PubMed Health⌕ Search

Biomedical subjects

T D DuBose

Publications and source records attributed to T D DuBose.

At least 19 recordsLinked to original sources
PubMed2000

Molecular and pathophysiologic mechanisms of hyperkalemic metabolic acidosis.

In summary, hyperkalemia may have a dramatic impact on ammonium production and excretion. Chronic hyperkalemia decreases ammonium production in the proximal tubule and whole kidney, inhibits absorption of NH4+ in the mTALH, reduces medullary interstitial concentrations of NH4+ and NH3, and decreases entry of NH4+ and NH3 into the medullary collecting duct. The potential for development of a hyperchloremic metabolic acidosis is greatly augmented when renal insufficiency with associated reduction in functional renal mass coexists with the hyperkalemia, or in the presence of aldosterone deficiency or resistance. Such a cascade of events helps to explain, in part, the hyperchloremic metabolic acidosis and reduction in net acid excretion characteristic of several experimental models of hyperkalemic-hyperchloremic metabolic acidosis including: obstructive nephropathy, selective aldosterone deficiency, and chronic amiloride administration (7.9).

Acidosis↗
PubMed1999

The colonic H+,K+-ATPase functions as a Na+-dependent K+(NH4+)-ATPase in apical membranes from rat distal colon.

Recent studies have suggested that the colonic H+,K+-ATPase (HKalpha2) can secrete either Na+ or H+ in exchange for K+. If correct, this view would indicate that the transporter could function as either a Na+ or a H+ pump. To investigate this possibility a series of experiments was performed using apical membranes from rat colon which were enriched in colonic H+,K+-ATPase protein. An antibody specific for HKalpha2 was employed to determine whether HKalpha2 functions under physiological conditions as a Na+-dependent or Na+-independent K+-ATPase in this same membrane fraction. K+-ATPase activity was measured as [gamma-32P]ATP hydrolysis. The Na+-dependent K+-ATPase accounted for approximately 80% of overall K+-ATPase activity and was characterized by insensitivity to Sch-28080 but partial sensitivity to ouabain. The Na+-independent K+-ATPase activity was insensitive to both Sch-28080 and ouabain. Both types of K+-ATPase activity substituted NH4+ for K+ in a similar manner. Furthermore, our results demonstrate that when incubated with native distal colon membranes, the blocking antibody inhibited dramatically Na+-dependent K+-ATPase activity. Therefore, these data demonstrate that HKalpha2 can function in native distal colon apical membranes as a Na+-dependent K+-ATPase. Elucidation of the role of the pump as a transporter of Na+ versus H+ or NH4+ versus K+ in vivo will require additional studies.

Adenosine Triphosphatases↗
PubMed1999

Unexpected severe hypocalcemia during continuous venovenous hemodialysis with regional citrate anticoagulation.

Citrate is known to induce acute hypocalcemia in patients undergoing liver transplantation during the anhepatic phase. We describe the case of a 71-year-old woman with fulminant hepatic failure secondary to hepatitis A, who was started on continuous venovenous hemodialysis (CVVHD) for acute renal failure. Because anticoagulation with heparin was untenable, regional anticoagulation was accomplished by trisodium citrate (46.7%) infusion. Unfortunately, severe hypocalcemia developed when citrate accumulated because of impaired hepatic metabolism. Because of chelation by citrate, the ionized calcium concentration declined to values as low as 2.72 mg/dL (normal, 4.5 to 5.6 mg/dL), whereas the total calcium concentration remained in the normal range. With an unusually high calcium chloride infusion rate via a central line (up to 140 mL/h of 10 mEq/dL CaCl2) and additional boli of CaCl2 (for a total of 190 mEq), the ionized calcium concentration could be maintained at target levels. Nevertheless, the ionized calcium concentration was maintained in the normal range, and the total calcium concentration increased to a value as high as 15 mg/dL. Thus, the total to ionized calcium ratio was 3.5:1. After 24 hours of treatment, trisodium citrate infusion was gradually reduced from 15 mL/h to 7 mL/h, and the calcium chloride infusion was decreased to 50 mL/h. Nevertheless, persistence of the elevated total to ionized calcium ratio (3:1) indicated citrate accumulation likely secondary to decreased hepatic metabolism. Using this approach, the patient was successfully maintained on CVVHD with regional citrate anticoagulation for a total of 11 days without any additional complications. We conclude that CVVHD with regional citrate anticoagulation can be used in patients with acute hepatic failure if increased CaCl2 requirements are anticipated and if citrate is infused at a lower rate compatible with decreased citrate metabolism. Citrate accumulation should be suspected in patients with an elevated total to ionized Ca++ ratio during CVVHD with citrate anticoagulation.

