Microvillus Membrane Vesicles Research Articles

Inter-individual variations and modulators of MDR1 transport activity in human placenta

IntroductionATP binding cassette (ABC) membrane transporter multidrug resistance 1 (MDR1) is one of the most important efflux transporters in the human placenta protecting the fetus from exposure to xenobiotic toxicity. Recent studies have focused on placental MDR1 expression, but few studies have analyzed placental MDR1 transport activity. The purpose of this study was to investigate placental MDR1 transport activity using a relatively large sample size of human placentas. Furthermore, the effect of ABCB1 gene polymorphisms were investigated along with physiological factors including maternal age, times of pregnancy, BMI, delivery mode or pregnancy complications on placental MDR1 transport activity. MethodsA total of 252 human placentas were obtained after delivery. MDR1 transport activity was detected by N-methyl quinidine uptake in placental microvillus membrane vesicles (MVMVs). Nine common single nucleotide polymorphisms (SNPs) in ABCB1 genes were determined by Snapshot. The association between ABCB1 gene polymorphisms, maternal age, times of pregnancy, BMI, delivery mode or pregnancy complications, and transporter activity was investigated. ResultsInter-individual variations of MDR1 transport activity were observed among 252 subjects. The per unit protein activity was ranged from 0.05 to 0.15/mg. Nine SNPs in ABCB1 gene didn't exhibit significant association with transporter activity of MDR1. Likewise, neither age, times of pregnancy, delivery mode nor pregnancy complications showed any significant effect of placental MDR1 transport activity. But placental MDR1 transport activity in obese pregnant women was lower than those in non-obese pregnant women. ConclusionInter-individual variations of MDR1 transport activity existed in human placentas. This may contribute to variations in drug exposure to the fetus affecting clinical outcomes. Maternal age, times of pregnancy, delivery mode nor pregnancy complications included in this study maybe not significantly impact placental MDR1 transport activity, but maternal obese could inhibit placental MDR1 activity.

Glycol chitosan: A stabilizer of lipid rafts in the intestinal brush border

Chitosan is a polycationic polysaccharide consisting of β-(1–4)-linked glucosamine units and due to its mucoadhesive properties, chemical derivatives of chitosan are potential candidates as enhancers for transmucosal drug delivery. Recently, glycol chitosan (GC), a soluble derivative of chitosan, was shown to bind specifically to lipid raft domains in model bilayers. The small intestinal brush border membrane has a unique lipid raft composition with high amounts of glycolipids cross-linked by lectins, and the aim of the present work therefore was to study the interaction of FITC-conjugated GC (FITC-GC) with the small intestinal epithelium. Using organ culture of pig jejunal mucosal explants as a model system, we observed widespread binding of luminal FITC-GC to the brush border. Only little uptake via constitutive endocytosis into apical early endosomes occurred, unless endocytosis was induced by the simultaneous presence of cholera toxin B subunit (CTB). Biochemically, GC bound to microvillus membrane vesicles and caused a change in the density profile of detergent resistant membranes (DRMs). Collectively, the results showed that FITC-GC binds passively to lipid raft domains in the brush border, i.e. without inducing endocytosis like CTB. Instead, and unlike CTB, FITC-GC seems to exert a stabilizing, detergent-protective effect on the lipid raft organization of the brush border.

