The association between Alzheimer-associated neuronal thread protein (AD7c-NTP) and plasma inflammatory cytokine levels in individuals with cognitive impairment and demographically-matched controls

Abstract

Background

Alzheimer-associated neuronal thread protein (AD7c-NTP) has emerged as a potential diagnostic biomarker for Alzheimer's disease (AD) and mild cognitive impairment (MCI). Growing research indicates neuroinflammatory mechanisms contribute to AD pathogenesis.

Objective

To investigate the relationship between AD7c-NTP level and inflammatory biomarkers.

Methods

This cross-sectional study enrolled 112 participants comprising 72 cognitively impaired individuals (CI group) and 40 demographically matched controls with cognitively normal (CN group). Comprehensive physical evaluations and standardized neuropsychological assessments were administered. Urinary AD7c-NTP concentrations were quantified through enzyme-linked immunosorbent assay (ELISA), while plasma levels of twelve inflammatory markers—IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, TNF-α, IFN-α, and IFN-γ—were analyzed via flow cytometry-based immunofluorescence techniques.

Results

Concentrations of IL-2, IFN-α, and IFN-γ showed marked elevation in CI patients relative to CN controls (p = 0.005, 0.008, and 0.010, respectively). Strong positive associations emerged between AD7c-NTP concentrations and both IL-2 (r = 0.492, p < 0.001) and IFN-α (r = 0.492, p < 0.001). The four-marker panel (AD7c-NTP combined with IL-2, IFN-α, and IFN-γ) achieved optimal diagnostic accuracy for cognitive impairment, yielding an AUC value of 0.9774 with 94% sensitivity and 75% specificity.

Conclusions

Integrating IL-2, IFN-α, and IFN-γ measurements with AD7c-NTP detection could constitute a superior diagnostic framework for early-stage cognitive decline. The observed correlations between AD7c-NTP and these cytokine profiles may indicate previously unrecognized metabolic pathways relevant to AD pathology.

Keywords

AD7c-NTP cytokines Alzheimer's disease cognitive impairment

Introduction

Alzheimer's disease (AD), recognized as a leading cause of dementia, involves progressive deterioration of brain neurons, primarily manifesting through cognitive impairments, including memory decline and learning deficits. The disease's pathological signature features extracellular neuritic plaques and intracellular neurofibrillary tangles, formed respectively by aggregated amyloid-β (Aβ) peptides and abnormally phosphorylated tau (p-tau) proteins.¹ Research indicates that AD-related pathological changes may initiate 15–20 years prior to clinical symptom manifestation.^2,3 Growing experimental data highlights neuroinflammatory mechanisms as critical contributors to AD progression.^4–7 Within the central nervous system, microglial cells and astrocytic networks serve as principal regulators of inflammatory responses, orchestrating both protective and detrimental processes throughout disease development. In AD patients, these immune cells are commonly observed clustering near neurofibrillary tangles and amyloid plaques.^8,9 Activated microglial cells secrete various inflammatory mediators such as TNF-α and interleukin family members (IL-1β, IL-6, IL-8), which demonstrate significant correlations with neuronal degeneration and pathological features of AD. Current research indicates that cytokine involvement in AD pathogenesis presents a multifaceted interaction pattern, with particular uncertainties persisting regarding their roles in initial phases of cognitive deterioration.

Alzheimer-associated neuronal thread protein (AD7c-NTP), a novel cDNA encoding a phosphorylated transmembrane protein of approximately 41 kDa, was first characterized by de la Monte and colleagues in 1996.¹⁰ Over three decades of research, this neuronal thread protein has emerged as a potential early diagnostic indicator, demonstrating consistent elevation in specific brain regions, cerebrospinal fluid, and urinary samples from AD and mild cognitive impairment (MCI) patients.^10–15 While current clinical guidelines do not endorse AD7c-NTP as a definitive AD biomarker, its strong correlation with neurodegenerative processes positions it as a valuable non-invasive detection tool, especially during the initial phases of AD pathogenesis.^16,17

Postmortem analyses have revealed the co-localization of AD7c-NTP with p-tau and phosphorylated neurofilament proteins within neuronal cell bodies, dystrophic neurites, and axonal swellings observed in frontal and temporal lobe specimens from AD patients.¹⁰ Experimental evidence from transfected post-mitotic rat neuronal cultures demonstrates that AD7c-NTP gene overexpression induces neuronal apoptosis and abnormal neurite outgrowth.^18,19 Building upon the documented interplay between tau pathology and inflammatory mediators,²⁰ we postulated a potential link between AD7c-NTP and neuroinflammatory mechanisms. This investigation sought to examine the correlation between AD7c-NTP levels and systemic inflammatory biomarkers across cognitive impairment stages, while evaluating whether integrating AD7c-NTP measurements with plasma inflammatory profiles could improve early detection of cognitive deterioration.