Acute Kidney Injury↗
PubMed1999

H+,K+-ATPase.

The H+,K+-ATPases comprise a group of integral membrane proteins that belong to the X+,K+-ATPase subfamily of P-type cation-transporting ATPases. Although these H+,K+-ATPase isoforms share approximately 60-70% amino acid identity, they exhibit discrete kinetic and pharmacological properties when expressed in heterologous systems. HK alpha2 has been categorized by its insensitivity to Sch-28080, an inhibitor of the gastric H+,K+-ATPase, and partial sensitivity to ouabain, an inhibitor of the Na+,K+-ATPase. This functional profile contrasts with the pharmacological sensitivities ascribed to HK alpha2 in transport studies in rat isolated medullary collecting ducts perfused in vitro and in mouse medullary collecting duct cell lines. HK alpha2 mRNA and protein abundance appears to be both tissue and site-specifically upregulated in response to chronic hypokalemia. This regulatory response has been localized to the outer and inner medulla. To reconcile these expressed sensitivities to those reported in vitro in isolated tubules and cells in culture, it would be necessary to invoke modification of the pharmacologic insensitivity of the colonic H+,K+-ATPase to Sch-28080. Although a 'unique' beta-subunit has been reported recently, this beta-subunit (beta(c)) is identical at the amino acid level to the recently cloned beta3-Na+,K+-ATPase. Moreover, while HK alpha2 can assemble indiscriminately with any X+,K+-ATPase beta-subunit, HK alpha2 has been reported to assemble stably with beta1-Na+,K+-ATPase in the renal medulla and in the distal colon. It remains conceivable that subunit assembly could be tissue specific and might respond to different physiological and pathophysiological stimuli. Furthermore, recent studies have suggested that the H+,K+-ATPase is both Na+-dependent and localized to the apical membrane in the distal colon. Therefore, future studies will need to resolve these discrepancies by determining if a unique, yet undiscovered H+,K+-ATPase isoform exists in kidney, or if post-translational modifications of the alpha- and/or beta-subunits could account for these functional diversities.

Animals↗
PubMed1999

Contrasting functional and regulatory profiles of the renal H+,K+-ATPases.

The H+,K+-ATPases belong to the X+,K+-ATPase subfamily of P-type cation-transporting ATPases. While these H+,K+-ATPase isoforms share approximately 60%-70% amino acid identity, they exhibit discrete kinetic and pharmacological properties. The colonic alpha isoform (HKalpha2) is insensitive to Sch-28080, an inhibitor of the gastric H+,K+-ATPase, and is sensitive to high concentrations of ouabain. This profile contrasts with the sensitivities attributed to HKalpha2 in transport studies. HKalpha2 mRNA and protein abundance appear to be both site-specifically upregulated in response to chronic hypokalemia, and have been localized to the outer and inner medulla. To reconcile expressed sensitivities with those reported in vitro in isolated tubules and cells in culture, it requires transformation of the expressed insensitivity of the colonic H+,K+-ATPase to Sch-28080. Although a "unique" beta subunit has been reported recently, this beta subunit ("betac"), is identical at the amino acid level to the recently cloned beta3-Na+,K+-ATPase. Moreover, while HKalpha2 can assemble indiscriminately with any X+,K+-ATPase beta subunit, HKalpha2 has been reported to assemble stably with beta1-Na+,K+-ATPase in the renal medulla and in the distal colon. It is conceivable that subunit assembly could be tissue-specific and might respond to different physiological and pathophysiological stimuli. Recent studies have suggested that the H+,K+-ATPase is both Na+-dependent and localized to the apical membrane in the distal colon. Future studies will be needed to resolve these discrepancies by determining if a unique, yet undiscovered H+,K+-ATPase isoform exists in the kidney, or if posttranslational modifications of the alpha and/or beta-subunits could account for these functional diversities.

