Effects of a new lipid-based drug delivery system on the absorption of low molecular weight heparin (Fragmin) through monolayers of human intestinal epithelial Caco-2 cells and after rectal administration to rabbits

The effects of a novel drug delivery system, consisting of a lipid matrix and a drug, on the permeability, morphology and drug transport across monolayers of human intestinal epithelial cells were investigated. A 1:3 mixture (w/w) of chromatographically purified soybean phosphatidylcholine and medium chain monoacylglycerol was chosen as lipid matrix (PC:MG) and mannitol (mol. wt. 182) and Fragmin (low molecular weight heparin; mean mol. wt. 5000) were chosen as hydrophilic model drugs. PC:MG had an immediate and dose-dependent effect on the permeability of the cell monolayers, and on epithelial morphology (as seen under transmission electron microscopy). PC:MG (4–10 mM) dose-dependently enhanced the transport rate of mannitol and Fragmin, causing an approximately 10-fold increase in the transport rate of Fragmin at concentrations of 6–8 mM. The increase was independent of the dose of Fragmin and was reversible in concentrations up to 6 mM. At higher doses, clear effects on the apical cell membranes were observed although the tight junctions remained intact. PC:MG also enhanced Fragmin absorption in vivo after rectal administration to New Zealand White rabbits. Co-administration of Fragmin with PC:MG (7 mM) resulted in an increase in the relative bioavailability (compared with s.c. administration) from < 1% (without PC:MG) to 21 ± 12%. A maximal increase in relative bioavailability to 90 ± 19% was obtained at a PC:MG concentration of 35 mM. Thus, PC:MG functions as an absorption enhancer both in the cell monolayer model and in vivo, in the same concentration range. The results indicate that as well as providing mechanistic information, studies of absorption enhancers in Caco-2 monolayers also provide information on suitable dosage regimens for in vivo studies.     

壳聚糖、羧甲基壳聚糖的降脂及抗氧化作用

目的：研究壳聚糖(Chitosan)、羧甲基壳聚糖(Carboxymethyl chitosan，CMCS)对高脂血症的新西兰白兔的降血脂及体内抗氧化作用。方法：40只新西兰白兔随机分为5组：正常对照组(A)、高脂模型组(B)、1％壳聚糖组(C)、1．5％壳聚糖组(D)、1．5％羧甲基壳聚糖组(E)，分别以各组的饲料连续喂养6周。结果：壳聚糖、羧甲基壳聚糖降低血清甘油三酯(TG)、总胆固醇(TC)和低密度脂蛋白胆固醇(LDLc)，明显升高血清高密度脂蛋白胆固醇(HDLc)。结论：壳聚糖的降血脂活性比其衍生物羧甲基壳聚糖高。壳聚糖及羧甲基壳聚糖还具有抗脂质氧化的作用，表现为降低兔血清中的氧化型低密度脂蛋白(OX—LDL)及丙二醛(MDA)含量。     

Population analysis to assess the influence of age and body weight on pharmacokinetics and pharmacodynamics of dexmedetomidine in New Zealand White rabbits

The purpose of this work was i) to develop a population pharmacokinetic (PK) and pharmacodynamic (PD) model of dexmedetomidine (DEX) in New Zealand White rabbits, ii) to investigate the influence of the age and weight of the animals on the model parameters, and iii) to assess the linearity of DEX PKs in the examined dose range. This was a prospective, crossover study, using a total of 18 New Zealand White rabbits. DEX was administered as a single intravenous bolus injection in the doses from 25 to 300 μg kg⁻¹. Each New Zealand White rabbit was given the same dose of drug in its three developmental stages. To determine the DEX PK, seven blood samples were taken from each animal. The pedal withdrawal reflex was the PD response used to assess the degree of sedation. Nonlinear mixed effects modelling was used for the population PK/PD analysis. The typical value of elimination clearance was 0.061 L min⁻¹ and was 35% higher in younger New Zealand White rabbits compared with older animals. The PK of DEX was linear in the examined concentration range. Age-related changes in sensitivity to DEX were not detected. The results suggest that due to the pharmacokinetics, younger animals will have lower DEX concentrations and a shorter duration of sedation than older animals given the same doses of DEX per kg of body weight.     

