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lncRNA six3os1 diagnoses acute stroke, predicts disease severity, and predicts post-stroke cognitive impairment

BMC Neurology volume 24, Article number: 491 (2024) Cite this article

Abstract

Background

Stroke is the main cause of death and disability. Post-stroke cognitive impairment (PSCI) is one of the most severe complications of stroke, which lacks effective biomarkers for its early detection.

Objective

This study evaluated the significance of lncRNA SIX3OS1 in acute stroke and PSCI aiming to identify a novel biomarker.

Patients and methods

The study enrolled 138 patients with acute stroke and 80 healthy individuals. By comparing the serum SIX3OS1 in acute stroke and healthy individuals, the significance of SIX3OS1 in diagnosing acute stroke, assessing disease severity, and predicting the risk of PSCI was revealed.

Results

Significant upregulation of SIX3OS1 in acute stroke was observed, which discriminated patients with acute stroke from healthy individuals and indicated severe disease conditions of patients. There were 72 acute stroke patients who had PSCI accounting for 52.17% that showed a higher serum SIX3OS1 level than post-stroke cognitive normal patients. The increasing serum SIX3OS1 level was also identified as a risk factor for PSCI and could distinguish PSCI patients. Additionally, SIX3OS1 showed a negative correlation with the MoCA score of PSCI patients.

Conclusion

Serum SIX3OS1 level can be considered a biomarker for screening acute stroke and a predictor for PSCI.

Peer Review reports

Introduction

Stroke has become one of the top disabling and fatal diseases in China and seriously threatens human health [1]. Stroke is a focal lesion of the central nervous system and neurological dysfunction caused by the interruption of the blood supply to the brain. Recently, with the development of public health and medical services, the survival of stroke has been greatly improved, and an increasing number of studies have paid attention to the sequela of stroke [2, 3]. Post-stroke cognitive impairment (PSCI) is the main sequela of stroke. It has been reported that about 1/3 of stroke patients experience PSCI, which would lead to spatial disorders, executive dysfunction, memory disorders, attention disorders, and other disorders in activities of daily life [4]. The relationship between stroke and PSCI is complex involving various mechanisms, including impaired areas of cognitive function, increased degeneration caused by hypoxia, neurodegeneration, endothelial dysfunction, damaged blood-brain barrier, and neuroinflammation [5]. Compared with neurodegeneration diseases, PSCI showed a rapid development and finally developed into irreversible nerve injury. The early detection of PSCI would not only benefit the clinical management of PSCI, but it would also improve the prognosis of stroke patients [6].

Long non-coding RNAs (lncRNAs), the major member of the ncRNA family, have attracted special attention in recent epigenetics studies. Previously, studies have identified abnormally expressed lncRNA in neurodegeneration diseases, such as Alzheimer’s disease, and demonstrated their dysregulation in PSCI [7, 8]. Therefore, it was hypothesized that functional molecules in neurodegeneration diseases might also play roles in PSCI [9]. lncRNA SIX3OS1 (SIX3OS1) was suggested to regulate cell autophagy and participate in the development of Alzheimer’s disease [10]. SIX3OS1 was also demonstrated to mediate the protective effect of geniposide on depression disorder via modulating oxidative stress [11], indicating the potential of SIX3OS1 in regulating PSCI.

In this study, the expression and significance of SIX3OS1 in distinguishing stroke patients and predicting the risk of PSCI were investigated. The predictive value of SIX3OS1 in disease severity and progression was also assessed through the enrollment of stroke patients with or without PSCI and healthy individuals.

