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Thiol protein defect in sodium-lithium countertransport in subset of essential hypertension
P Mead1, R Wilkinson, T H Thomas
1Department of Medicine (Nephrology), University of Newcastle-on-Tyne, Newcastle-on-Tyne, UK.
Insights
Abnormal erythrocyte sodium-lithium countertransport (Na/Li CT) is linked to hypertension. Modifying thiol proteins in red blood cells mimics this abnormality, identifying a specific patient subgroup with altered membrane function.
Area of Science:
- Biochemistry
- Cardiovascular Physiology
- Membrane Transport
Background:
- Essential hypertension (EHT) likely has diverse causes.
- Abnormal erythrocyte sodium-lithium countertransport (Na/Li CT) is observed in a subset of patients with EHT and a family history of hypertension and cardiovascular disease (EHT-FH).
Purpose of the Study:
- To investigate if altering a membrane thiol protein can replicate the abnormal Na/Li CT seen in EHT-FH patients.
- To better understand the mechanism of abnormal Na/Li CT for identifying homogeneous patient subgroups.
Main Methods:
- Determined Na/Li CT kinetics in untreated erythrocytes.
- Performed thiol group alkylation using N-ethylmaleimide (NEM) on erythrocytes.
- Compared kinetic patterns between normal controls and EHT-FH patients before and after NEM treatment.
Main Results:
- EHT-FH erythrocytes exhibited a low K(m) and high Vmax/K(m) ratio for Na/Li CT compared to controls.
- NEM treatment reproduced this kinetic pattern in normal erythrocytes.
- NEM treatment in EHT-FH erythrocytes caused a marked increase in Vmax, indicating increased transporter turnover, unlike the increased affinity seen in normal cells.
- Approximately 75% of EHT-FH patients showed abnormal kinetic changes with NEM.
Conclusions:
- A specific, NEM-reactive thiol group in Na/Li CT is implicated in abnormal transport in a subgroup of hypertensive patients.
- This thiol group may serve as a valuable intermediate phenotype for a specific disease group within EHT.
- Standard Na/Li CT flux assays are insufficient for discriminating this subgroup; identifying the specific thiol protein is crucial for understanding altered membrane function.
Abstract:
There is probably a heterogeneous etiology for essential hypertension (EHT), and abnormal erythrocyte sodium-lithium countertransport (Na/Li CT) is common in a subgroup of patients with a strong family history of hypertension and cardiovascular disease (EHT-FH patients). The aim of this study was to test the hypothesis that altering a membrane thiol protein could mimic the abnormal Na/Li CT observed in the patients and that a more refined understanding of the mechanism of abnormal Na/Li CT would facilitate a clearer identification of a subgroup of patients with a homogeneous biochemical abnormality. Na/Li CT kinetics were determined in untreated erythrocytes and after thiol group alkylation with N-ethylmaleimide (NEM). Compared with normal control erythrocytes, untreated erythrocytes from EHT-FH patients had a low K(m) of Na/Li CT, with a high ratio of maximum velocity to K(m). This kinetic pattern was reproduced in normal erythrocytes by treatment with NEM in sodium-free medium. The same treatment in EHT-FH erythrocytes caused a markedly abnormal effect with an increase in maximum velocity, indicating an increase in transporter turnover in contrast to the increase in sodium affinity seen in normal control erythrocytes. Frequency distributions of these kinetic changes showed a subgroup of approximately 75% of EHT-FH patients with abnormal kinetic changes with NEM. Therefore, the key Na/Li CT thiol group that is very reactive to NEM and causes the abnormal Na/Li CT in a subgroup of hypertensive patients may be a useful intermediate phenotype for a disease group within the syndrome of EHT. The single flux assay of Na/Li CT at 140 mmol/L sodium poorly discriminates this group. Identification of the thiol protein involved may lead to a molecular explanation of the altered membrane function in this subgroup of patients.