1.Prognostic significance of TRIM28 elevation in non-M3 acute myeloid leukemia
Siqi GONG ; Cong LI ; Mengmeng FAN ; Huiping WANG ; Wanqiu ZHANG ; Xue LIANG ; Qianshan TAO ; Qiang HONG ; Zhimin ZHAI
Acta Universitatis Medicinalis Anhui 2026;61(2):301-308
ObjectiveTo clarify the expression of TRIM28 in non-M3 acute myeloid leukemia (AML) and its correlation with clinical indicators and prognosis, and to further explore the effect of TRIM28 expression levels on the proliferation and apoptosis of AML cells using small interfering RNA. MethodsThe GSE34577 dataset was analyzed using R software to compare TRIM28 expression between healthy controls and non-M3 acute myeloid leukemia (AML) patients. Clinical samples from non-M3 AML patients were collected, with TRIM28 expression levels measured using real-time quantitative PCR (qPCR). The analysis focused on correlations between TRIM28 expression and various clinical indicators, treatment efficacy, and patient prognosis. Furthermore, small interfering RNA (siRNA) technology was employed to downregulate TRIM28 expression in human primary AML cells (HL60 cell line). The effects on cell proliferation and apoptosis were then assessed through CCK-8 assays and flow cytometry, respectively. ResultsThe results showed that TRIM28 was up-regulated in non-M3 AML of both online database GSE34577 and clinical samples (P<0.000 1), TRIM28 expression of new diagnosis group and relapsed refractory group was higher than iron deficiency anemia group (P<0.01), and there was no significance between different French-American-British classification systems subtype. TRIM28 expression was higher in non-M3 AML patients with a poor genetic prognosis stratified as moderate than in the good prognosis group, and TRIM28 expression was associated with NPM1 combined with the FLT3-ITD mutation, positively correlated with age, bone marrow blast, peripheral blood blast and white blood cell, negatively correlated with hemoglobin. In addition, interference TRIM28 greatly inhibited cell proliferation and promoted cell apoptosis. ConclusionThis study reveals that TRIM28 is highly expressed in non-M3 AML and associated with prognosis, and plays a key role in the proliferation and apoptosis of AML cells, suggesting that TRIM28 may serve as a novel therapeutic target for non-M3 AML.
2.WANG Xixing's Clinical Experience in Treating Immune Checkpoint Inhibitor-Related Pneumonitis Based on the Theory of "Cough Attributed to the Five Zang (脏) Organs"
Xue QI ; Xi YANG ; Xinyue WANG ; Dongxin ZHANG ; Yuxing MAO ; Yuankun HAN ; Wenbo ZHAI ; Boyang LYU ; Yifang LI ;
Journal of Traditional Chinese Medicine 2026;67(5):477-481
This paper summarizes Professor WANG Xixing's clinical experience in treating immune checkpoint inhibitor-related pneumonitis (CIP) based on the theory of "cough attributed to the five zang (脏) organs". Cough is a common predominant symptom of CIP. According to the theory of "cough attributed to the five zang organs", drug toxicity triggers cancer toxin, leading to disharmony among the five zang organs, and then lung failing to diffuse and govern descent as the core pathogenesis. Therefore, treatment should focus on harmonizing the five zang organs to restore the normal function of lung qi to diffuse and govern descent. In clinical practice, CIP can be classified into four syndrome patterns, including lung yin depletion, deficiency of both the lung and the spleen with phlegm-dampness, liver fire harassing the lung, and lung-kidney yin deficiency. Correspondingly, Chaimai Jinluo Runfei Decoction (柴麦金络润肺汤) is used to nourish yin and moisten the lung; Qigui Peitu Huayin Decoction (芪桂培土化饮汤) is used to fortify the spleen and tonify the lung, resolve dampness and dispel phlegm; Chaidan Shuyu Runjin Decoction (柴丹疏郁润金汤) is used to drain liver and clear the lung; and Dimai Jinshui Xiangsheng Decoction (地脉金水相生汤) is used to nourish the kidney and moisten the lung.
3.A practice guideline for therapeutic drug monitoring of mycophenolic acid for solid organ transplants.
