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U Hopfer

Publications and source records attributed to U Hopfer.

At least 19 recordsLinked to original sources
PubMed2000

Regulation of NHE3 activity by G protein subunits in renal brush-border membranes.

NHE3 activity is regulated by phosphorylation/dephosphorylation processes and membrane recycling in intact cells. However, the Na(+)/H(+) exchanger (NHE) can also be regulated by G proteins independent of cytoplasmic second messengers, but the G protein subunits involved in this regulation are not known. Therefore, we studied G protein subunit regulation of NHE3 activity in renal brush-border membrane vesicles (BBMV) in a system devoid of cytoplasmic components and second messengers. Basal NHE3 activity was not regulated by G(s)alpha or G(i)alpha, because antibodies to these G proteins by themselves were without effect. The inhibitory effect of D(1)-like agonists on NHE3 activity was mediated, in part, by G(s)alpha, because it was partially reversed by anti-G(s)alpha antibodies. Moreover, the amount of G(s)alpha that coimmunoprecipitated with NHE3 was increased by fenoldopam in both brush-border membranes and renal proximal tubule cells. Furthermore, guanosine 5'-O-(3-thiotriphosphate) but not guanosine 5'-O-(2-thiodiphosphate), the inactive analog of GDP, increased the amount of G(s)alpha that coimmunoprecipitated with NHE3. The alpha(2)-adrenergic agonist, UK-14304 or pertussis toxin (PTX) alone had no effect on NHE3 activity, but UK-14304 and PTX treatment attenuated the D(1)-like receptor-mediated NHE3 inhibition. The ability of UK-14304 to attenuate the D(1)-like agonist effect was not due to G(i)alpha, because the attenuation was not blocked by anti-G(i)alpha antibodies or by PTX. Anti-Gbeta(common) antibodies, by themselves, slightly inhibited NHE3 activity but had little effect on D(1)-like receptor-mediated NHE3 inhibition. However, anti-Gbeta(common) antibodies reversed the effects of UK-14304 and PTX on D(1)-like agonist-mediated NHE3 inhibition. These studies provide concrete evidence of a direct regulatory role for G(s)alpha, independent of second messengers, in the D(1)-like-mediated inhibition of NHE3 activity in rat renal BBMV. In addition, beta/gamma dimers of heterotrimeric G proteins appear to have a stimulatory effect on NHE3 activity in BBMV.

Adrenergic alpha-Agonists↗
PubMed1999

Carnitine transport and its inhibition by sulfonylureas in human kidney proximal tubular epithelial cells.

The kidney plays an important role in the homeostasis of carnitine by its ability to reabsorb carnitine almost completely from the glomerular filtrate. The transport process responsible for this reabsorption has been investigated thus far only in laboratory animals. Here we report on the characteristics of carnitine uptake in a proximal tubular epithelial cell line derived from human kidney. The uptake process was found to be obligatorily dependent on Na+ with no involvement of anions. The process was saturable, with a Michaelis-Menten constant of 14 +/- 1 microM. The Na+:carnitine stoichiometry was 1:1. The same process also was found to be responsible for the uptake of acetylcarnitine and propionylcarnitine, two acyl esters of carnitine with potential for therapeutic use in humans. The uptake process was specific for carnitine and its acyl esters. Betaine, a structural analog of carnitine, interacted with the uptake process to a significant extent. The present studies also showed that sulfonylureas, oral hypoglycemic agents currently used in the management of type 2 diabetes, inhibited the carnitine uptake system. Among the sulfonylureas tested, glibenclamide was the most potent inhibitor. The inhibition was competitive. Glibenclamide inhibited the uptake not only of carnitine but also of acetylcarnitine and propionylcarnitine. The inhibition most likely was the result of direct interaction of the compound with the carnitine transporter because the inhibition could be demonstrated in purified rat kidney brush border membrane vesicles.

Administration, Oral↗
PubMed1999

Effects of pH on kinetic parameters of the Na-HCO3 cotransporter in renal proximal tubule.

