Acidification Of Blood Research Articles

O2-Filled Swimbladder Employs Monocarboxylate Transporters for the Generation of O2 by Lactate-Induced Root Effect Hemoglobin

The swimbladder volume is regulated by O2 transfer between the luminal space and the blood In the swimbladder, lactic acid generation by anaerobic glycolysis in the gas gland epithelial cells and its recycling through the rete mirabile bundles of countercurrent capillaries are essential for local blood acidification and oxygen liberation from hemoglobin by the “Root effect.” While O2 generation is critical for fish flotation, the molecular mechanism of the secretion and recycling of lactic acid in this critical process is not clear. To clarify molecules that are involved in the blood acidification and visualize the route of lactic acid movement, we analyzed the expression of 17 members of the H+/monocarboxylate transporter (MCT) family in the fugu genome and found that only MCT1b and MCT4b are highly expressed in the fugu swimbladder. Electrophysiological analyses demonstrated that MCT1b is a high-affinity lactate transporter whereas MCT4b is a low-affinity/high-conductance lactate transporter. Immunohistochemistry demonstrated that (i) MCT4b expresses in gas gland cells together with the glycolytic enzyme GAPDH at high level and mediate lactic acid secretion by gas gland cells, and (ii) MCT1b expresses in arterial, but not venous, capillary endothelial cells in rete mirabile and mediates recycling of lactic acid in the rete mirabile by solute-specific transcellular transport. These results clarified the mechanism of the blood acidification in the swimbladder by spatially organized two lactic acid transporters MCT4b and MCT1b.

The mechanism of local blood acidification in the swimbladder by spatially organized monocarboxylate transporters

The gas in the swimbladder of teleost fish is composed primarily of O2, and the swimbladder volume is regulated by O2 transfer between the luminal space of the swimbladder and the blood. In the swimbladder, lactic acid generation by anaerobic glycolysis in the gas gland epithelial cells and its recycling through the rete mirabile bundles of countercurrent capillaries are essential for local blood acidification and oxygen liberation from hemoglobin by the “Root effect.” To clarify molecules that are involved in the blood acidification and visualize the route of lactic acid movement, we analyzed the expression of 16 members of the H+/monocarboxylate transporter (MCT) family in the fugu genome and found that only MCT1b and MCT4b are highly expressed in the fugu swimbladder. Immunohistochemistry demonstrated that (i) MCT4b expresses in gas gland cells together with the glycolytic enzyme GAPDH at high level and mediate lactic acid secretion by gas gland cells, and (ii) MCT1b expresses in arterial, but not venous, capillary endothelial cells in rete mirabile and mediates recycling of lactic acid in the rete mirabile by solute‐specific transcellular transport. Electrophysiological analyses demonstrated that MCT1b is a high‐affinity lactate transporter whereas MCT4b is a low‐affinity/high‐conductance lactate transporter. These results clarified the mechanism of the blood acidification in the swimbladder by spatially organized two lactic acid transporters MCT4b and MCT1b.

Histological demonstration of glucose transporters, fructose-1,6-bisphosphatase, and glycogen in gas gland cells of the swimbladder: Is a metabolic futile cycle operating?

Luminal surface of the swimbladder is covered by gas gland epithelial cells and is responsible for inflating the swimbladder by generating O2 from Root-effect hemoglobin that releases O2 under acidic conditions. Acidification of blood is achieved by lactic acid secreted from gas gland cells, which are poor in mitochondria but rich in the glycolytic activity. The acidic conditions are locally maintained by a countercurrent capillary system called rete mirabile. To understand the regulation of anaerobic metabolism of glucose in the gas gland cells, we analyzed the glucose transporter expressed there and the fate of ATP generated by glycolysis. The latter is important because the ATP should be immediately consumed otherwise it strongly inhibits the glycolysis rendering the cells unable to produce lactic acid anymore. Expression analyses of glucose transporter (glut) genes in the swimbladder of fugu (Takifugu rubripes) by RT-PCR and in situ hybridization demonstrated that glut1a and glut6 are expressed in gas gland cells. Immunohistochemical analyses of metabolic enzymes demonstrated that a gluconeogenesis enzyme fructose-1,6-bisphosphatase (Fbp1) and a glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (Gapdh) are highly expressed in gas gland cells. The simultaneous catalyses of glycolysis and gluconeogenesis reactions suggest the presence of a futile cycle in gas gland cells to maintain the levels of ATP low and to generate heat that helps reduce the solubility of O2.