Materials and methods

Study subjects

The study enrolled a total of 138 patients who were diagnosed with acute stroke at The Affiliated Changsha Central Hospital, University of South China and 80 healthy individuals receiving regular physical examinations at the physical examination center of our hospital from April 2019 to March 2022. The included 138 acute stroke patients were composed of 70 patients with ischemic stroke and 68 patients with hemorrhagic stroke. The outcomes of patients with different subtypes showed no significant difference. The clinical records of acute stroke patients were completed, and patients were without any cognitive and communication disorders before their diagnosis. Patients with a history of stroke or other nervous system diseases were excluded. This study had been approved by the Ethics Committee of The Affiliated Changsha Central Hospital, University of South China (No. 0018083) and had obtained informed consent from all participants.

The NIHSS and activities of Daily living of acute stroke patients were employed to evaluate the neural function and daily living ability of acute stroke patients, while the cognitive function of acute stroke patients was evaluated by the MoCA scale after 6 months of their enrollment.

Sample collection

Fasting venous blood samples were collected from all study subjects at the time of their admission (for patients with acute stroke) or the time of receiving physical examination (for healthy individuals) and centrifugated at 1500 g for 10 min to isolate serum. Isolated serum was stored at -80 °C for the following analyses.

Real-time quantitative PCR

Total RNA was extracted from serum samples using TRIZOL reagent (Invitrogen, USA) and evaluated by the ratio of OD260/280 using NanoDrop 2000 (Nanodrop Technologies, USA). Extracted RNA was reverse transcribed into cDNA using a high-performance cDNA reverse Transcription kit (Applied Biosystem, USA) and further amplified on the 7900 PCR system (Applied Biosystem, USA) with the help of SYBR Green kit (Invitrogen, USA). The relative expression level was calculated the 2− deltdeltCT method with GADPH as the internal reference.

Statistical analyses

The baseline information between the two groups was compared with the student’s t-test (for continuous variables) or Chi-square test (for counting data). The significance of SIX3OS1 in distinguishing acute stroke patients and predicting PSCI was evaluated by ROC and logistic regression analysis, respectively. Additionally, the correlation of SIX3OS1 with the severity of acute stroke patients and the progression of PSCI was assessed by Pearson correlation analysis. P < 0.05 indicates statistically significant.

Results

Baseline information of study subjects

Healthy individuals and acute stroke patients showed matched age and gender composition. Healthy individuals were composed of 44 males and 36 females with an average age of 54.71 ± 11.31 years. The average age of acute stroke patients was 56.05 ± 10.69 years including 76 males and 62 females. The percentage of smokers and drinkers in healthy individuals and acute stroke patients also showed no significant difference. Acute stroke patients showed a higher incidence of diabetes (37.5% vs. 55.8%) and hypertension (40.0% vs. 59.4%) than healthy individuals (Table 1).

Table 1 Baseline information of study subjects
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Dysregulation of SIX3OS1 and its significance in acute stroke

Compared with healthy individuals, patients with acute stroke showed a higher serum SIX3OS1 level (Fig. 1a), which discriminated acute stroke patients with a sensitivity of 80.43% and specificity of 87.50% (AUC = 0.904, Fig. 1b). The severity of acute stroke patients was assessed by NIHSS, and the serum SIX3OS1 level of acute stroke patients showed a significantly positive correlation with NIHSS, indicating its close association with the disease severity of acute stroke patients (Fig. 1c).

Fig. 1
figure 1

SIX3OS1 was upregulated in acute stroke and showed significant diagnostic value. (a) serum expression of SIX3OS in healthy individuals and acute stroke patients. (b) ROC evaluating the diagnostic potential of SIX3OS1 in acute stroke. (c) correlation between serum SIX3OS1 level and NIHSS score in acute stroke patients. ****P < 0.0001

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Predictive value of SIX3OS in PSCI of acute stroke patients