Methods

Participants

The study cohort comprised individuals who completed cognitive function evaluations at Xuanwu Hospital's Health Management Center from June to December 2024. Researchers enrolled seventy-two confirmed cases of cognitive impairment (CI) alongside forty cognitively intact controls. Diagnostic parameters for CI required education-corrected Montreal Cognitive Assessment (MoCA) scores below 26 points, while not meeting the diagnostic criteria of early AD. The CN group enrolled healthy subjects with an education-adjusted MoCA score of ≥ 26 and matched the demographic data during the same period. Exclusion parameters encompassed: (1) documented instances of major infections, traumatic injuries, or surgical procedures occurring within the four weeks preceding enrollment; (2) diagnosis of significant systemic illnesses, particularly acute unstable conditions (including active malignancies, decompensated cardiovascular conditions, or unmanaged thyroid dysfunction); (3) prior mental health diagnoses (such as depressive disorders, anxiety conditions, or schizophrenia spectrum disorders), or extended usage of sleep-regulating pharmacotherapies (including benzodiazepines, Z-drugs, tricyclic antidepressants, or neuroleptic agents); (4) pre-existing neurodegenerative conditions affecting cognition; and (5) presence of the APOE ε4 genetic variant.

Disease definitions

Demographic and clinical parameters such as sex, age, marital status, educational attainment, body composition metrics (body mass index, skeletal muscle content, adipose tissue proportion, visceral adiposity index, and waist-hip ratio) were collected through standardized health screenings. Hypertension classification followed JNC 7 guidelines, identifying individuals with systolic readings ≥140 mmHg and/or diastolic pressures ≥90 mmHg, documented medical history of hypertension, or active antihypertensive treatment.²¹ Metabolic disorder diagnosis adhered to ADA standards, applying to subjects presenting with fasting plasma glucose ≥7.0 mmol/L, prior diabetes diagnosis, or ongoing glucose-lowering therapy.²² Fatty liver refers to metabolic dysfunction-associated fatty liver disease (MAFLD), diagnosed based on imaging findings showing ≥ 5% macrovesicular or mixed hepatic steatosis, with or without nonspecific liver inflammation.²³

Questionnaires

The Self-Rating Anxiety Scale (SAS) and Self-Rating Depression Scale (SDS) each comprise 20 items scored on a 1–4 scale, yielding a total raw score spanning 20–80. Standardized scores are derived by multiplying raw values by 1.25, producing final scores between 25 and 100, with clinical anxiety/depression indicated at ≥50 standardized points.²⁴ The Pittsburgh Sleep Quality Index (PSQI), developed by Buysse et al.,²⁵ employs a 0–21 scoring system where scores exceeding 5 demonstrate high sensitivity and specificity in identifying sleep disturbances. The Aging and Dementia–8 (AD8) screening tool, also termed the Washington University Dementia Screening Interview, serves to distinguish normal cognitive aging from early dementia using a threshold score of 2.²⁶

Plasma inflammatory cytokine quantification

The plasma inflammatory cytokines, including interleukin-1β (IL-1β), interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-10 (IL-10), interleukin-12p70 (IL-12p70), interleukin-17A (IL-17A), tumor necrosis factor-α (TNF-α), interferon-α (IFN-α), and interferon-γ (IFN-γ), were detected by immunofluorescence assay. Experimental procedures strictly followed the manufacturer's guidelines from the multiplex cytokine detection system (Saiji Biological Technology Co., Ltd, Nanchang, China) implemented on a DxFLEX flow cytometer (Beckman Coulter, Inc., California, USA). Briefly, capture microspheres were thoroughly mixed with microsphere buffer and vortexed to ensure homogeneity. The mixture was then incubated at room temperature in the dark for 15–30 min. Subsequently, 25 μL of the prepared capture microsphere mixture was added to each test tube. Next, 25 μL of either the sample to be analyzed, a gradient-diluted calibrator, or a quality control sample was added to each respective test tube, followed by vortexing to ensure uniform mixing. Then, 25 μL of fluorescent detection reagent was added to each tube, and the mixtures were incubated at room temperature in the dark for 2.5 to 3 h. Finally, 1 mL of phosphate-buffered saline (PBS) solution was added to each test tube. The samples were then centrifuged at 200 × g for 5 min, after which the supernatant was carefully removed. Thereafter, 100 μL of fresh PBS was added to each tube prior to instrumental analysis. The concentrations of cytokines were determined based on the standard curve.

