Blood Of Uremic Patients Research Articles

Citrate dialysate does not induce oxidative stress or inflammation in vitro as compared to acetate dialysate

Increased acetataemia during haemodialysis sessions has been associated with a number of abnormalities, including increased oxidative stress, pro-inflammatory cytokines and nitric oxide synthesis. Citric acid may play an alternative role to acetate as a dialysate stabilizer given that the effect of citrate on complement and leukocyte activation is different to that of acetate. The purpose of this study was to compare the inflammatory effect in immunocompetent blood cells of acetate dialysate and citrate dialysate. Materials and methodsThe effect of acetate and/or citrate was investigated in the whole blood of uremic patients and in healthy in vitro samples. Four types of dialysate were tested: dialysate 1, acetate-free with 1mmol/L of citrate; dialysate 2, with 0.8mmol/L of citrate and 0.3mmol/L of acetate; dialysate 3, citrate-free with 3mmol/L of acetate; and dialysate 4, citrate-free with 4mmol/L of acetate. The cell types used were: human monocyte culture (THP-1); and peripheral blood mononuclear cells (PBMCs) from healthy subjects and uremic patients on haemodialysis. ICAM-1 was determined and levels of reactive oxygen species and total microvesicles were quantified. ResultsUnlike the citrate dialysates, the dialysates with acetate (dialysate 3 and dialysate 4) induced increased ICAM-1 expression density in THP-1 cells; an increase in ICAM-1 expression was observed in the immunocompetent cells of healthy subjects with acetate dialysate (dialysate 3 and dialysate 4) but not with citrate dialysate (dialysate 1 and dialysate 2). No significant ICAM-1 differences were found between the different dialysates in the cells of haemodialysed patients. Reactive oxygen species expression and the number of microvesicles increased significantly with acetate dialysate but not with citrate dialysate in the cells of both healthy subjects and haemodialysed patients. ConclusionAt the concentrations in which it is generally used in clinical practice, acetate-based dialysate increases oxidative stress and the total number of microvesicles and may induce another pro-inflammatory stimuli in uremic patients on haemodialysis. Citrate dialysates do not induce this activation, which could make them a suitable alternative in clinical practice.

Influence of nanoporous poly(ether imide) particle extracts on human aortic endothelial cells (HAECs).

Accumulated uremic toxins like indoxyl sulphate, hippuric acid and p-cresyl sulphates in renal failure patients stimulate proinflammatory effects, and consequently kidney and cardiovascular diseases. Low clearance rate of these uremic toxins from the blood of uremic patients by conventional techniques like hemodialysis is due to their strong covalent albumin binding (greater than 95%) and hydrophobic nature, which led to alternatives like usage of hydrophobic adsorber's in removing these toxins from the plasma of kidney patients. Polymers like polyethylene, polyurethane, polymethylmethacrylate, cellophane and polytetrafluoroethylene were already in use as substitutes for metal devices as dialysis membranes. Among new synthetic polymers, one such ideal adsorber material are highly porous microparticles of poly(ether imide) (PEI) with diameters in the range from 50-180μm and a porosity around 88±2% prepared by a spraying and coagulation process.It is essential to make sure that these synthetic polymers should not evoke any inflammatory or apoptotic response during dialysis. Therefore in our study we evaluated in vitro effect of PEI microparticle extracts in human aortic endothelial cells (HEACs) concerning toxicity, inflammation and apoptosis. No cell toxicity was observed when HAECs were treated with PEI extracts and inflammatory/apoptotic markers were not upregulated in presence of PEI extracts. Our results ensure biocompatibility of PEI particles and further hemocompatibility of particles will be tested.

Divergent behavior of hydrogen sulfide pools and of the sulfur metabolite lanthionine, a novel uremic toxin, in dialysis patients

Dialysis patients display a high cardiovascular mortality, the causes of which are still not completely explained, but are related to uremic toxicity. Among uremic toxins, homocysteine and cysteine are both substrates of cystathionine β-synthase and cystathionine γ-lyase in hydrogen sulfide biosynthesis, leading to the formation of two sulfur metabolites, lanthionine and homolanthionine, considered stable indirect biomarkers of its production. Hydrogen sulfide is involved in the modulation of multiple pathophysiological responses. In uremia, we have demonstrated low plasma total hydrogen sulfide levels, due to reduced cystathionine γ-lyase expression. Plasma hydrogen sulfide levels were measured in hemodialysis patients and healthy controls with three different techniques in comparison, allowing to discern the different pools of this gas. The protein-bound (the one thought to be the most active) and acid-labile forms are significantly decreased, while homolanthionine, but especially lanthionine, accumulate in the blood of uremic patients. The hemodialysis regimen plays a role in determining sulfur compounds levels, and lanthionine is partially removed by a single dialysis session. Lanthionine inhibits hydrogen sulfide production in cell cultures under conditions comparable to invivo ones. We therefore propose that lanthionine is a novel uremic toxin. The possible role of high lanthionine as a contributor to the genesis of hyperhomocysteinemia in uremia is discussed.