Bioavailability of diazepam from aqueous-organic solution, submicron emulsion and solid lipid nanoparticles after rectal administration in rabbits.

The bioavailability of diazepam in rabbits after rectal administration of three formulations: organic-aqueous Relsed rectal solution (containing ethanol, benzyl alcohol and propylene glycol), submicron emulsion and solid lipid nanoparticles (SLN), was studied. Submicron emulsion contained MCT oil (20% w/w), egg lecithin and poloxamer; SLN were prepared with cetyl palmitate 10% w/v and non-ionic emulsifying agent, Plantacare. All formulations contained 4 mg/ml of diazepam and the dose administrated to rabbits was 2 mg/kg. In both submicron preparations nearly the same mean size of the dispersed particles (201-206 nm) and the fraction of the free drug in aqueous phase (0.9-1.5%) was determined. Besides very moderate prolongation of drug release, the submicron emulsion as a vehicle did not alter pharmacokinetics of diazepam when compared with the solution: the mean C(max) was 48.9+/-24.0 and 49.5+/-17.0 ng/ml, and area under the curve was 134.0+/-42.3 and 186.8+/-59.8 ng h/ml, for solution and emulsion, respectively. The low relative bioavailability, 47% compared to the solution, was observed after administration of SLN. Transmission electron microscopy pictures revealed that some of diazepam is present on the surface of the SLN and this fraction was immediately absorbed, while the diffusion of the drug in the solid core was not efficient enough to allow a complete release. It may be concluded that submicron emulsion may be a good choice of an ethanol-free drug formulation, but lipid matrix, which is solid at body temperature, is not advantageous system for diazepam rectal delivery, even if delivered as a submicron dispersion.     

Substrate specificity of the di/tripeptide transporter in human intestinal epithelia (Caco-2): identification of substrates that undergo H(+)-coupled absorption.

1. pH-dependent transepithelial transport and intracellular accumulation of the hydrolysis-resistant dipeptide glycylsarcosine (Gly-Sar) have been demonstrated in the model human intestinal epithelial cell line, Caco-2. 2. Experiments with BCECF (2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein)-loaded Caco-2 cells demonstrated that dipeptide (Gly-Sar) transport across the apical membrane is coupled to proton flow into the cell. 3. A range of postulated substrates for the intestinal di/tripeptide carrier were tested for their abilities to: (a) inhibit pH-dependent [14C]Gly-Sar apical-to-basal transport and intracellular accumulation and (b) stimulate H+ flow across the apical surface of BCECF-loaded Caco-2 cell monolayers. 4. A range of compounds (including Gly-Gly, Leu-Leu, Gly-Gly-Gly, cefadroxil and cephalexin) caused marked acidification of intracellular pH when perfused at the apical surface of Caco-2 cell monolayers. In contrast leucine and D-Leu-D-Leu failed to induce proton flow. The ability to induce proton-flow across the apical surface by these compounds, in this intestinal epithelium, was directly correlated to the relative inhibitory effects on [14C]-Gly-Sar transport and accumulation. 5. The determination of substrate-induced intracellular pH change in the Caco-2 cell system may provide a useful rapid screen for candidate substrates for absorption via H(+)-coupled transport mechanisms such as the intestinal di/tripeptide carrier in an appropriate physiological context.     

The Transepithelial Transport of a G-CSF-Transferrin Conjugate in Caco-2 Cells and Its Myelopoietic Effect in BDF1 Mice

