On-line production of ultrapure substitution fluid reduces TNF-alpha- and IL-6 release in patients on hemodiafiltration therapy.

On-line hemodiafiltration (HDF) has been introduced into clinical practice in the last few years. The most important technical and regulatory challenges were the safety and microbiological quality of ultrafiltrated substitution/replacement fluid. The application of ultrafilters in a different technical arrangement in the fluid path based on polysulfone or polyamide membranes should prevent patient contact with endotoxins and other pyrogenic or bacteria-derived substances. After resolving these problems and providing clinically safe and technically robust product solutions, increasing numbers of patients have been treated, especially those with severe clinical conditions, e.g., diabetes, hypo- or hypertension. The benefit for patients was brought about by the increase of substitution rate in hemodiafiltration and enhancing convective mass transfer. The impact of highly convective therapy modes on the state of immunomodulation towards the syndrome of microinflammation has not been investigated in a systematic prospective manner. In this study, 8 patients undergoing bag-HDF treatment with lactate buffered solution were investigated before on-line HDF treatment with commercially available whole blood stimulation assays testing for TNF-alpha and IL-6 release. Both assays are based on phytohemagglutinine (for TNF) and lipo-polysaccharide stimulation (for IL-6). Thereafter the patients were switched to on-line production of substitution fluid. After a wash-out period of 2 sessions the whole blood stimulation assays were applied to the same patients. The Wilcoxon test (for paired analysis) was done, revealing a statistically significant lower release of proinflammtory cytokines from patients' blood upon stimulation with PHA or LPS. The reduction of IL-6 and TNF concentration and release capacity in whole blood may be attributed to the use of high quality ultrapure substitution fluid and dialysate in on-line treatment instead of lactate buffer bag solution. These results indicate that not only an increase of convective mass transfer by higher volume exchange, but also a decrease in unspecific activation of immunocompetent cells may have advantages for HDF-treated patients.

The measurement of cytokine production capacity during dialysis--a new dynamic method for the evaluation of biocompatibility?

The activation of monocytes and other immunocompetent cells during hemodialysis can be attributed to their contact with immunogenic structures such as membranes, blood lines, and endotoxins. The simple measurement of cytokines in blood cannot completely describe the whole dimension of this event. Stimulation of monocytes and other immunocompetent cells in whole blood with lipopolysaccharides (LPS) for IL-6 and phytohemagglutinine (PHA) for TNF at the start and end of dialysis may make it possible to better analyze cellular response during dialysis. Ten healthy volunteers and 10 patients suffering from chronic renal failure were tested with the commercial whole-blood stimulation assays "Dynamix"-IL-6-DIA and -TNF-alpha-DIA (Biosource Diagnostics, Ratingen, Germany). Then 24 patients undergoing hemodialysis with hemophane (n=12) and polyamide (n=12) membranes were examined before and after dialysis treatment. The unpaired Wilcoxon t- test was used for statistical analysis. Healthy volunteers and patients with chronic renal failure showed no statistical differences in concentrations of TNF-alpha and IL-6 before or after whole blood stimulation (WBS). In comparison to patients with chronic renal failure, pre-WBS concentrations of both cytokines (p<0.034) were increased in patients of each membrane group before dialysis. After whole blood stimulation, no differences were observed. At the end of dialysis treatment, the pre- and post-WBS IL-6 values were both significantly higher in the hemophane group (p=0.049 and p=0.0038, respectively) TNF-alpha concentrations were unchanged. No significant differences in the polyamide group were found between the start and end of treatment for either cytokine. A comparison of these membrane groups showed that only the pre-WBS IL-6 concentration in the hemophane group was elevated (p=0.022) after dialysis. In conclusion, the presence of uremia alone could not influence the cytokine production and release capacity. In our patients, dialysis elevated pre-WBS concentrations of TNF-alpha and IL-6, and increased IL-6 release from immunocompetent cells after whole blood stimulation in the hemophane group. The use of polyamide membranes decreased the action of monocytes and other immunocompetent cells, but could not completely prevent this phenomenon. The whole blood stimulation assays for measurement of TNF-alpha and IL-6 may represent a new, dynamic method for evaluating biocompatibility.