Role of Parathyroid Hormone in the Downregulation of Liver Cytochrome P450 in Chronic Renal Failure

Chronic renal failure (CRF) is associated with a decrease in drug metabolism secondary to a decrease in liver cytochrome P450 (P450). The predominant theory to explain this decrease is the presence of factors in the blood of uremic patients. This study tested the hypothesis that parathyroid hormone (PTH) could be this factor. The objectives of this study were to determine (1) the role of PTH in the downregulation of hepatocyte P450 induced by rat uremic serum, (2) the role of PTH in the downregulation of liver P450 in rats with CRF, and (3) the effects of PTH on P450 in hepatocytes. For this purpose, (1) hepatocytes were incubated with serum from rat with CRF that was depleted with anti-PTH antibodies or with serum from parathyroidectomized (CRF-PTX) rat with CRF, (2) the effect of PTX on liver P450 was evaluated in rats with CRF, and (3) the effects of PTH on P450 in hepatocytes were determined. The depletion of PTH from CRF serum completely reversed the downregulating effect of CRF serum on P450 in hepatocytes. Addition of PTH (10(-9) M) to depleted CRF serum induced a decrease in P450 similar to nondepleted CRF serum. The serum of CRF-PTX rats had no effect on P450 in hepatocytes compared with CRF serum. Adding PTH to CRF-PTX serum induced a similar decrease in P450 as obtained with CRF serum. Finally, PTX prevented the decrease of liver P450 in rats with CRF. In summary, PTH is the major mediator implicated in the downregulation of liver P450 in rats with CRF.

Generation of superoxide anions during the reaction of guanidino compounds with methylglyoxal.

Uremic toxins are accumulated in the blood of patients with chronic renal failure (CRF), although alteration of the toxicity by the interaction of various uremic retention products has not been precisely clarified. In this study, we found that cytochrome c added to incubation mixtures containing guanidino compounds and methylglyoxal in phosphate buffer solution (pH 7.4) resulted in reduction of cytochrome c. Superoxide anions were generated from incubation mixtures of each guanidino compound with methylglyoxal, because the reduction was inhibited by the addition of superoxide dismutase. The incubation mixture containing each guanidino compound and methylglyoxal had different rates of generation of the superoxide anion from other mixtures. A relatively higher superoxide anion formation rate was observed in the incubation mixture containing Arg and methylglyoxal (7.9 +/- 0.5nmol x m(-1) x min(-1)), or in the incubation mixture containing methylguanidine and methylglyoxal (6.3 +/- 0.6 nmol x ml(-1) min(-1)). These findings suggest that interactions of various uremic retention products which accumulate in the blood of uremic patients may generate reactive oxygen species and may be involved in the oxidative stress observed in CRF patients. The addition of aminoguanidine, which is known to inhibit the formation of advanced glycation end products, to a mixture of guanidino compounds and methylglyoxal inhibited reactions between guanidino compounds and methylglyoxal.

Increase in Resting Levels of Superoxide Anion in the Whole Blood of Uremic Patients on Chronic Hemodialysis

Recently, we developed a new method to measure the resting level of superoxide anion in whole blood using an ultrasensitive chemiluminescence analyzer and lucigenin amplification. The advantage of this method is that the assay system can be performed in the absence of leukocyte isolation and stimulant administration. In this study, we applied this method to measure the blood resting levels of superoxide anion in 104 uremic patients on chronic hemodialysis (CHD) and 98 sex- and age-matched healthy controls to clarify the influence of HD on blood levels of superoxide anion. Simultaneously, the plasma levels of copper, zinc superoxide dismutase (Cu,Zn-SOD), glutathion peroxidase (GPX), myeloperoxidase (MPO) and lactoferrin (Lacto-F) were measured. The results showed that the basal blood levels of superoxide anion, Cu,Zn-SOD, and MPO in CHD patients were significantly greater than those of healthy controls. However, there was no difference in the basal plasma levels of Lacto-F and GPX between CHD patients and healthy controls. One session of HD further increased the blood levels of superoxide anion, MPO, Lacto-F and Cu,Zn-SOD but not GPX. These results suggest that the blood levels of superoxide anion are higher in CHD patients and further increase after one session of HD. This mechanism should be studied further.