Adaptation, Physiological↗
PubMed1998

The alpha-subunit of the colonic H+,K+-ATPase assembles with beta1-Na+,K+-ATPase in kidney and distal colon.

Previous experiments from our laboratory (Codina, J., Kone, B. C., Delmas-Mata, J. T., and DuBose, T. D., Jr. (1996) J. Biol. Chem. 271, 29759-29763) demonstrated that the alpha-subunit of the colonic H+, K+-ATPase (HKalpha2) requires coexpression with a beta-subunit to support H+/K+ transport in a heterologous expression system (Xenopus laevis oocytes). In these studies, HKalpha2 formed stable and functional alpha.beta complexes when coexpressed with either the rat beta1-subunit of the Na+,K+-ATPase or the beta-subunit of the gastric H+,K+-ATPase, suggesting that different beta-subunits may interact with HKalpha2. The present studies tested this hypothesis by development and application of a specific antibody against HKalpha2 peptide. Subsequently, immunoprecipitation experiments were performed to determine if HKalpha2 co-precipitates with the same beta-subunit in organs known to express HKalpha2 protein. The data demonstrate that HKalpha2 assembles with beta1-Na+,K+-ATPase in the renal medulla and in distal colon.

Animals↗
PubMed1998

Expression of HKalpha2 protein is increased selectively in renal medulla by chronic hypokalemia.

Our laboratory has demonstrated by Northern analysis that chronic hypokalemia increases HKalpha2 (i.e., alpha-subunit of the colonic H+-K+-ATPase) mRNA abundance in the rat. To determine whether the increase in mRNA correlated with an increase in HKalpha2 protein, an antibody was raised against a synthetic peptide derived from amino acids 686-698 of the HKalpha2 sequence. The anti-HKalpha2 antibody hybridized to rat distal colon membranes which migrated at approximately 100 kDa (expected mobility of HKalpha2). HKalpha2 protein was not detected in plasma membranes from rat whole kidney or stomach (100 microg) derived from control animals. The antibody was then used to investigate changes in expression of HKalpha2 in renal cortex, renal medulla, and distal colon in two pathophysiological conditions: 1) chronic hypokalemia (LK) and 2) chronic metabolic acidosis (CMA). In LK rats there was a marked, but selective, increase in the abundance of HKalpha2 protein in membranes prepared from renal medulla. Nevertheless, a corresponding increase in HKalpha2 protein abundance was not observed in membranes prepared from the distal colon of LK rats. HKalpha2 protein abundance in CMA was indistinguishable from controls. Moreover, chronic hypokalemia had no effect on expression of alpha1-Na+-K+-ATPase or HKalpha1 in kidney or distal colon under any experimental condition. Therefore, HKalpha2 protein is tissue- and site-specifically upregulated in response to chronic hypokalemia but not by CMA. Furthermore, this regulatory response is localized to the renal medulla.

Animals↗
PubMed1998

Dietary K+ restriction upregulates total and Sch-28080-sensitive bicarbonate absorption in rat tIMCD.

