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P J Gaillard

Publications and source records attributed to P J Gaillard.

At least 19 recordsLinked to original sources
PubMed2007

Drug targeting to the brain.

The central nervous system (CNS) is a sanctuary site and is protected by various barriers. These regulate brain homeostasis and the transport of endogenous and exogenous compounds by controlling their selective and specific uptake, efflux, and metabolism in the brain. Unfortunately, potential drugs for the treatment of most brain diseases are therefore often not able to cross these barriers. As a result, various drug delivery and targeting strategies are currently being developed to enhance the transport and distribution of drugs into the brain. Here we discuss briefly the biology and physiology of the blood-brain barrier (BBB) and the blood-cerebro-spinal-fluid barrier (BCSFB), and, in more detail, the possibilities for delivering large-molecular-weight drugs by local and global delivery and by viral and receptor-mediated nonviral drug delivery to the (human) brain.

Biological Transport↗
PubMed2006

Blood-brain barrier dysfunction and recovery.

In this paper we discuss the importance of the blood-brain barrier (BBB) as an interface between blood and brain. Many (brain) diseases change the functionality and integrity of the BBB. Mostly this results in increased BBB permeability. Therefore we have studied de various signal transduction routes that are influenced by inflammatory stimuli. The radical scavenger N-acetylcysteine was able to protect the BBB against inflammatory stimuli. Similar results were found following application of glucocorticoids. In addition, it was observed that glucocorticoids and interferon-alpha,beta increased the tightness of the in vitro BBB and when given together a potentiating effect was seen.

Animals↗
PubMed2002

The role of drug transporters at the blood-brain barrier.

The blood-brain barrier (BBB) is a dynamic interface between the blood and the brain. It eliminates (toxic) substances from the endothelial compartment and supplies the brain with nutrients and other (endogenous) compounds. It can be considered as an organ protecting the brain and regulating its homeostasis. Until now, many transport systems have been discovered that play an important role in maintaining BBB integrity and brain homeostasis. In this review, we focus on the role of carrier- and receptor-mediated transport systems (CMT, RMT) at the BBB. These include CMT systems, such as P-glycoprotein, multidrug-resistance proteins 1-7, nucleoside transporters, organic anion transporters, and large amino-acid transporters; RMT systems, such as the transferrin-1 and -2 receptors; and the scavenger receptors SB-AI and SB-BI.

Biological Transport, Active↗
PubMed2001

Glucocorticoid and type 1 interferon interactions at the blood-brain barrier: relevance for drug therapies for multiple sclerosis.

The pharmacological effect of glucocorticoids and type 1 interferons (IFNs), simultaneously used as therapeuticals for multiple sclerosis (MS), on the (inflamed) blood-brain barrier (BBB) was investigated in vitro. Although both drugs additively decreased BBB permeability, they did not prevent the increase in BBB permeability induced by lipopolysaccharide (LPS), which served as a pro-inflammatory stimulus. The beneficial clinical effect of glucocorticoid and IFN therapy for MS seems there- fore not to be mediated through a direct action at the level of the BBB. Most strikingly, however, pretreatment with type 1 IFNs (alpha and beta) potentiated the effect of glucocorticoids by two orders of magnitude. This lead us to hypothesize that type 1 IFNs may restore the dysfunctional T-helper 1 (Th1)/Th2 balance associated with MS, by a mechanism that involves an increased sensitivity for glucocorticoids.

Animals↗
PubMed2001

Endothelial cell hypertrophy induced by vascular endothelial growth factor in the retina: new insights into the pathogenesis of capillary nonperfusion.

OBJECTIVE: To investigate the mechanism leading to capillary nonperfusion of the retina in a monkey model of vascular endothelial growth factor A (VEGF)-induced retinopathy in which capillary closure occurs in a late stage after VEGF treatment. METHODS: Two monkeys received 4 intravitreous injections of 0.5 microg of VEGF in one eye and of phosphate-buffered saline in the other eye and were killed at day 9. After perfusion and enucleation, retinal samples were snap frozen for immunohistochemical analysis with the panendothelial cell marker CD31 or were fixed for morphometric analysis at the light and electron microscopic level. RESULTS: At the light microscopic level, all capillaries in the retina of VEGF-injected eyes displayed hypertrophic walls with narrow lumina. In a quantitative analysis of the deep capillary plexus in the inner nuclear layer, VEGF-injected eyes had a significant 5- to 7-fold decrease in total capillary luminal volume. CD31 staining showed that this decrease was not accompanied by a change in the number of capillaries. Electron microscopy revealed that the luminal volume of individual capillaries of the inner nuclear layer of VEGF-injected eyes was significantly decreased due to a 2-fold hypertrophy of the endothelial cells. CONCLUSIONS: Luminal narrowing caused by endothelial cell hypertrophy occurs in the deep retinal capillary plexus in VEGF-induced retinopathy in monkeys. This suggests a causal role of endothelial cell hypertrophy in the pathogenesis of VEGF-induced retinal capillary closure. A similar mechanism may operate in retinal conditions in humans associated with ischemia and VEGF overexpression. CLINICAL RELEVANCE: Capillary nonperfusion occurs in diabetic retinopathy and other ischemic diseases associated with overexpression of VEGF. In addition, VEGF-induced endothelial cell hypertrophy may be causative for capillary closure in these diseases.