The effects of pH on cotransporter kinetics were studied in renal proximal tubule cells. Cells were grown to confluence on permeable support, mounted in an Ussing-type chamber, and permeabilized apically to small monovalent ions with amphotericin B. The steady-state, dinitrostilbene-disulfonate-sensitive current (DeltaI) was Na+ and HCO3- dependent and therefore was taken as flux through the cotransporter. When the pH of the perfusing solution was changed between 6.0 and 8.0, the conductance attributable to the cotransporter showed a maximum between pH 7.25 and pH 7.50. A similar profile was observed in the presence of a pH gradient when the pH of the apical solutions was varied between 7.0 and 8.0 (basal pH lower by 1), but not when the pH of the basal solution was varied between 7.0 and 8.0 (apical pH lower by 1 unit). To delineate the kinetic basis for these observations, DeltaI-voltage curves were obtained as a function of Na+ and HCO3- concentrations and analyzed on the basis of a kinetic cotransporter model. Increases in pH from 7.0 to 8.0 decreased the binding constants for the intracellular and extracellular substrates by a factor of 2. Furthermore, the electrical parameters that describe the interaction strength between the electric field and substrate binding or charge on the unloaded transporter increased by four- to fivefold. These data can be explained by a channel-like structure of the cotransporter, whose configuration is modified by intracellular pH such that, with increasing pH, binding of substrate to the carrier is sterically hindered but electrically facilitated.

Animals↗
PubMed1999

Purinergic and cholinergic agonists induce exocytosis from the same granule pool in HT29-Cl.16E monolayers.

Several secretagogues induce mucin secretion in epithelial monolayers, as determined by measuring released granule contents. To assess whether different agonists act on the same granule pool, capacitance changes in intact monolayers of the goblet cell line HT29-Cl.16E were measured by a novel impedance method. Apical ATP (purinergic agonist) and basolateral carbachol (cholinergic agonist) induce rapid exocytosis with maximal capacitance changes within 3 min. The maximal levels of exocytosis that can be induced by optimal concentrations of either agonist are the same and produce a 30-40% increase in total monolayer capacitance. When ATP and carbachol are applied simultaneously, the magnitude of exocytosis is unchanged from the single-secretagogue level. The recovery of capacitance to baseline (endocytosis) is significantly faster after ATP stimulation than after carbachol stimulation. When ATP and carbachol are applied sequentially at doses that give maximal exocytosis, the magnitude of the capacitance increase produced by the second secretagogue is less than or equal to that of the capacitance decrease during the recovery period. Together, these data suggest that purinergic and cholinergic agonists act on the same granule pool.

Adenosine Triphosphate↗
PubMed1999

Renal proximal tubular AT2 receptor: signaling and transport.

Previous studies have documented that the vasoactive agonist angiotensin II (AngII) directly affects proximal tubular sodium-bicarbonate reabsorption in a biphasic manner, whereby picomolar concentrations promote reabsorption and nanomolar concentrations have the converse effect. Although it is generally agreed that the AT1 receptor subtype mediates AngII-induced sodium-bicarbonate reabsorption primarily through adenylate cyclase, the receptor subtype mediating natriuresis is less well defined. Using mouse proximal tubular cells, this study documents AT1-dependent enhancement (candesartan-inhibitable) of bicarbonate reabsorption and AT2-induced (PD123319- and CGP42112A-inhibitable) decrement of bicarbonate absorption. The signaling mechanisms were examined in rabbit proximal tubule cells in culture. The AT2 signaling involves G protein beta- and gamma-mediated phospholipase A2 activation, arachidonic acid release, and downstream events linked to Shc/Grb2/Sos and p21ras rather than protein kinase C as reported previously for AngII receptors. These observations provide a novel mechanism for AngII-AT2 receptor-mediated transport modulation.

Angiotensin II↗
PubMed1998

Role of hypoxia-induced Bax translocation and cytochrome c release in reoxygenation injury.