Urine AD7C-NTP detection assay

Urinary AD7c-NTP concentrations were quantified through an enzyme-linked immunosorbent assay (ELISA) procedure employing commercial kits from Anqun Biological Technology Co., Ltd (Shenzhen, China), adhering strictly to the protocol specifications. The operational procedures are consistent with previously published research.²⁷

Statistical analysis

Clinical characteristics categorized as nominal variables were expressed as frequency counts and percentage values (n(%)) and analyzed with either Pearson's chi-square test or Fisher's exact probability test based on data suitability. Metric variables underwent initial normality assessment using statistical tests. Quantities demonstrating Gaussian distribution were reported as arithmetic mean ± standard deviation (x̄ ± s) and evaluated through an independent-samples t-test for group differences. Non-normally distributed measurements were characterized by median values with interquartile ranges [M (Q1-Q3)] and assessed using the nonparametric Mann-Whitney U test. Predictive relationships between variables were examined through general linear modeling. Diagnostic performance metrics, including discriminative power, were quantified using receiver operating characteristic (ROC) analysis with corresponding area under the curve (AUC) values, while optimal classification thresholds were established through Youden's index maximization.

Results

Comparison of clinical characteristics between the CI and CN groups

The CI cohort comprised 72 participants (64.3%), while the CN group included 40 individuals (35.7%). Table 1 details the comparative demographic and clinical characteristics across both cohorts. Statistical analysis revealed no significant intergroup variations regarding gender distribution, age range, marital status, educational attainment, body composition metrics (body mass index, skeletal muscle mass, body fat percentage, visceral adipose tissue, waist-hip ratio), or comorbid conditions, including hypertension, hepatic steatosis, and diabetes mellitus (all p > 0.05). Similarly, psychological assessment scores (SAS, SDS, PSQI, AD8) showed comparable results between groups. Notably, marked disparities emerged in cognitive evaluation outcomes, with the CI group demonstrating significantly lower Mini-Mental State Examination (MMSE) (p < 0.001) and MoCA (p < 0.001) scores compared to the CN group.

Table 1.

Demographic and clinical characteristics of the CI and CN groups.

Characteristics	CI group (n = 72)	CN group (40)	Z/х²/F	p
Gender, males, n (%)	36 (50.0)	20 (50.0)	NA	NA
Age, y	61.0 (56.0, 68.5)	57.5 (54.0, 67.8)	−1.086	0.278^a
Married, n (%)	66 (91.8)	39 (97.5)	1.493	0.222^b
Education, y	12.0 (9.0, 15.0)	15.0 (9.0, 15.0)	−1.604	0.109^a
Body mass index	25.9 ± 3.7	25.7 ± 3.5	0.185	0.668^c
Skeletal muscles, kg	25.1 (19.6, 30.5)	24.6 (19.0, 30.2)	−0.036	0.971^a
Body fat percentage, %	33.3 ± 6.6	32.6 ± 7.8	1.697	0.195^c
Visceral fat area, cm²	115.7 ± 32.3	113.6 ± 40.4	3.685	0.057^c
Waist-hip rate	0.9 (0.8, 0.9)	0.9 (0.8, 0.9)	−0.036	0.971^a
Hypertension, n (%)	22 (30.6)	12 (30.0)	0.004	0.951^b
Fatty liver, n (%)	24 (33.3)	15 (37.5)	0.197	0.657^b
Diabetes, n (%)	8 (11.1)	5 (12.5)	NA	0.526^d
SAS	37.0 (33.0, 40.0)	36.5 (32.0, 41.0)	−0.195	0.845^a
SDS	42.0 (37.0, 44.0)	37.5 (32.3, 45.0)	−1.655	0.098^a
PSQI	6.0 (4.0, 7.0)	5.0 (4.0, 8.5)	−0.191	0.849^a
AD8	1.0 (1.0, 2.0)	1.0 (1.0, 2.0)	−0.048	0.962^a
MMSE	25.0 (22.0, 28.0)	29.0 (28.0, 29.0)	−5.993	<0.001^a
MoCA	20.0 (16.0, 22.0)	27.0 (26.0, 28.8)	−8.768	<0.001^a

SAS: Self-Rating Anxiety Scale; SDS: Self-Rating Depression Scale; PSQI: Pittsburgh Sleep Quality Index; AD8: Aging and dementia−8; MMSE: Mini-Mental State Examination; MoCA: Montreal Cognitive Assessment; p^a indicates Mann-Whitney U test; p^b indicates Chi-squared test; p^c indicates Independent-samples t test; p^d indicates Fisher's exact test.

Comparison of glucose and lipid metabolism indicators between the CI and CN groups

The analysis revealed no notable variations in glucose and lipid metabolism parameters between CI and CN groups, encompassing fasting glucose, hemoglobin A1c (GH), triglyceride levels (TG), total cholesterol (TC), high- and low-density lipoproteins (HDL/LDL), and homocysteine concentrations (HCY), with all measurements showing p-values exceeding 0.05 as detailed in Table 2.

Table 2.

The comparison of glucose and lipid metabolism indicators between the CI and CN groups.

