1.In Vitro Anti-psoriatic Effect of Kangfuxin Liquid via Inhibiting Cell Proliferation and Migration Ability and Blocking JAK3/STAT3 Signaling Pathway
Shuai LI ; Xuan LIU ; Wenyan TANG ; Zhenqi WU ; Chunhui CHEN ; Dadan QIU ; Yi XU ; Chenggui ZHANG ; Jianquan ZHU ; Jiali ZHU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):123-133
ObjectiveThis paper aims to explore the in vitro anti-psoriasis activity and potential mechanism of Kangfuxin liquid (KFX liquid), providing experimental evidence for the anti-psoriasis effect of KFX liquid. MethodsFirstly, the uninduced human immortalized keratinocyte cells (HaCaT cells) were divided into seven groups, namely the control group and KFX liquid groups with different doses (5, 10, 20, 40, 80, 160 g·L-1). After being treated with different concentrations of KFX liquid, the effect of KFX liquid on the normal cell proliferation was detected by using the cell counting kit-8 (CCK-8) method. Secondly, the uninduced HaCaT cells were divided into six groups, namely the control group and recombinant human interleukin-7A (rh-IL-7A) groups with different doses (5, 10, 50, 100, 120 g·L-1). After being treated with different concentrations of recombinant human interleukin-17A (rh IL-17A) liquid, the effect of rh IL-17A on cell proliferation was detected. The optimal induction concentration was screened. Then, normal HaCaT cells were divided into a control group and KFX liquid groups with different doses (5, 10, 20, 40, 80, 160 g·L-1). Except for the control group, the other groups established psoriasis cell models with the optimal induction concentration of rh IL-17A. After being treated with different concentrations of KFX liquid, the effects of KFX liquid on the psoriasis-like HaCaT cell proliferation were investigated. Finally, the uninduced HaCaT cells were divided into six groups, namely the control group, rh IL-17A group, methotrexate (MTX) group, and KFX liquid groups with different doses (20, 40, 80 g·L-1). Except for the control group, the other groups used the optimal induction concentration of rh IL-17A to establish psoriasis cell models. After being treated with different drugs, the cell migration levels were detected through scratch assays, and real-time quantitative polymerase chain reaction (Real-time PCR) was used to detect the relative mRNA expression levels of Ki-67 antigen (Ki67), S100 calcium-binding protein A7 (S100A7), S100 calcium-binding protein A8 (S100A8), and S100 calcium-binding protein A9 (S100A9), thereby comprehensively evaluating the in vitro anti-psoriasis activity of KFX liquid. By detecting the relative mRNA expression levels of interleukin-1β (IL-1β), interleukin-6 (IL-6), and chemokine-20 (CXCL-20) inflammatory-related factors in psoriasis-like HaCaT cells and the protein expression levels of Janus kinase 3 (JAK3), phosphorylated Janus kinase 3 (p-JAK3), signal transducer and activator of transcription 3 (STAT3), and phosphorylated signal transducer and activator of transcription 3 (p-STAT3), the mechanism was explored. ResultsCompared with that of control group, when treated with 80 g·L-1 KFX liquid for 72 h (P<0.05) and at different times with 160 g·L-1 KFX liquid, the HaCaT cell proliferation activity was significantly affected (P<0.01), while the other concentrations of KFX liquid had no significant differences in cell morphology and cell proliferation activity at different times, indicating that the KFX liquid is relatively safe for HaCaT cells and has no obvious toxic side effects. Compared with that of control group, when treated with different concentrations of rh IL-17A for 24 h, the HaCaT cell proliferation activity was significantly enhanced, and the cell activity was the strongest when the concentration was 100 μg·L-1 (P<0.05), with a density close to 100% and intact cell morphology, indicating that 100 μg·L-1 is the optimal concentration for inducing HaCaT cell proliferation. The results of the KFX liquid treatment on rh IL-17A-induced psoriasis-like cells show that the KFX liquid not only effectively inhibits the rh IL-17A-induced psoriasis-like HaCaT cell proliferation activity (P<0.01), but also significantly reduces the migration ability of rh IL-17A-induced psoriasis-like HaCaT cells (P<0.01), and the relative mRNA expression levels of Ki67, S100A7, S100A8, and S100A9 (P<0.01). Moreover, the KFX liquid can significantly reduce the relative mRNA expression levels of IL-1β, IL-6, and CXCL-20 in rh IL-17A-induced psoriasis-like cells (P<0.01), and significantly inhibit the phosphorylation levels of JAK3 and STAT3 proteins (P<0.05, P<0.01). ConclusionThe KFX liquid has no obvious toxicity to uninduced HaCaT cells. It can inhibit rh IL-17A-induced psoriasis-like HaCaT cell proliferation, reduce the cell migration ability, and has good in vitro anti-psoriasis activity. Its action mechanism may be related to downregulating the expression levels of inflammation-related cytokines in the JAK3/STAT3 signaling pathway and inhibiting the phosphorylation levels of JAK3 and STAT3 proteins.