We investigated mechanisms of cell death during hypoxia/reoxygenation of cultured kidney cells. During glucose-free hypoxia, cell ATP levels declined steeply resulting in the translocation of Bax from cytosol to mitochondria. Concurrently, there was cytochrome c release and caspase activation. Cells that leaked cytochrome c underwent apoptosis after reoxygenation. ATP depletion induced by a mitochondrial uncoupler resulted in similar alterations even in the presence of oxygen. Moreover, inclusion of glucose during hypoxia prevented protein translocations and reoxygenation injury by maintaining intracellular ATP. Thus, ATP depletion, rather than hypoxia per se, was the cause of protein translocations. Overexpression of Bcl-2 prevented cytochrome c release and reoxygenation injury without ameliorating ATP depletion or Bax translocation. On the other hand, caspase inhibitors did not prevent protein translocations, but inhibited apoptosis during reoxygenation. Nevertheless, they could not confer long-term viability, since mitochondria had been damaged. Omission of glucose during reoxygenation resulted in continued failure of ATP production, and cell death with necrotic morphology. In contrast, cells expressing Bcl-2 had functional mitochondria and remained viable during reoxygenation even without glucose. Therefore, Bax translocation during hypoxia is a molecular trigger for cell death during reoxygenation. If ATP is available during reoxygenation, apoptosis develops; otherwise, death occurs by necrosis. By preserving mitochondrial integrity, BCL-2 prevents both forms of cell death and ensures cell viability.

Adenosine Triphosphate↗
PubMed1998

Purinergic receptor-induced changes in paracellular resistance across cultures of human cervical cells are mediated by two distinct cytosolic calcium-related mechanisms.

In human cervical (CaSki) cells, extracellular adenosine triphosphate (ATP) induces an acute decrease in the resistance of the lateral intercellular space (RLIS), phase I response, followed by an increase in tight junctional resistance (RTJ), phase II response. ATP also stimulates release of calcium from intracellular stores, followed by augmented calcium influx, and both effects have similar sensitivities to ATP (EC50 of 6 microM). The objective of the study was to determine the degree to which the changes in [Ca2+]i mediate the responses to ATP. 1,2-bis (2-aminophenoxy) ethane-N,N,N1,N1-tetraacetic acid (BAPTA) abrogated calcium mobilization and phase I response; in contrast, nifedipine and verapamil inhibited calcium influx and attenuated phase II response. Barium, La3+, and Mn2+ attenuated phase I response and attenuated and shortened the ionomycin-induced phase I-like decrease in RLIS, suggesting that store depletion-activated calcium entry was inhibited. Barium and La3+ also inhibited the ATP-induced phase II response, but Mn2+ had no effect on phase II response, and in the presence of low extracellular calcium it partly restored the increase in RTJ. KCl-induced membrane depolarization stimulated an acute decrease in RLIS and a late increase in RTJ similar to ATP, but only the latter was inhibited by nifedipine. KCl also induced a nifedipine-sensitive calcium influx, suggesting that acute increases in [Ca2+]i, regardless of mobilization or influx, mediate phase I response. Phase II-like increases in RTJ could be induced by treatment with diC8, and were not affected by nifedipine. Biphasic, ATP-like changes in RTE could be induced by treating the cells with ionomycin plus diC8. We conclude that calcium mobilization mediates the early decrease in RLIS, and calcium influx via calcium channels activates protein kinase C and mediates the late increase in RTJ.

Adenosine Triphosphate↗
PubMed1998

Voltage and cosubstrate dependence of the Na-HCO3 cotransporter kinetics in renal proximal tubule cells.