Indicators	CI group (n = 72)	CN group (40)	Z	p
Glucose	5.34 (4.98, 5.70)	5.41 (5.04, 6.00)	−0.489	0.625
GH	5.70 (5.30, 6.00)	5.60 (5.40, 6.05)	−0.192	0.848
TG	1.31 (0.96, 1.95)	1.43 (1.08, 1.83)	−0.437	0.662
TC	4.99 (4.34, 5.76)	5.27 (4.86, 5.74)	−1.208	0.227
HDL	1.36 (1.17, 1.65)	1.39 (1.16, 1.55)	−0.304	0.761
LDL	2.93 (2.16, 3.56)	3.21 (2.87, 3.73)	−1.266	0.205
HCY	10.15 (8.90, 12.88)	9.70 (7.73, 12.15)	−1.867	0.062

GH: glycosylated hemoglobin; TG: triglycerides; TC: total cholesterol; HDL: high-density lipoprotein; LDL: low-density lipoprotein; HCY: homocysteine; Mann-Whitney U test.

Comparisons of inflammatory blood cell markers between the CI and CN groups

No significant disparities across multiple parameters: leukocyte count (WBC), differential counts (LYM, MON, NEU, EOS, BAS), and their respective proportional values (LYM%, MON%, NEU%, EOS%, BAS%), with statistical analysis confirming non-significant differences (p > 0.05) as presented in Supplemental Table 1.

Comparison of AD7c-NTP levels between the CI and CN groups

AD7c-NTP concentrations in CI patients [0.87 (0.69, 1.40) ng/mL] demonstrated marked elevation compared to CN controls [0.47 (0.38, 0.56) ng/mL] (p < 0.001), as illustrated in Figure 1.

Figure 1.

Comparison of AD7c-NTP levels between the CI and CN groups. AD7c-NTP: Alzheimer-associated neuronal thread protein; Mann-Whitney U test.

Correlation between AD7c-NTP and cognitive function scores

Notable inverse associations emerged between AD7c-NTP concentrations and cognitive assessments, with MoCA (r = −0.496, p < 0.001) and MMSE (r = −0.423, p < 0.001) scores both showing strong negative correlations, detailed in Figure 2.

Figure 2.

Correlation between AD7c-NTP and cognitive function scores. (A) Correlation between AD7c-NTP and MoCA score; (B) Correlation between AD7c-NTP and MMSE score. AD7c-NTP: Alzheimer-associated neuronal thread protein; MoCA: Montreal Cognitive Assessment; MMSE: Mini-Mental State Examination.

Comparison of inflammatory cytokines between the CI and CN groups

Analysis revealed comparable concentrations of multiple immune markers (IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, TNF-α; p > 0.05) across groups. Notably, CI subjects exhibited substantially increased IL-2 (p = 0.005), IFN-α (p = 0.008), and IFN-γ (p = 0.010) levels versus controls, as presented in Table 3.

Table 3.

Comparison of inflammatory cytokines between the CI and CN groups.

Indicators	CI group (n = 72)	CN group (40)	Z	p
IL-1β	0.73 (0.48, 1.42)	0.73 (0.21, 0.96)	−1.740	0.082
IL-2	0.91 (0.47, 1.42)	0.47 (0.13, 0.80)	−2.838	0.005
IL-4	0.29 (0.15, 0.72)	0.51 (0.29, 0.72)	−1.142	0.254
IL-5	0.47 (0.42, 0.58)	0.47 (0.43, 0.57)	−0.540	0.589
IL-6	2.57 (1.59, 3.26)	2.27 (1.50, 3.09)	−1.267	0.205
IL-8	5.38 (3.66, 7.11)	4.80 (4.23, 6.53)	−0.415	0.678
IL-10	1.68 (1.09, 2.54)	1.68 (1.09, 2.25)	−0.646	0.518
IL-12p70	1.39 (0.90, 1.83)	1.54 (1.23, 1.83)	−1.606	0.108
IL-17A	2.50 (2.08, 3.73)	2.92 (1.63, 3.73)	−0.183	0.855
TNF-α	0.76 (0.37, 1.16)	0.62 (0.39, 0.89)	−1.520	0.128
IFN-α	0.53 (0.21, 0.86)	0.21 (0.12, 0.53)	−2.657	0.008
IFN-γ	0.51 (0.26, 0.78)	0.40 (0.19, 0.55)	−2.590	0.010

IL-1β: Interleukin-1β; IL-2: Interleukin-2; IL-4: Interleukin-4; IL-5: Interleukin-5; IL-6: Interleukin-6; IL-8: Interleukin-8; IL-10: Interleukin-10; IL-12p70: Interleukin-12p70; IL-17A: Interleukin-17A; TNF-α: Tumor necrosis factor-α; IFN-α,:Interferon-α; IFN-γ: Interferon-γ; Mann-Whitney U test.

Relationship between AD7c-NTP and inflammatory cytokines

Strong positive associations emerged between AD7c-NTP concentrations and both IL-2 (r = 0.492, p < 0.001) and IFN-α (r = 0.492, p < 0.001). Conversely, statistical analysis revealed no meaningful relationship between AD7c-NTP measurements and IFN-γ levels (p > 0.05), as illustrated in Figure 3.

