Search

Commercially available ready to use syringes (Radiometer, Sarstedt, Ciba-Corning and Portex), containing heparinate as an anticoagulant have been tested to evaluate the magnitude of induced preanalytical errors. Tonometered serum pools adjusted to four Ca2+ concentrations were sampled anaerobically.Ca2+ and pH (ICA2 with 3 digits, Radiometer Medical A/S, Denmark; Heparin: anti-Xa factor activity on a chromogenic substrate. Results were expressed as means of 10 measurements and as percentages of the reference values. Sarstedt syringes, (Li-heparinate), yielded a negative bias (-3%). However for 0.5 or 1 mL samples the bias reached -4% to -6%. Radiometer syringes (QS50 and QS90; calcium titrated heparinate) demonstrated biases below -2%. The bias in the Ciba-Corning (Gas-Lyte) syringe was below 2%. Portex (Pulsator) syringes showed biases above +4% even for nominal sampling volumes. All syringes (except Pulsator) released anticoagulant amounts corresponding to the expected values. Radiometer and Ciba-Corning were the only recommendable devices.

The angiotensin I converting enzyme inhibitor (ACEI) perindopril (2 mg/kg body weight), the peripheral vasodilator dihydralazine (DHL) (25 mg/kg body weight) or distilled water was given daily from birth to day 14 to neonatal rats. Blood pressure, plasma creatinine, plasma renin activity (PRA), substrate (PRS) and concentration (PRC) and renin content of kidney tissue sections were evaluated on days 14 and 28. By day 14, a high mortality in the ACEI group was observed. ACEI, but not DHL, led to a significant fall (P < 0.01) in blood pressure, 57 +/- 11 versus 89 +/- 25 in the DHL group and 103 +/- 24 mmHg in controls, and to a dramatic increase in plasma creatinine. PRA and PRS were undetectable in ACEI-treated rats; in contrast, PRC and renal staining with anti-renin antibody were significantly increased in ACEI rats. On day 28, the blood pressure was normal in all groups and plasma creatinine returned to the normal range in ACEI rats. PRA, PRS and PRC were not significantly different in the ACEI group and controls. These results suggest that the renin-angiotensin system (RAS) plays a major postnatal role in the neonatal rat. Inhibition of the RAS during the first 2 weeks of life leads to high mortality, severe hypotension, reversible renal failure and a defect in circulating angiotensinogen.

Dopamine decreases tubular sodium reabsorption, attributed in part to Na-K-ATPase inhibition in the proximal convoluted tubule (PCT). Because the final regulation of sodium excretion occurs in the collecting duct, where specific dopamine DA1 binding sites have been demonstrated, we examined the effects of dopamine, as well as of DA1 and DA2 receptor agonists on Na-K-ATPase activity and on the number of units in Madin-Darby canine kidney (MDCK) cells, which retain differentiated properties of the renal cortical collecting tubule epithelium. Dopamine (10(-5) M) inhibited pump activity (by 50%) and reduced the number of units. This effect was reproduced by the DA1 agonist SKF 38393, which inhibited pump activity in a dose- and time-dependent manner (maximum, 10(-5) M). The DA2 agonist quinpirole hydrochloride was without effect, either alone or in combination with SKF 38393. Inhibition of pump activity by dopamine was totally abolished by H7 (100 microM), an inhibitor of protein kinase (PK), but partially by 2',5'-dideoxy-adenosine (DDA) and H4, respective inhibitors of cAMP production and PKA, which suggests that the dopamine effect on Na-K-ATPase activity may be linked to activation of both PKC and PKA. In these cells, amiloride addition during preincubation did not alter the effect of dopamine on Na-K-ATPase activity; in contrast, furosemide increased further the inhibitory effect of dopamine on the enzyme activity. Monensin addition (10(-3) M) reversed the inhibitory effect of dopamine after a 30-min preincubation.(ABSTRACT TRUNCATED AT 250 WORDS)