In tubules from the terminal segment of the inner medullary collecting duct (tIMCD) from rats with chronic metabolic acidosis, our laboratory has shown that bicarbonate absorption (JtCO2) is inhibited by removal of K+ from the luminal fluid or by the addition of Sch-28080 to the perfusate. The present study asked whether total and/or Sch-28080-sensitive JtCO2 is regulated by changes in systemic K+ homeostasis. Rat tIMCD tubules were perfused in vitro in symmetrical, HCO-3/CO2-buffered solutions containing 10 mM KCl + 6 mM NH4Cl. Total and Sch-28080-sensitive JtCO2 were measured in rats with varying K+ intake. In K+-replete rats, baseline JtCO2 was 2.1 +/- 0.3 pmol . mm-1 . min-1 (n = 6). In rats fed a K+-deficient diet for 3 days, JtCO2 was 5.4 +/- 0.7 pmol . mm-1 . min-1 (n = 16, P < 0. 05). To determine the mechanism for the increase in HCO-3 absorption observed with K+ restriction, the Sch-28080-sensitive component of JtCO2 was measured in each treatment group. Following the addition of Sch-28080 (10 microM) to the perfusate, a 40% reduction in JtCO2 was observed in K+-restricted rats. JtCO2 was not reduced following the addition of Sch-28080 in rats with normal K+ intake. Because Sch-28080-sensitive JtCO2 was increased in K+-restricted rats, Sch-28080-sensitive JtCO2 was studied further in tIMCD tubules from rats in this treatment group. In K+-restricted rats, JtCO2 decreased by 20% following the addition of 5 mM ouabain to the perfusate. This ouabain-induced decline in JtCO2 was observed both in the presence and in the absence of Sch-28080. We conclude that total and Sch-28080-sensitive net acid secretion is increased with dietary K+ restriction. However, since approximately 50% of JtCO2 is insensitive to both Sch-28080 and ouabain, future studies will be necessary to define other mechanisms of luminal acidification in the rat tIMCD.

Absorption↗
PubMed1997

Acute renal failure in the 21st century: recommendations for management and outcomes assessment.

Acute renal failure (ARF) remains a common and potentially devastating disorder affecting as many of 5% of all hospitalized patients, with a higher prevalence in patients in critical care units. ARF is more frequently observed in the setting of multiorgan dysfunction syndrome (MODS) and in elderly patients with complex disease, where mortality is high. Numerous technical advances have not yet impacted favorably on this high mortality rate. This report summarizes recommendations from participants at the National Institutes of Health Conference: "Acute Renal Failure in the 21st Century," May 6 to 8, 1996, in Bethesda, MD. The focus is on categorizing recent clinically relevant developments in the field and on identification of new research initiatives to transfer a new body of knowledge derived from fundamental studies and laboratory investigation to the management of ARF in the new millennium. The development of a multicenter database through cooperative multicenter studies is advocated. Future studies should define the appropriate outcome measures to assess and emphasize the impact of hemodynamic monitoring, adjunctive agents, and adequacy and modality of renal replacement therapy on outcomes in ARF.

Acute Kidney Injury↗
PubMed1997

Adaptation to low-K+ media increases H(+)-K(+)-ATPase but not H(+)-ATPase-mediated pHi recovery in OMCD1 cells.

Studies in rat and rabbit outer medullary collecting duct of inner stripe origin (OMCDis) suggest that both H(+)-ATPase and H(+)-K(+)-ATPase participate in H+ secretion. However, the relative contributions of these transporters, and, in particular, that of H(+)-K(+)-ATPase to K+ absorption have not been defined precisely. The present study was designed to delineate more clearly the response of these two transporters to hypokalemia and acidosis in a newly developed mouse OMCD1 cell line. In cells grown in normal K+ (5 mM) media, intracellular pH (pHi) recovery was similar either in the presence or absence of K+ in the perfusate (delta pHi/min = 0.014 +/- 0.001 vs. 0.017 +/- 0.003, not significant). The inhibitory effects of Sch-28080 (10 microM) and bafilomycin A1 (10 nM) on pHi recovery were evident only in the presence and absence of K+ in the perfusate, respectively. In cells grown in low-K+ (2.5 mM) media to simulate chronic hypokalemia, pHi recovery was significantly faster than in cells grown in normal K+ media (delta pHi/min = 0.045 +/- 0.01 vs. 0.014 +/- 0.001, P < 0.01) and was inhibited specifically by Sch-28080, not by bafilomycin A1. In contrast, in cells preconditioned to low pH (7.0) to simulate chronic acidosis, the enhanced pHi recovery was abolished by bafilomycin A1 but not by Sch-28080. 86Rb+ uptake, when used as a K+ congener, was inhibited by Sch-28080. The K(m) for 86Rb+ uptake (H(+)-K(+)-ATPase activity) and the 50% inhibitory concentration for Sch-28080 were 270 and 5.0 microM, respectively. These studies provide evidence that, in morphologically homogeneous OMCD1 cells, 1) both H(+)-K(+)-ATPase and H(+)-ATPase participate in pHi regulation, 2) the H(+)-K(+)-ATPase is selectively upregulated by preconditioning in low-K+ media, and 3) conversely, preconditioning in low-pH media stimulates only the H(+)-ATPase. Thus, in OMCDis, the H(+)-K(+)-ATPase and H(+)-ATPase respond selectively and independently to chronic hypokalemia and acidosis, respectively.

