Application of biomarker in preparation of product for early diagnosis of cognitive impairment of sleep apnea patient

Abstract

The invention relates to the technical field of biomedicine, in particular to application of a biomarker in preparation of a product for early diagnosis of cognitive impairment of a sleep apnea patient. The biomarker is a combination of pho-IRS-1 and pS312-IRS-1, can accurately diagnose cognitive impairment of a sleep apnea patient in an early stage, and provides a more timely clinical strategy.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technologies, and in particular to the application of biomarkers in the preparation of products for early diagnosis of cognitive impairment in patients with sleep apnea. Background Art

[0002] Obstructive sleep apnea (OSA) is the most common sleep-related breathing disorder, which is characterized by recurrent upper airway closure during sleep. The overall prevalence of OSA in the population is 9%-38%, and the prevalence is higher in men. OSA has a relatively high prevalence in patients with mild cognitive impairment (MCI). Intermittent hypoxia, sleep fragmentation, and inflammatory activation are the main pathophysiological mechanisms by which OSA ultimately leads to MCI symptoms and even Alzheimer's disease (AD) pathology.

[0003] Multiple studies have shown that OSA is independently associated with reduced insulin sensitivity and increased insulin resistance, and significantly affects the brain and memory formation, toxin excretion, hormone function, and cognitive function. In patients with OSA, the occurrence of sleep apnea leads to a decrease in blood oxygen saturation and tissue hypoxia. Insulin resistance is an adaptive response to tissue hypoxia, which develops under conditions of limited tissue oxygen supply and plays an important role in the pathogenesis of OSA. Insulin resistance, inflammation, and metabolic syndrome increase with the severity of OSA. Animal studies have shown that intermittent hypoxia alters plasma exosome load, induces adipocyte dysfunction (increased insulin resistance), and improves with physical activity.

[0004] The pathophysiological process of hypoxia-induced insulin resistance includes the upregulation of hypoxia-inducible factor-1 and the downregulation of peroxisome proliferator-activated receptor-gamma (PPARγ), and the latter is involved in the regulation of peroxisome proliferator-activated receptor-gamma coactivator-1-alpha (PGC1α). The molecular activity driving insulin signal transduction is controlled by the tyrosine phosphorylation of the insulin receptor β-subunit (IRβ) and insulin receptor substrates (IRS)-1 and IRS-2. It is currently believed that insulin resistance is caused by reduced insulin receptor signal transduction, attributed to its abnormal association with more phospho-serine-IRS-1 and less phospho-tyrosine-IRS-1. Phosphorylation of serine site 312 of the first node IRS-1 (pS312-IRS-1) is a classical insulin signaling pathway, which can affect glucose and lipid homeostasis, and the levels of downstream messengers pS312-IRS-1 or pS616-IRS-1 are indicators of insulin resistance.

[0005] Although many studies have explored insulin resistance in OSA, there is a lack of assessment based on central insulin resistance. Neuron-derived exosomes (NDEs), as a non-invasive method for the diagnosis of many central nervous system diseases, indicate pathological changes in the central nervous system and can be used as biomarkers for cognitive impairment. In this study, we aimed to evaluate the relationship between OSA cognitive dysfunction and peripheral and central insulin resistance through the triglyceride-glucose (TyG) index and the NDE levels of PGC1α, phospho-IRS-1, and pS312-IRS-1.

[0006] References:

[0007] [1] Berry RB, Albertario CL, Harding SM, et al. The AASM Manual for the Scoring of Sleep and Associated Events: Rules, Terminology and Technical Specifications. Version 2.5. Darien, IL:

[0008] American Academy of Sleep Medicine.2018。

[0009] [2]Li M,Sun H,Shen T,et al.Increased serum levels of cyclophilin aand matrixmetalloproteinase-9are associated with cognitive impairment inpatients with obstructivesleep apnea.Sleep Med.2022;93:75-83。

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[0011] [4]Tahapary DL,Pratisthita LB,Fitri NA,et al.Challenges in thediagnosis of insulin resistance:

[0012] Focusing on the role of HOMA-IR and Tryglyceride / glucoseindex.Diabetes Metab Syndr.

