Osmotic gradient ektacytometry - a novel diagnostic approach for neuroacanthocytosis syndromes

Abstract

Introduction: The unique red blood cell (RBC) properties that characterize the rare neuroacanthocytosis syndromes (NAS) have prompted the exploration of osmotic gradient ektacytometry (Osmoscan) as a diagnostic tool for these disorders. In this exploratory study, we assessed if Osmoscans can discriminate NAS from other neurodegenerative diseases.

Methods: A comprehensive assessment was conducted using Osmoscan on a diverse group of patients, including healthy controls (n = 9), neuroacanthocytosis syndrome patients (n = 6, 2 VPS13A and 4 XK disease), Parkinson's disease patients (n = 6), Huntington's disease patients (n = 5), and amyotrophic lateral sclerosis patients (n = 4). Concurrently, we collected and analyzed RBC indices and patients' characteristics.

Results: Statistically significant changes were observed in NAS patients compared to healthy controls and other conditions, specifically in osmolality at minimal elongation index (O_min), maximal elongation index (EI_max), the osmolality at half maximal elongation index in the hyperosmotic part of the curve (O_hyper), and the width of the curve close to the osmolality at maximal elongation index (O_max-width).

Discussion: This study represents an initial exploration of RBC properties from NAS patients using osmotic gradient ektacytometry. While specific parameters exhibited differences, only O_hyper and O_max-width yielded 100% specificity for other neurodegenerative diseases. Moreover, unique correlations between Osmoscan parameters and RBC indices in NAS versus controls were identified, such as osmolality at maximal elongation index (O_max) vs. mean cellular hemoglobin content (MCH) and minimal elongation index (EI_min) vs. red blood cell distribution width (RDW). Given the limited sample size, further studies are essential to establish diagnostic guidelines based on these findings.

Keywords: Osmoscan; RBC deformability; VPS13A disease; XK disease; acanthocytes; ektacytometry; neurodegeneration.

Figure 1

Ektacytometry data. Panel (A) shows representative Osmoscan curves of a NAS patient sample compared to a healthy control donor. In addition, characteristic parameters of the curve are annotated. Panel (B) provides the Osmoscan curves of both VPS13A disease patients and all 4 XK disease patients, showing their similarity and justifying to pool these patients in a NAS group for statistical analysis. Panel (C) depicts representative Osmoscan curves for both, VPS13A and XK mutation carriers showing their similarity. Panel (D) plots all Osmoscan curves of patients from all investigated neurodegenerative diseases except NAS to indicate the lack of difference between control curves and the neurodegenerative disorders HD, PD and ALS. In panels (A–D) each curve represents an individual patient or a healthy volunteer. Panels (E–L) show the statistical analysis of the characteristic parameters annotated in panel A. These are EI_min, O_min, EI_max, O_max, AUC, O_min-width, O_hyper and O_max-width, respectively. Plotted are the individual values with indication of the mean and the standard error of mean (SEM). Significance was checked with an ordinary one-way ANOVA test, and 1–4 stars correspond to p-values lower than 0.05, 0.01, 0.001, and 0.0001, respectively.

Figure 2

RBC indices of the different patient groups and mutation carriers and correlations with the ektacytometry parameters. Panels (A–I) present the RBC number per blood volume, the hemoglobin concentration of the blood, the hematocrit, the mean RBC volume (MCV), the mean RBC hemoglobin content, the mean RBC hemoglobin concentration, the RBC distribution width (RDW), the percentage of reticulocytes and the mean reticulocyte hemoglobin content, respectively. Data are based on 2 VPS13A disease patients, 4 XK disease patients, 3 VPS13A mutation carriers, 4 XK mutation carriers, 6 PD patients, 5 HD patients, 4 ALS patients and 9 controls. The NAS patients as well as the NAS mutation carriers are based on pooled respective subgroups (VPS13A and XK disease). Plotted are the mean values with the standard error of mean (SEM). Significance was checked with an Ordinary one-way ANOVA test, and 1–4 stars correspond to p-values lower than 0.05, 0.01, 0.001, and 0.0001, respectively. Panel (J) is a color-coded correlation matrix for healthy controls (HC) and pooled NAS patients of the values presented in Figures 1E–L and panels (A–G). Selected correlation blots for EI_min vs. RDW and O_max vs. MCH are provided in panels K and L, respectively.