The voltage dependence of the kinetics of the sodium bicarbonate cotransporter was studied in proximal tubule cells. This electrogenic cotransporter transports one Na+, three HCO3-, and two negative charges. Cells were grown to confluence on a permeable support, mounted on a Ussing-type chamber, and permeabilized apically to small monovalent ions with amphotericin B. The steady-state, di-nitro-stilbene-di-sulfonate-sensitive current was shown to be sodium and bicarbonate dependent and therefore was taken as flux through the cotransporter. Voltage-current relations were measured as a function of Na+ and HCO3- concentrations between -160 and +160 mV under zero-trans and symmetrical conditions. The kinetics could be described by a Michaelis-Menten behavior with a Hill coefficient of 3 for HCO3- and 1 for Na+. The data were fitted to six-state ordered binding models without restrictions with respect to the rate-limiting step. All ordered models could quantitatively account for the observed current-voltage relationships and the transinhibition by high bicarbonate concentration. The models indicate that 1) the unloaded transporter carries a positive charge; 2) the binding of cytosolic bicarbonate to the transporter "senses" 12% of the electric field in the membrane, whereas its translocation across the membrane "senses" 88% of the field; 3) the binding of Na+ to the cotransporter is voltage independent.

Amphotericin B↗
PubMed1998

System for dynamic measurements of membrane capacitance in intact epithelial monolayers.

Dynamic measurements of exocytosis have been difficult to perform in intact epithelial monolayers. We have designed a system that estimates with +/-1% accuracy (99% confidence) the total membrane capacitance of monolayers represented by a lumped model. This impedance measurement and analysis system operates through a conventional transepithelial electrophysiology clamp, performing all signal measurements as frequently as every 5 s. Total membrane capacitance (the series combination of apical and basolateral membranes) is the inverse of one of three unique coefficients that describe the monolayer impedance. These coefficients are estimated using a weighted, nonlinear, least-squares algorithm. Using the estimated coefficients, solution ranges for individual membrane parameters are calculated, frequently providing results within +/-20% of true values without additional electrophysiological measurements. We determined the measurement system specifications and statistical significance of estimated parameters using 1) analytical testing with circuit simulation software and equation-generated data; 2) a system noise analysis combined with Monte Carlo simulations; and 3) analog model circuits for calibration of the electronic system and to check equation-generated results. Finally, the time course of capacitance changes associated with purinergically stimulated mucin exocytosis are quantified in monolayers of the colonic goblet cell-like cell line HT29-CI.16E.

Algorithms↗
PubMed1998

Therapeutic concentrations of cyclosporine A, but not FK506, increase P-glycoprotein expression in endothelial and renal tubule cells.

BACKGROUND: The immunosuppressive drugs cyclosporine A (CsA) and tacrolimus (FK506) are extruded from cells by the multidrug resistance P-glycoprotein (P-gp), an efflux pump for drugs and xenobiotics, which may limit their therapeutic effectiveness and/or incidence of toxic side effects. In the present study, we investigated the effect of therapeutic concentrations of CsA and FK506 on the expression of P-gp in cultured endothelial and proximal tubule cells. METHODS: P-gp expression in human arterial endothelial (HAEC) and rat proximal tubule cells (RPTC) was determined by immunoblotting and immunocytochemistry, and correlated with P-gp-mediated transport by measuring the intracellular accumulation of the fluorescent probe calcein. RESULTS: Following incubation of HAEC with therapeutic concentrations of 0.1 to 1.6 microM CsA up to seven days, P-gp expression increased in a time- and concentration-dependent manner, maximally to 291 +/- 42% of controls with 0.8 microM CsA for seven days. Similar effects of CsA were observed in RPTC. In contrast, therapeutic concentrations of FK506 (0.01 to 0.2 microM up to 7 days) did not change P-gp expression in either cell type, though at higher, supratherapeutic concentrations of FK506 (0.6 to 1.2 microM) P-gp expression was also increased. Immunocytochemistry revealed increased P-gp expression in the plasma membrane of HAEC and RPTC treated with 0.8 microM CsA, which was reflected by a decrease of P-gp-mediated accumulation of calcein in both cell types. CONCLUSIONS: The data suggest that the induction of P-gp expression in HAEC and RPTC at concentrations of CsA or FK506 above 0.5 microM is part of the protective answer of cells to toxic concentrations of the drugs and could therefore interfere with the therapeutic effectiveness of CsA in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Sodium space and intravascular volume: dietary sodium effects in cystic fibrosis and healthy adolescent subjects.