The elderly frequently suffer long lengths of hospital stay (LOS). These long stays are often associated with long social care stays which occur when patients no longer require acute care and are awaiting post-discharge services. In this study, actual acute care LOS and social care LOS were studied specifically in hospitalized frail elderly. Our data demonstrate that frail elderly receiving only acute care do not suffer markedly prolonged total LOS (TLOS). However, in hospitalized frail elderly patients who experience acute care and social care stays, social care LOS accounts for over half of all hospital days. When patients were grouped and studied according to the type of post-discharge services being sought by the health care team, significant differences in acute LOS and social care LOS were noted. Subgroups of patients were also identified among the various groups which differed significantly in their LOS parameters. Patients who required more than one discharge plan during the course of hospitalization experienced the longest hospital stays of all groups, and spent almost 70% of these days receiving non-acute social care. In a study of the relationship between the intensity of social work intervention and social care LOS in the frail elderly, a statistically significant relationship was noted between the timing and frequency of social work intervention and the actual length of social care stays. Early and frequent social work interventions were associated with significantly shorter social care LOS. We conclude that the study of TLOS should include acute LOS and social care LOS to obtain a reliable measure of the course and cost of hospital care for the frail elderly. The study of social care subgroups may facilitate future investigations to define the social care problems which contribute most to TLOS, and the patient populations which should be most heavily targeted for early and intensive social work intervention.

To evaluate the influence of protein kinase C (PKC) activation on Na/K-ATPase activity in MDCK cells, we studied the effect of phorbol myristate acetate (PMA) and two diacylglycerol analogues, oleoylacetylglycerol and dioctanoylglycerol, on the enzyme activity. Na/K-ATPase activity was determined by cytochemistry. PMA induced a time- and dose-dependent inhibition of Na/K-ATPase activity and at 100 ng/ml decreased the enzyme activity by 55% of the initial value. These effects were mimicked by oleoylacetylglycerol and dioctanoylglycerol, and were abolished by two inhibitors of PKC, 1-(5-isoquinolinylsulphonyl)-2-methylpiperazine (H7) and sphingosine. A phorbol ester that does not activate PKC, 4 alpha-phorbol 12,13-didecanoate, did not inhibit Na/K-ATPase activity. PMA inhibition persisted in the presence of cycloheximide and actinomycin D but not in the presence of amiloride. Dopamine (10 microM) inhibition of Na/K-ATPase activity was abolished in a dose-dependent manner by sphingosine. Results suggest that in MDCK cells Na/K-ATPase is an effector protein for PKC and that dopamine inhibition of its activity may be mediated by PKC.

Na + ,K + -adenosine triphosphatase (Na + ,K + -ATPase) is an integral protein usually located in the basolateral membrane [1, 2] that actively pumps Na + out and K + in to the cell, through ATP hydrolysis [3]. The activity of Na + ,K + -ATPase is essential for the existence of water and solute transport by epithelial cells [4,5]. Measurement of Na + ,K + -ATPase activity is usually done by biochemical assays. Cytochemical methods [6] allow cellular enzyme localization and may offer an alternative approach for determination of Na + ,K + -ATPase activity in disrupted cells. We have used a cytochemical method, based on measurements of inorganic phosphate generated during ATP hydrolysis by ATPases during the incubation period and precipitated by a lead ammonium citrate acetate complex (LACA) as lead phosphate [Pb 3 (PO 4 ) 2 ]. The colorless lead phosphate was converted into a brown precipitate of lead sulphide (PbS) by treatment with ammonium sulphide. We chose to evaluate Na + ,K + -ATPase activity in MDCK (Madin-Darby canine kidney) cells because they retain many differentiated properties characteristic of distal tubule epithelia [4, 7]. In these cells, apical to basolateral salt and water transport was shown to be inhibited by ouabain and amiloride [7–10]. We show that this method is a simple, convenient, and could be easily performed to measure Na + ,K + -ATPase activity and its modulation under physiological conditions.