PURPOSE: The purpose of this study was to investigate the transferrin-receptor (TfR)-mediated transepithelial transport of G-CSF-transferrin (Tf) conjugate in cultured enterocyte-like Caco-2 monolayers and the myelopoietic effect of subcutaneously and orally administered G-CSF-Tf in BDF1 mice. METHODS: Caco-2 monolayers exhibiting a minimum transepithelial electrical resistance of 500 ohms-cm2 and BDF1 mice were used as in vitro and in vivo models, respectively. TfR-mediated transcytosis wa measured by using 125I-G-CSF-Tf and analyzing the downstream compartment by gamma counter. The efficacy of subcutaneously and orally administered G-CSF-Tf was determined by performing daily absolute neutrophil counts. RESULTS: Transport experiments in Caco-2 cells revealed that the monolayers that received 125I-G-CSF-Tf exhibited significantly higher apical-to-basolateral transport rates compared to the monolayers that received 125I-G-CSF. Inclusion of 100-fold excess unlabeled Tf reduced the extent of 125I-G-CSF-Tf transport by 80%. Chromatographic and bioactivity assays revealed that the protein recovered from the basolateral compartment was the intact conjugate, and it retained full ability to stimulate the proliferation of the granulocyte-colony stimulating factor (G-CSF) dependent cell line, NFS-60, upon reduction. Subcutaneous administration of G-CSF-Tf in BDF1 mice demonstrated that the conjugate is able to elicit a statistically significant enhancement in therapeutic effect relative to filgrastim, which includes a longer duration of action with higher absolute neutrophil counts. Oral administration of G-CSF-Tf in BDF1 mice demonstrated that G-CSF-Tf is able to elicit a significant, and apparently dose-dependent, increase in absolute neutrophil counts whereas filgrastim had no effect. CONCLUSIONS: Our data indicate that G-CSF-Tf is transported across Caco-2 monolayers by TfR-specific processes at a rate that is significantly higher than the nonspecific flux of G-CSF. G-CSF-Tf is also able to elicit a prolonged myelopoietic effect relative to filgrastim when administered subcutaneously or orally in BDF1 mice. The development of an orally bioavailable G-CSF has the potential to provide great benefit to patients under sustained G-CSF dosing regimes.     

Evaluation of the impact of altered lipoprotein binding conditions on halofantrine induced QTc interval prolongation in an anaesthetized rabbit model

Halofantrine has been observed to cause QT interval prolongation in susceptible patients and the effect has most commonly been observed after post-prandial administration. Halofantrine-induced QT prolongation occurs in conjunction with a significant increase in plasma halofantrine concentrations and an increase in halofantrine association with post-prandial plasma lipoproteins. The increased association of halofantrine with post-prandial lipoproteins is accompanied by a marked change in drug distribution between the different plasma lipoprotein fractions. This study was designed to evaluate the putative role of myocardium-based lipoprotein receptor-mediated uptake of lipoproteins as a possible contributing factor to the observed effect of halofantrine on QT intervals. The extent of QT interval prolongation following intravenous halofantrine administration (10 mg kg(-1)) to normolipidaemic (fasted) or hyperlipidaemic (induced with Intralipid infusion) anaesthetized New Zealand White rabbits (n = 6) was determined, as was the distribution of halofantrine between the plasma lipoprotein classes. The results, however, were in contrast to the suggested hypothesis since the QT interval was reduced (and not increased) after halofantrine administration to hyperlipidaemic rabbits relative to fasted rabbits. Therefore, it is unlikely that lipoprotein-based uptake of halofantrine into the myocardium is a major contributor to the previously observed increase in QT prolongation after post-prandial administration of halofantrine.     

Effects of capsaicin on cellular damage and monolayer permeability in human intestinal Caco-2 cells

Recent studies suggest that capsaicin (Cap), a major constituent of hot pepper, may affect the function and permeability of the intestinal mucosa in vitro. However, the relationships between the dose of Cap and the barrier and/or transporter functions on intestinal epithelial cells are unknown. The aim of this study was to investigate whether Cap initiates cellular injury and alter epithelial permeability in Caco-2 cells. Cellular toxicity, as measured using a lactate dehydrogenase release assay, was not observed at high concentrations of Cap (up to 300 microM). When cell viability was measured by a WST-1 assay (tetrazolium salt-based assay), damage to Caco-2 monolayers was observed at doses of 200 and 300 microM of Cap. The barrier function of tight junctions was assessed by measuring transepithelial electrical resistance (TEER) in Caco-2 cells. Treatment of Caco-2 cells with Cap at doses above 100 microM significantly decreased the TEER compared to treatment with buffer alone for 2 h (p<0.05). We next examined the effects of Cap on the activity of P-glycoprotein (P-gp) found on transcellular transporters. At doses of 100 and 200 microM, Cap inhibited the transport of rhodamine 123 by P-gp-mediated efflux in Caco-2 cells. Cap thus exhibited inhibitory effects on P-gp. The results of this study indicate that Cap, a dietary phytochemical, causes functional and structural changes in Caco-2 cell monolayers at noncytotoxic doses (less than 100 microM of Cap). The concomitant administration of Cap with drugs that are substrates of P-gp might increase the plasma concentrations of such drugs.     

