Increase In Plasma Potassium Levels Research Articles

PS-BPB10-4: CREATION AND CHARACTERIZATION OF THE UBIQUITIN LIGASE KLHL3-S433A KNOCK-IN MICE

Background and aim: Kelch-like 3 (KLHL3) is a component of an E3 ubiquitin ligase complex that regulates blood pressure and electrolyte homeostasis. Mutation and inactivation of KLHL3 are known to cause hypertension and hyperkalemia mainly due to the impaired with-no-lysine (WNK) degradation and increased Na-Cl cotransporter (NCC) activity in the kidney. Previously, we have reported that angiotensin II (Shibata et al. PNAS 2014), potassium deficiency, and impaired glucose tolerance (Ishizawa et al. BBRC 2016 and JASN2019) can inactivate KLHL3 by altering phosphorylation at S433 in the Kelch-domain. We and others have also shown that decreased KLHL3 expression is also involved in the pathology of hypertension and hyperkalemia. The current study was designed to demonstrate the physiological importance of S433 in KLHL3 function in vivo in mice. Method: Using CRISPR/Cas9, S433 of KLHL3 was substituted to alanine in C57BL/6 mice (KLHL3-S433A Knock in mice: KI). Creation of KI was confirmed by DNA sequencing and RFLP analysis. Blood pressure was evaluated by telemetry, and blood gas was analyzed by iSTAT. Levels of KLHL3, WNK1/4, and NCC in the kidney were determined by Western blot analysis. Gene expression of KLHL3, WNK1 and WNK4 was evaluated by real time RT-PCR. Results: On a normal salt diet, blood pressure levels were similar between WT and KI; however, KI mice showed significant increase in response to a high salt diet compared with WT on the same diet. KI mice showed a significant increase in plasma potassium levels and a significant decrease in bicarbonate levels already at baseline, confirming that S433 in KLHL3 is also critical in regulating potassium and acid/base homeostasis. Urinalysis indicated that the increase in plasma potassium levels is mainly attributable to the decreased potassium excretion. Of interest, we found that total KLHL3 abundance was drastically reduced by Western blot analysis, which was accompanied by the increase in KS-WNK1 and WNK4 and an increase in NCC both. Real time RT-PCR showed that gene expression of Klhl3, as well as those of Wnk1 and Wnk4, was unchanged, indicating that S433A substitution promotes degradation of KLHL3, leading to hyperkalemia and increased salt sensitivity. Conclusion: The study confirmed the critical importance of S433 in KLHL3 in controlling the ubiquitin ligase function and electrolyte homeostasis in vivo. It also indicates that the modification of KLHL3-S433 influences the protein abundance without affecting gene transcription.

Storage related haematological and biochemical changes in Plasmodium falciparum infected and sickle cell trait donor blood

BackgroundIn sub-Saharan Africa where sickle cell trait (SCT) and malaria is prevalent, significant proportions of blood donors may be affected by one or more of these abnormalities. The haemato-biochemical properties of SCT and asymptomatic malaria in donor blood have not been evaluated. This study evaluated the haemato-biochemical impact of SCT and asymptomatic malaria infections in citrate-phosphate-dextrose-adenine (CPDA-1) stored donor blood units.MethodsFifty-milliliters of sterile CPDA-1 anti-coagulated blood were drained into the sample pouch attached to the main blood bag. Ten units each of sickle cell/malaria negative, sickle cell and malaria positive blood were analyzed. Baseline and weekly haematological profiling and week 1, 3 and 5 concentrations of plasma haemoglobin, % haemolysis, sodium, potassium and chloride and lactate dehydrogenase (LDH) were assayed. Differences between baseline and weekly data were determined using one-way analysis of variance (ANOVA) and Kruskal-Wallis test, whereas differences between baseline parameters and week 1–3 data pairs were determined using paired t-test. P-value < 0.05 was considered statistically significant.ResultsStorage of SCT and malaria infected blood affected all haematological cell lines. In the SCT donors, red blood cells (RBC) (4.75 × 1012/L ± 1.43baseline to 3.49 × 1012/L ± 1.09week-5), haemoglobin (14.45 g/dl ± 1.63baseline to 11.43 g/dl ± 1.69week-5) and haematocrit (39.96% ± 3.18baseline to 33.22% ± 4.12week-5) were reduced. In the asymptomatic malaria group, reductions were observed in RBC (5.00 × 1012/L ± 0.75baseline to 3.72 × 1012/L ± 0.71week-5), haemoglobin (14.73 g/dl ± 1.67baseline to 11.53 g/dl ± 1.62week-5), haematocrit (42.72% ± 5.16baseline to 33.38% ± 5.80week-5), mean cell haemoglobin concentration (35.48 g/dl ± 1.84baseline to 35.01 g/dl ± 0.64week-5) and red cell distribution width coefficient of variation (14.81% ± 1.54baseline to 16.26% ± 1.37week-5). Biochemically, whereas plasma LDH levels significantly increased in asymptomatic malaria blood donors (319% increase at week 5 compared to baseline), SCT blood donors had the most significant increase in plasma potassium levels at week 5 (382% increase). Sodium ions significantly reduced in SCT/malaria negative and sickle cell trait blood at an average rate of 0.21 mmol/L per day. Moreover, elevations in lymphocytes-to-eosinophils and lymphocytes-to-neutrophils ratios were associated with SCT and malaria positive blood whilst elevation lymphocytes-to-basophils ratio was exclusive to malaria positive blood.ConclusionSevere storage lesions were significant in SCT or malaria positive donor blood units. Proper clinical evaluation must be done in prospective blood donors to ensure deferral of such donors.