Placental Glucose Transporter GLUT1 Expression in Pre-eclampsia

Placental Glucose Transporter (GLUT1) Expression in Pre- Eclampsia. INTRODUCTION: Glucose is the most important substrate for fetal growth. Indeed, there is no significant de novo glucose synthesis in the fetus and the fetal up-take of glucose rely on maternal supply and transplacental transport. Therefore, a defective placental transporter system may affect the intrauterine environment compromising fetal as well as mother well-being. On this line, we speculated that the placental glucose transport system could be impaired in pre-eclampsia (PE). METHODS: Placentae were obtained after elective caesarean sections following normal pregnancies and pre-eclamptic pregnancies. Syncytial basal membrane (BM) and apical microvillus membrane (MVM) fractions were prepared using differential ultra-centrifugation and magnesium precipitation. Protein expression was assessed by western blot. mRNA levels were quantified by quantitative real-time PCR. A radiolabeled substrate up-take assay was established to assess glucose transport activity. FACS analysis was performed to check the shape of MVM. Statistical analysis was performed using one way ANOVA test. RESULTS: GLUT1 protein levels were down-regulated (70%; P<0.01) in pre-eclamptic placentae when compared to control placentae. This data is in line with the reduced glucose up-take in MVM prepared from preeclamptic placentae. Of note, the mRNA levels of GLUT1 did not change between placentae affected by PE and normal placentae, suggesting that the levels of GLUT1 are post-transcriptionally regulated. FACS analysis on MVM vesicles from both normal placentae and pre-eclamptic placentae showed equal heterogeneity in the complexes formed. This excluded the possibility that the altered glucose up-take observed in pre-eclamptic MVM was caused by a different shape of these vesicles. CONCLUSIONS: Protein and functional studies of GLUT1 in MVM suggest that in pre-eclampsia the glucose transport between mother and fetus might be defective. To further investigate this important biological aspect we will increase the number of samples obtained from patients and use primary cells to study trans epithelial transport system in vitro.

Staphylococcus aureus enterotoxins A− and B: binding to the enterocyte brush border and uptake by perturbation of the apical endocytic membrane traffic

Enterotoxins of Staphylococcus aureus are among the most common causes of food poisoning. Acting as superantigens they intoxicate the organism by causing a massive uncontrolled T cell activation that ultimately may lead to toxic shock and death. In contrast to our detailed knowledge regarding their interaction with the immune system, little is known about how they penetrate the epithelial barrier to gain access to their targets. We therefore studied the uptake of two staphylococcal enterotoxins (SEs), SEA and SEB, using organ cultured porcine jejunal explants as model system. Attachment of both toxins to the villus surface was scarce and patchy compared with that of cholera toxin B (CTB). SEA and SEB also bound to microvillus membrane vesicles in vitro, but less efficiently than CTB, and the binding was sensitive to treatment with endoglycoceramidase II, indicating that a glycolipid, possibly digalactosylceramide, acts as cell surface receptor at the brush border. Both SEs partitioned poorly with detergent resistant membranes (DRMs) of the microvillus, suggesting a weak association with lipid raft microdomains. Where attachment occurred, cellular uptake of SEA and SEB was also observed. In enterocytes, constitutive apical endocytosis normally proceeds only to subapical early endosomes present in the actomyosin-rich "terminal web" region. But, like CTB, both SEA and SEB penetrated deep into the cytoplasm. In conclusion, the data show that after binding to the enterocyte brush border SEA and SEB perturb the apical membrane trafficking, enabling them to engage in transcytosis to reach their target cells in the subepithelial lamina propria.

Placental sulphate transport: A review of functional and molecular studies

Sulphate is required by the feto-placental unit for a number of important conjugation and biosynthetic pathways. Functional studies performed several decades ago established that sulphate transport in human placental microvillus and basal membrane vesicles was mainly via a DIDS-sensitive anion-exchange mechanism. In contrast, no evidence was found for Na+-dependent transport. Studies performed using isolated human placental tissue confirmed anion-exchange as the main mechanism. More recently, molecular studies have established the presence of anion-exchange proteins which could play a role in transplacental sulphate movement. However, the presence of transcripts for NaS2 has been reported and has prompted the suggestion that Na+-sulphate cotransport may play an important role in maternal-fetal sulphate transport. This article reviews our present knowledge of placental sulphate transport, both functional and molecular, and attempts to form a model based on the available evidence.