Shuang LIU ; Hongsheng CHEN ; Zaiwei SONG ; Qi GUO ; Xianglin ZHANG ; Bingyi SHI ; Suodi ZHAI ; Lingli ZHANG ; Liyan MIAO ; Liyan CUI ; Xiao CHEN ; Yalin DONG ; Weihong GE ; Xiaofei HOU ; Ling JIANG ; Long LIU ; Lihong LIU ; Maobai LIU ; Tao LIN ; Xiaoyang LU ; Lulin MA ; Changxi WANG ; Jianyong WU ; Wei WANG ; Zhuo WANG ; Ting XU ; Wujun XUE ; Bikui ZHANG ; Guanren ZHAO ; Jun ZHANG ; Limei ZHAO ; Qingchun ZHAO ; Xiaojian ZHANG ; Yi ZHANG ; Yu ZHANG ; Rongsheng ZHAO
Journal of Zhejiang University. Science. B 2025;26(9):897-914
Mycophenolic acid (MPA), the active moiety of both mycophenolate mofetil (MMF) and enteric-coated mycophenolate sodium (EC-MPS), serves as a primary immunosuppressant for maintaining solid organ transplants. Therapeutic drug monitoring (TDM) enhances treatment outcomes through tailored approaches. This study aimed to develop an evidence-based guideline for MPA TDM, facilitating its rational application in clinical settings. The guideline plan was drawn from the Institute of Medicine and World Health Organization (WHO) guidelines. Using the Delphi method, clinical questions and outcome indicators were generated. Systematic reviews, Grading of Recommendations Assessment, Development, and Evaluation (GRADE) evidence quality evaluations, expert opinions, and patient values guided evidence-based suggestions for the guideline. External reviews further refined the recommendations. The guideline for the TDM of MPA (IPGRP-2020CN099) consists of four sections and 16 recommendations encompassing target populations, monitoring strategies, dosage regimens, and influencing factors. High-risk populations, timing of TDM, area under the curve (AUC) versus trough concentration (C0), target concentration ranges, monitoring frequency, and analytical methods are addressed. Formulation-specific recommendations, initial dosage regimens, populations with unique considerations, pharmacokinetic-informed dosing, body weight factors, pharmacogenetics, and drug-drug interactions are covered. The evidence-based guideline offers a comprehensive recommendation for solid organ transplant recipients undergoing MPA therapy, promoting standardization of MPA TDM, and enhancing treatment efficacy and safety.
Mycophenolic Acid/administration & dosage*
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Drug Monitoring/methods*
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Humans
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Organ Transplantation
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Immunosuppressive Agents/administration & dosage*
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Delphi Technique
4.Thalidomide alleviates ferroptosis and oxidative stress in diabetic nephropathy by upregulating nuclear factor E2-related factor 2/heme oxygenase 1/glutathione peroxidase 4 signaling pathway
Lu ZHAI ; Xiaoxiao XUE ; Xiaomei LIU ; Yuxiao MA ; Hongxia ZHANG
Chinese Journal of Nephrology 2025;41(4):276-281
To investigate the potential protective effect of thalidomide (THD) on diabetic nephropathy (DN) and its underlying mechanisms. Twenty-four C57BL/6J mice were randomly divided into control, DN and DN+THD200 groups by random number table method. The DN mouse model was established via intraperitoneal injection of streptozotocin. The DN+THD200 group received THD treatment (200 mg·kg -1·d -1) for 8 weeks. Blood and urine biochemical parameters, as well as renal histopathological changes, were compared among the three groups. For in vitro experiments, a high glucose (HG)-induced injury model was established in mouse glomerular podocytes (MPC5). Cells were divided into control (NG), HG, HG+DMSO, HG+THD100 (100 μg/ml), and HG+THD200 (200 μg/ml) groups. THD-treated cells were exposed to THD for 24 h. Western blotting and real-time quantitative PCR were performed to respectively detect protein and mRNA expression levels of ferroptosis-related molecules, including nuclear factor erythroid 2-related factor 2 (NRF2), heme oxygenase 1 (HO-1), and glutathione peroxidase 4 (GPX4). Immunofluorescence was used to evaluate the expression of solute carrier family 7 member 11 (SLC7A11) and 4-hydroxynonenal (4-HNE). The results showed that, compared with control group, DN group exhibited significantly lower blood urea nitrogen, serum creatinine and 24 h urinary albumin levels (all P<0.05). Compared with DN group, DN+THD200 group exhibited significantly lower blood urea nitrogen, serum creatinine and 24 h urinary albumin levels (all P<0.05). Histopathological examination revealed glomerular expansion, mesangial widening, and basement membrane thickening in DN group compared to control group, which were markedly ameliorated by THD treatment. In vitro, HG group showed significantly decreased protein and mRNA expression levels of GPX4, NRF2 and HO-1 compared to NG group. Both HG+THD100 and HG+THD200 groups exhibited upregulated expression levels of these proteins and corresponding mRNA compared to HG group (all P<0.05). Immunofluorescence demonstrated HG group had enhanced 4-HNE fluorescence intensity and reduced SLC7A11 fluorescence intensity, which were reversed by THD treatment. THD alleviates renal injury in DN mice and mitigates HG-induced ferroptosis in MPC5 cells, potentially via activation of NRF2-HO-1-GPX4 signaling pathway.