Hémodialyse sans acétate : qu’est-ce vraiment ?

Substituting bicarbonate by acetate in dialysis fluids has been proposed for avoiding precipitation of calcium and magnesium carbonates. However, acetate hemodialysis has been abandoned because of deleterious effects of acetate. Conventional bicarbonate hemodialysis is not totally acetate-free, because 3 to 7 mEq/l of acetic acid are added to the dialysate. Acetate-free hemodialysis is possible with another acid (chlorhydric acid or citric acid) or without acid by using some techniques of low-efficiency hemodiafiltration, as acetate-free biofiltration, which avoids the deleterious effect of blood acidification into the dialyzer. In this paper, advantages and disadvantages of different techniques of acetate-free hemodialysis are discussed.

Postmortem Acidification of Blood/Organs Induces an Increase in Flecainide Concentration in Cardiac Blood and the Contribution of the Lungs to This Increase

Postmortem acidification of blood and the contribution of this phenomenon to increased flecainide concentrations in cardiac blood were evaluated in rabbits. Flecainide was administered intravenously, antemortem peripheral blood was collected 15 min after administration, then rabbits were sacrificed. Blood and organs were collected immediately or 24 h after death, or immediately or 24 h after performance of cardiac massage. Postmortem left/right cardiac blood and organs showed lower pH than antemortem blood, and flecainide concentrations in all postmortem blood samples were higher than those in antemortem blood. Increased flecainide concentrations in cardiac blood were enhanced by postmortem cardiac massage and postmortem interval. In perfusion experiments using rabbit lung and heart, even if the flecainide concentration in inflow was kept constant, outflow concentrations were 2- to 3-fold higher than in inflow when inflow pH changed from 7.4 to 5.5. In contrast, flecainide concentration in outflow decreased immediately and then remained low when pH of perfusate changed from 5.5 to 7.4. These results demonstrate that flecainide accumulates in the lungs before death, and this accumulated flecainide releases into blood following postmortem acidification of blood/organs.

Blood acidification enhances carbon dioxide removal of membrane lung: an experimental study

Extracorporeal CO2 removal is an effective procedure to allow a protective ventilatory strategy in ARDS patients, but it is technically challenging due to the high blood flow required. Increasing the CO2 transfer through the membrane lung (ML) may lower the demand of extracorporeal blood flow and consequently allow for a wider clinical application of this technique. Since only the dissolved CO2 (5% of the total CO2 content) is easily removed by the ML, we tested whether acidifying the blood entering the ML to convert bicarbonate ions towards dissolved CO2 could enhance the CO2 transfer though the ML. Six pigs were connected to an extracorporeal circuit comprising a ML. The extracorporeal blood flow was 500 ml/min, while the gas flow was 10 l/min. A 15-min continuous infusion of 0.5 N lactic acid was added to the extracorporeal blood flow before the ML at a rate of 1, 2 and 5 mEq/min. Between steps we waited for a reequilibration time of at least 30 min. Acid infusion at 0, 1, 2 and 5 mEq/min increased pCO2 (56.19 +/- 7.92, 68.24 +/- 11.73, 84.28 +/- 11.17 and 136.66 +/- 18.46 mmHg, respectively) and decreased pH (7.39 +/- 0.05, 7.30 +/- 0.05, 7.20 +/- 0.05 and 6.91 +/- 0.05, respectively). ML CO2 removal increased 11, 23 and 70% during acid infusion at 1, 2 and 5 mEq/min, respectively. Blood acidification at the inlet of a ML with infusion of 1, 2 and 5 mEq/min of lactic acid can increase the CO2 removal capacity of the ML up to 70%.