[0013] 2022,16(8):102581。

[0014] [5]Nasreddine ZS,Phillips NA,Bédirian V,et al.The Montreal CognitiveAssessment,MoCA:abrief screening tool for mild cognitive impairment.J AmGeriatr Soc.2005;53(4):695-699。

[0015] [6]Winblad B, Palmer K, Kivipelto M, et al. Mild cognitive impairment--beyond controversies, towards a consensus: report of the International Working Group on Mild Cognitive Impairment. J Intern Med. 2004;256(3):240-246.

[0016] [7]Nogueras-Ortiz CJ, Eren E, Yao P, Calzada E, et al. Single-extracellular vesicle (EV) analyses validate the use of L1 Cell Adhesion Molecule (L1CAM) as a reliable biomarker of neuron-derived EVs. J Extracell Vesicles. 2024;13(6):e12459.

[0017] [8]Li M, Sun Z, Sun H, et al. Paroxysmal slow wave events are associated with cognitive impairment in patients with obstructive sleep apnea. Alzheimers Res Ther. 2022,14(1):200.

[0018] [9]Li M, Shen T, Yao R, et al. Mitochondrial dysfunction is associated with cognitive impairment in adults with OSA without dementia. Sleep Med. 2024,119:234-243. Summary of the Invention

[0019] The present invention provides the use of a biomarker in the preparation of a product for early diagnosis of cognitive impairment in patients with sleep apnea, which can accurately diagnose cognitive impairment in patients with sleep apnea at an early stage, provide more timely clinical strategies, and solve the problems existing in the prior art.

[0020] The technical solution adopted by the present invention is:

[0021] Use of a biomarker in the preparation of a product for early diagnosis of cognitive impairment in patients with sleep apnea, wherein the biomarker is a combination of phospho-IRS-1 and pS312-IRS-1.

[0022] Furthermore, the biomarker phospho-IRS-1 and pS312-IRS-1 are derived from neurogenic exosomes.

[0023] Furthermore, by detecting the expression levels of the above biomarkers, it is possible to early diagnose whether a patient with sleep apnea has mild cognitive impairment due to sleep apnea.

[0024] Furthermore, the expression levels of phospho-IRS-1 and pS312-IRS-1 in patients with mild cognitive impairment due to sleep apnea are significantly higher than those of these two indicators in patients without mild cognitive impairment due to sleep apnea.

[0025] Furthermore, a high expression level of phospho-IRS-1 and pS312-IRS-1 means that the average CD63-normalized NDE level of phospho-IRS-1 is not lower than 13.22 ± 3.38 mg / ml and the average CD63-normalized NDE level of pS312-IRS-1 is not lower than 5.08 ± 1.74 U / ml.

[0026] Furthermore, for the above application, by simultaneously detecting the expression levels of phospho-IRS-1 and pS312-IRS-1, a statistical analysis was performed on the value of predicting mild cognitive impairment in patients with sleep apnea. The efficacy of the combined detection is higher, and the sensitivity reaches 81.2% on the premise of ensuring the specificity.

[0027] Furthermore, the product includes a chip or a kit.

[0028] Furthermore, the kit is a PCR kit for detecting phospho-IRS-1 and pS312-IRS-1.

[0029] Advantages of the present invention:

[0030] The present invention can accurately diagnose mild cognitive impairment in patients with sleep apnea at an early stage and provide more timely clinical strategies. By quantitatively measuring the NDE levels of PGC1α, phospho-IRS-1, and pS312-IRS-1 in patients in the OSA+MCI group, OSA-MCI group, and control group, and evaluating the relationship between OSA cognitive dysfunction and peripheral and central insulin resistance using the TyG index, phospho-IRS-1 and pS312-IRS-1 were identified as combined markers capable of accurately diagnosing mild cognitive impairment in patients with sleep apnea. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 Exosome concentrations and TyG index of PGC1α, Phospho-IRS-1, and pS312-IRS-1 in the cross-sectional OSA+MCI group (n = 117), OSA-MCI group (n = 111), and control group (n = 56) of the present invention;

[0032] Figure 2 Receiver operating characteristic (ROC) curve of the present invention.

[0033] Figure 3 Changes in NDE levels of PGC1α, Phospho-IRS-1, and pS312-IRS-1 in OSA patients (n = 45) under different treatment modes of the present invention.

[0034] Among them, Figure 1 A in shows that the level of PGC1α in neuron-derived exosomes in the OSA+MCI group is higher than that in the OSA-MCI group and the control group; B shows that the level of Phospho-IRS-1 in neuron-derived exosomes in the OSA+MCI group is higher than that in the OSA-MCI group and the control group; C shows that the levels of pS312-IRS-1 in neuron-derived exosomes in the OSA+MCI group, OSA-MCI group, and control group gradually decrease; D shows that the TyG index in the OSA+MCI group is higher than that in the OSA-MCI group and the control group.