5.Interpretation of the WHO′s “Ethics and Governance of Artificial Intelligence for Health: Guidance on Large Multi-Modal Models” and its implications for China
Yao YANG ; Cui Victor YU ; Yuting WANG ; Peng XUE ; Xiaomei ZHAI ; Youlin QIAO
Chinese Journal of Preventive Medicine 2025;59(6):960-969
With the rapid advancement and iterative development of new artificial intelligence technologies, there remains a regulatory vacuum in corresponding governance measures among governments worldwide. Simultaneously, a technological and governance gap exists between developing countries and developed economies. In response, the World Health Organization (WHO) has released "Ethics and Governance of Artificial Intelligence for Health: Guidance on Large Multi-Modal Models" to assist governments in strengthening governance capabilities in this field. This paper provides an in-depth analysis of the Guidance, aiming to identify challenges and risks associated with the application of multimodal large models in healthcare. Guided by ethical principles for advancing health through artificial intelligence, the paper examines the three-tier governance framework and recommendations outlined in the Guidance. Additionally, it evaluates the current state of AI governance in China, offering insights and reference points for improving AI governance in China′s healthcare sector.
6.Expert consensus on intraoperative repositioning for patients with spine fracture and dislocation (version 2025)
Dongmei BIAN ; Ke SUN ; Ningbo CHEN ; Caixia BAI ; Miao WANG ; Yafeng QIAO ; Fei WANG ; Hong WANG ; Feng TIAN ; Mei YAN ; Meng BAI ; Linjuan ZHANG ; Liyan ZHAO ; Yaqing CUI ; Xue JIANG ; Leling FENG ; Ning NING ; Junqin DING ; Lan WEI ; Yonghua ZHAI ; Yu ZENG ; Zengmei ZHANG ; Jiqun HE ; Fenggui BIE ; Hong CHEN ; Zengyan WANG ; Li LI ; Li ZHANG ; Yaying ZHOU ; Bing SHAO ; Ying WANG ; Caixia XIE ; Yanfeng YAO ; Jingjing AN ; Wen SHI ; Xiongtao LIU ; Xiaoyan AN ; Ning NAN ; Lan LI ; Xiaohui GOU ; Qiaomei LI ; Xiuting WU ; Yuqin ZHANG ; Jing LIU ; Fusen XIANG ; Xu XU ; Na MEI ; Jiao ZHOU ; Shan FAN ; Qian WANG ; Shuixia LI
Chinese Journal of Trauma 2025;41(2):138-147
Spine fracture and dislocation are common traumatic spinal conditions that often require surgical intervention due to compromised spinal stability. Surgical approaches include anterior, posterior, and combined anterior-posterior spinal procedures. According to the specific surgical requirements, patients may be placed in the prone position or repositioned between prone and supine positions during surgery. Intraoperative repositioning has become an essential step in patient positioning. However, during repositioning, patients with spinal fracture and dislocation are at increased risk for complications such as hemodynamic instability, nerve injury, and pressure injuries to the skin and soft tissue. Notably, due to the instability of the spinal cord, even minor manipulations can further exacerbate the damage, potentially leading to severe outcomes like paraplegia. Although the current clinical guidelines provide instructive recommendations for standard position, there remains no specific protocols for intraoperative repositioning in patients with spine fracture and dislocation. With a concern for the lack of clinical studies on positioning techniques, risk prevention, and operational norms for special patients, no applicable guidelines or standards are available. A consensus was required to provide clinical reference, meet the requirements of surgical treatment, and minimize the safety risks of patients caused by improper placement of positions. Professional Committee of Operating Room Nursing of Shaanxi Nursing Association organized experts in nursing management and operating room nursing from major hospitals across China to formulate Expert consensus on intraoperative repositioning for patients with spinal fracture and dislocation ( version 2025). The consensus provides 11 recommendations covering pre-repositioning preparation, intraoperative maneuvers, and post-repositioning observation, aiming to provide references for clinical standardization of the intraoperative repositioning process and protection of patients′ safety.