A comparison of commonly used polyethoxylated pharmaceutical excipients on their ability to inhibit P-glycoprotein activity in vitro

P-glycoprotein (P-gp), a multidrug resistance (MDR) protein encoded by the MDR1 gene in humans, is responsible for the efflux of structurally diverse drugs. Previous studies in our laboratory have shown that excipients such as poly(ethylene)glycol (PEG)-300, Cremophor EL, and Tween 80 inhibit P-gp activity in Caco-2 cell monolayers. The objective of this study was to determine the effects of these excipients in an MDR1- transfected Madin Darby Canine Kidney (MDR1-MDCK) cell line and to compare the results with those obtained from Caco-2 cells. The results presented herein show that PEG-300 (20%, v/v) causes almost complete inhibition of P-gp activity in both Caco-2 and MDR1-MDCK cell monolayers, whereas Cremophor EL (0.1%, w/v) and Tween 80 (0.05%, w/v) only partially inhibit P-gp activity in Caco-2 cells. Cremophor EL (0.1%, w/v) and Tween 80 (0.05%, w/v) were inactive as P-gp inhibitors in MDR1-MDCK cell monolayers. This inability of Tween 80 and Cremphor EL to inhibit P-gp activity in MDR1-MDCK cells may be related to differences in the interactions of the surfactants with these different cell membranes. PEG-induced changes in P-gp activity are probably related to changes in the fluidity of the polar head group regions of cell membranes.     

Timing of implantation and closure of the palatal shelf in New Zealand white and Japanese white rabbits

Two specific developmental events, namely implantation and palatal shelf closure, are of specific interest because they define, respectively, the beginning and the end of the treatment period in embryo-fetal developmental toxicity studies for pharmaceutical products. Thus, a detailed evaluation of the timing of implantation and closure of the hard palate is necessary to assure use of the proper exposure window in developmental toxicity studies in rabbits, the nonrodent species most commonly evaluated in regulatory developmental toxicology studies. The purpose of this study was to determine the timeline for implantation and closure of the hard palate in the New Zealand White rabbit, and to determine if this timeline differed in the Japanese White rabbit. To describe the timing of implantation, the uteri from does of the New Zealand White rabbit and the Japanese White rabbit were examined on gestation days (GDs) 5 through 8 for macroscopic evidence of implantation. To assess palatal shelf closure, fetuses were removed on GDs 17, 18, and 19 and fixed in Bouin's solution. The fetuses were then categorized into five stages of palatal shelf closure: open (Stage I); approach of the palatal shelves (Stage II); partial closure of the hard palate (Stage III); full closure of the hard palate (Stage IV); and full closure of the soft palate (Stage V). In both the New Zealand White and Japanese White rabbit strains, implantation was initiated on GD 6.5 and was completed on GD 7. Partial closure of the palate began on GD 17.5, and by GD 19, closure of the hard palate was completed in all fetuses, and closure of the soft palate was completed in 75-96% of the fetuses. The timing of implantation and palatal shelf closure were comparable between the New Zealand White rabbit and the Japanese White rabbit. Therefore, treatment beginning on GD 7 and continuing until GD 19 encompasses the period of major organogenesis and is considered appropriate for use in developmental toxicity studies using either of these two strains of rabbits.     

Gamma-Aminobutyric acid (GABA) transport across human intestinal epithelial (Caco-2) cell monolayers

1. Transintestinal absorption of gamma-aminobutyric acid (GABA) via a pH-dependent mechanism is demonstrated in the model human intestinal epithelial cell line Caco-2. 2. Experiments with BCECF [2',7',-bis(2-carboxyethyl)-5(6)- carboxyfluorescein]-loaded Caco-2 cells demonstrate that GABA transport across the apical membrane is coupled to proton flow into the cell. 3. Short-circuit current (ISC) measurements using Caco-2 cell monolayers under voltage-clamped conditions demonstrate that pH-dependent GABA transport is a rheogenic process even in the absence of extracellular Na+, consistent with H+/GABA symport. 4. A range of GABA analogues were tested for their abilities to: (a) inhibit pH-dependent [3H]GABA uptake across the apical membrane; (b) stimulate H+ flow across the apical surface of BCECF-loaded Caco-2 cell monolayers; (c) increase inward ISC across voltage-clamped Caco-2 cell monolayers. 5. Nipecotic acid, isonipecotic acid, D,L-beta-aminobutyric acid, and 3-amino-1-propanesulphonic acid each caused a marked acidification of intracellular pH and an increase in ISC when superfused at the apical surface of Caco-2 cell monolayers. In contrast L-alpha-amino-n-butyric acid failed to induce proton flow or ISC. The ability of these compounds to induce proton or current flow across the apical surface of this intestinal epithelium was closely related to the relative inhibitory effects on [3H]GABA uptake. 6. These observations demonstrate H+/GABA symport and suggest that this transport mechanism may be accessible as a route for oral absorption of therapeutically-useful GABA analogues.     