2.In Vitro Anti-psoriatic Effect of Kangfuxin Liquid via Inhibiting Cell Proliferation and Migration Ability and Blocking JAK3/STAT3 Signaling Pathway
Shuai LI ; Xuan LIU ; Wenyan TANG ; Zhenqi WU ; Chunhui CHEN ; Dadan QIU ; Yi XU ; Chenggui ZHANG ; Jianquan ZHU ; Jiali ZHU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):123-133
ObjectiveThis paper aims to explore the in vitro anti-psoriasis activity and potential mechanism of Kangfuxin liquid (KFX liquid), providing experimental evidence for the anti-psoriasis effect of KFX liquid. MethodsFirstly, the uninduced human immortalized keratinocyte cells (HaCaT cells) were divided into seven groups, namely the control group and KFX liquid groups with different doses (5, 10, 20, 40, 80, 160 g·L-1). After being treated with different concentrations of KFX liquid, the effect of KFX liquid on the normal cell proliferation was detected by using the cell counting kit-8 (CCK-8) method. Secondly, the uninduced HaCaT cells were divided into six groups, namely the control group and recombinant human interleukin-7A (rh-IL-7A) groups with different doses (5, 10, 50, 100, 120 g·L-1). After being treated with different concentrations of recombinant human interleukin-17A (rh IL-17A) liquid, the effect of rh IL-17A on cell proliferation was detected. The optimal induction concentration was screened. Then, normal HaCaT cells were divided into a control group and KFX liquid groups with different doses (5, 10, 20, 40, 80, 160 g·L-1). Except for the control group, the other groups established psoriasis cell models with the optimal induction concentration of rh IL-17A. After being treated with different concentrations of KFX liquid, the effects of KFX liquid on the psoriasis-like HaCaT cell proliferation were investigated. Finally, the uninduced HaCaT cells were divided into six groups, namely the control group, rh IL-17A group, methotrexate (MTX) group, and KFX liquid groups with different doses (20, 40, 80 g·L-1). Except for the control group, the other groups used the optimal induction concentration of rh IL-17A to establish psoriasis cell models. After being treated with different drugs, the cell migration levels were detected through scratch assays, and real-time quantitative polymerase chain reaction (Real-time PCR) was used to detect the relative mRNA expression levels of Ki-67 antigen (Ki67), S100 calcium-binding protein A7 (S100A7), S100 calcium-binding protein A8 (S100A8), and S100 calcium-binding protein A9 (S100A9), thereby comprehensively evaluating the in vitro anti-psoriasis activity of KFX liquid. By detecting the relative mRNA expression levels of interleukin-1β (IL-1β), interleukin-6 (IL-6), and chemokine-20 (CXCL-20) inflammatory-related factors in psoriasis-like HaCaT cells and the protein expression levels of Janus kinase 3 (JAK3), phosphorylated Janus kinase 3 (p-JAK3), signal transducer and activator of transcription 3 (STAT3), and phosphorylated signal transducer and activator of transcription 3 (p-STAT3), the mechanism was explored. ResultsCompared with that of control group, when treated with 80 g·L-1 KFX liquid for 72 h (P<0.05) and at different times with 160 g·L-1 KFX liquid, the HaCaT cell proliferation activity was significantly affected (P<0.01), while the other concentrations of KFX liquid had no significant differences in cell morphology and cell proliferation activity at different times, indicating that the KFX liquid is relatively safe for HaCaT cells and has no obvious toxic side effects. Compared with that of control group, when treated with different concentrations of rh IL-17A for 24 h, the HaCaT cell proliferation activity was significantly enhanced, and the cell activity was the strongest when the concentration was 100 μg·L-1 (P<0.05), with a density close to 100% and intact cell morphology, indicating that 100 μg·L-1 is the optimal concentration for inducing HaCaT cell proliferation. The results of the KFX liquid treatment on rh IL-17A-induced psoriasis-like cells show that the KFX liquid not only effectively inhibits the rh IL-17A-induced psoriasis-like HaCaT cell proliferation activity (P<0.01), but also significantly reduces the migration ability of rh IL-17A-induced psoriasis-like HaCaT cells (P<0.01), and the relative mRNA expression levels of Ki67, S100A7, S100A8, and S100A9 (P<0.01). Moreover, the KFX liquid can significantly reduce the relative mRNA expression levels of IL-1β, IL-6, and CXCL-20 in rh IL-17A-induced psoriasis-like cells (P<0.01), and significantly inhibit the phosphorylation levels of JAK3 and STAT3 proteins (P<0.05, P<0.01). ConclusionThe KFX liquid has no obvious toxicity to uninduced HaCaT cells. It can inhibit rh IL-17A-induced psoriasis-like HaCaT cell proliferation, reduce the cell migration ability, and has good in vitro anti-psoriasis activity. Its action mechanism may be related to downregulating the expression levels of inflammation-related cytokines in the JAK3/STAT3 signaling pathway and inhibiting the phosphorylation levels of JAK3 and STAT3 proteins.