[0035] Figure 3 A in is the change in NDE levels of PGC1α, Phospho-IRS-1, and pS312-IRS-1 before and after CPAP treatment (n = 23) in OSA patients; B is the change in NDE levels of PGC1α, Phospho-IRS-1, and pS312-IRS-1 before and after weight loss (n = 22) in OSA patients. DETAILED DESCRIPTION OF THE INVENTION

[0036] To clearly illustrate the technical features of the present solution, the present invention will be described in detail below through specific embodiments in conjunction with the accompanying drawings.

[0037] I. Clinical Trials

[0038] This invention recruited patients who visited the Sleep Medicine Center of Weihai Municipal Hospital from October 2019 to September 2022 and were clinically considered to have sleep-disordered breathing. Matched controls in terms of age and gender were recruited from the community. Before study enrollment, all participants gave informed consent. This study was approved by the institutional ethics committee.

[0039] Inclusion criteria: (1) Aged 35 - 80 years old; (2) Untreated; (3) Mini-mental state examination (MMSE) score ≥ 24 points; (4) No major surgery or inflammatory diseases (such as pneumonia, gastroenteritis, arthritis, connective tissue inflammation, etc.) within the past 3 months.

[0040] Exclusion criteria: (1) Severe neurological diseases, such as acute stroke, intracranial infection, neurodegenerative diseases, epilepsy, brain trauma, brain tumor, etc.; (2) Severe psychological disorders, such as severe depression, schizophrenia, etc.; (3) Other systemic diseases, such as hyperthyroidism, liver failure, chronic renal failure, autoimmune diseases, systemic steroid treatment, hormone replacement therapy; (4) Long-term use of drugs that interfere with cognition or sleep (such as antihistamines, benzodiazepines, tricyclic antidepressants, donepezil, etc.); (5) Pregnancy; (6) Central sleep apnea; (7) Rapid eye movement (REM) sleep behavior disorder (RBD) or REM sleep without dystonia (RSWA) and restless legs syndrome.

[0041] Demographics, polysomnography (PSG), and cognitive scale measurements were completed. The subjects were divided into three groups: OSA + MCI group (apnea hypopnea index (AHI) > 5 times / hour, Montreal Cognitive Assessment (MoCA) < 26), OSA - MCI group (AHI > 5 times / hour, MoCA ≥ 26), and control group (cognitively normal, non - OSA).

[0042] 1. Polysomnography

[0043] All-night PSG (Grael, Compumedics, Australia) was used and conducted in a sleep laboratory from 9 p.m. to 6 a.m. the next morning. The PSG consisted of a continuous polysomnograph, and the monitored contents included electroencephalogram (F3-M2, F4-M1, C3-M2, C4-M1, T3-M2, T4-M1, O1-M2, O2-M1), electrooculogram, electromyogram of the masseter muscle, electromyogram of the anterior tibial muscle, electrocardiogram, thermistor probe, nasal pressure sensor, thoracic and abdominal respiratory movement recording, finger pulse oximetry sensor, microphone, and sleep position. The standard leads recommended by version 2.5 of the American Academy of Sleep Medicine (AASM) were used.

[0044] The PSG variables analyzed included sleep latency, total sleep time (TST), sleep efficiency, proportion of sleep stages (stage 1 of nonrapid eye movement (NREM) sleep (N1), stage 2 of NREM sleep (N2), stage 3 of NREM sleep (N3), REM sleep (R)), AHI, oxygen desaturation index (ODI), mean oxygen saturation during sleep (meanSaO2), minimum oxygen saturation during sleep (minSaO2), arousal index, percentage of sleep time with oxygen saturation <90% (T90). The definition of OSA was based on our published research [1、2] . The severity of OSA was classified as mild (AHI ≥ 5 to <15), moderate (AHI ≥ 15 to <30), and severe (AHI ≥ 30) [3] .

[0045] In the morning after PSG, blood pressure was measured in the supine position when waking up at 6 - 7 a.m. Then venous blood was taken to measure total cholesterol, triglycerides, low-density lipoprotein cholesterol (LDL-C), and glucose. The TyG index was used as a surrogate biomarker for peripheral insulin resistance [4] , and the calculation formula was ln[fasting triglycerides (mg / dl) × fasting blood glucose (mg / dl) / 2].

