Barrier: A Role of Vascular Endothelial Growth Receptor 2.

BACKGROUND: Cerebral complications in preeclampsia are leading causes of maternal mortality worldwide but pathophysiology is largely unknown and a challenge to study. Using an in vitro model of the human blood-brain barrier (BBB), we explored the role of vascular endothelial growth factor receptor 2 (VEGFR2) in preeclampsia. METHODS: The human brain endothelial cell line (hCMEC/D3) cultured on Tranwells insert was exposed (12 hours) to plasma from women with preeclampsia (n = 28), normal pregnancy (n = 28), and nonpregnant (n = 16) controls. Transendothelial electrical resistance (TEER) and permeability to 70 kDa fluorescein isothiocyanate (FITC)-dextran were measured for the assessment of BBB integrity. We explored possible underlying mechanisms, with a focus on the expression of tight junction proteins and phosphorylation of 2 tyrosine residues of VEGFR2, associated with vascular permeability and migration (pY951) and cell proliferation (pY1175). Plasma concentrations of soluble FMS-like tyrosine kinase-1 (sFlt-1) and placental growth factor (PlGF) were also measured. RESULTS: hCMEC/D3 exposed to plasma from women with preeclampsia exhibited reduced TEER and increased permeability to 70 kDa FITC-dextran. These cells upregulated the messenger ribonucleic acid (mRNA) levels of VEGFR2, and pY951-VEGFR2, but reduced pY1175-VEGFR2 (P < 0.05 in all cases). No difference in mRNA expression of tight junction protein was observed between groups. There was no correlation between angiogenic biomarkers and BBB permeability. CONCLUSIONS: We present a promising in vitro model of the BBB in preeclampsia. Selective tyrosine phosphorylation of VEGFR2 may participate in the increased BBB permeability in preeclampsia irrespective of plasma concentrations of angiogenic biomarkers.

The constitutive androstane receptor and pregnane X receptor in the brain.

Since their discovery, the orphan nuclear receptors constitutive androstane receptor (CAR;NR1I3) and pregnane X receptor (PXR;NR1I2) have been regarded as master regulators of drug disposition and detoxification mechanisms. They regulate the metabolism and transport of endogenous mediators and xenobiotics in organs including the liver, intestine and brain. However, with proposals of new physiological functions for NR1I3 and NR1I2, there is increasing interest in the role of these receptors in influencing brain function. This review will summarise key findings regarding the expression and function of NR1I3 and NR1I2 in the brain, hereby highlighting the need for further research in this field.

Basal Sodium-Dependent Vitamin C Transporter 2 polarization in choroid plexus explant cells in normal or scorbutic conditions.

Vitamin C is incorporated into the cerebrospinal fluid (CSF) through choroid plexus cells. While the transfer of vitamin C from the blood to the brain has been studied functionally, the vitamin C transporter, SVCT2, has not been detected in the basolateral membrane of choroid plexus cells. Furthermore, it is unknown how its expression is induced in the developing brain and modulated in scurvy conditions. We concluded that SVCT2 is intensely expressed in the second half of embryonic brain development and postnatal stages. In postnatal and adult brain, SVCT2 is highly expressed in all choroidal plexus epithelial cells, shown by colocalization with GLUT1 in the basolateral membranes and without MCT1 colocalization, which is expressed in the apical membrane. We confirmed that choroid plexus explant cells (in vitro) form a sealed epithelial structure, which polarized basolaterally, endogenous or overexpressed SVCT2. These results are reproduced in vivo by injecting hSVCT2wt-EYFP lentivirus into the CSF. Overexpressed SVCT2 incorporates AA (intraperitoneally injected) from the blood to the CSF. Finally, we observed in Guinea pig brain under scorbutic condition, that normal distribution of SVCT2 in choroid plexus may be regulated by peripheral concentrations of vitamin C. Additionally, we observed that SVCT2 polarization also depends on the metabolic stage of the choroid plexus cells.

Drug Transport at the Brain and Endothelial Dysfunction in Preeclampsia: Implications and Perspectives.