Highly effective separation and highly sensitive detection for clinical chemistry and biochemistry

Highly effective separation and highly sensitive detection reagents for clinical chemistry and biochemistry were developed and their applications were investigated. The sensitive detection of carboxylic acids was accomplished using 9-anthryldiazomethane (ADAM) which gave intensely fluorescent derivatives from carboxylic acids without catalysts or heating. 1-Pyrenyldiazomethane was then synthesized and proved to react also readily with carboxylic acid and more sensitive than ADAM. The optical resolution of amino acid enantiomers was achieved using 2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyl isothiocyanate (GITC). GITC derivatized enantiomeric amino acids under mild conditions to give highly hydrophobic diastereomers which could be resolved on conventional reversed phase columns and easily detected by the absorption based on the thiourea structure. Then we devised an o-phthalaldehyde-N-acetylcystein reagent (OPA/NAcCys) giving diastereomers which were also resolved on a reversed phase column and detected fluorometrically with excellent sensitivity. OPA/NAcCys was useful for the assay of D-amino acids in the blood of uremic patients. The hypochlorite-thiamine method for the assay of proteins and peptides was established providing sensitive fluorometry, which well reflected the number of peptide groups in the molecule. This principle was applied to the assay using N-chlorodansylamide, designed for the fluorometry of peptides. The alkaline ninhydrin method was applied to the detection of guanidino compounds in the blood of uremic patients. Several fluorometric methods for the simultaneous detection of reducing and non-reducing carbohydrates, and guanidines were found to be useful reagents for this purpose because these compounds were resistant to the oxidation with periodate. Then protamine-bound columns were prepared for the separation of carbohydrates on HPLC, which showed excellent recovery of reducing carbohydrates in comparison with conventional alkylamino columns.

腎糸球体ろ過 血液透析からみた人工透析膜のあり方

Recent advance of materials and design for dialyser membrane made possible to achieve high dialysis efficacy and high flux. The purpose of dialysis membrane now and in the future is to refine blood biocompatibility and to get the character to remove the special substances from the blood of uremic patients. Because of the difficulties to achieve the complete biocompatibility by artificial polymers now used, it is important for us to change the idea for invention of new dialyser material of the next generation.

Inadequate ability of T-lymphocytes from chronic uremic subjects to stimulate the in vitro growth of committed erythroid progenitors (BFU-E).

The growth of normal burst-forming units (BFU-E) is known to depend on a burst-promoting activity (BPA) produced by T-lymphocytes. Few BFU-E colonies have been observed in cultures of blood mononuclear cells (MNC) of uremic patients. The aim of the present study was to examine the concentration of BFU-E in the blood of uremic patients and to evaluate the ability of uremic T-lymphocytes to produce BPA. We have studied 6 chronic uremic patients treated with maintenance hemodialysis. When 5 X 10(5) blood MNC depleted of T-lymphocytes of uremic subjects were stimulated by 1 X 10(6) normal T-lymphocytes in a methylcellulose culture system we observed the growth of a number of BFU-E colonies that did not differ significantly from normal (29 +/- 11.8 colonies). On the contrary, when 5 X 10(5) blood MNC depleted of T-lymphocytes of normal subjects were stimulated by 1 X 10(6) T-lymphocytes of uremic patients, the number of BFU-E colonies obtained was significantly lower than normal (1.9 +/- 3.1 colonies). These data show that the decreased number of BFU-E colonies obtained from blood MNC of uremic patients is due to a defect of uremic T-lymphocytes. The impairment of T-lymphocytes can be due to inhibitors of T-lymphocyte function or to variations in T cell subsets, leading to a decrease in the OKT4/OKT8 cell ratio. In any case it is a significant pathogenetic mechanism contributing to anemia in chronic uremia.

Effects of hemodialysis on guanidinopropionic acid metabolism.