Ion-selective electrodes (ISEs) respond to ion-activity and therefore do not sense substance concentration directly. However, it is recognized that sodium and potassium in plasma will continue to be expressed for clinical purposes in terms of substance concentration (mmol/l). A convention is proposed whereby for routine clinical purposes results of ISE measurements of sodium and potassium in undiluted plasma should be reported in terms of substance concentration (mmol/l). In specimens with normal concentrations of plasma water, total CO2, lipids, protein and pH, the values will concur with the total substance concentration as determined for example by flame atomic emission spectrometry (FAES) or ISE measurements on diluted samples. In specimens with abnormal concentrations of plasma water, the results will differ. However, under these circumstances, measurements of sodium and potassium by ISE in the undiluted sample will more appropriately reflect the activity of sodium and potassium and are therefore clinically more relevant than the determination in diluted samples. Detailed recommendations are made about practical procedures to achieve this. The recommended name for this quantity is the substance concentration of ionized sodium or ionized potassium in plasma, as opposed to total sodium or total potassium determined by, e.g. FAES, or ISE measurements on diluted samples.

The present study was designed to investigate the effects of chronic administration of the arginine analogue L-Name (50 mg/kg body weight), the angiotensin converting enzyme inhibitor, perindopril (2 mg/kg body weight), and perindopril (2 mg/kg) plus L-Name (50 mg/kg) on blood pressure, plasma renin activity, plasma angiotensinogen, and hepatic angiotensinogen mRNA levels in young and adult rats. The drugs were given daily from birth to day 21 to puppies and for 15 days to adults. Analytical procedures were performed on day 21 for the puppies and at 10 weeks for the adults. In puppies, blood pressure did not change with L-Name, it decreased to 45% of control values (P

Osteocalcin (Oc) is a 49-amino-acid noncollagenous protein synthesized by osteoblasts. Because of the presence of three γ-carboxyglutamic acid (GLA) residues, newly synthesized Oc will be primarily bound to hydroxyapatite present in bone matrix, only a small fraction being released into the circulation (1). Although serum Oc concentration is considered a valid marker of bone turnover in most situations (2), it reflects specifically bone formation when resorption and formation are uncoupled as observed in corticoid-induced osteoporosis (3). Several reports have stressed recently that absolute Oc concentrations obtained with various in-house (4) or commercial (5) assays cannot be compared directly. An important cause for the discrepancies between Oc assays is the variabily of the anti-Oc antibody specificity for the circulating Oc fragments described by Garnero et al. (6). The intact Oc molecule (1–49 Oc) is highly susceptible to tryptic proteolysis because of the presence of 2 Arg-Arg sites in positions 19–20 and 43–44 (7). When serum samples are not rapidly frozen, 1–49 Oc will be degraded quickly, the largest degradation product being a big N-terminal mid-fragment (1–43 Oc). This fragment is accumulated in patients with chronic renal failure (CRF) (6). This problem of degradation is a major drawback in terms of specificity with an Oc assay that recognizes only 1–49 Oc. On the contrary, an Oc assay recognizing both 1–49 Oc and 1–43 Oc with equal affinity will be less sensitive to the effect of proteolysis (8) but of very limited value in CRF patients, as even those with adynamic bone disease will appear to have high serum Oc concentrations (9). Antibody specificity is thus an important variable to be taken into account when interpreting serum Oc concentrations obtained with a given commercially available assay. The objective of the present study was to assess the specificity …

Diabetic patients receiving insulin therapy generally develop anti-insulin antibodies that must be eliminated, usually by extraction with polyethylene glycol (PEG), before determining the concentration of free (active) insulin in plasma. We describe a new method for removing such antibodies, with the use of Protein A coupled to Sepharose microspheres. The results correlate well with those by the PEG method, although values are systematically higher or lower for given samples, according to the initial titer of the antibody measured in terms of binding capacity. Further studies are required to clarify this observation.