Role of cyclooxygenase-2 in the prolonged regulation of renal function.

The role of cyclooxygenase-2 (COX-2) in the prolonged regulation of renal function was evaluated during changes in sodium intake and reduction of NO synthesis. It was evaluated in conscious dogs by administering a selective inhibitor (nimesulide) during 8 consecutive days. Nimesulide administration to dogs with normal or high sodium load did not modify glomerular filtration rate but reduced renal blood flow (16%; P<0.05). The vasoconstriction elicited by COX-2 inhibition was greater when NO production was inhibited because glomerular filtration rate decreased by >25% when nimesulide was administered to dogs with a reduced NO synthesis. During low sodium intake, COX-2 inhibition elicited a decrease (P<0.05) of both glomerular filtration rate (34%) and renal blood flow (31%). Sodium excretion only decreased (P<0.05) during the first day of COX-2 inhibition in dogs with normal or high sodium load. The increase in plasma potassium levels elicited by COX-2 inhibition was greater in dogs with low sodium intake and was enhanced when NO production was inhibited. This change in potassium was not secondary to a decrease in plasma aldosterone levels. The results of this study suggest that COX-2-derived metabolites (1) play a more important role in the long-term regulation of renal hemodynamic when sodium intake is low, (2) protect the renal vasculature from the vasoconstriction secondary to a reduction in NO, (3) are only acutely involved in regulating urinary sodium excretion, and (4) play a more important role in regulating plasma potassium concentration when NO synthesis is reduced.

Role of alpha-adrenoceptors in control of plasma potassium in conscious rabbits.

1. In conscious fed rabbits the intravenous infusion of amidephrine (10 micrograms kg-1 min-1), an alpha 1-adrenoceptor agonist, caused a significant increase in plasma potassium levels that was blunted by prazosin (50 micrograms kg-1, s.c.). Idazoxan failed to modify this response. 2. Clonidine (2 micrograms kg-1 min-1, i.v.), an alpha 2-adrenoceptor agonist, also evoked a hyperkalaemic response which was antagonized by idazoxan (1 microgram kg-1, s.c.), yohimbine (0.45 mg kg-1, s.c.) and prazosin (50 micrograms kg-1, s.c.). Apamin (40 micrograms kg-1, i.v. bolus) also suppressed the clonidine-mediated hyperkalaemia. 3. Verapamil (5 micrograms kg-1, s.c.) prevented both alpha 1- and alpha 2-adrenoceptor-mediated increase in plasma potassium levels. 4. It is concluded that in conscious fed rabbits both alpha 1- and alpha 2-adrenoceptor stimulation induce hyperkalaemia by activation of hepatic Ca(2+)-activated K(+)-channels.

Effect of a simultaneous potassium and carbohydrate load on extrarenal K homeostasis in end-stage renal failure.

