The data used in the present study were from the online database of the Alzheimer’s Disease Neuroimaging Initiative (ADNI).¹ Healthy individuals (as control 54), patients with mild cognitive impairment (345), and patients with AD (96) were followed up for 1 year, and data on dynamic changes in GSTα protein levels in plasma and primary information data (including clinical assessment, neuropsychological tests) were collected.

Recruitment methods and diagnostic approaches for ADNI have been reported previously (Allen et al., 2012). Individuals diagnosed with mild AD were required to fulfill the criteria established by the National Institute of Neurological and Communicative Disorders and Stroke–Alzheimer’s Disease and Related Disorders Association for probable Alzheimer’s Disease (Kiely et al., 2018).

Written consent from participants was acquired in every source study by the Declaration of Helsinki and with the approval of local Institutional Review Boards. Comprehensive information regarding ethics approval, study design, participant recruitment, and clinical assessments can be found at adni-info.org.

The Kolmogorov–Smirnov test was employed to assess the normality of the data. Data that adhered to a normal distribution were reported as the mean ± standard deviation. For comparisons between two groups, the two-sample t-test was applied, while a one-way ANOVA was utilized for comparisons involving three groups. In cases where the data did not meet the assumptions of normality, the median was reported alongside the first and third quartiles; in these instances, the Mann–Whitney U test was used for two-group comparisons, and the Kruskal-Wallis test was applied for three groups. Categorical data were represented as percentages, with the χ² test implemented for analysis. Pearson’s correlation coefficient was used for normally distributed data, whereas Spearman’s correlation analysis was applied to examine relationships among non-normally distributed parameters. Receiver operating characteristic (ROC) curves were generated to evaluate GSTα protein levels in the context of mild cognitive impairment (MCI) and Alzheimer’s disease (AD), and the area under the curve (AUC) was computed. All hypothesis tests were conducted as two-tailed, with a significance threshold set at p < 0.05. Data processing and graphical representation were performed using SPSS version 26.0 and Origin 2021 software.

A total of 495 subjects were included in this study, and the essential information is shown in Table 1. There were significant differences in MMSE and CDR scores and the type of APOE genes carried by CN, MCI, and AD before follow-up (p < 0.05), and there were no significant differences in gender, age, years of education, and GSTα, (p > 0.05).

The first measurement plasma GSTα protein levels at follow-up were not statistically different among the three groups (p > 0.05, Figure 1A). A year later, plasma protein levels of GSTα in the Alzheimer’s Disease (AD) group were significantly lower than those in the Cognitively Normal (CN) group (0.94 vs. 1.05, p = 0.04) and the Mild Cognitive Impairment (MCI) group (0.94 vs. 1.09, p < 0.001) (Figure 1B). The difference in GSTα changes in plasma protein levels after 1 year of follow-up was minimal when comparing the MCI group to the AD group (0.04 vs. 0.33, p < 0.001) (Figure 1C). Additionally, plasma GSTα protein levels were lower than baseline after 1 year of follow-up in the CN group (1.21 vs. 0.88, p = 0.04) (Figure 1D), in the MCI group (1.12 vs. 1.06, p = 0.03) (Figure 1E), and in the AD group (1.11 vs. 0.78, p < 0.001) (Figure 1F).

We analyzed the relationship between GSTα protein levels changes and baseline in the plasma three populations, with a non-significant correlation between plasma GSTα protein levels changes and baseline levels in the CN group (r = 0.18, p = 0.20) (Figure 2A) and the MCI group plasma GSTα changes were positively correlated with baseline levels (r = 0.50, p < 0.001) (Figure 2B), and in the AD group (r = 0.57, p < 0.001) (Figure 2C). At the same time, Plasma GSTα protein level changes were positively correlated with altered MMSE levels in MCI and AD patients (r = 0.09, p = 0.04) (Figure 2D).

To evaluate the ability of plasma GSTα to predict MCI and AD, we plotted ROC curves, and after 1 year of follow-up, the AUC of the area under the curve of plasma GSTα for MCI prediction (95% CI) was 0.63 (0.54–0.71), p = 0.02, (Figure 3A), and that of the area under the curve of plasma GSTα for AD prediction (95% CI) was 0.74 (0.69–0.80), p < 0.001 (Figure 3B), while we plotted ROC curves for the difference in plasma GSTα changes after 1 year of follow-up, and the results showed that plasma GSTα alteration on MCI predicted area under the curve AUC (95% CI) 0.76 (0.696–0.83), p < 0.001, (Figure 3C), plasma GSTα alteration on AD predicted area under the curve AUC (95% CI) 0.75 (0.69–0.80), p < 0.001 (Figure 3D), and plasma GSTα alteration had diagnostic value and accurate risk measurement ability for MCI and AD.

We statistically analyzed the difference in plasma GSTα protein level changes by gender at 1-year follow-up. The results showed that plasma GSTα changes were less than MCI changes in men than in the AD group (0.03 vs. 0.36, p < 0.001) (Figure 4A), and plasma GSTα changes were less than MCI changes in women than in the AD group (0.05 vs. 0.30, p < 0.001) (Figure 4D). The ROC curves were plotted, and the results showed that plasma GSTα alteration in males had an AUC (95% CI) of 0.76 (0.69–0.83), p < 0.001, for the area under the prediction curve for MCI (Figure 4B), and plasma GSTα alteration had an AUC (95% CI) of 0.72 (0.63–0.80), p < 0.001, for the area under the prediction curve for AD (Figure 4C). Plasma GSTα alterations in women had an AUC (95% CI) of 0.72 (0.62–0.83), p < 0.001, for the area under the prediction curve for MCI (Figure 4E), and plasma GSTα alterations had an AUC (95% CI) of 0.78 (0.70–0.86), p < 0.001, for the area under the prediction curve for AD (Figure 4F), and plasma GSTα alterations in different sexes had the same diagnostic value and accurate risk-measurement ability for MCI and AD.