A randomized trial of minidose warfarin for the prevention of late malfunction in tunneled, cuffed hemodialysis catheters.

BACKGROUND: Minidose warfarin (1 mg/day) has been associated with a 74% reduction in the thrombosis rate of central venous catheters used in oncology patients. To determine the efficacy of minidose warfarin on late malfunction caused by thrombosis or fibrin sheath formation in tunneled, cuffed catheters (TCC) used for hemodialysis (HD), we performed a randomized, placebo-controlled trial. METHODS: One hundred five chronic HD patients with TCCs were initially randomized. Of these, 85 (warfarin 41 and placebo 44) completed the first two weeks of the protocol and were followed for the first year of TCC life or until TCC removal. RESULTS: Sixteen TCCs failed with late TCC malfunction, eight in each group. In a multivariate analysis, there was no significant effect of warfarin on thrombosis-free TCC survival or time to the first urokinase (UK) instillation for incipient thrombosis. The presence of a low hemoglobin (Hgb; <10.5 g/dL) or a low international normalized ratio (INR; <1.00) was significantly associated with a higher risk of late TCC malfunction (RR 5.2 and 4.0, respectively), a higher risk of incipient TCC thrombosis requiring UK (RR 2.0 and 2.8, respectively), and higher rates of UK dosing. Diabetics had a 3.6-fold higher risk of late TCC malfunction and a twofold higher risk of incipient thrombosis requiring UK, although these findings were not statistically significant. Aspirin use, race, age, number of hospitalizations, erythropoietin dose, intradialytic heparin dose, serum albumin, and the number of episodes of TCC-associated infection were not significantly associated with late TCC malfunction. CONCLUSIONS: Thrombosis prophylaxis using fixed minidose warfarin is not efficacious in TCCs used for HD. However, the present data suggest improved TCC survival in patients with an INR> 1.00. Patients with diabetes and those with a low Hgb or INR have a higher risk of late TCC malfunction.

A basolateral lactate/H+ co-transporter in Madin-Darby Canine Kidney (MDCK) cells.

Monolayers of Madin-Darby Canine Kidney (MDCK) cells grown on permeable filters generated lactate aerobically and accumulated it preferentially in the basolateral compartment, suggesting the presence of a lactate carrier. The mechanism of lactate transport across apical and basolateral membranes was examined by determining intracellular pH (pHi) microspectrofluorimetrically after addition of lactate to the extracellular solutions and by measuring uptake of [14C]lactate. Addition of 20 mM lactate to the apical compartment produced no change in pHi, whereas lactate added to the basolateral compartment rapidly and reversibly lowered pHi. Pyruvate produced similar results. Inhibitors of lactate/H+ co-transporters, alpha-cyano-4-hydroxycinnamate (CnCN) and quercetin, partially inhibited the fall in pHi produced by basolateral lactate. In contrast, the disulphonic stilbene. DIDS (4,4'-di-isothiocyanostilbene-2,2'-disulphonic acid) produced no inhibition at 0.5 mM. Kinetic analysis was performed by applying basolateral lactate at various concentrations and measuring the rate of entry (delta pHi/min) in the presence and absence of CnCN. Lactate flux was shown to occur by both non-ionic diffusion and a alpha-cyano-4-hydroxycinnamate-sensitive component (carrier). The latter has a Km of approximately 7 mM for the lactate anion. Propionate, but not formate, lowered pHi to the same degree as did equimolar lactate, but the propionate effect was not inhibited by CnCN. Influx of [14C]lactate was substantially greater across the basolateral membrane than across the apical membrane and occurred in the absence of Na+. We conclude that MDCK cells grown on permeable filters generate lactate aerobically and transport it across the basolateral membrane by way of a lactate/H+ cotransporter.

Imaging of filter-grown epithelial cells: MDCK Na(+)-H+ exchanger is basolateral.

We report a simple method for growing epithelial cells on permeable supports and for imaging the cells from the apical side using an inverted microscope. Madin-Darby canine kidney (MDCK) cells were either seeded onto the conventional side of Millipore-CM filters or onto "inverted" filters. The peak transepithelial resistance of confluent monolayers was the same with either system. Cells on inverted filters that were stained with various dyes and imaged by epifluorescence exhibited more distinct intercellular spaces, cell margins, nuclei, and subapical vesicles. We also perfused both sides of inverted filters with HCO3/CO2-free saline and measured intracellular pH (pHi) using 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF) and digital imaging. The intrinsic buffer capacity of MDCK cells increased exponentially as pHi decreased. After an NH4Cl load, the H+ extrusion rate (JH+) in control saline was 2.42 +/- 0.62 mM/min. JH+ was completely blocked by 1 mM basolateral amiloride. In contrast, 1 mM apical amiloride had no effect. We conclude that 1) growth of epithelial cells on an inverted filter system is useful for the microspectrofluorimetric determination of pHi in single cells and for the imaging of apical/subapical structures, and 2) the Na(+)-H+ exchanger of MDCK cells is functionally polarized to the basolateral membrane.