3.Effect of heterologous expression of Scenedesmus quadricauda malic enzyme gene SqME on photosynthetic carbon fixation and lipid accumulation in tobacco leaves.
Yizhen LIU ; Mengyuan LI ; Zhanqian LI ; Yushuang GUO ; Jingfang JI ; Wenchao DENG ; Ze YANG ; Yan SUN ; Chunhui ZHANG ; Jin'ai XUE ; Runzhi LI ; Chunli JI
Chinese Journal of Biotechnology 2025;41(7):2829-2842
Microalgae possess high photosynthetic efficiency, robust adaptability, and substantial biomass, serving as excellent biological resources for large-scale cultivation. Malic enzyme (ME), a ubiquitous metabolic enzyme in living organisms, catalyzes the decarboxylation of malate to produce pyruvate, CO2, and NAD(P)H, playing a role in multiple metabolic pathways including energy metabolism, photosynthesis, respiration, and biosynthesis. In this study, we identified the Scenedesmus quadricauda malic enzyme gene (SqME) and its biological functions, aiming to provide excellent target genes for the genetic improvement of higher plants. Based on the RNA-seq data from S. quadricauda under the biofilm cultivation mode with high CO2 and light energy transfer efficiency and small water use, a highly expressed gene (SqME) functionally annotated as ME was cloned. The physicochemical properties of the SqME-encoded protein were systematically analyzed by bioinformatics tools. The subcellular localization of SqME was determined via transient transformation in Nicotiana benthamiana leaves. The biological functions of SqME were identified via genetic transformation in Nicotiana tabacum, and the potential of SqME in the genetic improvement of higher plants was evaluated. The ORF of SqME was 1 770 bp, encoding 590 amino acid residues, and the encoded protein was located in chloroplasts. SqME was a NADP-ME, with the typical structural characteristics of ME. The ME activity in the transgenic N. tabacum plant was 1.8 folds of that in the wild-type control. Heterologous expression of SqME increased the content of chlorophyll a, chlorophyll b, and total chlorophyll by 20.9%, 26.9%, and 25.2%, respectively, compared with the control. The transgenic tobacco leaves showed an increase of 54.0% in the fluorescence parameter NPQ and a decrease of 30.1% in Fo compared with the control. Moreover, the biomass, total lipids, and soluble sugars in the transgenic tobacco leaves enhanced by 20.5%, 25.7%, and 9.5%, respectively. On the contrary, the starch and protein content in the transgenic tobacco leaves decreased by 22.4% and 12.2%, respectively. Collectively, the SqME-encoded protein exhibited a strong enzymatic activity. Heterologous expressing of SqME could significantly enhance photosynthetic protection, photosynthesis, and biomass accumulation in the host. Additionally, SqME can facilitate carbon metabolism remodeling in the host, driving more carbon flux towards lipid synthesis. Therefore, SqME can be applied in the genetic improvement of higher plants for enhancing photosynthetic carbon fixation and lipid accumulation. These findings provide scientific references for mining of functional genes from S. quadricauda and application of these genes in the genetic engineering of higher plants.