Animals↗
PubMed2001

Establishment and functional characterization of an in vitro model of the blood-brain barrier, comprising a co-culture of brain capillary endothelial cells and astrocytes.

OBJECTIVE: The aim was to establish a flexible, abundantly available, reproducible and functionally characterized in vitro model of the blood-brain barrier (BBB). METHODS: In a first step, bovine brain capillaries and newborn rat astrocytes were isolated. Subsequently, a co-culture of primary brain capillary endothelial cells (BCEC) on semi-permeable filter inserts, with astrocytes on the bottom of the filter was established. The cell material was characterized on the basis of specific cell-type properties and (functional expression of) specific BBB properties. RESULTS: BCEC displayed: (1) characteristic endothelial cell morphology; (2) expression of endothelial cell markers (i.e., CD51, CD62P, CD71 and cadherin 5); (3) marginal F-actin localization; (4) tight junction formation between the cells; (5) expression of gamma-glutamyl-transpeptidase (gamma-GTP); (6) expression of P-glycoprotein (Pgp); (7) functional transendothelial transferrin transport and uptake; (8) restriction of paracellular transport; and (9) high transendothelial electrical resistance (TEER). Astrocytes displayed characteristic astrocyte morphology and expressed glial fibrillary acidic protein (GFAP). Co-culture with astrocytes increased TEER and decreased paracellular transport. In addition, expression of the glucocorticoid receptor (GR) was demonstrated in the endothelial cells of the BBB, while no expression of the mineralocorticoid receptor (MR) was found. CONCLUSIONS: A high quality and mass-production in vitro BBB model was established in which experiments with physiological (e.g., regulation of BBB permeability), pharmacological (e.g., pharmacokinetics and pharmacodynamics) and pathophysiological (e.g., disease influence on BBB permeability) objectives can be reproducibly performed.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

P-glycoprotein inhibition leads to enhanced disruptive effects by anti-microtubule cytostatics at the in vitro blood-brain barrier.

PURPOSE: To investigate whether P-glycoprotein (Pgp) protects the in vitro BBB against the cytotoxic effects of anti-tumour drugs. METHODS: In an in vitro BBB coculture model the influence of the anti-microtubule drugs vinblastine, colchicine, paclitaxel and the non-anti-microtubule drugs doxorubicin, fluorouracil and etoposide in the absence or presence of Pgp modulators on the trans-endothelial electrical resistance (TEER), which is an indicator for the integrity, was investigated. RESULTS: In the absence of Pgp modulators vinblastine, colchicine and paclitaxel dose dependently decreased TEER values to less than 20% of control. Non-anti-microtubule drugs did not affect TEER values. Following competitive inhibition of Pgp by various Pgp modulators and substrates, even low concentrations of vinblastine, colchicine and paclitaxel substantially decreased TEER. IC50 values of LY 335979, SDZ-PSC 833, cyclosporin A, and verapamil were 0.03, 0.25, 0.46, and 13.7 microM, respectively. CONCLUSIONS: These results indicate that Pgp normally protects the in vitro BBB against the disruptive effects of anti-microtubule drugs, but its integrity is lost when anti-microtubule drugs are used in combination with potent Pgp modulators. In addition, this procedure offers the possibility to characterize Pgp modulators and substrates with respect to their efficacy and to elucidate drug interactions at the level of Pgp.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Relationship between permeability status of the blood-brain barrier and in vitro permeability coefficient of a drug.