[0046] 2. Assessment of cognition

[0047] Each subject underwent a face-to-face standardized diagnostic interview and neurological examination by a trained neurologist using the MMSE and the Beijing version of the MoCA. The psychosocial status was evaluated using the Hamilton Anxiety Scale (HAMA) and the Hamilton Depression Scale (HAMD)-17. Daytime sleepiness was evaluated using the Epworth Sleepiness Scale (ESS).

[0048] The MoCA score ranges from 0 to 30 points and includes visual executive function, naming, attention, language, abstraction, delayed recall, and orientation. MCI was defined as MoCA < 26 points [5] and met the revised Mayo Clinic criteria [6] : (1) subjective cognitive impairment; (2) the presence of objective impairment in one or more cognitive domains; (3) mild impairment of complex instrumental activities of daily living, but preservation of independent activities of daily living; (4) no dementia.

[0049] 3. Isolation of NDEs and quantification of PGC1α, phospho-IRS-1, and pS312-IRS-1

[0050] Venous blood was collected at 6 - 7 am. The blood samples were centrifuged at 4000×g for 10 minutes within 30 minutes after collection and immediately stored at -80°C. Nogueras-Ortiz and colleagues provided evidence supporting the use of L1CAM as a target for isolating plasma NDEs and using it to identify biomarkers reflecting neuronal function [7] . Specific NDEs were isolated according to our published protocol [8、9] . The enzyme-linked immunosorbent serological assay kits for human PGC1α (CUSABIO, CSB-E11761h, China), PathScan Phospho-IRS-1 (Cell Signaling Technology, 7133C, USA), and IRS-1 [pS312] (Cell Signaling Technology, 7133C, USA) were used to quantify PGC1α, Phospho-IRS-1, and pS312-IRS-1. The protein levels of each sample were normalized to the exosome content labeled with the CD63 exosome marker (RayBio, ELH-CD63, Norcross, GA30092, USA).

[0051] 4. Intervention

[0052] The present invention also compared the effects of weight loss and continuous positive airway pressure (CPAP) on insulin sensitivity in patients with moderate to severe OSA among the aforementioned 228 patients. Patients with moderate to severe OSA who received CPAP treatment or weight loss treatment were followed up for 12 months. The inclusion criteria for the CPAP treatment group were as follows: (1) Good compliance with CPAP treatment, that is, at least 4 hours of use per night and at least 5 nights of use per week; (2) Effective CPAP treatment (AHI < 5 times / hour); (3) Adherence to CPAP treatment for at least 1 year; (4) No significant weight loss in the CPAP group. The inclusion criteria for the weight loss group were to achieve a 5-10% weight loss through a combination of physical activity and diet control.

[0053] Among patients with moderate to severe OSA who underwent weight loss and CPAP treatment, a part of them (n = 23) received CPAP treatment, and another part (n = 22) received weight loss treatment consisting of an individualized lifestyle. After 1 year, each subject completed a cognitive questionnaire and the levels of PGC1α, phospho-IRS-1, and pS312-IRS-1 were measured.

[0054] II. Results

[0055] 2.1 Clinical and polysomnography variables of different groups

[0056] A total of 344 subjects were recruited, and 43 subjects were excluded, including 35 subjects with contraindications and 8 subjects who refused to participate in the study. Among the 301 subjects included in the cross-sectional study, 17 subjects were excluded after PSG and cognitive assessments. Finally, 284 patients were included in this study, with an average age of (57.79 ± 9.62) years. There were 202 male patients (71.13%). The 228 OSA patients were divided into the OSA+MCI group (n = 117) and the OSA-MCI group (n = 111). The remaining 56 were regarded as the control group.