OBJECTIVE: To assess the physiologic response to salt depletion in subjects with cystic fibrosis (CF) and control male adolescents for sodium balance, sodium space, and stimulation of the renin-angiotensin-aldosterone axis. DESIGN: Seven subjects with CF and six controls received a salt-replete (150 or 290 mmol NaCl per day) diet and then a salt-deplete (10 mmol NaCl per day) diet while in a clinical research center. RESULTS: Space maintenance: CF subjects responded to salt depletion with a greater weight loss than did controls (1.9 vs 0.8 kg) and a decrease in 24Na+ space, whereas controls maintained 24Na+ space. Paired (Na-deplete/Na-replete) blood volumes decreased in subjects with CF, but not in controls. Renin-angiotensin-aldosterone axis stimulation: During salt repletion, subjects with CF had significantly higher aldosterone values than did controls in the afternoon, but not at 7:00 AM. During salt depletion, plasma renin activity and aldosterone increased significantly more in subjects with CF than in controls (renin, 35 vs 13 ng/mL/hour [9.7 vs 3.6 ng.L-1 s-1]; aldosterone: 181 vs 101 ng/dL [5021 vs 2802 pmol/L]). Furthermore, the angiotensin antagonist saralasin increased renin much more in subjects with CF (154 vs 36 ng/mL per hour [43 vs 10 ng.L-1 s-1]). Vasomotor functions: Mean arterial pressure was decreased in subjects with CF on both diets and decreased significantly more with low salt only in subjects with CF. During salt depletion, subjects with CF showed enhanced orthostatic tolerance (less heart rate increase with standing) compared with controls, thus obscuring their volume loss. The blood pressure response to an acute infusion of saralasin suggested that in salt-replete subjects with CF, but not in controls, angiotensin receptors were functional in maintaining vascular tone. During salt depletion, angiotensin was more important for maintenance of blood pressure in subjects with CF than in controls, because the saralasin-induced drop in blood pressure was 20%, ie, close to shock levels, in subjects with CF, and only 6% in controls. CONCLUSION: The data suggest that patients with CF are so successful in compensating for volume depletion by vigorous activation of the renin-angiotensin system that salt depletion/dehydration cannot be recognized easily by routine clinical measurements, eg, capillary refill, serum sodium levels, or tachycardia.

Adolescent↗
PubMed1997

Pharmacological evidence that calcium is not required for P2-receptor-stimulated Cl- secretion in HT29-Cl.16E.

Extracellular ATP at micro- to millimolar concentrations activates Cl- conductance and increases cytosolic calcium ([Ca]i) in many epithelial cells, including the colonic epithelial cell line HT29-Cl. 16E. Therefore, [Ca]i has been postulated to be the intracellular messenger for Cl- channel activation. HT29-Cl.16E is a highly differentiated cell line that forms confluent monolayers and secretes mucins and Cl-. The involvement of [Ca]i in the purinergically-stimulated Cl- secretion was investigated pharmacologically in this cell line by whole-cell patch-clamp and Ussing chamber techniques, as well as [Ca]i measurements in fura-2 loaded cells. The calmodulin inhibitors W13 (5 microm) and chlorpromazine (50 microm) abolished increases in ATP-stimulated [Ca]i-increases by 90% and 80%, respectively. However, these inhibitors had no effect on the ATP-stimulated Cl- conductance measured in either individual cells or confluent monolayers. As controls, the effects of W13 and chlorpromazine on Ca2+-ionophore stimulated Cl- conductance was measured. In this case, the two compounds inhibited whole cell Cl- conductance and monolayer Isc by 90% and 100%, respectively. These data demonstrate: (1) The purinergically-stimulated increase in Cl- current does not require an increase in [Ca]i, suggesting the involvement of either another signaling pathway or direct activation of Cl- channels by purinergic receptors. (2) A calmodulin or a calmodulinlike binding site that is sensitive to W13 and chlorpromazine participates in the regulation of the [Ca]i increase by purinergic receptors in HT29-Cl.16E.