Blood levels of guanidinopropionic acid (GPA), a putative uremic toxin, have been evaluated in 5 uremic patients before a dialytic session, at the end of it and during the following 68 h. GPA levels are markedly higher in uremic patients than in controls and are significantly reduced at the end of dialysis even if still higher than in controls. The clearance of GPA is similar to those of urea and creatinine, even if at the end of the dialysis session the percent decrease of GPA is significantly lower than that of urea. During the first 8 h after the end of dialysis GPA levels increase steeply; subsequently, the rate of accumulation of GPA in blood declines markedly remaining constant until the 68th hour. In conclusion GPA is markedly increased in blood of uremic patients and is significantly removed by dialysis. The evaluation of GPA increase per hour after the end of dialysis may provide an estimation of GPA production in uremic patients.

Thiol S-methylation in uremia: erythrocyte enzyme activities and plasma inhibitors.

Thiopurine methyltransferase (TPMT) catalyzes thiopurine S-methylation, an important metabolic pathway for drugs such as 6-mercaptopurine. Thiol methyltranferase (TMT) catalyzes the S-methylation of a variety of aliphatic sulfhydryl compounds. Erythrocyte (RBC) TPMT activity is elevated in the blood of uremic patients on maintenance hemodialysis, 15.83 +/- 0.90 U/ml RBCs (mean +/- SEM, n = 41), whereas in blood from randomly seleted nonuremic subjects it was 12.76 +/- 0.16 U/ml (n = 298, p < 0.001). RBC TPMT activity is not affected by hemodialysis. The plasma of uremic patients reversibly inhibits RBC TPMT activity to a greater extent than normal plasma does and contains higher concentrations of endogenous methyl acceptors than normal plasma. Plasma TPMT inhibitors are not removed by hemodialysis. There are large individual variations in inhibition of RBC TPMT by plasma from patients with renal failure. Inhibition varied from 1% to 93% in 20 microliters of plasma from each of 20 randomly selected uremic patients. There was a positive correlation between the inhibition of TPMT and the content of endogenous methyl acceptors in uremic plasma (r = 0.914, n = 20, p < 0.001), but there was no significant correlation between degree of inhibition and urea nitrogen, serum creatinine, or hematocrit. The ability of plasma from individual uremic patients to inhibit TPMT also correlated with its ability to inhibit two other drug metabolizing methyltransferases in the RBC, catechol-O-methyltransferase and phenol-O-methyltransferase, RBC TMT activity is not altered in patients with uremia. The results of these and other studies of methyl conjugation in renal failure focus attention on the accumulation of methyl acceptor substrates in some of these patients and on the possible effects of these methyl acceptors on a variety of methylation reactions.

“In vitro” studies on the fate of diphenylhydantoin in uremia

“In vitro” studies were undertaken to evaluate the fate of DPH in blood of uremic patients. DPH incubated for 18 h at 37°C disappears from plasma in the presence of erythrocytes and more when the blood underwent hemolysis. This phenomenon was more pronounced in uremic blood. These findings suggest not only that DPH is absorbed onto the erythrocyte membrane, but also that it is bound by erythrocyte contents. It could be possible that, because of a low binding capacity of the albumin carrier in uremic blood, the drug penetrates more easily the erythrocyte membrane and is bound there. An intraery throcyte enzymatic metabolism of DPH is not excluded.

Erythrocyte Transketolase Inhibition, Neuropathy and Uremia

The first clinically successful treatment of renal failure with the artificial kidney by W. J. Kolff in 1943 must be regarded as an empirical triumph. Although several normal solutes had been identified in abnormally high concentration in the blood of uremic patients, there was no information about whether reducing their concentrations would restore symptomatic normality. Cellophane was selected as the dialysis membrane with only a general awareness of its permeability characteristics. The dramatic reversal of symptomatic uremia by dialysis, coupled with the difficulty in assaying the pathophysiologic importance of solutes identified in uremic blood, has slowed work on the identification . . .

Oral cephalexin and ampicillin: antimicrobial activity, recovery in urine, and persistence in blood of uremic patients.

A new orally absorbed cephalosporin derivative (cephalexin) and potassium ampicillin were studied in subjects with normal and decreased renal function. A 1-week course of each drug gave adequate therapeutic blood and urinary concentrations, without evidence of cumulation or toxicity in normal subjects. Almost all of the cephalexin and about one half of the ampicillin were recovered in the urine. The same treatment in uremic patients was associated with elevated concentrations of both drugs in the serum with only modest accumulation over 1 week. Concentrations in urine progressively rose and were adequate for most urinary tract organisms against which these agents are active. One of five patients had a fall in creatinine clearance 2 weeks after the last dose of cephalexin, but renal function later improved, and the effect cannot be proved drug-related. Both cephalexin and ampicillin may be useful in treatment of urinary infections in moderately uremic patients whose creatinine clearance exceeds 9 to 10 ml/min.