Mechanisms mediating the diuretic and natriuretic actions of the incretin hormone glucagon-like peptide-1

Glucagon-like peptide-1 (GLP-1) is a gut incretin hormone considered a promising therapeutic agent for type 2 diabetes because it stimulates beta cell proliferation and insulin secretion in a glucose-dependent manner. Cumulative evidence supports a role for GLP-1 in modulating renal function; however, the mechanisms by which GLP-1 induces diuresis and natriuresis have not been completely established. This study aimed to define the cellular and molecular mechanisms mediating the renal effects of GLP-1. GLP-1 (1 μg·kg(-1)·min(-1)) was intravenously administered in rats for the period of 60 min. GLP-1-infused rats displayed increased urine flow, fractional excretion of sodium, potassium, and bicarbonate compared with those rats that received vehicle (1% BSA/saline). GLP-1-induced diuresis and natriuresis were also accompanied by increases in renal plasma flow and glomerular filtration rate. Real-time RT-PCR in microdissected rat nephron segments revealed that GLP-1 receptor-mRNA expression was restricted to glomerulus and proximal convoluted tubule. In rat renal proximal tubule, GLP-1 significantly reduced Na(+)/H(+) exchanger isoform 3 (NHE3)-mediated bicarbonate reabsorption via a protein kinase A (PKA)-dependent mechanism. Reduced proximal tubular bicarbonate flux rate was associated with a significant increase of NHE3 phosphorylation at the PKA consensus sites in microvillus membrane vesicles. Taken together, these data suggest that GLP-1 has diuretic and natriuretic effects that are mediated by changes in renal hemodynamics and by downregulation of NHE3 activity in the renal proximal tubule. Moreover, our findings support the view that GLP-1-based agents may have a potential therapeutic use not only as antidiabetic drugs but also in hypertension and other disorders of sodium retention.

Association of Na+-H+ Exchanger Isoform NHE3 and Dipeptidyl Peptidase IV in the Renal Proximal Tubule

In an attempt to identify proteins that assemble with the apical membrane Na(+)-H(+) exchanger isoform NHE3, we generated monoclonal antibodies (mAbs) against affinity-purified NHE3 protein complexes isolated from solubilized renal microvillus membrane vesicles. Hybridomas were selected based on their ability to immunoprecipitate NHE3. We have characterized in detail one of the mAbs (1D11) that specifically co-precipitated NHE3 but not villin or NaPi-2. Western blot analyses of microvillus membranes and immunoelectron microscopy of kidney sections showed that mAb 1D11 recognizes a 110-kDa protein highly expressed on the apical membrane of proximal tubule cells. Immunoaffinity chromatography was used to isolate the antigen against which mAb 1D11 is directed. N-terminal sequencing of the purified protein identified it as dipeptidyl peptidase IV (DPPIV) (EC ), which was confirmed by assays of DPPIV enzyme activity. We also evaluated the distribution of the NHE3-DPPIV complex in microdomains of rabbit renal brush border. In contrast to the previously described NHE3-megalin complex, which principally resides in a dense membrane population (coated pits) in which NHE3 is inactive, the NHE3-DPPIV complex was predominantly in the microvillar fraction in which NHE3 is active. Serial precipitation experiments confirmed that anti-megalin and anti-DPPIV antibodies co-precipitate different pools of NHE3. Taken together, these studies revealed an unexpected association of the brush border Na(+)-H(+) exchanger NHE3 with dipeptidyl peptidase IV in the proximal tubule. These findings raise the possibility that association with DPPIV may affect NHE3 surface expression and/or activity.