Development of a rectal nicotine delivery system for the treatment of ulcerative colitis.

The aims of this investigation were: i. to develop a rectal nicotine delivery system with bioadhesives for the treatment of ulcerative colitis and ii. to evaluate nicotine transport and cytotoxicity of the delivery system using Caco-2 cell culture systems. Rectal nicotine suppository formulations were prepared in semi-synthetic glyceride bases (Suppocire AM and AI, Gattefosse Inc.) by fusion method. The in vitro release of nicotine was carried out in modified USP dissolution apparatus 1. Differential scanning calorimetry (DSC) and powder X-ray diffraction were used to study the polymorphic changes if any in the formulations. An LC method was used for the assay of nicotine. The effect of bioadhesives (glyceryl monooleate (GMO), and Carbopol) on the nicotine flux was evaluated using Caco-2 cell permeability studies and Caco-2 cell viability was determined using the MTT toxicity assay. In vitro release studies indicated that the low melting AI base was superior to that of the AM base. Presence of GMO in the formulation enhanced the release of nicotine whereas Carbopol showed an opposite effect. The enhanced release of nicotine in the presence of GMO was found to be partly due to the melting point lowering effect of this compound. Caco-2 cell absorption studies showed that there was a decrease in the flux of nicotine in the presence of both the bioadhesives. The flux of the fluorescein marker which is used to study the integrity of the cell monolayers was found to be slightly higher only in the presence of 10% (w/w) Carbopol. Nicotine, Carbopol, and GMO do not have any cytotoxic effect on these cell monolayers within the concentration range used in the formulations. Rectal nicotine formulations containing bioadhesives were developed and characterized. Both in vitro release and cell culture studies have indicated that one can manipulate the nicotine release from these rectal delivery systems by incorporation of various bioadhesives or the use of different bases in the formulation. Nicotine concentration below 2% (w/v) and bioadhesive concentration below 10% (w/w) do not have any cytotoxic effect on Caco-2 cells.     

Modulation of matrix metalloproteinase-1, its tissue inhibitor, and nuclear factor-kappa B by losartan in hypercholesterolemic rabbits

Upregulation of angiotensin II receptor, may be involved in the initiation and progression of atherosclerosis. To examine the contribution of AT1 receptor in the expression of matrix metalloproteinase-1 (MMP-1) and its tissue inhibitor (TIMP-2) in lipid-deposited arterial tissues, New Zealand white rabbits were given high-cholesterol chow (with losartan 25 mg/d or vehicle) for 10 weeks. Losartan reduced the areas of sudanophilia in the aorta of rabbits fed high-cholesterol diet (p < 0.01 vs. control). Losartan also significantly decreased the enhanced mRNA expression of MMP-1 and TIMP-2 in aortas of rabbits with high-cholesterol diet. Losartan-treated rabbits revealed a reduction in immunohistochemical expression of MMP-1, whereas TIMP-2 expression became localized to the intima. In addition, losartan treatment reduced the activation of NF-kappa B by inhibiting the degradation of its inhibitor I kappa-B alpha. These observations demonstrate that AT1 receptor blockade with losartan reduces lipid deposition and exerts potent inhibitory effects on NF-kappa B activation and modulates the expression of MMP-1 and TIMP-2 in hypercholesterolemic rabbits.     