7.Regulation of Tumor Immune Homeostasis by Programmed Cell Death and Intervention Effect of Traditional Chinese Medicine Under Theory of Regulating Qi and Resolving Toxins
Bingwei YANG ; Xue CHEN ; Chenglei WANG ; Haoyu ZHAI ; Weidong LI ; Baojin HUA
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(7):212-220
Tumor immune homeostasis is a dynamic equilibrium state in which the body removes abnormal mutated cells in time to prevent tumor development without damaging other normal cells under the surveillance of the immune system. It is an important concept to understand the process of tumor development. Programmed cell death (PCD) is a kind of regulable cell death including various forms such as apoptosis, autophagy, pyroptosis, necrosis, and ferroptosis. It is regarded as an important way for the body to remove abnormal or mutated cells. In recent years, modern research has found that PCD has a bi-directional regulatory effect on carcinogenesis and tumor development. In the early stage of tumor formation, PCD can control tumor development in time by playing a specific immune clearance role, while in the later tumorigenic stage, PCD can promote the growth and development of tumor cells by forming a tumor-specific microenvironment, resulting in carcinogenic effects. Therefore, PCD is regarded as an important way to maintain tumor immune homeostasis. Based on the idea of ''supporting the vital Qi and cultivating the root'' by professors Yu Guiqing and Piao Bingkui, the team proposed the theory of ''regulating Qi and resolving toxins'' and applied it to clinical tumor prevention and treatment. Based on the theory of ''regulating Qi and resolving toxins'', the research summarized the current progress of modern medical research on mechanisms related to PCD to explore the role of PCD in the regulation of tumor immune homeostasis. The article believed that the harmonious state of Qi movement was the basic condition for normal PCD to maintain tumor immune homeostasis, while the disorder of Qi movement and the evolution of tumor toxicity were the core processes of abnormal PCD and disorder of tumor immunity homeostasis, which led to the escape and development of tumor cells. Therefore, under the guidance of ''regulating Qi and removing toxins'', the idea of full-cycle prevention and treatment of tumors was proposed summarily. In the early stage of tumor formation, the method of ''regulating Qi movement and strengthening vital Qi'' was applied to reestablish tumor immune homeostasis and to promote the elimination of abnormal cells. In the late tumorigenic stage, the method of ''resolving toxins and dispelling evils'' was applied to reverse the specific microenvironment of tumors and inhibit the development of tumor cells, with a view to providing new theoretical support for the prevention and treatment of tumors through traditional Chinese medicine.