K(+)-permeability in diabetics and nondiabetics with and without renal insufficiency.

Looking for causes or consequences of primary hypertension much attention is drawn to the ion transport systems of the cellular membrane. The existence of endogeneous digitalis-like factors, that lower the activity of Na+/K(+)-ATPase and result in a complex change of electrolyte balance of cells are discussed as a reaction of the organism to salt and volume retention. The measurement of passive permeability of erythrocyte membranes for potassium is an easy and useful method for the detection of disturbances of Na+/K(+)-transport, especially for extensive screening investigations. We examined the potassium permeability of erythrocytes in healthy individuals (GR1, n = 48), patients with compensated renal insufficiency (GR2, n = 36) and diabetics (GR3, n = 25) as well as a group of diabetics with renal failure (GR4, n = 47). The relative change of potassium concentration in the whole blood, based on the efflux of potassium during a 4-hour-incubation at 37 degrees C, is defined as a measure for K(+)-permeability. K(+)-concentrations are determined every 60 minutes with ion sensitive electrodes. K(+)-permeability was significantly increased in patients with compensated renal insufficiency compared to the control group and to diabetics. Diabetics differed markedly in their erythrocyte reaction regarding K(+)-permeability. Whereas patients with renal insufficiency show an efflux of potassium during investigation there is a decrease of potassium in plasma in diabetics. The K(+)-permeability results of patients with both diseases are intermediate between the GR2- and GR3 results and are significantly different from the control group. When g-strophanthin is added to inhibit the sodium pump, the differences between the groups are abolished. The decreased K+permeability in diabetics compared to the control group could be explained by the increased supply of energy-rich substrates for the Na+/K(+)-ATPase.

Salt sensitivity and insulin resistance in normotensives.

Among hypertensive patients salt sensitivity and insulin resistance are commonly observed together. We investigated if a causal relationship already exists in young normotensive adults. With a standardized dietary regimen we determined salt sensitivity in 35 male volunteers by measuring diastolic blood pressure (24-hours-RR-recording). Insulin resistance was tested using hyperinsulinaemic-euglycaemic-clamp-technique by de Fronzo after a freely chosen diet, after 7 days of salt loading (260 mmol/d) and after 7 days of salt restriction (60 mmol/d). Data from euglycaemic-clamp technique were available from 27 subjects. 18 of them (67%) could be characterized as salt resistant; 9 persons (33%) were salt sensitive. Glucose infusion rate, mean glucose and insulin concentrations were measured in plasma, metabolic clearance rate (MCR) and indices of insulin sensitivity (ISI) were calculated. The results of MCR and ISI show large interindividual variances. There were no differences between the salt sensitive and salt resistant group regarding the mean insulin concentrations and also the mean glucose uptake in the steady state clamp period and also the calculated MCR and ISI. Comparing the periods of different salt intake, there were no differences between salt loading and salt restriction. Whereas salt sensitivity can already be shown in the normotensive state, with this experimental design a changed insulin sensitivity is not detectable. This supports the idea, that insulin resistance is not causally linked with salt sensitivity. It may be a secondary phenomenon of salt induced hypertension.

Prevalence of increased intracellular signal transduction in immortalized lymphoblasts from patients with essential hypertension and normotensive subjects.

OBJECTIVE: To determine the prevalence of enhanced signal transduction in immortalized B lymphoblasts from normotensive subjects and patients with essential hypertension. METHODS: We established Epstein-Barr virus-immortalized lymphoblast cell lines from 26 normotensive and 37 hypertensive subjects. Subsequently, we quantified rises in the cytosolic free Ca2+ concentration, [Ca2+]i, evoked by 0.1 micromol/l platelet-activating factor (PAF) in Fura-2-loaded cells. RESULTS: PAF-induced [Ca2+]i rises were independent of donor age in cells from normotensive and hypertensive subjects. Baseline values of [Ca2+]i were not significantly different in the two groups. Using the mean + 2SD of the PAF-evoked rises in [Ca2+]i above basal (110 nmol/l) as the upper normal value, we estimate that enhanced [Ca2+]i rises are distinctly more prevalent in hypertensive subjects (27%) than they are in normotensive subjects (4%). Similarly, upon definition of normal values by the 99% confidence interval (75 nmol/l), 19% of cells from normotensive versus 43% from hypertensive subjects display enhanced intracellular signaling. CONCLUSION: Enhanced intracellular signal transduction could be the primary defect in approximately one-third of the overall population with essential hypertension.