Based on the occurrence of PSCI, acute stroke patients were divided into the PSCI and post-stroke cognitive normality (PSCN) groups. It was found that the serum expression of SIX3OS1 was significantly higher in patients with PSCI relative to PSCN patients (Fig. 2a). The logistic regression analysis (sensitivity = 75.7%, specificity = 72.1%, positive predictive value = 73.6%, negative predictive value = 74.2%) showed that SIX3OS1 acted as a risk factor of PSCI in patients with acute stroke with the OR value of 4.673 (95% CI = 2.035–10.732) together with NIHSS (OR = 3.386, 95% CI = 1.171–9.795) and MoCA (OR = 0.313, 95% CI = 0.142–0.690, Table 2), which are major indicators for the severity of acute stroke and the metal states of patients. Consistently, increasing serum SIX3OS1 could effectively distinguish patients with PCSI with a sensitivity and specificity of 86.36 and 73.61%, respectively (AUC = 0.855, Fig. 2b). Additionally, SIX3OS1 showed a negative correlation with the MoCA score of patients with PSCI, which indicates severe cognitive impairment (Fig. 2c).

Fig. 2
figure 2

SIX3OS1 was elevated in acute stroke patients developing PSCI and showed significant diagnostic value. (a) serum expression of SIX3OS1 in acute stroke patients with or without PSCI. (b) ROC evaluating the diagnostic potential of SIX3OS1 in PSCI. (c) correlation between serum SIX3OS1 level and MoCA score in acute stroke patients with PSCI. ****P < 0.0001

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Table 2 Logistic regression analysis evaluating the risk factors for PSCI
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Discussion

Recent studies have paid special attention to the identification of serum biomarkers for brain injury, which is covert and always reaches a severe condition by the time of symptoms. The function of SIX3OS in acute stroke has not been confirmed, but it was demonstrated to play roles in nervous system diseases, such as Alzheimer’s disease, and mediate the protective effect of geniposide on depression disorder [10,11,12]. This study observed the elevating serum SIX3OS1 in acute stroke patients relative to healthy individuals, which discriminates acute stroke patients and healthy individuals. Additionally, serum SIX3OS1 levels in acute stroke patients showed a significantly positive correlation with patients’ NIHSS score, which was clinically employed to evaluate the severity of patients [13]. Hence, SIX3OS1 was considered a biomarker diagnosing acute stroke, and higher serum SIX3OS1 level indicated more severe disease conditions.

Due to the similar pathophysiological mechanisms between acute stroke and vascular cognitive impairment, cognitive dysfunction and dementia have become major complications in acute stroke [14, 15]. According to statistical data, over one-third of stroke survivors experienced some degree of PSCI [16]. Compared with the symptoms of physical disability, cognitive impairment is relatively unobvious and therefore is easy to be ignored by patients, families, and even clinicians. Among enrolled acute stroke patients, over half of patients developed PSCI, and patients who developed PSCI showed a higher serum SIX3OS1 level. SIX3OS1 was identified as a risk factor for PSCI in acute stroke patients, together with NIHSS and MoCA scores, two widely accepted scales assessing nervous injury and cognitive function. Consistently, increasing serum SIX3OS1 showed significant potential in distinguishing acute stroke patients developing PSCI. MoCA showed good validity and reliability in estimating cognitive impairment, and the lower MoCA score indicates poor cognitive function [17, 18]. A negative correlation of serum SIX3OS1 levels in PSCI patients with the MoCA score was observed in the present study. Hence, increasing serum SIX3OS1 was identified as an indicator of the occurrence of PSCI in acute stroke and worsened cognitive dysfunction. However, due to the limited sample size and research centers, the statistical results showed several limitations. Additionally, some studies and researchers also considered MoCA to be subject to various factors, which need complementary evaluation scales in future studies. The complications of diabetes and hypertension increase the risk of acute stroke. Also, patients with diabetes and hypertension easily develop into complex disease conditions and severe complications [19, 20]. The enrolled stroke patients also showed a higher percentage of diabetes and hypertension. Therefore, in theory, diabetes and hypertension are risk factors for the onset of PSCI, which has not been confirmed in the present study. Future studies should expand the sample size and include multiple centers to improve the clinical significance of SIX3OS1. Additionally, previous studies have demonstrated that the stroke subtypes and topography are closely associated with the prognosis of patients. Among the subtypes of stroke, lacunar stroke showed distinct characteristics and risk factors. Also, its prognosis was distinct from other subtypes. More importantly, lacunar stroke syndrome induced by lacunar or non-lacunar stroke plays a critical role in the prognosis of patients with stroke [21]. It was reported that the in-hospital mortality of patients with infarction in the territory of the posterior cerebral artery was lower, but their several clinical features were more frequent [22]. However, the significance of SIX3OS1 in both the above perspectives has not been completely revealed. Future investigations should deeply explore the difference in SIX3OS among various stroke subtypes and topography providing comprehensive evidence for the clinical potential of SIX3OS1.