Nicotiana/genetics*
;
Photosynthesis/physiology*
;
Malate Dehydrogenase/biosynthesis*
;
Plant Leaves/genetics*
;
Scenedesmus/enzymology*
;
Carbon Cycle/genetics*
;
Lipid Metabolism/genetics*
;
Plants, Genetically Modified/metabolism*
4.Clinical characteristics and contributors to diagnostic delay in autoimmune gastritis
Haofeng LI ; He MA ; Tao FU ; Xinyi HUANG ; Qing SHI ; Yan ZHENG ; Hanning LIU ; Hengqi LIU ; Yan GUO ; Chunhui LAN
Journal of Army Medical University 2025;47(19):2396-2404
Objective To analyze the diagnostic process and clinical characteristics of autoimmune gastritis(AIG)in order to improve the awareness and diagnostic proficiency of this disease.Methods A retrospective cohort study was conducted on 114 patients diagnosed with AIG in Army Medical Center of PLA between January 2021 and June 2024.Comprehensive statistical analysis was performed on clinical data,including demographic characteristics(age,sex),clinical symptoms,comorbidities,diagnostic process,Helicobacter pylori(H.pylori)infection and treatment history,laboratory indicators[results of routine blood test,anemia-related indices,thyroid function,anti-parietal cell antibody(APCA),intrinsic factor antibody(IFA)],and gastrointestinal endoscopic findings(frequency and endoscopic features).Results Among the 114 patients,males accounted for 28.1%(32/114)and females for 71.9%(82/114),and they were at a mean age of 56.3±8.4 years.Predominant symptoms included epigastric/upper abdominal pain(47.4%,54/114)and postprandial fullness(43.0%,49/114),while 24.6%(28/114)reported acid reflux or heartburn.Diagnostic delay occurred in 76.4%(87/114)of patients,with a median delay duration of 11.5 months.Primary diagnostic clues were endoscopic reverse gradient atrophy(significantly more severe mucosal atrophy in the gastric corpus/fundus versus antrum;53.5%,61/114)and repeated H.pylori eradication failure(≥2 attempts;22.8%,26/114).Positivity rate of thyroid peroxidase antibody(TPOAb)and thyroglobulin antibody(TgAb)was 56.9%(33/58)and 36.2%(21/58),respectively.APCA positive rate was 98.8%(81/82),IFA positive rate was 34.1%(28/82),and dual-antibody rate was 32.9%(27/82).Anemia was present in 25.7%(26/101)of the patients.Gastric neuroendocrine tumors(NET)were found in 12.2%(14/114),intraepithelial neoplasia in 5.3%(6/114),and gastric adenocarcinoma in 0.9%(1/114).Among colonoscopy-examined patients,tubular adenomas occurred in 25.0%(13/52)and colorectal malignancies in 3.4%(2/58).There were 18.4%(21/114)patients having gallbladder-related diseases,7.9%(9/114)having diabetes mellitus,and 1.8%(2/114)of subacute combined degeneration of the spinal cord.Conclusion AIG is frequently associated with diagnostic delay.The reverse pattern of atrophy on endoscopy serves as a critical diagnostic clue,necessitating enhanced recognition in endoscopists.Patients with recurrent H.pylori eradication failure(≥2 attempts)should be evaluated for AIG.
5.Acupuncture and moxibustion therapy in the management of Crohn's disease: opportunities and challenges
Chunhui BAO ; Jinrong ZHANG ; Xinyi ZHU ; Zhou HAO ; Luyi WU ; Huirong LIU ; Huan'gan WU
Chinese Journal of Inflammatory Bowel Diseases 2025;09(4):286-291
With the increasing global recognition of Traditional Chinese Medicine (TCM), acupuncture, as a non-pharmacological treatment, has demonstrated promising potential in alleviating symptoms, improving quality of life, and modulating immune and neurological system functions in patients with Crohn's disease (CD). This review summarizes the theoretical foundations and recent research progress on acupuncture for CD, and introduces a personalized "Three Discriminations" diagnostic and therapeutic strategy based on syndrome differentiation, meridian differentiation, and acupoint selection. It also identifies the target patient populations and clinical application value of acupuncture in CD treatment. Despite preliminary progress, acupuncture treatment for CD still faces multiple challenges, including limited high-quality clinical research evidence, uneven resource distribution, non-standardized efficacy evaluation systems, insufficiently standardized treatment strategies, and an incomplete understanding of its underlying mechanisms. Future efforts should focus on establishing high-quality evidence systems, optimizing integrative treatment strategies combining acupuncture with conventional medicine, and conducting deep exploration of multi-dimensional mechanisms, with the goal of promoting the standardized integration of acupuncture into the management of CD and providing patients with safer, more precise, and sustainable therapeutic options.