Patients with chronic renal failure (CRF) are continuously exposed to hyperkalemia. In these patients the extrarenal disposal of a potassium load may be very important to determine the plasma potassium levels. We studied the effect of a combined oral load of potassium (0.5 mEq/kg body weight) and carbohydrate (0.5 g/kg body weight) to mimic normal ingestion of potassium. Eight CRF patients and 5 control subjects were studied. The maximal increase in plasma potassium levels achieved was significantly higher in the patients (1.07 +/- 0.1 mEq/l) than in controls (0.39 +/- 0.05 mEq/l). Basal insulin levels were higher in the CRF patients and increased with the oral potassium and carbohydrate load in both controls and patients. In the CRF patients only 58.9 +/- 3% of the potassium load was translocated to the intracellular space compared to 81 +/- 6% in the controls. No correlation was found between the acid base status and maximal potassium increase. We conclude that patients with CRF exhibit an impaired extrarenal handling of potassium and that this abnormality does not appear to be related to insulin secretion or acid base status.

Effect of enalapril on ventricular arrhythmias in congestive heart failure

Twenty-four-hour Holter electrocardiographic recordings were used to measure the effects of a converting-enzyme inhibitor, enalapril, given for 12 weeks, on the frequency of cardiac arrhythmias in 10 patients with congestive heart failure (New York Heart Association functional class II to III) receiving maintenance therapy with digoxin and furosemide. Nine patients were given placebo, and both study groups were conducted in a double-blind, parallel manner. The placebo group had no change in the frequency of arrhythmias, whereas enalapril-treated patients showed a significant decrease in the frequency of premature ventricular complexes, ventricular couplets and ventricular tachycardia. A minor, nonsignificant reduction in atrial premature complexes was seen in patients who received enalapril. Compared with placebo patients, those who received enalapril had an increase in plasma potassium levels of 0.33 mmol/liter, a decrease in plasma digoxin, and decreases in pulmonary artery wedge, mean pulmonary artery and right atrial pressures. However, none of these indexes were correlated with the concomitant decline in cardiac arrhythmias. It is concluded that enalapril reduces the frequency of ventricular arrhythmias in congestive heart failure, although the underlying mechanisms are not known.

Glucagon-induced increase in plasma potassium levels in type 1 (insulin-dependent) diabetic subjects.

To investigate the hypothesis that in Type 1 (insulin-dependent) diabetes the increase in plasma potassium during decompensation may be due to a rise in glucagon concentrations, we have measured plasma glucose, potassium and glucagon levels in five diabetic patients during two tests with 0.154 mol/l saline or somatostatin (500 micrograms/h) performed on two successive days. The patients were maintained normoglycaemic overnight by means of a continuous insulin infusion. After insulin withdrawal during the saline infusion, glucose and potassium levels rose markedly (delta maximum: glucose, 12.0 +/- 1.5 mmol/l; potassium, 0.73 +/- 0.12 mmol/l), while glucagon showed a slight, but significant increment (delta maximum: 10.6 +/- 1.0 pmol/ml, p less than 0.05). The potassium increment was not mediated by a reduction in blood pH. Somatostatin abolished the rise in glucagon concentration and simultaneously markedly inhibited the rise in potassium and glucose levels. It is concluded that in acute insulin deficiency, glucagon could be one of the factors that contributes to hyperkalaemia.

Influence of saline drinking water on mineral balances in sheep

The influence of sodium chloride ingestion via the drinking water upon the mineral balance in sheep has been examined. Four Merino ewes were offered rainwater containing zero, 0.8, or 1.3 % sodium chloride as the only source of drinking water. After correction for variations in mineral intake, the data showed that the urinary excretion of calcium, magnesium, potassium, phosphorus, sodium, and chloride was increased by saline water ingestion. The faecal excretion of calcium, magnesium, and phosphorus was not affected by the inclusion of sodium chloride in the drinking water, but faecal potassium was decreased and sodium and chloride increased. The drinking of saline water by the sheep resulted in a decline in the calcium, phosphorus, and potassium balances, an increase in the sodium and chloride balances, and no change in the magnesium balance. The sodium balance was negative for the rainwater treatment, but all other balances were positive. Saline water ingestion also caused an increase in the glomerular filtration rate, a decline in plasma magnesium levels, and an increase in plasma potassium levels. It is concluded that saline water ingestion causes alterations in the pathways of excretion of minerals and the mineral balances in sheep, but the changes would appear to be of insufficient consequence to have detrimental effects.