Effect of arterial chemoreceptor stimulation with almitrine bismesylate on plasma renin activity, aldosterone, ACTH and cortisol in anaesthetized, artificially ventilated cats.

1. Changes in plasma renin activity (PRA) and in the plasma concentration of aldosterone, adrenocorticotrophic hormone (ACTH) and cortisol in response to an intravenous infusion of the chemoreceptor stimulant almitrine bismesylate (0.2 mg/kg) were studied in two groups of anaesthetized, paralysed and constantly ventilated cats. In one group, the peripheral arterial chemoreceptors remained innervated, whereas in the other they were denervated by bilateral cervical vagotomy and section of the carotid sinus nerves. 2. Animals with innervated chemoreceptors (n = 16) reacted to almitrine bismesylate with a significant (P <0.05) increase in both ACTH and cortisol. These responses were not present in cats in which the peripheral arterial chemoreceptors had been surgically denervated (n = 16). 3. Plasma renin activity and plasma aldosterone increased with time during experiments on both the chemoreceptor-intact and chemoreceptor-denervated cats. Almitrine did not affect the time course of the rise in PRA and plasma aldosterone in either group of animals. 4. These data indicate that, under the conditions of our experiments, almitrine induced arterial chemoreceptor reflex mechanisms stimulate ACTH and cortisol release, but has no chemoreceptor-dependent influence on PRA or plasma aldosterone.

Sodium kinetics in salt-sensitive and salt-resistant normotensive and hypertensive subjects.

OBJECTIVE: To test the hypotheses that sodium kinetics are not affected by blood pressure, salt sensitivity, salt resistance or race, and that the kinetics of sodium balance are not a first-order process. DESIGN, PARTICIPANTS AND INTERVENTIONS: Two studies were conducted. In the first, 18 normotensive and 36 hypertensive men and women were given sodium at 120 mmol/day for 6 days, followed by 10 mmol/day for 8 days, then 400 mmol/day for 8 more days. Salt sensitivity was defined as an increase in diastolic blood pressure from the 10 to the 400 mmol/day intake. Salt resistance was defined as no increase, or a decrease in diastolic blood pressure with the increased sodium intake. In the second study, 12 white and 12 black normotensive men ingested sodium at 10, 200 or 400 mmol/day in random order, each for 7 days. All urine was collected in both protocols. SETTING: Metabolic ward at the University of Greifswald (Greifswald, Germany; study 1), and Clinical Research Center (Indiana University, Indianapolis, Indiana, USA; study 2). MAIN OUTCOME MEASURE: In addition to conventional statistics, a pharmacokinetic analysis was carried out to determine the elimination rate constant and half-life. RESULTS: In the Greifswald study, when the sodium intake was decreased, a longer half-life was determined for the salt-sensitive than the salt-resistant hypertensive subjects. The half-life for the normotensive salt-sensitive and salt-resistant subjects did not differ. When the sodium intake was decreased, a monoexponential equation fitted the data for all subjects; when the sodium intake was increased, only data for half the subjects could be fitted to the same equation. In the Indianapolis study, black race had a significant influence upon urinary sodium excretion. Furthermore, the half-life for sodium elimination was dependent upon sodium intake; namely, the greater the intake, the longer the elimination half-life. CONCLUSIONS: The time required to reach sodium balance may increase following salt-sensitive increases in blood pressure rather than precede them. Race influences the time required to achieve salt balance. Sodium kinetics are not a first-order process.

Definitions and characteristics of salt-sensitivity and resistance of blood pressure: should the diagnosis depend on diastolic blood pressure?