Acidification of Blood Is Superior to Sodium Fluoride Alone as an Inhibitor of Glycolysis

Sodium fluoride is the preferred agent to inhibit glycolysis. Its action is not immediate, however, and complete inhibition is delayed for up to 4 hours. A more effective method is needed. Acidification of blood combined with the addition of NaF and EDTA appears to be such a method. We studied whether acidification was indeed more effective than NaF. We conducted 6 independent studies over a 10-month period at 3 Quest Diagnostics laboratory sites. In each study, we drew venous blood from 6-24 nonfasting employee volunteers into 3 or 4 different serum- or plasma-collection tubes, which were stored under different conditions and aliquoted at different times. We analyzed the aliquots in duplicate by means of a hexokinase-based enzymatic method. The mean glucose concentration decreased by 0.3% at 2 h and by 1.2% at 24 h when blood was drawn into tubes containing citrate buffer, NaF, and EDTA. In contrast, the mean glucose concentration decreased by 4.6% at 2 h and by 7.0% at 24 h when blood was drawn into tubes containing NaF and sodium oxalate. Acidification should replace NaF alone as the recommended method for obtaining an accurate glucose concentration. Diagnostic cut points based on blood samples collected into tubes containing NaF as the only inhibitor of glycolysis are likely to be too low.

Differential Effect of Extracellular Acidosis on the Release and Dispersal of Soluble and Membrane Proteins Secreted from the Weibel-Palade Body

Proteins secreted from Weibel-Palade bodies (WPBs) play important roles in regulating inflammatory and hemostatic responses. Inflammation is associated with the extracellular acidification of tissues and blood, conditions that can alter the behavior of secreted proteins. The effect of extracellular pH (pH(o)) on the release of von Willebrand factor (VWF), the VWF-propolypeptide (Proregion), interleukin-8, eotaxin-3, P-selectin, and CD63 from WPBs was investigated using biochemical approaches and by direct optical analysis of individual WPB fusion events in human endothelial cells expressing green or red fluorescent fusions of these different cargo proteins. Between pH(o) 7.4 and 7.0, ionomycin-evoked WPB exocytosis was characterized by the adhesion of VWF to the cell surface and the formation of long filamentous strands. The rapid dispersal of Proregion, interleukin-8, and eotaxin-3 into solution, and of P-selectin and CD63 into the plasma membrane, was unaltered over this pH(o) range. At pH(o) 6.8 or lower, Proregion remained associated with VWF, in many cases WPB failed to collapse fully and VWF failed to form filamentous strands. At pH(o) 6.5 dispersal of interleukin-8, eotaxin-3, and the membrane protein CD63 remained unaltered compared with that at pH(o) 7.4; however, P-selectin dispersal into the plasma membrane was significantly slowed. Thus, extracellular acidification to levels of pH(o) 6.8 or lower significantly alters the behavior of secreted VWF, Proregion, and P-selectin while rapid release of the small pro-inflammatory mediators IL-8 and eotaxin-3 is essentially unaltered. Together, these data suggest that WPB exocytosis during extracellular acidosis may favor the control of inflammatory processes.

Counterpoint: Lactic acid is not the only physicochemical contributor to the acidosis of exercise

to the editor: The acidification of blood during muscular exercise cannot be understood from an analysis of blood from a single site, nor from examination of only arterial (or arterialized) blood. Acidification of blood during muscular contraction occurs as a result of rapid and marked increases in

The acid nature of CO2‐evoked adenosine release in the CNS

The acid nature of CO₂‐evoked adenosine release in the CNS

Volume of RBCs, 24- and 48-hour posttransfusion survivals, and the lifespan of (51)Cr and biotin-X-N-hydroxysuccinimide (NHS)-labeled autologous baboon RBCs: effect of the anticoagulant and blood pH on (51)Cr and biotin-X-NHS elution in vivo.