Adaptation, Physiological↗
PubMed1997

Effect of chronic hypokalemia on H(+)-K(+)-ATPase expression in rat colon.

Although the kidney plays the major role in the regulation of systemic K+ homeostasis, the colon also participates substantively in K+ balance. The colon is capable of both K+ absorption and secretion, the magnitude of which can be modulated in response to dietary K+ intake. The H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase) has been proposed as a possible mediator of K+ absorption in distal colon, but inhibitor profiles obtained in recent studies suggest that two, and perhaps more, distinct H(+)-K(+)-ATPase activities may be present in mammalian distal colon. We have developed highly specific probes for the catalytic alpha-subunits of colonic and gastric H(+)-K(+)-ATPase, alpha 1-Na(+)-K(+)-ATPase, and beta-actin, which were used in Northern analysis of total RNA from whole distal colon and stomach obtained from one of three experimental groups of rats: 1) controls, 2) chronic dietary K+ depletion, and 3) chronic metabolic acidosis. The probe for the colonic but not the gastric H(+)-K(+)-ATPase alpha-isoform hybridized to distal colon total RNA in all groups. A significant increase in colonic H(+)-K(+)-ATPase mRNA abundance was observed in response to chronic dietary K+ depletion but not to chronic metabolic acidosis. The alpha 1-isoform of Na(+)-K(+)-ATPase, which is also expressed in distal colon, did not respond consistently to either chronic dietary K+ depletion or chronic metabolic acidosis. The gastric probe did not hybridize to total RNA from distal colon but, as expected, hybridized to total stomach RNA. However, the abundance of gastric H(+)-K(+)-ATPase or Na(+)-K(+)-ATPase in stomach was not altered consistently by either chronic dietary K+ depletion or metabolic acidosis. Under the conditions of this study, it appears that the mRNA encoding the colonic alpha-isoform is upregulated by chronic dietary K+ restriction, a condition shown previously to increase K+ absorption in the distal colon.

Acidosis↗
PubMed1996

Functional expression of the colonic H+,K+-ATPase alpha-subunit. Pharmacologic properties and assembly with X+,K+-ATPase beta-subunits.

The functional and pharmacological properties of the alpha-subunit of the colonic H+,K+-ATPase (alphaC) were studied in Xenopus laevis oocytes. alphaC was injected with different rat beta-subunits, the beta-subunit of the gastric H+,K+-ATPase (betaG, the only H+, K+-ATPase beta-subunit identified in rat), or the beta1-subunit of the Na+,K+-ATPase (beta1) (associated with the basolateral Na+, K+-ATPase, but also expressed in the epithelial apical membranes of rat distal colon) (Marxer, A., Stieger, B., Quarini, A., Kashgarian, M., and Hauri, H. P. (1989) J. Cell Biol. 109, 1057-1069). The effect of the different beta-subunits was studied by measuring 86Rb+ uptake (a K+ congener) in the presence or absence of Sch-28080 and ouabain. Significant Na+-independent 86Rb+ uptake was observed only when alphaC was coexpressed with one of the beta-subunits. The expressed alphaCbeta1 and alphaCbetaG complexes were not inhibited by Sch-28080, were only partially sensitive to ouabain (IC50 = 400-600 microM, in the presence of external 1 mM KCl), and exhibited comparable K+ activation kinetics. Coexpression of alphaC with epitope-tagged betaG or beta1, followed by immunopurification of the alphabeta complexes, confirmed stable assembly of alphaCbetaG and alphaCbeta1 complexes. Since the beta1-subunit, but not the alpha1-subunit, of Na+,K+-ATPase is expressed in the apical membrane of rat colonocytes, our data support the view that, in rat distal colon, the beta1-subunit may play a surrogate role as the beta-subunit for the colonic H+,K+-ATPase.

Animals↗
PubMed1996

Effective clearance of methotrexate using high-flux hemodialysis membranes.