Nephrotoxicity of Antibiotics

The potentially nephrotoxic antibiotics in current clinical use are neomycin, kanamycin, paromomycin, bacitracin, the polymyxins (polymyxin B, and colistin), and amphotericin B. 1-3 Nephrotoxicity was reported with early lots of streptomycin, but the drug now commercially available does not appear to have this property. Fortunately, the nephrotoxicity of each of these drugs appears to be reversible provided their administration is stopped soon enough. Patients with underlying renal disease appear to be particularly sensitive to most of these agents. This sensitivity may be attributable in part to the tendency of these drugs to accumulate rapidly in the blood of uremic patients given ordinary therapeutic doses and in part to the diminished renal functional reserve of uremic patients. Tetracycline may now be added to the group of potentially nephrotoxic drugs, because a reversible Fanconi-like syndrome produced by preparations containing degraded tetracycline has been observed. Tetracyclines.— A primary renal toxic effect of undegraded

Significance of circulating phenols in anemia of renal disease.

SummaryFree and conjugated ether-soluble phenols in blood of uremic patients have been determined by a specific and reliable method. Free phenol and cresol are not elevated in the great majority of these patients. Reasons are given for discounting the role of elevated conjugated phenols and free phenolic acids in the pathogenesis of the anemia of chronic renal disease.

The chlorides of the plasma in uremia

Previous investigations relating to the chlorid content of the blood or plasma in uremia have yielded conflicting results, some figures much lower than the lowest normal limit having been reported.2 It is known that the chlorid content of nephritic plasma is usually somewhat higher than that of the average normal plasma, but the findings in uremia have apparently so far not been explained.We have been able, in several cases, to make frequent observations of the chlorid content of the plasma of nephritic individuals during life, and up to the time of death in uremic coma. We have found a diminution of chlorids in the plasma to be the usual accompaniment of uremia, and we have found this decrease of chlorids in the plasma to accompany the increased H + ion concentration frequently observed in the blood of uremic patients shortly before death.That increased acidity of the blood causes a diminution of the plasma chlorids, and an increase in the chlorid content of the cells had been shown experimentally both i...

The isolation of a toxic substance from the blood of uremic patients

The analyses of bloods from cases of uremia have yielded a substance which is toxic. Control analyses of bloods from a wide variety of conditions not associated with uremia failed to discover a similar substance. Guinea-pigs were used as the test animal and enough material can be recovered from 200 c.c. of uremic blood to cause death. The isolation of the substance was effected by a combination of several methods in current use for the separation of animal bases.

On indican in the blood of uremic patients

A demonstration of the constant presence in uremic blood of an abnormal aromatic body such as indican would be highly significant since this would be suggestive of the nature of the abnormal metabolism. Hence a recent report1 to the effect that indican is present in the blood in uremia invited confirmation, and especially as the amounts present are stated to be sufficient to give the ordinary blue color with chloroform when only 10 c.c. of serum are used for the test. Considering the total volume of blood this would mean an enormous concentration.The method for detecting indican employed by Obermayer, was, in brief, to separate all of the proteins from the serum by means of alcohol, which after filtration is evaporated on the steam bath. The residue from the alcohol filtrate is taken up in water, freed of salts with lead acetate and the latter in turn removed by sodium phosphate. A water clear filtrate is the final result which is tested by the usual method employed for urine with Obermayer's reagent.In r...

AN ANALYTIC STUDY OF UREMIA.WITH SOME GENERAL CONCLUSIONS IN REGARD TO ITS CAUSES AND TREATMENT.

CRITIQUE OF CURRENT UREMIA THEORIES. If the current belief that uremia is due to renal inadequacy and the retention in the blood and tissues of excrementitious urinary bodies were correct, then (1) complete anuria should always produce uremia, and (2) the blood of uremic patients should always show an increase and the urine a corresponding decrease of urinary substances. Anuria Without Uremia. —Many cases of complete anuria, due to various causes, are, however, recorded, in some instances persisting for several weeks, in which none of the characteristic phenomena of uremia developed. Most of these patients for days complained of nothing more than a feeling of extreme lassitude, death resulting suddenly from heart failure without preceding coma or amaurosis, and without eclamptic spasms or typic changes of the blood pressure. On the other hand, uremia not infrequently occurs when the flow of urine is abundant and the excretion of urinary solids