Placental glucose transport and GLUT 1 expression in insulin-dependent diabetes

Objective: Altered transport functions in the placenta might contribute to adverse outcome of pregnancies in women with diabetes. Therefore we studied placental glucose transport in this pregnancy complication. Study Design: Syncytiotrophoblast microvillous membrane vesicles and basal membrane vesicles were isolated from women with uneventful pregnancies (control subjects, n = 21) and from women with pregnancies complicated by insulin-dependent diabetes mellitus, White class D (n = 7). Glucose uptake and GLUT 1 (glucose transporter 1) expression were studied by means of radiolabeled tracers and Western blot, respectively. Results: In the group with insulin-dependent diabetes mellitus, values for hemoglobin A 1c were moderately elevated in the first trimester (6.61 ± 0.35) but not later in pregnancy and 4 of the 7 neonates were large for gestational age. In the basal membrane vesicles, insulin-dependent diabetes mellitus was associated with a 40% increase in GLUT 1 expression and a 59% higher mediated uptake of d -glucose. No alterations could be demonstrated in microvillus membrane vesicles. Conclusion: Placental glucose transport capacity appears to be increased in insulin-dependent diabetes mellitus. These alterations might explain the occurrence of macrosomia despite well-controlled diabetes. (Am J Obstet Gynecol 1999;180:163-8.)

Glutamine transport in human and rat placenta

Glutamine plays an important role in fetal nutrition. This study explored the transport of [3H]glutamine into apical and basal predominant membrane vesicles derived from rat and human placenta. Na+-dependent glutamine transport was present in both apical and basal predominant vesicles derived from 20- and, to a lesser degree, 14-day gestation rat placenta. Amino-acid transport systems A, ASC-like, B(o,+) (in apical membrane vesicles) and, perhaps, y+L were involved in Na+-dependent glutamine transport. Na+-dependent glutamine uptake into human placental microvillus and basolateral membrane vesicles also occurred via several distinct transport activities. Glutamine transport via system N was not detected in either rat or human placental preparations. Na+-dependent glutamine transport in the rat was more pronounced in basal as compared to apical membrane vesicles. Conversely, in the human preparations, activity was significantly higher in microvillus as compared to basolateral membrane vesicles. It is concluded that Na+-dependent glutamine transport occurs through a variety of transport agencies in both the rat and human placenta. Transport varies with ontogeny and between species.

Mechanism of H(+)-coupled formate transport in rabbit renal microvillus membranes.

It has been proposed that a major fraction of Cl- absorption in the mammalian proximal tubule occurs by Cl-/formate exchange across the apical membrane with recycling of formate by nonionic diffusion. The purpose of this study was to characterize the mechanism of formate recycling in rabbit renal microvillus membrane vesicles. Formate uptake was stimulated by an inside-alkaline pH gradient. When external pH (pH alpha) was varied at constant internal pH (pHi), the initial rate of formate uptake was less than predicted for nonionic diffusion of formic acid at constant formic acid permeability. When pHi was varied at constant pHi, the initial rate of formate uptake exhibited cooperative and saturable kinetics with respect to pHi, in contrast to the pHi independence predicted for nonionic diffusion. pH gradient-stimulated [14C]formate uptake was stimulated by internal formate, indicating formate/formate exchange. pH gradient-stimulated formate influx was sensitive to inhibition by 1 mM 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid but not by furosemide or hydroxycinnamate. We conclude that pH gradient-stimulated formate uptake takes place by a carrier-mediated process of H(+)-formate cotransport, OH-/formate exchange, or facilitated formic acid diffusion, rather than solely by passive nonionic diffusion through the lipid bilayer.

P-17-5 - Regional cerebral perfusion by Tc-99m HMPAO SPECT before and after tacrine in alzheimer's disease