[0057] As shown in Table 1 below, the data of all subjects' demographics and polysomnography variables are presented. There were significant differences in BMI, hypertension, hyperlipidemia, systolic blood pressure, diastolic blood pressure, fasting blood glucose, HAMD score, ESS score, AHI, ODI, TST, sleep efficiency, arousal index, mean SaO2, min SaO2, T90, N1%, N3%, and R% among the three groups (P < 0.001, P = 0.016, P = 0.034, P = 0.028, P = 0.004, P = 0.002, P = 0.037, P < 0.001, P < 0.001, P < 0.001, P = 0.027, P = 0.018, P < 0.001, P < 0.001, P < 0.001, P < 0.001, P < 0.001, P = 0.001, P = 0.001). Among them, hypertension, fasting blood glucose, AHI, ODI, T90, and N1% in the OSA+MCI group were significantly higher than those in the OSA-MCI group and the control group (all P < 0.05). TST and mean SaO2 in the OSA+MCI group were significantly lower than those in the OSA-MCI group and the control group (all P < 0.05). There were no significant differences among the groups in terms of age, gender, education level, alcohol consumption, smoking, total cholesterol, triglyceride, LDL-C, HAMA score, sleep latency, N2%, etc.

[0058] Table 1 Baseline clinical characteristics of the OSA+MCI group, OSA-MCI group, and control group (n = 284)

[0059]

[0060]

[0061]

[0062] a: Compared with the OSA-MCI group, p < 0.05; b: Compared with the control group, p < 0.05. 1 mmHg = 0.133 kPa.

[0063] 2.2 Baseline TyG index and NDE levels of PGC1α, phospho-IRS-1, and pS312-IRS-1 in different groups

[0064] In this invention, the average CD63-normalized NDE levels and TyG indices of PGC1α, phospho-IRS-1, and pS312-IRS-1 in the OSA+MCI group were higher than those in the OSA-MCI group (PGC1α: 283.43±39.17 pg / ml vs 244.32±38.85 pg / ml, P<0.001; phospho-IRS-1: 13.22±3.38 mg / ml vs 10.38±1.66 mg / ml, P<0.001; pS312-IRS-1: 5.08±1.74 U / ml vs 2.63±1.26 U / ml, P<0.001; TyG: 8.92±0.60 vs 8.71±0.62, P = 0.010) and the control group (PGC1α: 283.43±39.17 pg / ml vs 238.03±33.65 pg / ml, P<0.001; phospho-IRS-1: 13.22±3.38 mg / ml vs 10.16±1.28 mg / ml, P<0.001; pS312-IRS-1: 5.08±1.74 U / ml vs 1.37±0.40 U / ml, P<0.001; TyG: 8.92±0.60 vs 8.70±0.53, P = 0.023). And the NDE level of pS312-IRS-1 in the OSA-MCI group was higher than that in the control group (P<0.001). However, there was no significant difference in PGC1α, phospho-IRS-1, and TyG index between the OSA-MCI group and the control group ( Figure 1 ).

[0065] Table 2 shows that after adjusting for age, gender, education level, vascular risk factors (hypertension, diabetes, hyperlipidemia), HAMD score, and AHI, the NDE levels of PGC1α, phospho-IRS-1, pS312-IRS-1, and TyG index were all independent influencing factors for cognitive dysfunction in OSA patients. The increase in PGC1α, phospho-IRS-1, pS312-IRS-1, and TyG index was significantly associated with an increased risk of cognitive impairment in OSA patients (OR: 1.028, 95% CIs: 1.019-1.037, P<0.001; OR: 1.693, 95% CIs:

[0066] 1.379-1.862, P<0.001; OR: 3.153, 95% CIs: 2.320-4.285, P<0.001; OR: 3.096, 95% CIs: 1.108-8.657, P = 0.031).

[0067] Table 2 Logistic regression analysis of OSA patients (n = 228)

[0068]

[0069]

[0070] In Table 2, Model 0: unadjusted. Model 1: adjusted for age, sex, BMI, education level. Model 2: adjusted for age, sex, education level, vascular risk factors (such as hypertension, diabetes, hyperlipidemia, etc.), HAMD score, AHI.

[0071] 2.3 Insulin resistance plays a mediating role between cognitive function and sleep characteristics in OSA patients

[0072] See Table 3. Preliminary correlation analysis showed the correlation between the NDE levels of phospho-IRS-1 and pS312-IRS-1 and the cognitive function of OSA patients. As shown in Table 3, overall cognition was significantly negatively correlated with the NDE levels of phospho-IRS-1 (r = -0.607, P < 0.001) and pS312-IRS-1 (r = -0.626, P < 0.001). Similar results were also found in the correlation analysis of cognitive subdomains, including visual executive function, naming, attention, language, abstraction, delayed recall, and orientation, especially delayed recall (phospho-IRS-1 = -0.493, P < 0.001; pS312-IRS-1 r = -0.570, P < 0.001).