Calcium↗
PubMed1997

Extracellular Ca2+ directly regulates tight junctional permeability in the human cervical cell line CaSki.

Lowering extracellular calcium concentration ([Ca2+]o) increases acutely and reversibly the transepithelial electrical conductance (G(TE)) and the epithelial permeability to pyranine (Ppyr) across CaSki cultures. Effects were already observed after lowering calcium from 1.2 to 1.0 mM and were maximal at 0.1 mM. The dose-response curves were sigmoidal (calcium concentration that produces half-maximal effect = 0.3 mM), and the time courses indicated simple exponential trends (time constants of 4-5 min). The effect of calcium was not mediated by mobilization of cytosolic calcium or altering calcium influx, and manganese was found to be a partial agonist to [Ca2+]o. The effects of [Ca2+]o, on permeability were additive to those of hypertonic conditions, indicating that calcium modulates junctional permeability. The experimental data were fitted to theoretical models that relate changes in G(TE) to the probability of assembled/disassembled tight junctions. The results suggest that calcium interacts directly and cooperatively at extracellular sites with junctional elements that are arranged in parallel, and it shifts the probability state of the junctions from "open" to "closed" state. Changes in extracellular calcium may affect the permeability of tight junctions of the cervical epithelium and may play a role in regulating production of cervical mucus.

Calcium↗
PubMed1997

Purinergic-mediated inhibition of Na+-K+-ATPase in proximal tubule cells: elevated cytosolic Ca2+ is not required.

The involvement of cytosolic Ca2+ concentration ([Ca2+]i) as messenger for the regulation of Na+-K+-ATPase activity was investigated in a renal cell line recently developed by immortalization of early proximal tubule primary cultures from the Wistar-Kyoto rat strain. Na+-K+-ATPase was measured as short-circuit current (Isc) in intact monolayers after permeabilization of the apical plasma membrane with amphotericin B. With symmetrical solutions, Isc quantitatively reflects Na+-K+-ATPase activity as judged by ouabain inhibition and dependence on Na+ and K+. Extracellular ATP (50% effective concentration = 0.32 mM) on the apical side produced acute inhibition of Na+-K+-ATPase-generated Isc of up to 50%. The inhibition peaked within 1 min and lasted approximately 5 min. The potency order was ATP > ADP >> beta,gamma-methyleneadenosine 5'-triphosphate = UTP, consistent with a P2y receptor. Extracellular ATP also stimulated a transient increase in [Ca2+]i. This increase had a similar time course as the inhibition of ATPase and reached a peak change of approximately 120 nM. However, the elevation of [Ca2+]i is not required in the purinergic inhibition of the Na+-K+-ATPase, since, first, increases in [Ca2+]i produced with a Ca2+ ionophore (ionomycin) failed to mimic the purinergic inhibition and, second, 1,2-bis(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid, which abolished the [Ca2+]i elevation, failed to block the purinergic inhibition.

Adenosine Triphosphate↗
PubMed1997

Purinergic agonists, but not cAMP, stimulate coupled granule fusion and Cl- conductance in HT29-Cl.16E.