On the other hand, the development of acute stroke involves various pathological processes, such as inflammation, the injury of gliocytes and the apoptosis of neurons. Previous studies have also focused on the regulatory effect of lncRNAs on these related processes to reveal the mechanism underlying functional lncRNAs [23,24,25,26]. The regulatory effect and functional role of SIX3OS1 on these processes also need deep investigations. Moreover, the assessment of PSCI patients by MoCA is relatively weak in the function of spatial perception, visual movement, and thought operation, and the assessment scale is influenced by the experience and skills of the testers [27]. Hence, different assessing scales should be performed in future studies to evaluate the cognitive function of patients from various perspectives more accurately. Moreover, different sources of lncRNAs always showed distinct function. The diagnostic cutoff for different sourced lncRNAs would also be various. It is a limitation that the source of SIX3OS1 has not been validated in the present study, which is a critical problem that should be solved in further investigations.

Taken together, this study confirmed the clinical significance of increasing serum SIX3OS1 in acute stroke patients, which diagnosed patients, predicted the risk of PSCI, and indicated the severity of acute stroke and PSCI. SIX3OS1 can be considered a biomarker for acute stroke and PSCI, which needs deep and wide clinical validations.

Data availability

The data used and analyzed can be obtained from the corresponding author under a reasonable request.

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Acknowledgements

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Funding

The Natural Science Foundation of Hunan Province (2024JJ9493) and the Science and Technology Plan Project of Changsha (kq2403175).

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Author notes

  1. Yan Liu and Rui Wang contributed equally to this work.

Authors and Affiliations

  1. Department of Neurology, The Affiliated Changsha Central Hospital, University of South China, No. 161 Shaoshan South Road, Changsha, 410004, Hunan, China

    Yan Liu, Junsheng Zeng, Wei Zhao, Yanqiao Xiao, Heng Jiang & Te Wang

  2. Department of Neurology, Liaocheng People’s Hospital, Liaocheng, 252000, Shandong, China

    Rui Wang

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Contributions

Yan Liu and Rui Wang carried out the research design and conception; Rui Wang and Junsheng Zeng and Wei Zhao analyzed and interpreted the data regarding; Yanqiao Xiao and Heng Jiang performed the examination of sample; Te Wang and Wei Zhao and Yanqiao Xiao contributed essential reagents or tools; Yan Liu and Te Wang authors wrote and revised the manuscript. All authors read and approved the final manuscript.

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Correspondence to Te Wang.

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Ethics approval and consent to participate

The experimental procedures were all in accordance with the guideline of the Ethics Committee of The Affiliated Changsha Central Hospital, University of South China and has approved by the Ethics Committee of The Affiliated Changsha Central Hospital, University of South China. This study complies with the Declaration of Helsinki. A signed written informed consent was obtained from each patient.

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The authors declare no competing interests.

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Liu, Y., Wang, R., Zeng, J. et al. lncRNA six3os1 diagnoses acute stroke, predicts disease severity, and predicts post-stroke cognitive impairment. BMC Neurol 24, 491 (2024). https://doiorg.publicaciones.saludcastillayleon.es/10.1186/s12883-024-04003-5

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BMC Neurology

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