We report the first series demonstrating effective clearance of methotrexate using acute intermittent hemodialysis with a high-flux dialyzer. The study was performed on six patients, two females and four males aged 13 to 72 years. All were patients at M.D. Anderson Cancer Center. Patients were dialyzed for 4 to 6 hours daily using a Fresenius F-80 membrane (Fresenius Inc, Walnut Creek, CA). Following the initiation of dialysis, there was a reduction in arterial and venous serum concentration of methotrexate with time. Mean plasma clearance of methotrexate during dialysis in these six patients was 92.1 +/- 10.3 mL/min. One patient who was nearly functionally anephric was studied in detail. In this patient, following a high dose of methotrexate (7.2 g/m2), approximately 63% of this dose was cleared with 6 hours of hemodialysis. With subsequent dialysis performed daily for 6 hours, the drug was cleared completely in 5.6 +/- 0.3 days (n = 7 separate methotrexate treatments). A reduction in plasma methotrexate concentration from 1,733 +/- 40 micromol/L 1 hour postinfusion to less than 0.3 micromol/L in 5 to 6 days was observed for these seven separate treatments. We conclude that significant clearance of methotrexate can be achieved with high-flux dialyzers, making methotrexate therapy a viable treatment option in patients with responsive malignancies despite the presence of renal failure.

Adolescent↗
PubMed1996

H,K-ATPase.

At least four unique H,K-ATPases have been cloned and are expressed in several tissues. Recent findings have enhanced our appreciation of the roles of H,K-ATPases in the kidney with respect to their molecular identities, functional properties, segmental and intrarenal distribution, and regulatory features. The major role of the H,K-ATPases expressed in mammalian kidney, the gastric and colonic H,K-ATPases, is to regulate potassium and bicarbonate absorption in collecting duct segments.

Absorption↗
PubMed1996

Role of H(+)-K(+)-ATPase in pHi regulation in inner medullary collecting duct cells in culture.

Studies in inner medullary collecting duct (IMCD) cells in primary culture have proposed two mechanisms for Na(+)-independent hydrogen ion transport: an H(+)-adenosinetriphosphatase (H(+)-ATPase) and an H(+)-K(+)-ATPase. In the present study, we have employed two sources of IMCD cells, cells in primary culture derived from the terminal papilla of the Munich-Wistar rat (IMCDp) and an established murine cell line (mIMCD-3), to define the predominant mechanism(s) of Na(+)-independent intracellular pH (pHi) recovery in the IMCD. In confluent monolayers of IMCDp and mIMCD-3 cells, pHi was measured using the pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF) following addition and withdrawal of NH4Cl. Removal of K+ completely abolished Na(+)-independent pHi recovery in both IMCDp (delta pHi/min = 0.039 +/- 0.006 to 0.005 +/- 0.003; P < 0.001) and in mIMCD-3 (delta pHi/min = 0.055 +/- 0.009 to -0.003 +/- 0.002; P < 0.001) cells, respectively. In mIMCD-3 cells, K(+)-dependent pHi recovery was abolished by either of two specific inhibitors of the H(+)-K(+)-ATPase, Sch-28080 (5 or 10 microM) or A-80915A (10 microM). In contrast, bafilomycin A1 (2.5 and 10 nM), an inhibitor of the H(+)-ATPase, failed to attenuate K(+)-dependent pHi recovery. Moreover, sequence verified mouse gastric and colonic alpha-H(+)-K(+)-ATPase probes hybridized to total RNA from mIMCD-3 cells. Based on these findings, we conclude that Na(+)-independent pHi recovery from an acid load in both IMCDp and mIMCD-3 cells in critically dependent on extracellular K(+)-That K(+)-dependent pHi recovery was inhibited by both Sch-28080 and A-80915A but not by bafilomycin A1 suggests that the predominant mechanism by which Na(+)-independent pHi recovery is accomplished in IMCD is through the H(+)-K(+)-ATPase. Expression of both gastric and colonic alpha-H(+)-K(+)-ATPase mRNA in mIMCD-3 cells suggests that one or both of these H(+)-K(+)-ATPases may be responsible for proton secretion in the IMCD.

Animals↗