Figure 3.

Correlation between AD7c-NTP and inflammatory cytokines in the CI and CN groups. (A) AD7c-NTP and IL-2 ; (B) AD7c-NTP and IFN-α; (C) AD7c-NTP and IFN-γ; AD7c-NTP: Alzheimer-associated neuronal thread protein; IL-2: Interleukin-2; IFN-α: Interferon-α; IFN-γ: Interferon-γ.

Diagnostic performance of AD7c-NTP and inflammatory cytokines

The ability of AD7c-NTP to distinguish between CI and CN cases, either independently or when paired with individual inflammatory cytokines, showed satisfactory diagnostic accuracy. When evaluating discriminant validity, the combinations ranked from highest to lowest were: AD7c-NTP combined with IFN-γ, followed by AD7c-NTP with IL-2, then AD7c-NTP with IFN-α, and finally AD7c-NTP alone, achieving AUC values of 0.9271, 0.9076, 0.9066, and 0.9007, respectively (Figure 4A). Integrating AD7c-NTP with any inflammatory biomarker (IFN-γ, IL-2, or IFN-α) yielded superior diagnostic outcomes compared to using AD7c-NTP in isolation. The most effective discrimination was observed when combining all four markers (AD7c-NTP, IL-2, IFN-α, and IFN-γ), which produced an AUC of 0.9774 along with 94% sensitivity and 75% specificity (Figure 4B).

Figure 4.

Receiver operating characteristic (ROC) curve analyses of AD7c-NTP and inflammatory cytokines in the CI and CN groups. (A) Single biomarker (AD7c-NTP) and dual composite biomarkers (AD7c-NTP + IL-2, AD7c-NTP + IFN-α, AD7c-NTP + IFN-γ) were used as predictors; (B) Four-component composite biomarker (AD7c-NTP + IL-2 + IFN-α + IFN-γ) was used as a predictor; AD7c-NTP: Alzheimer-associated neuronal thread protein; IL-2: Interleukin-2; IFN-α: Interferon-α; IFN-γ: Interferon-γ.

Discussion

It is widely recognized that AD-related neuropathology initiates decades before the manifestation of measurable cognitive deficits, often reaching irreversible stages before clinical detection. Consequently, scientific investigations have redirected efforts toward postponing dementia onset in preclinical populations. MCI, characterized by measurable deterioration in cognitive performance from baseline levels through both subjective reporting and standardized assessment, serves as the intermediary phase between regular age-related cognitive changes and diagnosable dementia.²⁸ Researchers and clinicians consider MCI a pivotal transitional phase where therapeutic interventions might effectively decelerate dementia progression.²⁹ Current clinical practice and research protocols predominantly utilize the MMSE and MoCA as primary screening instruments for cognitive dysfunction.^30–32 Our investigation employed comprehensive clinical evaluations to stratify participants into cognitively impaired (CI) and cognitively normal (CN) cohorts.

AD7c-NTP, a quantifiable urinary biomarker, functions as a supplementary diagnostic tool for AD.^17,33 Our investigation demonstrated markedly elevated AD7c-NTP concentrations in cognitively impaired subjects relative to cognitively normal controls. Longitudinal investigations have documented comparable biomarker distribution patterns between MCI patients and healthy populations.¹⁴ Research by Li and colleagues indicated the limited diagnostic utility of urinary AD7c-NTP in cases of subjective cognitive deterioration.³⁴ Conversely, Zhang's team revealed this biomarker demonstrated 91.7% specificity (95% CI: 59.8–99.6%) and 72.2% sensitivity (95% CI: 46.4–89.3%) for predicting Aβ accumulation in cognitively compromised individuals.¹⁵ Notably, our analysis identified an inverse relationship between AD7c-NTP concentrations and neuropsychological assessment results (MoCA/MMSE), corroborating existing scientific literature.^35,36

AD is extensively acknowledged as a neurological condition linked to neuroinflammatory processes.^5,37 Functioning as central elements within the immune network, cytokines act as vital intermediaries in neuroinflammatory responses and hold substantial importance in AD progression. Research indicates that astrocyte and microglial activation by Aβ triggers the release of multiple inflammatory mediators, including TNF-α, various interleukins (IL-1β, IL-2, IL-6, IL-12), and IFN-γ, which collectively drive AD-related inflammatory cascades.^38,39 Nevertheless, the precise mechanisms through which these cytokines influence neural degeneration or cognitive deficits, along with their interplay with AD's pathological components, require further elucidation. Following an extensive analysis of existing literature exploring AD7c-NTP's connection with inflammatory responses, our investigation quantified plasma concentrations of twelve inflammatory markers. The CI group exhibited marked elevations in three specific cytokines—IL-2, IFN-α, and IFN-γ—while the remaining nine parameters (IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17A, and TNF-α) showed no statistically significant variations across study groups.