[0073] Table 3 Correlation analysis of the NDE levels of Phospho-IRS-1, pS312-IRS-1, and TyG index with cognitive function in OSA patients (n = 228)

[0074]

[0075]

[0076] 2.4 Diagnostic efficacy of neurogenic exosome phospho-IRS-1 and pS312-IRS-1 for MCI in OSA patients

[0077] Figure 2Receiver operating characteristic (ROC) curve analysis showed that the area under the curve (AUC) for phospho-IRS-1 and pS312-IRS-1 in predicting MCI in OSA patients was 0.780 (95% CIs: 0.718 - 0.841, P < 0.001) and 0.874 (95% CIs: 0.830 - 0.918, P < 0.001), respectively.

[0078] Statistical analysis was performed on the value of the above single markers and the combination of two markers in predicting MCI in OSA patients, as shown in Table 4.

[0079] Table 4 AUC of biomarkers in neuronal-derived exosomes of OSA patients

[0080]

[0081] Cut-off value of phospho-IRS-1: 12.34, sensitivity 59%, specificity 90.1%.

[0082] Cut-off value of pS312-IRS-1: 4.02, sensitivity 73.5%, specificity 88.3%.

[0083] Combined sensitivity 81.2%, specificity 83.8%.

[0084] As shown in the results of Table 4, compared with single markers in predicting MCI in OSA patients, the combination of two markers in predicting MCI in OSA patients has greater value and improves the accuracy of clinical diagnosis.

[0085] 2.5 Effects of weight loss and CPAP treatment on the levels of PGC1α, phospho-IRS-1 and pS312-IRS-1 in OSA patients

[0086] After 12 months of intervention treatment, the levels of PGC1α (254.77±43.39 pg / ml vs 259.10±45.45 pg / ml, P = 0.041), phospho-IRS-1 (11.16±2.68 mg / ml vs 11.83±2.87 mg / ml, P = 0.011) and pS312-IRS-1 (3.38±1.87 U / ml vs 3.75±2.03 U / ml, P = 0.009) in the weight loss group decreased significantly, and MoCA (23.00±3.81 vs 22.64±3.94, P = 0.042) increased significantly. In the CPAP group, except for PGC1α (269.09±45.22 pg / ml vs 273.68±48.90 pg / ml, P = 0.045) and MoCA (25.22±2.70 vs 24.87±2.85, P = 0.029), the levels of phospho-IRS-1 and pS312-IRS-1 did not change significantly( Figure 3 ). The above research found that compared with CPAP treatment, weight loss is more effective in improving central insulin resistance of phospho-IRS-1 and pS312-IRS-1. However, there was no significant difference in the changes of PGC1α, phospho-IRS-1, and pS312-IRS-1 NDE levels between the weight loss group before and after treatment and the CPAP group.

[0087] The above specific implementation manners cannot be used as a limitation on the protection scope of the present invention. For those skilled in the art of this technology, any alternative improvement or transformation made to the implementation manner of the present invention falls within the protection scope of the present invention.

[0088] Matters not described in detail in the present invention are all well-known technologies to those skilled in the art of this technology.

Claims

1. Use of a biomarker in the preparation of a product for early diagnosis of cognitive impairment in patients with sleep apnea, characterized in that, The biomarker is a combination of phospho-IRS-1 and pS312-IRS-1.

2. The application according to claim 1, characterized in that, The sources of the biomarkers phospho-IRS-1 and pS312-IRS-1 are neurogenic exosomes.

3. The application according to claim 1, characterized in that, By detecting the expression levels of the biomarkers, early diagnose whether patients with sleep apnea have mild cognitive impairment due to sleep apnea.

4. The application according to claim 3, characterized in that The expression levels of phospho-IRS-1 and pS312-IRS-1 in patients with mild cognitive impairment due to sleep apnea are significantly higher than those of these two indicators in patients without mild cognitive impairment due to sleep apnea.

5. The application according to claim 4, wherein, High expression levels of phospho-IRS-1 and pS312-IRS-1 mean that the average CD63-normalized NDE level of phospho-IRS-1 is not lower than 13.22 ± 3.38 mg / ml and the average CD63-normalized NDE of pS312-IRS-1 is not lower than 5.08 ± 1.74 U / ml.

6. The application according to claim 1, characterized in that, The cognitive impairment is mild cognitive impairment.

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