The RBC injury that occurs during collection of the first few milliliters of blood into the pH 5.0 ACD (NIH, Formula A) is referred to as the lesion of collection. The RBC injury was evaluated by labeling the ACD RBCs with (51)Cr and measuring the 24-hour posttransfusion survival. The effect of the acidification of ACD blood on the in vivo elution of (51)Cr from the RBC has not been reported. Baboon blood was collected in heparin and in ACD and CP2D at different ratios of blood to anticoagulant. ACD blood with a pH of 5.7 to 6.9 was labeled with (51)Cr. Heparinized blood with a pH of 7.4 was labeled with biotin-X-N-hydroxysuccinimide (NHS). ACD blood with a pH of 5.9 was labeled with both (51)Cr and biotin-X-NHS. The RBC volumes, 24- and 48-hour posttransfusion survivals, and lifespans (T50) were measured. The RBC volume of ACD blood with a pH ranging from 5.7 to 6.9 was not affected by (51)Cr labeling. (51)Cr-labeled ACD blood with a pH of 6.9 had an RBC volume that was significantly greater than that seen in heparinized blood with a pH of 7.4 labeled with biotin-X-NHS. In vivo elution of (51)Cr from the RBCs prepared from the ACD blood with a pH of 5.7 to 6.9 and in vivo elution of biotin-X-NHS from RBCs prepared from ACD blood with a pH of 5.9 were associated with reductions in 24- and 48-hour posttransfusion survivals and T50. The anticoagulant and the pH of the medium in which the RBCs were labeled with (51)Cr or biotin-X-NHS affected in vivo elution of the label from the RBCs and may reduce posttransfusion survival values.

The generation of hyperbaric oxygen tensions in fish.

Surprising inventiveness in the molecular interactions in fish hemoglobins that express the Root effect (decreased oxygen-carrying capacity at low pH) and in metabolic adaptations in swim bladder gas gland cells and retinal tissues causes local acidification of blood and generates hyperbaric oxygen tensions that drive oxygen into the swim bladder (regulating buoyancy) and ensures the oxygen supply to the avascularized retinae.

Determination of Physiological Levels of Blood Cyanide without Interference by Thiocyanate.

Acidification of blood, which is a general pretreatment procedure for cyanide analysis, caused artifactual cyanide formation from thiocyanate. A method for the determination of trace levels of blood cyanide has been developed using headspace gas chromatography with a nitrogen-phosphorus detector and a GS-Q column. By the addition of 30 mmol/l of ascorbic acid, this method eliminated thiocyanate interference. The detection limit was roughly 0.05μmol/l blood (S/N=3). Using this established method, blood cyanide levels were determined for transfusion blood samples from healthy donors, and also plasma thiocyanate levels were determined by the Konig colorimetric method. The artifactually formed cyanide levels, which were obtained by subtraction of the cyanide levels determined in the presence of ascorbic acid from those in its absence, were in proportion to the thiocyanate levels, indicating that plasma thiocyanate is oxidized to cyanide through oxyhemoglobin acidification.

Roles of Buffering Capacity and Pentose Phosphate Pathway Activity in the Gas Gland of the Gulf Toadfish Opsanus Beta

ABSTRACT The teleost gas gland is truly remarkable in its abilities to secrete gases into the swim bladder of physoclistous fish. The physiological and metabolic adaptations of this tissue have been elegantly summarized in a recent review article by Pelster and Scheid (1992). There are two key contributors to the function of the gland. First, a specialized metabolism of the swim bladder, involving copious and simultaneous production of lactate and CO2 from anaerobic glycolysis and the pentose phosphate pathway (also known as the hexose monophosphate shunt), respectively, contributes to gas exchange through pH and salting-out effects on the oxygen-carrying capacity of the blood. Second, a countercurrent multiplier system (i.e. a rete mirabile) enables gas tensions to be elevated further by back diffusion. Several features of metabolism and acid–base physiology remain unclear. First, despite the remarkable ability of this tissue to produce acid, it is not clear if or how intracellular pH (pHi) is regulated. Since ultimately the blood must be acidified, one would predict that the pHi of the tissue would be well regulated via high rates of membrane exchange of protons and/or high tissue buffering capacity. Second, although the functioning of the pentose phosphate pathway has been strongly inferred from measurements of enzyme activities (Boström et al. 1972; Pelster and Scheid, 1991), and from measurements of enhanced rates of CO2 excretion relative to the rates of oxygen uptake (Pelster et al. 1989), direct evidence for the existence of the shunt is lacking. Lastly, although the pentose phosphate pathway is expected to produce CO2, and thus contribute to the acidification of blood entering the gland, the pathway may have a different primary, or perhaps a dual, role, namely to maintain high tissue levels of NADPH for protection against oxygen radical damage to cells (Pelster and Scheid, 1992). The composition of the gas stored in the swim bladder can approach pure oxygen in some species, so it is not surprising that the teleost gas gland contains substantial levels of the enzymes catalase, superoxide dismutase and glutathione peroxidase, which scavenge deleterious radicals of oxygen and related harmful compounds (Morris and Albright, 1984). Noteworthy is glutathione peroxidase, which requires a constant supply of NADPH (presumably from the shunt) to maintain glutathione in a reduced state. Reduced glutathione is then used in a variety of oxygen radical detoxification mechanisms. If the pentose phosphate pathway has a role in oxygen detoxification, one would predict that flux rates through the pathway would increase with increased oxygen levels.