Microvillus membrane vesicles isolated from renal cortex have subsequently served as a useful model system for characterizing the kinetic and biochemical features of a Na+–H+ exchanger whose properties are largely representative of plasma membrane Na+–H+ exchangers in general. This chapter reviews studies on the molecular properties of the renal microvillus membrane Na+–H+ exchanger. Additional transport systems for the transfer of acids or bases exist in parallel with the Na+–H+ exchanger on the luminal membrane of proximal tubule cells. The chapter describes the kinetics of the renal Na+–H+ exchanger. As the imidazolium ring of histidine is the principal ionic group in proteins that is titratable at near-neutral pH values, in the study described in the chapter, this helped to evaluate the effect of histidine-specific reagents on the activity of the renal microvillus membrane Na+–H+ exchanger. The carboxyl-activating reagent N-ethoxycarbonyl-2-ethoxy- 1,2-dihydroquinoline (EEDQ) inhibits Na+–H + exchange in renal microvillus vesicles. Also, The observations that N,N'-dicyclohexyl-carbodiimide (DCCD) inactivates the renal Na+–H+ exchanger irreversibly and that amiloride protects against this inactivation suggested a strategy for covalently labeling the transport protein or one of its subunits. The renal Na+–H+ exchanger helps in facilitating anion transport by HCO−3 reabsorption, organic anion reabsorption, Cl− reabsorption, and anion transport and maintaining intracellular pH.

Pathways for Oxalate Transport in Rabbit Renal Microvillus Membrane Vesicles

Recent evidence suggests that apical membrane Cl--oxalate exchange plays a major role in mediating Cl- absorption in the renal proximal tubule. To sustain steady-state Cl- absorption by a mechanism of exchange for intracellular oxalate requires the presence of one or more pathways for recycling oxalate from lumen to cell. Accordingly, we evaluated the mechanisms of oxalate transport in luminal membrane vesicles isolated from the rabbit renal cortex. We found that transport of oxalate by Na+ cotransport is negligible compared to the transport of sulfate. In contrast, we demonstrated that oxalate shares the electroneutral pathway mediating Na+-independent sulfate-carbonate exchange. We also demonstrated the presence of OH--oxalate exchange (indistinguishable from H+-oxalate cotransport). The process of OH--oxalate exchange was electrogenic and partially inhibited by Cl-, indicating that it occurs, at least in part, as a mode of the Cl--oxalate exchanger described previously. An additional component of OH--oxalate exchange was insensitive to inhibition by either Cl- or sulfate, suggesting that it takes place by neither the Cl--oxalate exchanger nor the sulfate-carbonate exchanger. We conclude that multiple anion exchange mechanisms exist by which oxalate can recycle from lumen to cell to sustain Cl- absorption occurring via apical membrane Cl--oxalate exchange in the renal proximal tubule.

Inhibition of choline uptake in syncytial microvillus membrane vesicles of human term placenta: Specificity and nature of interaction

The potency and nature of the inhibitory effect of various cationic drugs on the transport of choline across the placental syncytial microvillus membrane was investigated. Tetraethylammonium, a model substrate for organic cation transport, was a poor inhibitor. Enlarging the degree of alkylation of the quaternary ammonium increased the inhibitory effect, in proportion with increasing lipophilicity. Log concentration vs % control uptake curves showed marked differences in inhibitory potency for the different cationic drugs. Hemicholinium-3 inhibited mediated choline uptake in the micromolar range, whereas atropine and mepiperphenidol were less potent. The H 2-receptor antagonists cimetidine, ranitidine, and famotidine inhibited choline uptake in the millimolar ranges. Dixon analysis revealed a competitive nature of inhibition for hemicholinium-3 and atropine ( K i = 40 μM and 1.2 mM, respectively). Cimetidine interacted noncompetitively ( K i = 3.4 mM). Since relatively high concentrations were needed to reach half maximal inhibition, impairment of fetal choline supply due to maternal drug use during pregnancy is not to be expected.