Elevated concentrations of AD7c-NTP exhibited marked positive associations with IL-2 and IFN-α measurements, while TNF-α displayed comparable patterns of co-variation. As an anti-inflammatory cytokine, IL-2 adopts a four-helical bundle structure that interacts with receptor components including IL-2Rα (CD25), IL-2Rβ (CD122), and the standard gamma chain (γc/CD132).⁴⁰ The interferon (IFN) cytokine family has emerged as a potential therapeutic candidate for managing neurodegenerative disorders.⁴¹ Research indicates these molecules mediate biological responses through interferon α/β receptor engagement, initiating JAK-STAT cascade activation that triggers phosphorylation events in STAT1 and STAT2.⁴² Pathological alterations in JAK-STAT signaling have been associated with AD pathogenesis, possibly influencing synaptic function and neuronal viability.⁴³ While IL-2 and IFN-γ function as characteristic Th1-associated cytokines, IFN-α maintains crucial functions in antiviral responses and immunological homeostasis. Notably, our investigation revealed elevated circulating levels of IL-2, IFN-α, and IFN-γ in patients with cognitive impairment versus cognitively normal controls. Longitudinal cohort research indicates that IL-2 in blood plasma shows remarkable diagnostic potential for detecting cognitive impairment, particularly in amnestic mild cognitive impairment cases, where it achieved an 85.7% AUC score.⁴⁴ Investigations by Bermejo et al. revealed increased IFN-α concentrations in MCI patients relative to healthy controls, though comparable levels were noted between MCI and AD groups.⁴⁵ Separate findings corroborate our observations, demonstrating that heightened IFN-γ levels correlate with reduced cognitive deterioration rates independent of Aβ pathology.⁴⁶ Mirroring prior research,^14,47 our analysis confirms AD7c-NTP's standalone effectiveness in differentiating cognitively impaired individuals from normal controls (AUC = 0.9007). While no solitary biomarker comprehensively tracks AD progression, integrating AD7c-NTP with IL-2 and interferon subtypes enhances the detection of cognitive impairment (AUC = 0.9274).

This investigation represents the first exploration of the interaction between AD7c-NTP and cytokine profiles. Building upon prior cellular and animal research, the identified connections with IL-2, IFN-α, and IFN-γ could indicate previously unrecognized biochemical pathways relevant to AD pathogenesis. These findings underscore the necessity for expanded investigations into these molecular interactions.

Limitations

Several constraints should be acknowledged. First, the research was constrained by a relatively small cohort size, which precluded stratified analyses according to inflammatory status. Second, incorporating more core biomarkers of AD into the inclusion and exclusion criteria will significantly enhance the rigor of this research. Third, while demonstrating significant associations, this work establishes correlational rather than causal links between AD7c-NTP and cytokine levels, necessitating mechanistic studies for validation.

Conclusions

This research explored the interplay between AD7c-NTP and cytokine biomarkers, proposing that the combined evaluation of IL-2, IFN-α, and IFN-γ with AD7c-NTP could form an enhanced supplementary diagnostic framework for detecting early-stage cognitive decline.

Supplemental Material

sj-docx-1-alz-10.1177_13872877251389239 - Supplemental material for The association between Alzheimer-associated neuronal thread protein (AD7c-NTP) and plasma inflammatory cytokine levels in individuals with cognitive impairment and demographically-matched controls

Supplemental material, sj-docx-1-alz-10.1177_13872877251389239 for The association between Alzheimer-associated neuronal thread protein (AD7c-NTP) and plasma inflammatory cytokine levels in individuals with cognitive impairment and demographically-matched controls by He Jin, Jing He, Wei Zhang, Yuxin Wang, Yanchuan Wu, Xuemin Wang, Jing Zhao, Xi Chu and Rong Wang in Journal of Alzheimer's Disease

Footnotes

Acknowledgements

We thank all participants of this study.

Ethical considerations

The institutional ethics committee rigorously evaluated and authorized all research protocols (Ethics Approval Code: LINYANSHEN [2024] 136–001).

Consent to participate

Written informed consent was obtained from each participant following a detailed explanation of the study procedures.

Consent for publication

All participants provided written consent for publication.

Author contribution(s)

He Jin: Conceptualization; Funding acquisition; Investigation; Writing – original draft.

Jing He: Data curation; Methodology.

Wei Zhang: Formal analysis; Software.

Yuxin Wang: Conceptualization; Writing – review & editing.

Yanchuan Wu: Methodology.

Xuemin Wang: Conceptualization; Writing – review & editing.

Jing Zhao: Methodology.

Xi Chu: Investigation; Project administration; Resources.

Rong Wang: Conceptualization; Writing – review & editing.