Transport of drugs across biological barriers has been a subject of study for decades. The discovery and characterization of proteins that confer the barrier properties of endothelia and epithelia, including tight junction proteins and membrane transporters belonging to the ATP-binding cassette (ABC) and Solute Carrier (SLC) families, represented a significant step forward into understanding the mechanisms that govern drug disposition. Subsequently, numerous studies, including both pre-clinical approaches and clinical investigations, have been carried out to determine the influence of physiological and pathological states on drug disposition. Importantly, there has been increasing interest in gaining a better understanding of drug disposition during pregnancy, since epidemiological and clinical studies have demonstrated that the use of medications by pregnant women is significant and this condition embodies a series of significant anatomical and physiological modifications, particularly at excretory organs and barrier sites (e.g., placenta, breast) expressing transporter proteins which influence pharmacokinetics. Currently, most of the research in this field has focused on the expression profiling of transporter proteins in trophoblasts and endothelial cells of the placenta, regulation of drug-resistance mechanisms in disease states and pharmacokinetic studies. However, little attention has been placed on the influence that the cerebrovascular dysfunction present in pregnancy-related disorders, such as preeclampsia, might exert on drug disposition in the mother's brain. This issue is particularly important since recent findings have demonstrated that preeclamptic women suffer from long-term alterations in the integrity of the blood-brain barrier (BBB). In this review we aim to analyze the available evidence regarding the influence of pregnancy on the expression of transporters and TJ proteins in brain endothelial cells, as well the mechanisms that govern the pathophysiological alterations in the BBB of women who experience preeclampsia. Future research efforts should be focused not only on achieving a better understanding of the influence of preeclampsia-associated endothelial dysfunction on drug disposition, but also in optimizing the pharmacological treatments of women suffering pregnancy-related disorders, its comorbidities and to develop new therapies aiming to restore the integrity of the BBB.

Pro-inflammatory and anti-inflammatory compounds exert similar effects on P-glycoprotein in blood-brain barrier endothelial cells.

OBJECTIVES: The effects of anti-inflammatory glucocorticoids dexamethasone (DX) and hydrocortisone (HC), pro-inflammatory cytokine interleukin-1ß (IL-1ß) and dietary long-chain polyunsaturated fatty acids (PUFAs) on expression and activity of the ATP-binding cassette transporter P-glycoprotein (P-GP) were studied in porcine brain endothelial cells (PBECs). METHODS: Primary PBECs were treated for 24 h with glucocorticoids, IL-1ß and long-chain PUFAs. P-GP activity was determined by measuring intracellular calcein accumulation and P-GP expression by Western blotting. The effect of PUFAs on membrane fluidity was assessed by fluorescence recovery after photobleaching (FRAP). KEY FINDINGS: Dexamethasone, HC and IL-1ß significantly increased P-GP expression and activity. The effect of IL-1ß was attenuated by the IL-1 receptor antagonist (IL-1RA). This is the first report of the combined actions of IL-1ß and IL-1RA on P-GP expression and the first evidence of glucocorticoid-mediated P-GP up-regulation in PBECs. Arachidonic acid (AA), docosahexaenoic acid (DHA) and eicosapentenoic acid (EPA) significantly decreased P-GP activity without affecting expression or membrane fluidity. AA, DHA and EPA counteracted IL-1ß-mediated increases in P-GP activity, while AA and EPA, but not DHA, counteracted glucocorticoid-mediated increase in P-GP activity. CONCLUSIONS: While glucocorticoids and IL-1ß possess opposing actions in inflammation, they demonstrate functional consistency by increasing P-GP expression and activity in PBECs.

Are the Cognitive Alterations Present in Children Born From Preeclamptic Pregnancies the Result of Impaired Angiogenesis? Focus on the Potential Role of the VEGF Family.

Evidence from clinical studies has proposed that children born from preeclamptic women have a higher risk of suffering neurological, psychological, or behavioral alterations. However, to date, the mechanisms behind these outcomes are poorly understood. Here, we speculate that the neurodevelopmental alterations in the children of preeclamptic pregnancies result from impaired angiogenesis. The pro-angiogenic factors vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) are key regulators of both vascular and neurological development, and it has been widely demonstrated that umbilical blood of preeclamptic pregnancies contains high levels of soluble VEGF receptor type 1 (sFlt-1), a decoy receptor of VEGF. As a consequence, this anti-angiogenic state could lead to long-lasting neurological outcomes. In this non-systematic review, we propose that alterations in the circulating concentrations of VEGF, PlGF, and sFlt-1 in preeclamptic pregnancies will affect both fetal cerebrovascular function and neurodevelopment, which in turn may cause cognitive alterations in post-natal life.

Development, characterization and in vitro evaluation of biodegradable rhein-loaded microparticles for treatment of osteoarthritis.