Cl- conductance and capacitance were simultaneously measured in the mucin-secreting cell line, HT29-Cl.16E (Cl.16E), and its sister cell line, HT29-Cl.19A (Cl.19A), which lacks mucin granules. Purinergic stimulation by extracellular ATP transiently increased Cl- conductance in both cell lines with similar peak increases of 0.92 and 1.00 nS/pF in Cl.16E and Cl.19A cells, respectively (baseline of 0.08 nS/pF). Cell capacitance increased only in Cl.16E cells (17% above baseline of 22 pF in Cl.16E and 1% above baseline of 18 pF in Cl.19A cells). Wortmannin inhibited the purinergically activated Cl- conductance and capacitance increases in Cl.16E by 50 and 80%, respectively, but had no effects in Cl.19A cells. In Cl.16E cells, adenosine 3',5'-cyclic monophosphate (cAMP) signaling increased Cl- conductance from 0.08 to 0.52 nS/pF without changing capacitance. Cl- secretion in Cl.16E monolayers was additive in response to supramaximal stimulation of purinergic receptors and adenylyl cyclase, even though granule fusion is nine times greater with purinergic than adenylyl cyclase stimulation. In conclusion, 1) wortmannin does not directly inhibit activation of Cl- conductance, 2) at least 50% of purinergically activated Cl- conductance in Cl.16E is associated with granule fusion, and 3) cAMP-activated Cl- conductance is not associated with granules.

Adenosine Triphosphate↗
PubMed1997

Dipeptide-induced Cl- secretion in proximal tubule cells.

During a survey of dipeptides that might be transported by the renal PEPT2 transporter in proximal tubule cells, we discovered that acidic dipeptides could stimulate transient secretory anion current and conductance increases in intact cell monolayers. The stimulatory effect of acidic dipeptides was observed in several proximal tubule cell lines that have been recently developed by immortalization of early proximal tubule primary cultures from the Wistar-Kyoto and spontaneously hypertensive rat strains and humans, suggesting that this phenomenon is a characteristic of proximal tubule cells. The electrical current induced in intact monolayers by Ala-Asp, a representative of these acidic dipeptides, must represent Cl- secretion rather than Na+ or H+ absorption, because 1) it was Na+ independent, 2) it showed a pH dependence different from that of the PEPT2 cotransporter, and 3) it correlated with an Ala-Asp-induced increase in Cl- conductance of the apical membrane in basolaterally amphotericin B-permeabilized monolayers. The secretory current could be inhibited by stilbene disulfonates, but not diphenylamine-2-carboxylates, suggesting a non-cystic fibrosis transmembrane conductance regulator type of Cl- conductance. The effect of Ala-Asp was dose dependent, with an apparent 50% effective concentration of approximately 1 mM. Ala-Asp also produced intracellular acidification, suggesting that acidic dipeptides are also substrates for an H(+)-peptide cotransporter.

Animals↗
PubMed1997

Effect of TGF-beta 1 and TNF-alpha on the plasminogen system of rat proximal tubular epithelial cells.

Rat proximal tubular epithelial cells derived from Wistar-Kyoto and spontaneously hypertensive rats were grown to confluency on semipermeable tissue culture inserts, and the plasminogen system of these cells was analyzed using enzyme assays, Western analysis, zymography, and reverse transcriptase-polymerase chain reaction. The tubular epithelial cells are capable of activating exogenous plasminogen to plasmin by endogenous plasminogen activators. The cells produce tissue-plasminogen activator, urokinase-plasminogen activator, plasminogen activator inhibitor-1, and urokinase-plasminogen activator receptor. These cells also produce the Heymann nephritis autoantigen, gp330 (megalin), and an associated protein of 45 kd (RAP). Incubation with transforming growth factor-beta 1 resulted in a decrease in plasminogen activation, primarily because of an increase in plasminogen activator inhibitor-1 RNA and protein and a decrease in u-PA RNA as noted by quantitative reverse transcriptase-polymerase chain reaction, Western analysis, and zymography. Incubation of these cells with tumor necrosis factor-alpha resulted in an increase in plasminogen activating ability, presumably through an increase in urokinase. Gp330 and the associated 45-kd protein (RAP) RNA were decreased in cells treated with tumor necrosis factor-alpha. The data presented indicates that these transformed proximal tubular epithelial cells may be used to study changes that may occur during Heymann nephritis with respect to the plasminogen system and the autoantigen gp330.

Animals↗