Funding

The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This article was supported by the National Natural Science Foundation of China (grant no. 82160258), Guangxi Natural Science Foundation (grant no. 2023GXNSFBA026173), Guilin technology application and promotion plan (grant no. 202201397-4), and National Key Research and Development Program of China (grant no. 2022YFC2403504).

Declaration of conflicting interests

The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Data availability statement

The data supporting the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions.

Supplemental material

Supplemental material for this article is available online.

References

Sengoku

. Aging and Alzheimer's disease pathology. Neuropathology 2020; 40: 22–29.

Vos

Xiong

Visser

, et al. Preclinical Alzheimer's disease and its outcome: a longitudinal cohort study. Lancet Neurol 2013; 12: 957–965.

Jia

Ning

Chen

, et al. Biomarker changes during 20 years preceding Alzheimer's disease. N Engl J Med 2024; 390: 712–722.

Heneka

van der Flier

Jessen

, et al. Neuroinflammation in Alzheimer disease. Nat Rev Immunol 2025; 25: 321–352.

Rajesh

Kanneganti

. Innate immune cell death in neuroinflammation and Alzheimer's disease. Cells 2022; 11: 1885.

Leng

Edison

. Neuroinflammation and microglial activation in Alzheimer disease: where do we go from here? Nat Rev Neurol 2021; 17: 157–172.

Thakur

Dhapola

Sarma

, et al. Neuroinflammation in Alzheimer's disease: current progress in molecular signaling and therapeutics. Inflammation 2023; 46: 1–17.

Wang

Zong

Cui

, et al. The effects of microglia-associated neuroinflammation on Alzheimer's disease. Front Immunol 2023; 14: 1117172.

Singh

. Astrocytic and microglial cells as the modulators of neuroinflammation in Alzheimer's disease. J Neuroinflammation 2022; 19: 206.

10.

Monte

Ghanbari

Frey

, et al. Characterization of the AD7C-NTP cDNA expression in Alzheimer's disease and measurement of a 41-kD protein in cerebrospinal fluid. J Clin Invest 1997; 100: 3093–3104.

11.

Ghanbari

Beheshti

, et al. Biochemical assay for AD7C-NTP in urine as an Alzheimer's disease marker. J Clin Lab Anal 1998; 12: 285–288.

12.

Munzar

Levy

Rush

, et al. Clinical study of a urinary competitve ELISA for neural thread protein in Alzheimer disease. Neurol Clin Neurophysiol 2002; 2002: 2–8.

13.

Goodman

Golden

Flitman

, et al. A multi-center blinded prospective study of urine neural thread protein measurements in patients with suspected Alzheimer's disease. J Am Med Dir Assoc 2007; 8: 21–30.

14.

Chen

Wang

, et al. The level of Alzheimer-associated neuronal thread protein in urine may be an important biomarker of mild cognitive impairment. J Clin Neurosci 2015; 22: 649–652.

15.

Zhang

, et al. Urine AD7c-NTP predicts amyloid deposition and symptom of agitation in patients with Alzheimer's disease and mild cognitive impairment. J Alzheimers Dis 2017; 60: 87–95.

16.

Jack

Jr Andrews

Beach

, et al. Revised criteria for diagnosis and staging of Alzheimer's disease: Alzheimer's Association Workgroup. Alzheimers Dement 2024; 20: 5143–5169.

17.

Jin

Wang

. Alzheimer-associated neuronal thread protein: research course and prospects for the future. J Alzheimers Dis 2021; 80: 963–971.

18.

de la Monte

Wands

. Neurodegeneration changes in primary central nervous system neurons transfected with the Alzheimer-associated neuronal thread protein gene. Cell Mol Life Sci 2001; 58: 844–849.

19.

de la Monte

Wands

. Alzheimer-associated neuronal thread protein-induced apoptosis and impaired mitochondrial function in human central nervous system-derived neuronal cells. J Neuropathol Exp Neurol 2001; 60: 195–207.

20.

Chen

. Tau and neuroinflammation in Alzheimer's disease: interplay mechanisms and clinical translation. J Neuroinflammation 2023; 20: 165.

21.

Chobanian

Bakris

Black

, et al. The seventh report of the joint national committee on prevention, detection, evaluation, and treatment of high blood pressure: the JNC 7 report. JAMA 2003; 289: 2560–2572.

22.

American Diabetes Association . Standards of medical care in diabetes–2011. Diabetes Care 2011; 34: S11–S61.

23.

Eslam

Newsome

Sarin

, et al. A new definition for metabolic dysfunction-associated fatty liver disease: an international expert consensus statement. J Hepatol 2020; 73: 202–209.

24.

Guo

Huang

. Hospital anxiety and depression scale exhibits good consistency but shorter assessment time than Zung self-rating anxiety/depression scale for evaluating anxiety/depression in non-small cell lung cancer. Medicine (Baltimore) 2021; 100: e24428.

25.

Zitser

Allen

Falgàs

, et al. Pittsburgh Sleep Quality Index (PSQI) responses are modulated by total sleep time and wake after sleep onset in healthy older adults. PLoS One 2022; 17: e0270095.