Rhein is an active metabolite of the drug diacerein, whose anti-inflammatory properties have been demonstrated in both in vitro and in vivo models. However, the low oral bioavailability of rhein has limited its utility as a potential treatment of osteoarthritis (OA), a chronic inflammatory disease. In order to overcome this limitation, the aim of this work was the development of a drug delivery system intended for intra-articular administration of rhein, based on polymeric biodegradable PLGA microparticles (MPs) loaded with the drug. The MPs, prepared by the emulsion-solvent evaporation technique were characterized in terms of several parameters including morphology, encapsulation efficiency, molecular interactions between components of the formulation and in vitro release profiling. Furthermore, cell-based in vitro studies were performed to evaluate the cytotoxicity of the formulations and their effect on the release of inflammatory markers including pro-inflammatory cytokines and reactive oxygen species (ROS). Scanning electron microscopy demonstrated that the prepared MPs exhibited an almost spherical shape with smooth surface. The size distribution of the prepared MPs ranged between 1.9 and 7.9µm, with mean diameter of 4.23±0.87µm. The optimal encapsulation efficiency of rhein was 63.8±3.0%. The results of powder X-ray diffraction and differential scanning calorimetry studies demonstrated that the active ingredient is partially the crystalline state, dispersed in the polymer matrix. This outcome is somewhat reflected in the release kinetics of rhein from the MPs. The cytotoxicity evaluation, carried out in macrophages derived from THP-1 cells, showed that both rhein-loaded MPs and unloaded MPs did not significantly affect the cell viability at MP concentrations up to 13.8µM. In lipopolysaccharide-activated macrophages, the rhein-loaded MPs significantly decreased the production of interleukin-1ß (IL-1ß) and (ROS), when compared to the unloaded MPs. In conclusion, the results of this preliminary study suggest that an MP-based formulation of rhein could be tested in animal models of inflammation, aiming for an injectable commercial product capable of providing a therapeutic solution to patients suffering from chronic joint diseases.

Jugos de fruta y productos herbáceos como perpetradores de interacciones farmacocinéticas mediadas por enzimas metabolizadoras y transportadores de membrana. Relevancia en clínica / Pharmacokinetic interactions of fruit juices and herbal preparations

Drug disposition in the human body is strongly influenced by transporters and metabolizing enzymes expressed in key organs including intestine, liver and kidney. Since drugs and chemicals present in foods such as fruit juices and herb-based products are substrates of the above-mentioned proteins, there is a high probability of pharmacokinetic interactions. Findings from preclinical and clinical studies helped to characterize the mechanisms by which the components of fruit juices and herbs act as perpetrators of pharmacokinetic interactions. The aim of this review is to provide an overview of pharmacokinetic fruit juice- and herb-drug interactions that could be relevant in the clinical setting.

Capreomycin oleate microparticles for intramuscular administration: Preparation, in vitro release and preliminary in vivo evaluation.

Capreomycin sulfate (CS) is a second-line drug used for the treatment of multidrug-resistant tuberculosis (MDR-TB). The adverse effects profile and uncomfortable administration scheme of CS has led to the development of formulations based on liposomes and polymeric microparticles. However, as CS is a water-soluble peptide that does not encapsulate properly into hydrophobic particulate matrices, it was necessary to reduce its aqueous solubility by forming the pharmacologically active capreomycin oleate (CO) ion pair. The aim of this research was to develop a new formulation of CO for intramuscular injection, based on biodegradable microparticles that encapsulate CO in order to provide a controlled release of the drug with reduced local and systemic adverse effects. The CO-loaded microparticles prepared by spray drying or solvent emulsion-evaporation were characterized in their morphology, encapsulation efficiency, in vitro/in vivo kinetics and tissue tolerance. Through scanning electron microscopy it was confirmed that the microparticles were monodisperse and spherical, with an optimal size for intramuscular administration. The interaction between CO and the components of the microparticle matrix was confirmed on both formulations by X-ray powder diffraction and differential scanning calorimetry analyses. The encapsulation efficiencies for the spray-dried and emulsion-evaporation microparticles were 92% and 56%, respectively. The in vitro kinetics performed on both formulations demonstrated a controlled and continuous release of CO from the microparticles, which was successfully reproduced on an in vivo rodent model. The results of the histological analysis demonstrated that none of the formulations produced significant tissue damage on the site of injection. Therefore, the results suggest that injectable CO microparticles obtained by spray drying and solvent emulsion-evaporation could represent an interesting therapeutic alternative for the treatment of MDR-TB.