OBJECTIVE: The aim was to test the hypothesis that the assessment of basal and drug-induced changes in permeability of the blood-brain barrier (BBB) during in vitro drug transport assays is essential for an accurate estimation of the permeability coefficient of a drug. METHODS: An in vitro BBB model was used, comprising of brain capillary endothelial cells (BCEC) and astrocytes co-cultured on semi-permeable filter inserts. Experiments were performed under control and challenged experimental circumstances, induced to simulate drug effects. The apparent BBB permeability coefficient for two markers for paracellular drug transport, sodium fluorescein (P(app,FLU), M(w) 376 Da) and FITC-labeled dextran (P(app,FD4), M(w) 4 kDa), was determined. Transendothelial electrical resistance (TEER) was used to quantify basal and (simulated) drug-induced changes in permeability of the in vitro BBB. The relationship between P(app) and TEER was determined. Drug effects were simulated by exposure to physiologically active endogenous and exogenous substances (i.e., histamine, deferroxamine mesylate, adrenaline, noradrenaline, bradykinin, vinblastine, sodium nitroprusside and lipopolysaccharide). RESULTS: P(app,FLU) and P(app,FD4) in control experiments varied from 1.6 up to 17.6 (10(-6)cm/s) and 0.3 up to 7. 3 (10(-6)cm/s), respectively; while for individual filters P(app, FLU) was 4 times higher than P(app,FD4) (R(2)=0.97). As long as TEER remained above 131.Omega cm(2) for FLU or 122.Omega cm(2) for FD4 during the transport assay, P(app) remained independent from the basal permeability of the in vitro BBB. Below these TEER values, P(app) increased exponentially. This nonlinear relationship between basal BBB permeability and P(app) was described by a one-phase exponential decay model. From this model the BBB permeability status independent permeability coefficients for FLU and FD4 (P(FLU) and P(FD4)) were estimated to be 2.2+/-0.1 and 0.48+/-0.03 (10(-6)cm/s), respectively. In the experimentally challenged experiments, a reliable indication for P(FLU) and P(FD4) could be estimated only after the (simulated) drug-induced change in BBB permeability was taken into account. CONCLUSIONS: The assessment of basal BBB permeability status during drug transport assays was essential for an accurate estimation of the in vitro permeability coefficient of a drug. To accurately extrapolate the in vitro permeability coefficient of a drug to the in vivo situation, it is essential that drug-induced changes in the in vitro BBB permeability during the drug transport assay are determined.

Animals↗
PubMed2000

Astrocytes increase the functional expression of P-glycoprotein in an in vitro model of the blood-brain barrier.

PURPOSE: To investigate the influence of astrocytes on P-glycoprotein (Pgp) expression and intracellular accumulation of Pgp substrates, separate from their net transcellular transport across the blood-brain barrier (BBB). METHODS: An in vitro BBB model was used, comprising of brain capillary endothelial cells (BCEC) monolayers or BCEC co-cultured with astrocytes. RESULTS: BCEC+astrocyte co-cultures seemed to express a higher level of Pgp compared to BCEC monolayers. Inhibition of Pgp results in an increased intracellular accumulation of Pgp substrates in both BCEC monolayers and BCEC+astrocyte co-cultures, and increased the sensitivity for vinblastine mediated disruption of the in vitro BBB (called the vinblastine exclusion assay). BCEC monolayers were more sensitive to vinblastine mediated disruption compared to BCEC+astrocyte co-cultures. In the latter, but not in BCEC monolayers, an inhibitable polar transport of Pgp substrates was only found from the brain to the blood side of the filter. CONCLUSIONS: Astrocytes increase the functional expression of Pgp in our in vitro BBB model. These results also illustrate that an important role for Pgp on the BBB is to protect the barrier against intracellular accumulation of cytotoxic BBB disrupting compounds.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Differences in striatal extracellular amino acid concentrations between Wistar and Fischer 344 rats after middle cerebral artery occlusion.

We hypothesized that the interstrain difference between Wistar and Fischer-344 (F344) rats in cerebral infarction volume after proximal middle cerebral artery (MCA) occlusion might be explained by differences in excitotoxicity between both rat strains. Using microdialysis we measured during a 5 h period after MCA occlusion the release of aspartate, glutamate and taurine in the cerebral cortex and the striatum. The volume of striatal infarction was comparable in Wistar and F344 rats. We found, however, in Wistar rats a significantly higher striatal efflux of aspartate and glutamate than in F344 rats, whereas the striatal taurine efflux was of a similar magnitude in the two strains. Because of the (variably) smaller volume of cortical infarction in Wistar rats (than that in F344), the location of the microdialysis probe-membrane with respect to the area of cortical infarction differed between Wistar rats. Hence, a reliable comparison between the quantitative amount of amino acids in the dialysate from the cortical probes of both rat strains could not be made. These results, demonstrating differences in striatal excitotoxicity between Wistar and F344 rats after MCA occlusion, are the first to show interstrain differences in striatal pathophysiology of focal ischemia between these normotensive rat strains. They do however not explain why MCA occlusion results in a significantly different volume of cortical infarction between Wistar and F344 rats. The F344 strain will probably show in a more sensitive way, as compared to Wistar rats, neuroprotective effects of agents that diminish excitotoxic damage during focal cerebral ischemia.

Amino Acids↗
PubMed1982

Regulatory mechanisms in the development of bone and cartilage: the use of tissue culture techniques in the study of the development of embryonic bone and cartilage: a perspective.