26.

Usarel

Dokuzlar

Aydin

, et al. The AD8 (Dementia Screening Interview) is a valid and reliable screening scale not only for dementia but also for mild cognitive impairment in the Turkish geriatric outpatients. Int Psychogeriatr 2019; 31: 223–229.

27.

Jin

Yang

Chen

, et al. Effects of hepatorenal function on urinary Alzheimer-associated neuronal thread protein: a laboratory-based cross-sectional study among the older Chinese population. J Alzheimers Dis 2024; 100: 911–921.

28.

Albert

DeKosky

Dickson

, et al. The diagnosis of mild cognitive impairment due to Alzheimer's disease: recommendations from the National Institute on Aging-Alzheimer's Association workgroups on diagnostic guidelines for Alzheimer's disease. Alzheimers Dement 2011; 7: 270–279.

29.

Winblad

Amouyel

Andrieu

, et al. Defeating Alzheimer's disease and other dementias: a priority for European science and society. Lancet Neurol 2016; 15: 455–532.

30.

Arevalo-Rodriguez

Smailagic

Roqué-Figuls

, et al. Mini-Mental State Examination (MMSE) for the early detection of dementia in people with mild cognitive impairment (MCI). Cochrane Database Syst Rev 2021; 7: CD010783.

31.

Zhuang

Yang

Gao

. Cognitive assessment tools for mild cognitive impairment screening. J Neurol 2021; 268: 1615–1622.

32.

Jia

Wang

Huang

, et al. A comparison of the Mini-Mental State Examination (MMSE) with the Montreal Cognitive Assessment (MoCA) for mild cognitive impairment screening in Chinese middle-aged and older population: a cross-sectional study. BMC Psychiatry 2021; 21: 485.

33.

Wang

Han

, et al. Development of a novel urine Alzheimer-associated neuronal thread protein ELISA kit and its potential use in the diagnosis of Alzheimer's disease. J Clin Lab Anal 2016; 30: 308–314.

34.

Kang

Wang

, et al. Urinary Alzheimer-associated neuronal thread protein is not elevated in patients with subjective cognitive decline and patients with depressive state. J Alzheimers Dis 2019; 71: 1115–1123.

35.

Guo

Kang

Hui

, et al. The association between cognition of obstructive sleep apnea patients and urinary AD7c-NTP level: investigation and application. J Alzheimers Dis 2022; 90: 1215–1231.

36.

Dai

Wei

, et al. Urinary AD7c-NTP evaluates cognition impairment and differentially diagnoses AD and MCI. Am J Alzheimers Dis Other Demen 2022; 37: 15333175221115247.

37.

Mangalmurti

Lukens

. How neurons die in Alzheimer's disease: implications for neuroinflammation. Curr Opin Neurobiol 2022; 75: 102575.

38.

Khemka

Ganguly

Bagchi

, et al. Raised serum proinflammatory cytokines in Alzheimer's disease with depression. Aging Dis 2014; 5: 170–176.

39.

Sun

Wei

, et al. Plasma β-amyloid, tau, neurodegeneration biomarkers and inflammatory factors of probable Alzheimer's disease dementia in Chinese individuals. Front Aging Neurosci 2022; 14: 963845.

40.

Abbas

Trotta

R Simeonov

, et al. Revisiting IL-2: biology and therapeutic prospects. Sci Immunol 2018; 3: eaat1482.

41.

Hui

BSM

Zhi

Retinasamy

, et al. The role of interferon-α in neurodegenerative diseases: a systematic review. J Alzheimers Dis 2023; 94: S45–S66.

42.

Schreiber

Piehler

. The molecular basis for functional plasticity in type I interferon signaling. Trends Immunol 2015; 36: 139–149.

43.

Nicolas

Amici

Bortolotto

, et al. The role of JAK-STAT signaling within the CNS. JAKSTAT 2013; 2: e22925.

44.

Liang

Tsai

Lin

, et al. Better identification of cognitive decline with interleukin-2 than with amyloid and tau protein biomarkers in amnestic mild cognitive impairment. Front Aging Neurosci 2021; 13: 670115.

45.

Bermejo

Martín-Aragón

Benedí

, et al. Differences of peripheral inflammatory markers between mild cognitive impairment and Alzheimer's disease. Immunol Lett 2008; 117: 198–202.

46.

Yang

Zhang

Carlyle

, et al. Plasma IL-12/IFN-γ axis predicts cognitive trajectories in cognitively unimpaired older adults. Alzheimers Dement 2022; 18: 645–653.

47.

Kim

, et al. Comparison of urinary Alzheimer-associated neural thread protein (AD7c-NTP) levels between patients with amnestic and nonamnestic mild cognitive impairment. Am J Alzheimers Dis Other Demen 2020; 35: 1533317519880369.

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