To test prolactin (PRL) action on osmoregulation in mammals, we evaluated in the rat the effect of this hormone on a major enzyme in renal regulation of water and electrolyte: renal Na-K-ATPase. Enzyme activity was determined by cytochemistry in medullary ascending limb (MAL) and distal convoluted tubule (DCT) from rats treated either by bromocriptine, or by PRL. Three hours after a bromocriptine injection (0.1 mg/100 g IP) a significant decrease of Na-K-ATPase activity is observed in both MAL (80% of control values, p less than 0.001) and DCT (78% p less than 0.01). Reciprocally, a significant (p less than 0.001) increase in enzyme activity is induced 3 h after a single PRL injection (140 micrograms/100 g IM), in both segments (MAL: 165%, DCT: 172% of control activities) and persists 6 h after the injection (MAL: 130%, DCT: 118%). Na-K-ATPase activity was correlated to plasma PRL levels (r = 0.78 in DCT, r = 0.89 in MAL). A direct effect of PRL on the tubule is suggested by results from experiments in which PRL, at various concentrations, is added in vitro on renal slices before Na-K-ATPase activity measurements. The increase in Na-K-ATPase activity exhibits a log-dose dependency with PRL concentration (p less than 0.01) and is still observed when AVP antagonist is added before PRL incubation, ruling out the possible role of AVP contamination of PRL. These results suggest a direct effect of PRL on renal Na-K-ATPase in MAL and DCT.

Parathyroid response to aluminum in vitro: ultrastructural changes and PTH release. The endocrine response of porcine parathyroid gland tissue slices in vitro to aluminum was studied by electron microscopy and radioimmunoassay of PTH. Medium aluminum concentrations were 20 to 500 ng/ml covering the range corresponding to concentrations reported in the plasma of aluminum-intoxicated hemodialyzed patients. Aluminum inhibited iPTH-release and caused severe cell alterations. This inhibition was incomplete and there was an aluminum–insensitive iPTH-release capacity. This phenomenon seemed to be due to heterogeneous parathyroid cell population as regards aluminum sensivity, perhaps linked to the spontaneous asynchronous cyclic parathyroid cell changes. Sensitivity to aluminum was modulated by the extra–cellular calcium concentration. Sensitivity to extra–cellular calcium concentration variations persisted in aluminum intoxicated tissues. The severity of the observed cell lesions induced by high concentrations of aluminum suggested that the recovery of an iPTH-release capacity when parathyroid tissue was withdrawn from a toxic environment and switched to aluminum–free media is more likely to be due to activation of a “less-sensitive to aluminum” cell pool than to a true reversibility of the toxic effect.

Plasma renin activity (PRA) has been measured by microassay on capillary and venous blood sampled simultaneously in 21 subjects; there is no significant difference between these two groups of PRA values. PRA values in normal infants, children and adults have been measured with this microassay and the results are similar to those previously published by authors using different radioimmunological assays.

In a preliminary in vitro study of the direct toxic effects of aluminum on PTH release from incubated parathyroid gland slices [1], we observed that addition of fetal calf serum (FC) to the incubation medium introduced some variability into the final results and that this effect was accounted for by variations of the ionized calcium concentration in the medium. We also noted that the addition of aluminum produced cloudiness during medium preparation [21. Questioning the possibility of changes or instability in the medium composition, we investigated systematically the effects of aluminum salt and fetal calf serum addition on the electrolyte composition of the medium.

As a preliminary step in a study of the effects of calcium ligands on the pH standardization of ionized calcium (Ca2+) measurements in blood, the slope of logCa2+ = f(pH) linear relationship characterizing the pH-sensitive calcium buffer capacity of the specimen was investigated in 12 serum pools on three different instruments. The pH 7.40 correction line should be horizontal. This was the case for the ICA-2 but not for the ICA-1 and the NOVA-8. The discrepancy was caused by an incorrect setting of the built-in slope correction factor in the ICA-2; coincidentally, its value was close to the effective slopes of the serum pools used in the study. Thus, the 'abnormal' behaviour of the ICA-1 and the NOVA-8 was caused by an inadequacy of the built-in algorithm to the characteristics of our serum pools. These findings lead us to reconsider the use of a fixed and constant correction factor to normalize actual ionized calcium values.