8.Regulation of Tumor Immune Homeostasis by Programmed Cell Death and Intervention Effect of Traditional Chinese Medicine Under Theory of Regulating Qi and Resolving Toxins
Bingwei YANG ; Xue CHEN ; Chenglei WANG ; Haoyu ZHAI ; Weidong LI ; Baojin HUA
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(7):212-220
Tumor immune homeostasis is a dynamic equilibrium state in which the body removes abnormal mutated cells in time to prevent tumor development without damaging other normal cells under the surveillance of the immune system. It is an important concept to understand the process of tumor development. Programmed cell death (PCD) is a kind of regulable cell death including various forms such as apoptosis, autophagy, pyroptosis, necrosis, and ferroptosis. It is regarded as an important way for the body to remove abnormal or mutated cells. In recent years, modern research has found that PCD has a bi-directional regulatory effect on carcinogenesis and tumor development. In the early stage of tumor formation, PCD can control tumor development in time by playing a specific immune clearance role, while in the later tumorigenic stage, PCD can promote the growth and development of tumor cells by forming a tumor-specific microenvironment, resulting in carcinogenic effects. Therefore, PCD is regarded as an important way to maintain tumor immune homeostasis. Based on the idea of ''supporting the vital Qi and cultivating the root'' by professors Yu Guiqing and Piao Bingkui, the team proposed the theory of ''regulating Qi and resolving toxins'' and applied it to clinical tumor prevention and treatment. Based on the theory of ''regulating Qi and resolving toxins'', the research summarized the current progress of modern medical research on mechanisms related to PCD to explore the role of PCD in the regulation of tumor immune homeostasis. The article believed that the harmonious state of Qi movement was the basic condition for normal PCD to maintain tumor immune homeostasis, while the disorder of Qi movement and the evolution of tumor toxicity were the core processes of abnormal PCD and disorder of tumor immunity homeostasis, which led to the escape and development of tumor cells. Therefore, under the guidance of ''regulating Qi and removing toxins'', the idea of full-cycle prevention and treatment of tumors was proposed summarily. In the early stage of tumor formation, the method of ''regulating Qi movement and strengthening vital Qi'' was applied to reestablish tumor immune homeostasis and to promote the elimination of abnormal cells. In the late tumorigenic stage, the method of ''resolving toxins and dispelling evils'' was applied to reverse the specific microenvironment of tumors and inhibit the development of tumor cells, with a view to providing new theoretical support for the prevention and treatment of tumors through traditional Chinese medicine.
9.Mechanism of Shaoyaotang in Modulating MDSCs-related Immunosuppressive Microenvironment in Prevention and Treatment of Colitis-associated Carcinogenesis
Xue CHEN ; Chenglei WANG ; Bingwei YANG ; Haoyu ZHAI ; Ying WU ; Weidong LI
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(1):10-19
ObjectiveTo explore the mechanism of Shaoyaotang in the prevention and treatment of colitis-associated carcinogenesis (CAC) based on myeloid-derived suppressor cells (MDSCs)-related immunosuppressive microenvironment. MethodsA total of 140 six-week-old SPF FVB male mice were randomly divided into seven groups: Blank group, Shaoyaotang without model group (7.12 g·kg-1), model group, sulfasalazine group (0.52 g·kg-1), Shaoyaotang low-dose group (3.56 g·kg-1), Shaoyaotang medium-dose group (7.12 g·kg-1) and Shaoyaotang high-dose group (14.24 g·kg-1), with 20 mice in each group. The blank control group and the Shaoyaotang without model group received a single intraperitoneal injection of physiological saline (10 mg·kg-1), while the other five groups were given a single intraperitoneal injection of azoxymethane (AOM) (10 mg·kg-1). After 1 week, the mice were given drinking water containing 2% dextran sulfate sodium (DSS) for 1 week, followed by normal drinking water for 2 weeks. This cycle was repeated three times over a total period of 14 weeks to establish the CAC mouse model. Each group was administered gavage once daily for 2 weeks starting on the 14th day of the experiment, followed by three times a week until the end of the experiment. The body weight of the mice was recorded weekly. Mice were sacrificed on the 28th and 98th days of the experiment. After dissection, the colon length, colon weight, spleen weight, tumor size, and tumor number were measured. Hematoxylin and eosin (HE) staining was used to assess the pathological morphology of colon tumor tissue. Flow cytometry was used to detect MDSCs, regulatory T cells (Tregs), CD4+ T cells, CD8+ T cells, and the CD4+/CD8+ T cell ratio in the spleen. Immunohistochemistry was used to detect the expression levels of programmed cell death protein-1 (PD-1), programmed cell death ligand 1 (PD-L1), phosphorylated AMP-activated