Timing of Implantation and Closure of the Palatal Shelf in New Zealand White and Japanese White Rabbits

Two specific developmental events, namely implantation and palatal shelf closure, are of specific interest because they define, respectively, the beginning and the end of the treatment period in embryo-fetal developmental toxicity studies for pharmaceutical products. Thus, a detailed evaluation of the timing of implantation and closure of the hard palate is necessary to assure use of the proper exposure window in developmental toxicity studies in rabbits, the nonrodent species most commonly evaluated in regulatory developmental toxicology studies. The purpose of this study was to determine the timeline for implantation and closure of the hard palate in the New Zealand White rabbit, and to determine if this timeline differed in the Japanese White rabbit.

To describe the timing of implantation, the uteri from does of the New Zealand White rabbit and the Japanese White rabbit were examined on gestation days (GDs) 5 through 8 for macroscopic evidence of implantation. To assess palatal shelf closure, fetuses were removed on GDs 17, 18, and 19 and fixed in Bouin's solution. The fetuses were then categorized into five stages of palatal shelf closure: open (Stage I); approach of the palatal shelves (Stage II); partial closure of the hard palate (Stage III); full closure of the hard palate (Stage IV); and full closure of the soft palate (Stage V).

In both the New Zealand White and Japanese White rabbit strains, implantation was initiated on GD 6.5 and was completed on GD 7. Partial closure of the palate began on GD 17.5, and by GD 19, closure of the hard palate was completed in all fetuses, and closure of the soft palate was completed in 75–96% of the fetuses.

The timing of implantation and palatal shelf closure were comparable between the New Zealand White rabbit and the Japanese White rabbit. Therefore, treatment beginning on GD 7 and continuing until GD 19 encompasses the period of major organogenesis and is considered appropriate for use in developmental toxicity studies using either of these two strains of rabbits.     

A study of the nephrotoxicity of three cephalosporins in rabbits using 1H NMR spectroscopy

Male New Zealand White rabbits received a single intravenous injection of 125 mg/kg cephaloridine, 500 mg/kg cefoperazone or 1000 mg/kg cephalothin. Histological examination of kidneys at 48 h post-dose confirmed the presence of bilateral necrosis of the proximal convoluted tubules in the cephaloridine-treated animals. 1 H-NMR urinalysis of cephaloridine-treated rabbits detected drug-related resonances, decreased hippurate and increased glucose at 0–24 h post-dose accompanied by elevated levels of lactate, glycine, citrate, glutamine/glutamate and alanine at 24–48 h postdose. No histopathological changes were observed following administration of cefoperazone or cephalothin. 1 HNMR spectra of urine collected from these animals showed drug-related resonances and decreased hippurate levels at 0–24 h post-dose, and increased glucose levels at 24–48 h post-dose. Analysis of urine by conventional clinicalchemistry failed to reveal any statistically significant differences between the treatment groups. Under the conditions of this study, the nephrotoxic effects of cephaloridine and the minimal effects of cefoperazone and cephalothin could be clearly distinguished by 1 H-NMR urinalysis but not by conventional urinalysis.     

Epithelial transport of drugs in cell culture. VII: Effects of pharmaceutical surfactant excipients and bile acids on transepithelial permeability in monolayers of human intestinal epithelial (Caco-2) cells.

The effects of anionic (sodium dodecyl sulfate and sodium dioctyl sufosuccinate) and nonionic (polysorbate 80 and polyoxyl 40 hydrogenated castor oil) synthetic surfactants and bile acids (sodium taurocholate, sodium taurodeoxycholate, and sodium taurodihydrofusidate) on epithelial integrity were studied in monolayers of human intestinal epithelial (Caco-2) cells grown on microporous polycarbonate filters. The effects of the surfactants on intracellular enzyme activity, cell monolayer permeability, and morphology were studied. The effects on permeability were studied by two methods: measurements of transport of marker molecules (mannitol and polyethylene glycol) and measurements of transepithelial electrical resistance. All surfactants demonstrated concentration-dependent effects on intracellular enzyme activities, permeability, and morphology. The effects of the anionic surfactants were more pronounced than those of the nonionic surfactants. The effects on transepithelial electrical resistance correlated with intracellular dehydrogenase activity. Fluxes of marker molecules were the most sensitive measure of epithelial integrity. The results indicate that the hydrophilic marker molecules permeate the epithelial monolayers through different pathways at different concentrations of the surfactants. The effects of the surfactants were reversible at intermediate concentrations, even though the morphology of the monolayers had changed. The results agree with published data obtained with experimental animals and indicate that Caco-2 cells can be used to study the concentration-dependent effects of surfactants and other pharmaceutical additives on intestinal epithelial permeability.     