The aim of our study was to evaluate the function and growth of parathyroid tissue autografted into the forearm of hemodialysis patients using several presently available methods. In a dynamic study, the secretory function of autografted tissue was evaluated in seven patients using either zero calcium dialysate or calcium infusion. In an additional prospective study, seven patients had repeated determinations of plasma immunoreactive parathyroid hormone (iPTH) concentration on samples from both forearms, a radionuclide evaluation of autograft function using thallium-201 chloride, and real time ultrasonography. Light microscopy analysis was performed in two patients. The dynamic study demonstrated that induction of hypocalcemia was followed by an increase, and induction of hypercalcemia by a decrease, in circulating iPTH in both forearms using three different radioimmunoassays, similar to what has been reported for normal parathyroid tissue. A significant gradient (ie, greater than 2.0) of plasma iPTH concentration in samples from both forearms was observed in only three out of the seven patients of the prospective study. Two of these patients disclosed an increased uptake of 201TI chloride at the site of autografted tissue and had an echographically detectable mass. In both, hyperplastic parathyroid tissue was removed. At present, the remaining third patient does not have other features of recurrent hyperparathyroidism. In conclusion, autotransplanted parathyroid tissue of hemodialysis patients shows an adequate response to physiologic stimuli such as hypo- and hypercalcemia. Dynamic tests, therefore, appear to be a useful tool in the assessment of its function. In addition, radionuclide and echographic studies may be reliable adjuncts in the detection of marked parathyroid autograft hyperplasia.

uation o f this therapy in hospital did not obviate the need for peri toneal dialysis. Although abnormalities of coagulation have been demonstrated rarely in the HUS, 7 there is usually no evidence of ongoing disseminated intravascular coagulation in this disorder? Accordingly, there is little factual basis for anticoagulant therapy. As recommended by othersr we agree that a definite role for the use of inhibitors of platelet aggregation in the therapy of this disease must await more definitive studies clearly demonstrat ing that platelet consumption is a primary noxious event in the evolution of this syndrome.

The radio- and chemoprotective agent, S-2 (3-aminopropylamino) ethyl-phosphorothioic acid (WR-2721) has been reported to lower hypercalcaemia in patients with cancer, probably by increased renal calcium excretion and decreased parathyroid hormone (PTH) secretion and bone calcium resorption. The present study reports the first clinical use of WR-2721 in an anuric haemodialysis patient with severe secondary hyperparathyroidism. The drug was administered intravenously at different doses, i.e. 150, 300, and 500 mg/m2. The infusion was followed by a striking decrease of plasma immunoreactive (i) PTH within 30 min. The nadir of the iPTH decrease was reached at 60 min and was followed by a steady return to previous values. Serum ionised calcium decreased more progressively from 1.55 mmol/l initially to 1.30 mmol/l at 4 h after the 300-mg dose, remained at that level at 24 h, but rose again to pre-infusion values after 48 h. The extent and duration of the decrease in plasma iPTH and ionised calcium were dose-dependent. The circulating iPTH at 24 h was inversely related to the corresponding plasma ionised calcium concentration and had risen above preinfusion values at that time. Plasma concentrations of three other hormones, i.e. renin, insulin, and prolactin, were not affected by the administration of WR-2721. In conclusion, WR-2721 can induce a decrease in serum ionised calcium in the absence of any excretory kidney function. The rapid effect of the drug on circulating iPTH supports the notion of an interference with PTH secretion or catabolism.

The relative effect of glucose or lipids on whole body protein metabolism was investigated by a randomised crossover study in 5 infants (age 3-8 months) steadily growing on parenteral feeding, with constant protein (2,8 + 0,2 g/kg.d) and energy (103 + 3 kcal/kg.d) intakes. Energy was provided as glucose and as an isocaloric glucose lipid mixture, with 50 mg/kg.d L carnitine. Net glucose and fat oxidation rates were measured by indirect calorimetry (IC) for 6 hours. After 120 mn, a primed-constant (Ll13C) leucine infusion was started. Breath CO2 and plasma samples were obtained at 0, 60, 120 and every 30 mn. Enrichments of plasma ketoisocaproate and leucine were measured by mass spectrometry (MS) and 13CO2 production rate by using IC and MS data at steady state enrichment. Lipogenesis (2 + 0,8 g/kg.d) was present with glucose, whole body protein synthesis (PS) and breakdown (PB) rates amounting to 8,3 + 0,6 and 8,1 + 0,9 g/kg.d, respectively resulting in a 0,2 + 0,7 g/kg.d net synthesis (PNS) rate; with the glucose-lipid mixture, fat oxidation rate was 3,8 + 0,6 g/kg.d (p