To elucidate the importance of diastolic blood pressure in the definition of salt-sensitive hypertension, we studied 54 male subjects, 36 of whom had untreated, mild essential hypertension. The subjects received a 120 mmol/d Na (as the chloride salt) diet for six days. Thereafter they received a 10 mmol/d Na diet for eight days followed by a 400 mmol/d Na diet for another 8 days. Blood pressure was measured hourly "around the clock" on the last day of each diet; the averaged systolic, diastolic and mean blood pressure values were compared. In 22 subjects diastolic blood pressure increased, when salt intake was increased from 10 to 400 mmol/d. In 18 of these 22 subjects systolic blood pressure increased as well. In 20 subjects, systolic blood pressure increased with salt loading while diastolic blood pressure decreased. In 13 subjects both systolic and diastolic blood pressure decreased with increased salt intake. We defined those subjects showing an increase in diastolic blood pressure as salt-sensitive. If mean blood pressure were used to define salt-sensitivity, 8 of our subjects would have been labeled as salt-sensitive who actually decreased their diastolic blood pressure with salt loading. We suggest that consideration of systolic and diastolic blood pressure responses gives better insight into identifying volume and resistance-related phenomena in salt-sensitive hypertension, than does the consideration of mean blood pressure alone. The definition of salt-sensitivity may require reassessment.

[Opioid peptides in hemorrhagic shock]. / Opioidpeptide im hämorrhagischen Schock.

Regarding the treatment of vitally endangered intensive care patients the neuroendocrine regulation of the post-aggression metabolism is important. The role of opioid peptides in this system is investigated in the animal experiment "Haemorrhagic Shock in the Dog". It was shown that the opioid peptide beta-endorphin and metenkephalin rise grossly in connection with pathological and endocrinological alterations in shock. Based on the literature the role of increased concentration of opioids in haemorrhagic shock is discussed and conclusions for therapeutic measures are presented.

Hormonal and renal responses to arterial chemoreceptor stimulation by almitrine in healthy and normotensive men.

Healthy and normotensive men (n = 11) were hospitalized and kept under controlled fluid and sodium intake (120 mequ/d) for 5 days. Their systemic arterial blood pressures as well as heart and breathing rates were measured, and venous blood and urine samples were collected at intervals of 1-4 h. Diuresis was induced by scheduled drinking of tea (150 ml/h). Electrolytes, osmolality, and creatinine were determined in both plasma and urine samples. Aldosterone, cortisol, and vasopressin concentrations were measured only in the plasma. On the 2nd and 3rd day of the experiments the participants received orally either a placebo-pill or 100 mg almitrine bismesylate (Vectarion). Each subject was tested in a placebo- and an almitrine experiment. The subjects responded to the almitrine treatment with a suppression of the plasma aldosterone content, a transient rise of glomerular filtration rate, a natriuresis and an increase of renal concentrating ability. In the placebo-experiments, only the transient rise of filtration rate was significant. The data indicate that almitrine, by stimulating the peripheral arterial chemoreceptors, suppresses plasma aldosterone and inhibits renal proximal sodium reabsorption by so far unknown mechanisms. They also suggest that oral and intravenous almitrine administrations, respectively, might differently affect renal hemodynamics and excretory function.

Suppression of the plasma aldosterone to renin activity ratio in anaesthetized cats after pharmacological stimulation of the peripheral arterial chemoreceptors with almitrine bismesylate.

In chloralosed, non-vagotomized, spontaneously breathing cats the peripheral arterial chemoreceptors were stimulated by intravenous infusion of almitrine bismesylate (Vectarion, 0.20 mg/kg). Within 5 h after administration of the drug, a decline of both the mean systemic arterial blood pressure and the effective renal plasma flow, as well as an increase of the plasma renin activity (PRA) and the plasma aldosterone concentration (PAC) was observed. But as the PAC increase was less than that of PRA, a highly significant suppression of the PAC to PRA ratio was noted. The results indicate that not only whole body altitude hypoxia, but also stimulation of the peripheral arterial chemoreceptors in normoxic animals lowers the PAC-to-PRA ratio. It remains to be verified experimentally whether there exists a specific reflex influence of the peripheral arterial chemoreceptors on the renin-aldosterone relationship.