The changes in the endothelial expression of cell adhesion molecules and iNOS in the vessel wall after the short-term administration of simvastatin in rabbit model of atherosclerosis

Cell adhesion molecules P-selectin, VCAM-1 and ICAM-1 play an important role in the pathogenesis of atherosclerosis. High levels of nitric oxide (NO) produced by inducible NO synthase (iNOS) have been associated with atherosclerotic processes. Simvastatin is an HMG-CoA reductase inhibitor responsible for many clinical benefits. The aim of this study was to detect and quantify changes in endothelial expression of P-selectin, VCAM-1, ICAM-1 and iNOS in the vessel wall after the shortterm administration of simvastatin in a rabbit model of atherosclerosis. Eighteen New Zealand White rabbits were randomly divided into three groups (n=6). In the cholesterol group, rabbits consumed an atherogenic diet (0.4% cholesterol) for eight weeks. In the simvastatin group, rabbits consumed an atherogenic diet for six weeks and then consumed an atherogenic diet supplemented with simvastatin (10 mg kg(-1)) for two weeks. Biochemical analysis showed that administration of simvastatin led to an almost two-fold lowering of the total serum cholesterol, VLDL, LDL and HDL, but not triglycerides, compared with the cholesterol-fed rabbits only. Stereological analysis of the immunohistochemical staining revealed that administration of simvastatin (10 mg kg(-1) daily) in an atherogenic diet decreased the endothelial expression of P-selectin, ICAM-1 and iNOS in both aortic arch and carotid artery compared with the cholesterol fed-rabbits only. We conclude that simvastatin has beneficial effects on endothelial function by decreasing expression of P-selectin, ICAM-1 and iNOS in endothelial cells in the very early stages of atherogenesis.     

Convolution method to predict drug concentration profiles of 2,3,5,6-tetramethylpyrazine following transdermal application

The objective of this work is to predict the systemic drug concentration of 2,3,5,6-tetramethylpyrazine (TMP) following transdermal application in rabbits from the in vitro skin permeation data. The in vitro skin permeation was studied in Franz diffusion cells. Pharmacokinetic evaluation of TMP following transdermal application and bolus intravenous administration were carried out in New Zealand White (NZW) rabbits. Drug concentration-time curve following transdermal application was predicted via the convolution procedure using an in vitro skin permeation data as a weighting function, and the intravenous data as an unit impulse response. The results showed that the predicted drug concentration following transdermal application by convolution method was in good agreement with the observed drug absorption profiles. These findings indicated that in vitro skin permeation tests could be useful to predict in vivo drug absorption profiles following transdermal application.     

Apparent active transport of MDMA is not mediated by P-glycoprotein: a comparison with MDCK and Caco-2 monolayers

Amphetamines and their methylenedioxy derivatives generically display similar behavioral, physiologic and toxic effects. Inconsistent pharmacokinetic and toxicity data for methylenedioxymethamphetamine (MDMA) may suggest that active drug transporters are interacting with these compounds, and thus altering drug absorption and tissue distribution. In vitro models of CNS accumulation and intestinal drug transport were used to assess efflux transport of MDMA. Madin-Darby kidney cell epithelial (MDCK) monolayers displayed a 4-fold increase in accumulation in the basolateral to apical orientation relative to the apical to basolateral orientation, although no differential accumulation was noted between MDCK-WT and MDCK-MDR1 monolayers. Caco-2 monolayers demonstrated an approximate 2-fold increase in accumulation of MDMA. Exposure of various inhibitors of active drug transporters demonstrated mixed results; ritonavir, progesterone and indomethacin produced an approximately 50% reduction of MDMA transport, while verapamil, MK-571 and probenecid had no effect. Based on these data, it is concluded that MDMA efflux is mediated via the activity of a transporter distinct from P-glycoprotein. The possible inhibitory effects of amphetamines on rhodamine-123 transport were also assessed. MDMA, methylenedioxyamphetamine, amphetamine and methamphetamine, at physiologically relevant concentrations, did not significantly alter the transport of rhodamine-123 in Caco-2 monolayers or the LS180 cell line, suggesting that these compounds do not alter the function of P-glycoprotein.