Role of glucocorticoids in the maturation of renal cortical Na+/H+ exchanger activity during fetal life in sheep

We have studied the role of glucocorticoids in inducing the maturation in activity of the proximal tubule Na+/H+ exchanger that follows birth. Renal cortical microvillus membrane vesicles were prepared from 132-day gestation sheep fetuses (n = 8) that had received intraperitoneal cortisol (13 micrograms.kg-1.h-1) for the previous 48 h. Membrane vesicles were also obtained from sham-operated twin controls (n = 8). Amiloride-sensitive uptake of 22Na+ by these vesicles was measured, and Woolf-Augustinsson-Hofstee plots were used to determine the Michaelis constant (Km) and maximal velocity (Vmax). There was no significant difference in Km; however, the Vmax was 61% higher in cortisol-treated fetuses. Posttreatment circulating cortisol levels were significantly higher in the treated fetuses. Total RNA was collected from renal cortex of the eight pairs of twins when killed. Renal cortex Na+/H+ exchanger 3 (NHE3) mRNA levels were approximately fourfold higher in cortisol-treated than in control fetuses. Although proximal tubule Na+/H+ exchanger activity and renal cortex NHE3 mRNA levels increased significantly in cortisol-treated fetuses, cortisol infusion did not stimulate renal sodium reabsorption in the fetus but rather produced a natriuresis. These results demonstrate that glucocorticoids can induce an increase in both Na+/H+ exchanger activity and NHE3 mRNA levels during the last trimester of gestation in sheep. However, these changes are not associated with an increased ability of the fetal kidney to reabsorb sodium.

Maturation of proximal tubule Na+/H+ antiporter activity in sheep during transition from fetus to newborn

We have studied maturational changes in the kinetics of the proximal tubule Na+/H+ antiporter. Microvillus membrane vesicles were prepared from renal cortex of fetal and newborn lambs. Amiloride-sensitive uptake of 22Na+ by these vesicles was measured and Woolf-Augustinsson-Hofstee plots were used to determine the Michaelis constant (Km) and rate of maximal uptake (Vmax). Initial studies of fetal lambs at 130-132 days gestation (n = 5; term is 145 days) and 3- to 4-day-old lambs (n = 5) revealed no maturational change in Km (7.27 +/- 1.25 for fetuses and 9.01 +/- 1.03 mM for lambs); however, there was a 242% increase in Vmax (from 1.28 +/- 0.33 in the fetuses to 4.37 +/- 0.85 nmol.s-1.mg protein-1 in the lambs, P = 0.005). Further definition of the developmental change in Na+/H+ antiporter Vmax was obtained when 144-day-gestation fetuses (n = 5) were compared with 24-h-old sibling lambs (n = 5) that had been delivered by cesarean section at 144 days gestation. Again, no significant difference was seen in Na+/H+ antiporter Km (14.9 +/- 6.5 for fetuses and 12.5 +/- 3.4 mM for lambs); however, a significant increase in Na+/H+ antiporter Vmax occurred (from 1.41 +/- 0.51 in the fetuses to 3.32 +/- 0.37 nmol.s-1.mg protein-1 in the lambs, P < 0.01). This study shows that there is a maturational increase in renal cortical Na+/H+ antiporter Vmax during the transition from fetal to newborn life. This increase parallels the increase in renal tubular Na+ reabsorption that occurs at this time.

Specificity of choline uptake into syncytial microvillus membrane vesicles of human term placenta: Inhibition by organic cations

The clam Scrobicularia plana and the polychaete worm Nereis diversicolor were collected in several sites from a littoral enclosure in SW Spain. The aim of our study was to relate various biomarker responses in these species to a pollution gradient caused by untreated domestic discharges and to verify the adequacy of the selected species as sentinels in this habitat. The biomarkers selected were the antioxidant enzymes catalase (CAT), glutathione peroxidase (GPX) and DT-diaphorase (DT-D). In addition, the activities of cytochrome P450-dependent ethoxyresorufin O-deethylase (EROD) activity, the phase II detoxifying enzyme glutathione S-transferase (GST) and the neurotoxicity marker acetylcholinesterase (AChE) were measured. Metallothionein levels were selected as biomarkers of heavy metals exposure in both species. The results suggest a different response in the water filtering organism (clam) and the sediment eater (polychaete), probably as a consequent of different pollution exposure and that samples from the “Caño Sancti-Petri” were exposed to biologically active compounds that altered some of their biochemical responses. AChE was the most sensitive biomarker in both species and N. diversicolor proved to be a more robust sentinel in this ecosystem.