protein kinase (p-AMPK), phosphorylated nuclear factor-κB (p-NF-κB), and hypoxia-inducible factor 1α (HIF-1α) in the colon tissue. ResultsOn day 14, compared with the blank group, the body weight of the model group was significantly reduced (P<0.01), reaching its lowest point on day 28 (23.39 ± 0.95 ) g. On days 28 and 98, compared with the blank group, the colon length in the model group was significantly shortened (P<0.01), the colon index significantly increased (P<0.01), the spleen index significantly increased (P<0.01), and the tumor load significantly increased (P<0.01). HE staining showed that in the model group, tumor cells, a large number of inflammatory cell infiltrates, goblet cell disappearance, and crypt loss were observed. In each dose group of Shaoyaotang, the damage to the colonic mucosa, inflammatory cell infiltration, and crypt structure destruction were alleviated. Compared with the model group, the body weight of mice in each dose group of Shaoyaotang increased. On day 98, the colon length was significantly increased (P<0.01), the colon index significantly decreased (P<0.01), the spleen index significantly decreased (P<0.01), and the tumor burden significantly decreased (P<0.01) in each Shaoyaotang dose group. On days 28 and 98, MDSCs and Tregs in the spleen of the medium- and high-dose Shaoyaotang groups were significantly reduced (P<0.01), while CD4+ T cells and the CD4+/CD8+ T cell ratio were significantly increased (P<0.01). The proportion of CD8+ T cells in the spleen and the expression levels of PD-1 and PD-L1 in the colon tissues of mice in each Shaoyaotang dose group were significantly increased to varying degrees (P<0.05, P<0.01). On days 28 and 98, the expression of p-AMPK-positive cells in the colon tissue of the medium- and high-dose Shaoyaotang groups was significantly increased (P<0.01), while the expression of p-NF-κB and HIF-1α was significantly reduced (P<0.01). ConclusionShaoyaotang can regulate MDSC recruitment and modulate the immune function of T lymphocyte subsets to inhibit the occurrence and development of AOM/DSS-induced CAC in mice. The mechanism may be related to the activation of the AMPK/NF-κB/HIF-1α pathway.
10.Current status of cognitive frailty among the elderly in community
ZHAI Yujia ; ZHANG Tao ; GU Xue ; XU Le ; WU Mengna ; LIN Junfen ; WU Chen
Journal of Preventive Medicine 2025;37(8):762-766,772
Objective:
To investigate the current status and influencing factors for cognitive frailty among the elderly in community, so as to provide the evidence for early identification and prevention of cognitive frailty among the elderly.
Methods:
Residents aged 60 years and above with local household registration from 11 counties (cities, districts) in Zhejiang Province from 2021 to 2023 were selected as study participants using a multistage random sampling method. Demographic information, lifestyle, and health status were collected through questionnaire surveys. Depressive symptoms were assessed using the Patient Health Questionnaire. Cognitive frailty was evaluated using the FRAIL Scale and the Mini-Mental State Examination. Factors affecting cognitive frailty among the elderly in community were identified using a multivariable logistic regression model.
Results:
A total of 16 613 individuals were surveyed, including 7 465 males (44.93%) and 9 148 females (55.07%). The average age was (70.97±7.29) years. A total of 784 individuals were detected with depressive symptoms, with a detection rate of 4.72%. A total of 724 individuals were detected with cognitive frailty, with a detection rate of 4.36%. Multivariable logistic regression analysis showed that females (OR=1.419, 95%CI: 1.179-1.708), aged ≥70 years (70-<80 years old, OR=1.869, 95%CI: 1.490-2.345; ≥80 years old, OR=5.017, 95%CI: 3.935-6.398), without a spouse (OR=1.495, 95%CI: 1.234-1.810), sedentary (OR=2.420, 95%CI: 1.829-3.202), chronic diseases (1 type, OR=1.456, 95%CI: 1.175-1.804; ≥2 types, OR=1.639, 95%CI: 1.314-2.045), and depressive symptoms (OR=4.191, 95%CI: 3.361-5.225) were associated with a higher risk of cognitive frailty among the elderly in community. Conversely, a lower risk of cognitive frailty was seen among the elderly in community who had primary school or above (primary school, OR=0.512, 95%CI: 0.389-0.676; junior high school or above, OR=0.464, 95%CI: 0.354-0.608), engaged in physical exercise (OR=0.396, 95%CI: 0.291-0.539), and were reported average or good self-rated health status (average, OR=0.641, 95%CI: 0.475-0.866; good, OR=0.150, 95%CI: 0.109-0.208).
Conclusions
The detection rate of cognitive frailty among the elderly in community is relatively low and is influenced by demographic factors such as gender, age, education level, as well as lifestyle like sedentary and physical exercise, and health status. It is recommended to reduce the risk of cognitive frailty among the elderly through multidimensional interventions, including health education, promotion of healthy lifestyles, and enhanced mental health support.


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