EC CO2R: Oxygenator or Hemodialyzer? An in Vitro Study

In respiratory support of patients with acute respiratory distress syndrome (ARDS), the extracorporeal CO2 removal (EC CO2R) technique should be the earliest and easiest procedure so as to have the lowest blood flow rate. Extracorporeal circulation (ECC) can be achieved using an oxygenator for CO2 removal under the dry form (dissolved CO2) or a hemodialyser for CO2 removal under the wet form (bicarbonates). This study investigated different methods allowing an increase in CO2 transfer, using liquid flow rates up to 0.330 l/min. The experimental set-up employed heated (38 degrees C) aqueous polyelectrolytic solutions mimicking the venous blood (pH 7.20, PCO2 53 mmHg). Four in vitro methods were tested: Series I: a DIDECO D702 oxygenator without blood (= liquid) acidification, Series II: D702 oxygenator with inlet HCl acidification, Series III: a HOSPAL H10-10 hemodialyzer without dialysate alkalinisation, Series IV: H10-10 hemodialyzer with NaOH dialysate alkalinisation. Maximum gas flow in the oxygenator and dialysate rate in hemodialyzer were 5 and 0.55 l/min respectively. For the four series the CO2 transfer (TCO2) (mean +/- S.E. ml/min) and pH out were: [table: see text] The difference between the four series was statistically significant (t-test). Acidification using the oxygenator increased CO2 transfer by 80%, but CO2 elimination was better with hemodialysis.

Countercurrent Concentration and Gas Secretion in the Fish Swim Bladder

Many fish possess a gas-filled swim bladder, and the gas molecules enter the bladder by diffusion from its capillaries and epithelium. The high gas partial pressures necessary to drive diffusion are produced in two steps. First, in the blood perfusing the swim bladder capillaries the physical solubility of gases is reduced via the salting-out effect, and gases are released from a chemical binding site via blood acidification (e.g., Root effect for O₂; conversion of HCO₃⁻ to CO₂). These effects result in an increase in the gas partial pressure in blood. In a second step, this initial increase in partial pressure (the "single concentrating effect") is multiplied by back diffusion of gas molecules in the countercurrent system of the rete mirabile. For inert gases, the salting-out effect, induced by the release of lactic acid from the gas gland cells in the swim bladder tissue, results in a probably small reduction of solubility, while the acid-induced decrease in hemoglobin O₂-carrying capacity (Root effect) and the release of CO₂ from HCO₃ ⁻, combined with the release of anaerobically produced CO₂ from gas gland cells, result in a significant single concentrating effect for O₂ and CO₂. The relatively high permeability of the rete capillaries for gases allows for back diffusion, and thus for countercurrent concentration, of gases. The rete mirabile is, however, permeable not only for gases but also for metabolites and salt molecules, and the countercurrent concentration of solutes enhances the countercurrent concentration of gas molecules. This is particularly important for inert gases because their single concentrating effect is small. Back diffusion of CO₂ in the rete mirabile not only results in a countercurrent concentration of CO₂ but also enhances the partial pressure increase of O₂ by decreasing the hemoglobin O₂-carrying capacity within the arterial capillaries of the rete. This interdependence of O₂ and CO₂ may apply to countercurrent systems in other tissues as well, for example, to the skeletal muscle. Hence, the swim bladder rete mirabile may serve as a model tissue for studies of gas exchange and countercurrent blood flow in other organs. Furthermore, the regulation of countercurrent eficiency and gas secretion promises to be an interesting area of research, including also the regulation of metabolic activity and acid production in the gas gland cells.