Uptake of choline into syncytial micro villus membrane vesicles of human term placenta

The uptake of the quaternary ammonium compound choline was studied in syncytial microvillus membrane vesicles of human term placenta. Uptake was stimulated by an inside negative membrane potential and by loading the vesicles with unlabeled choline. Imposition of an inwardly directed Na + or outwardly directed H + gradient did not stimulate choline uptake. Several organic cations were able to inhibit choline transport in the following order: hemicholinium-3 ≥ choline ≥mepiperphenidol > cimetidine ≥ famotidine. The kinetics of uptake involved a saturable process for choline with high affinity ( K m = 550 μm). Our results confirm the presence of a carrier mediated transport system in human placental syncytial microvillus membranes. The system appears to be electrogenic, and able to transport choline efficiently from the maternal circulation into the placenta.

Inhibition of Ca-activated K+ channels from renal microvillus membrane vesicles by amiloride analogs

The effect of the K(+)-sparing diuretic amiloride and two of its hydrophobic analogs, methylisobutyl amiloride (MIA) and ethylisopropyl amiloride (EIPA), on Ca-activated K+ channels from renal microvillus membrane vesicles incorporated into planar lipid bilayers was investigated. Amiloride did not inhibit currents through Ca-activated K+ channels. MIA and EIPA, however, inhibited channel currents when added to both the internal and external solutions in concentrations between 10 and 250 microM. Furthermore, when dose-response data for channel inhibition were examined using Hill plots, Hill numbers of approximately 1.5 were found for both blockers from both sides, suggesting that the mechanism of block involves multiple inhibitory binding sites. A simple kinetic scheme is proposed that can account for the results.

Proline transport across the intestinal microvillus membrane may be regulated by membrane physical properties

There is now abundant evidence that integral membrane protein function may be modulated by the physical properties of membrane lipids. The intestinal brush border membrane represents a membrane system highly specialized for nutrient absorption and, thus, provides an opportunity to study the interaction between integral membrane transport proteins and their lipid environment. We have previously demonstrated that alterations in this environment may modulate the function of the sodium-dependent glucose transporter in terms of its affinity for glucose. In this communication we report that membrane lipid-protein interactions are distinctly different for the proline transport proteins. Maximal transport rates for l-proline by either the neutral brush border or amino transport systems are reduced 10-fold when the surrounding membrane environment is made more fluid over the physiological range that exists along the crypt-villus axis. Furthermore, in microvillus membrane vesicles prepared from enterocytes isolated from along the crypt villus axis a similar gradient exists in the functional activity of these transport systems. This would imply that either the functional activity of these transporters are regulated by membrane physical properties or that the synthesis and insertion of these proteins is coordinated in concert with membrane physical properties as the enterocyte migrates up the crypt-villus axis.

Salicylate inhibits human placental sulphate transport in vitro

The effect of salicylate on sulphate transport by the human placenta has been studied using isolated brush-border membrane vesicles and placental tissue slices. Sulphate uptake by isolated vesicles was inhibited in a dose dependent fashion ( K I ≈ 3 mM) by salicylate. It appears that this drug blocks sulphate accumulation in a non-competitive manner. Sulphate efflux from preloaded vesicles was also found to be markedly reduced by salicylate in a non-competitive fashion. Consistent with the vesicle studies salicylate inhibited sulphate transport by placental tissue slices. The results suggest that salicylate ingestion could compromise feto-placental sulphate homeostasis. In addition we have found that the aspirin-like drug, flufenamic acid, inhibits sulphate transport by isolated microvillus membrane vesicles.