Tissue culture techniques were used to study a number of factors and mechanisms which are important in the development and metabolism of bone tissue. As an example of an external factor influencing bone development, the importance of the composition of the gasphase which is in equilibrium with the fluid bathing osteoblasts and hypertrophic chondrocytes, was investigated in cultured metatarsal bone rudiments. In vivo, one expects the presence of an O2 gradient in the cartilaginous epiphyses of long bones: low O2 tension between the nonhypertrophic chondrocytes, high O2 tension in periosteum and hypertrophic zone, bordering the marrow cavity. The in vitro findings correlated with these expectations. High CO2 (5%) and high O2 (40%) tensions stimulated calcification; in air, calcification was severely inhibited. On the other hand, maturing chondrocytes were damaged by high O2 tensions. An important cellular mechanism in calcification is the intracellular accumulation of calcium (and phosphate) in osteoblasts and hypertrophic chondrocytes which can be demonstrated with the GBHA stain of Kashiwa [9]. The extracellular role of alkaline phosphatase (AP) present on the cell membranes of these cells was shown to be a less decisive factor in calcification. In the presence of AP inhibitor in a concentration high enough to inhibit AP activity to a large extent, calcification was shown to proceed normally. The effects of a number of hormones known to be important for the development and metabolism of bone tissue was studied using tissue culture (calvaria) as well as culture of different isolated bone cells. The parathyroid hormone (PTH) induced rise of the intracellular cAMP level was found to originate primarily from the osteoblasts not the osteoclasts. Isolated osteoblasts showed a high cAMP response after PTH addition. Cortisol was shown to inhibit PTH induced resorption but to potentiate PTH induced cAMP response in calvaria. Various PTH fragments (desamino 1-34, 2-34, 3-34) were shown to be active as stimulators of bone resorption (although they were less active in this respect than the intact molecule 1-84), but did not stimulate cAMP production in calvaria or isolated osteoblasts. The results obtained strengthened the hypothesis that cAMP is not the (only) mediator in PTH induced bone resorption.

Alkaline Phosphatase↗
PubMed1978

The effects of 1,25-dihydroxycholecalciferol on embryonic bone in vitro: a biochemical and histological study.

Explants from embryonic rat and mouse calvaria were cultivated in the presence of different concentrations of 1, 25-dihydroxycholecalciferol (1, 25-(OH)2D3). The bone resorbing effects of the vitamin D3 metabolite were evaluated by measuring the release of calcium, phosphate, lactate and citrate into the culture medium after 24 h of cultivation. The influence on bone morphology was studied using embryonic mouse radii, in which histological phenomena in the bony, cartilagenous and connective tissue compartments were observed. Both kinds of experiments show that 1, 25-(OH)2D3 has effects on embryonic bone which are typical for high concentrations of parathyroid hormone (PTH). However, the maximal effects on calcium release and on histology are 2.5 times less than those of PTH. In addition, some of the histological features such as the effects on epiphyseal cartilage and on osteoclasts were not observed. 1, 25-(OH)2D3 in concentrations up to 2 X 10(-9) M does not affect basal or PTH-stimulated cAMP levels in embryonic rat calvaria. In a concentration of 1 X 10(-8) M, however, a significant decrease in PTH-stimulated cAMP production was found. It is concluded from these in vitro experiments that (1) 1, 25-(OH)2D3 stimulates bone resorption, and (2) in 1, 25-(OH)2D3-induced bone resorption cAMP is apparently not involved as a second messenger.

Animals↗
PubMed1977

The effect of verapamil on the action of parathyroid hormone on embryonic bone in vitro.

Short term in vitro experiments showed that, added alone, verapamil inhibited both glycolysis and Ca uptake in embryonic chicken and rat bone cells. Added together with PTH, verapamil (0.02 MM) enhanced cAMP production, had no effect on lactate production, but significantly inhibited citrate, calcium and phosphate release from embryonic rat and mouse calvaria incubated under hypocalcemic conditions. The depression by verapamil of PTH-stimulated demineralization was confirmed histologically. It is concluded that in addition to cAMP, Ca plays a key role in the action of PTH on bone.

Animals↗
PubMed1976

Effects of bone in vitro of bovine parathyroid hormone and synthetic fragments representing residues 1-34, 2-34 and 3-34.

The biological activities of bovine parathyroid hormone (BPTH) and fragments comprising portions of its amino-terminal sequence have been compared in three different assay systems using embryonic rat bone in vitro. Whereas the 3-34 fragment was without significant activity the 1-34 fragment caused all the actions characteristic of BPTH 1-84, extending to bone previous evidence that the amino-terminal residues are sufficient for expression of the biological effects of intact parathyroid hormone. However, the relative potencies of the fragment and the intact hormone were different in the various systems. BPTH 1-34 showed relatively low osteolytic activity and induced anabolic effects in both osteoblasts and cartilage cells of cultivated embryonic mouse radii which were not evoked by the intact hormone. Further work is required to determine the mechanisms responsible for these interesting alterations in relative potency of fragment and native hormone.

Animals↗