Impact of immediate acidification of blood on measurement of plasma tissue plasminogen activator (tPA) activity in surgical patients.

Impact of immediate acidification of blood on measurement of plasma tissue plasminogen activator (tPA) activity in surgical patients.

Hemoglobin oxygen affinity in patients suffering from arterial occlusive disease of the legs.

Parameters characterizing the hemoglobin oxygen affinity were determined in blood of 12 male patients suffering from arterial occlusive disease (AOD) of the legs and compared with data obtained earlier from healthy human subjects (controls). Due to a COHb content of 4.8% +/- 2.2% in the cigarette-smoking AOD patients, the standard oxygen dissociation curve (ODC) was left-shifted, the half-saturation pressure (P50) amounted to 24.8 +/- 1.7 mmHg (3.30 +/- 0.23 kPa), although the 2,3-diphosphoglycerate concentration was increased to 15.3 +/- 1.7 mumol/g Hb. Correcting the effects of elevated COHb shifts the P50 to 26.3 mmHg (3.5 kPa) and increases the steepness of the ODC (Hill's "n") from 2.79 +/- 0.27 to about 2.99, which is significantly different from controls. The Bohr coefficients after acidification of blood with lactic acid (BCLac) show high values at low oxygen saturations of hemoglobin (-0.50 +/- 0.04 in AOD patients, -0.32 +/- 0.04 in controls; P less than 0.05 at 10% SO2). The cause of the alterations in hemoglobin oxygen affinity may be a reduced mean erythrocyte age, but also the influence of unknown factors generated, e.g., from anaerobic muscle metabolism in AOD.

Interactions of tissue-type plasminogen activator with plasma inhibitors and their pharmacologic implications.

To delineate interactions of infused tissue-type plasminogen activator (t-PA) with inhibitors in plasma and their impact on fibrinolytic activity, serial plasma samples from patients with acute myocardial infarction and from normal rabbits given infusions of t-PA were assayed for t-PA antigen, activity of "fast acting" plasminogen activator inhibitor (PAI-1), and the presence and nature of t-PA-inhibitor complexes. In patients, endogenous t-PA circulated predominantly as a 100 kilodalton (kDa) complex with PAI-1, as verified by immunoprecipitation. During infusions, t-PA circulated not only as free t-PA (55 kDa) but also in complexes with PAI-1 (100 kDa), alpha 2-antiplasmin (110 kDa), and C1-esterase inhibitor (170 kDa). After termination of infusions, levels of free t-PA declined, while inhibitor complexes remained prominent. Free PAI-1 activity, assayed spectrophotometrically, was markedly elevated in the 24 hr interval after infusion of t-PA in 47% of patients with infarction. The specific activity of t-PA during infusions was 0.4 IU/ng or greater. However, during the 3 hr interval after infusions in patients, specific activity declined in association with prominence of t-PA complexes, predominantly with PAI-1. Infusions of t-PA in normal rabbits did not result in reactive increases in PAI-1 activity or in the t-PA-PAI-1 complex. After infusions, t-PA was associated predominantly with alpha 2-antiplasmin and C1-esterase inhibitor rather than PAI-1. t-PA inhibitor complexes were seen despite immediate acidification of whole blood, indicating that they were present in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)

Ionic equilibria in red blood cells of rainbow trout ( Salmo gairdneri): Cl −, HCO 3− and H +

The equilibrium distributions of H +, Cl − between plasma and erythrocytes of rainbow trout ( Salmo gairdneri) blood were determined as a function of plasma pH (pHe, 7.0–8.2), temperature (5–15°C), and mode of blood acidification (CO 2, HCl, lactic acid). The relationship between red cell pH (pHi) and pHE was not affected significantly by temperature or mode of blood acidification. H +, Cl − and HCO 3 − were passively distributed across the red cell membrane as evidenced by the membrane's high permeability to Cl − and the similar Donnan ratios for H + and Cl − at any given pHe. The relationship between pHe and the Donnan ratio for bound CO 2 had a similar slope to the relationships between pHe and the Donnan H + and Cl − ratios but was shifted upwards, presumably due to formation of carbamino compounds inside the rd blood cell. This carbamino formation was enhanced at lower temperatures when blood pH was elevated.