6.Expert consensus on infection prevention and control of Creutzfeldt-Jakob disease in medical institutions
Tianxiang GE ; Yangyang JIA ; Chunhui LI ; Jianrong HUANG ; Xiujuan MENG ; Xiaodong GAO ; Jingping ZHANG ; Fu QIAO ; Lijuan XIONG ; Hui LIANG ; Wei LI ; Haiyan LOU ; Wenjuan WU ; Tianxin XIANG ; Jiansen CHEN ; Biao ZHU ; Kaijin XU ; Zhihui ZHOU ; Hongliu CAI ; Meihong YU ; Yan ZHANG ; Yanwan SHANGGUAN ; Haiting FENG ; Hangping YAO ; Lei GUO ; Tieer GAN ; Weihong ZHANG ; Jimin SUN ; Ye LU ; Qun LU ; Meng CAI ; Jin SHEN ; Yunsong YU ; Anhua WU ; Liu-yi LI ; Tingting QU
Chinese Journal of Infection Control 2025;24(4):437-450
Creutzfeldt-Jakob disease(CJD)is a rapidly progressive and fatal neurodegenerative disorder caused by prions,with certain infectivity and iatrogenic transmission risks.With the rapid progress and application of new dia-gnostic biomarkers and detection methods,as well as the construction and improvement of surveillance and reporting systems,the detection of CJD in patients domestically and internationally has shown an increasing trend year by year.Due to its long incubation period and heterogeneity of early symptoms,early identification and diagnosis of the disease is difficult,increasing the risk of transmission within medical institutions.Currently,there is a lack of con-sensus on the infection prevention and control of CJD.In order to timely identify and diagnose CJD as well as effec-tively block its transmission in medical institutions,this consensus summarizes 15 clinical concerns and formulates 24 specific recommendations based on the latest domestic and international research findings and clinical evidence,as well as combines with clinical practice,aiming to standardize healthcare-associated infection prevention and control measures for CJD and reduce its transmission risk in medical institutions.
7.The influence of preoperative gamma-glutamyl transferase to albumin ratio on the prognosis of patients with BCLC stage 0-A hepatocellular carcinoma after microwave ablation
Xiaolin LIU ; Feng XU ; Fanchuang KONG ; Yanhua HUANG ; Chunhui ZHOU ; Jing CHEN ; Xiaoguang WANG
Chinese Journal of Hepatobiliary Surgery 2025;31(1):17-22
Objective:To explore the effect of preoperative gamma-glutamyl transferase to albumin ratio (GAR) on prognosis of patients with Barcelona clinic liver cancer (BCLC)stage 0-A hepatocellular carcinoma (HCC) after microwave ablation (MWA).Methods:The clinical data of 201 patients with BCLC stage 0 to A who underwent MWA at two centers of Jiaxing Second Hospital and Shengjing Hospital of China Medical University from January 2011 to December 2018 were retrospectively analyzed, including 152 males and 49 females, aged (57.5±9.6) years. X-tile software was used to divide patients into a low GAR group ( n=84, GAR<0.9) and a high GAR group ( n=117, GAR≥0.9). Kaplan-Meier method was used for survival analysis, and log-rank test was used for survival comparison. Univariate and multivariate Cox regression analysis was used to evaluate the relationship between GAR and prognosis. Results:The cumulative survival rates at 1, 3 and 5 years postoperatively were 98.7%, 93.8%, 78.5% for the low GAR group, which were superior to that 97.2%, 87.1%, 70.2% for the high GAR group, and the difference was statistically significant ( χ2=11.89, P=0.001). The recurrence-free survival rates at 1, 3 and 5 years after surgery between the two groups was no significant difference ( χ2=1.70, P=0.192). Multivariate analysis revealed that high GAR ( HR=2.723, 95% CI: 1.508-4.914, P=0.001) was independent risk factors for overall survival after MWA in patients with BCLC stages 0 to A HCC. Male gender ( HR=1.959, 95% CI: 1.127-3.305, P=0.017) and tumor diameter ≥2 cm ( HR=1.547, 95% CI: 1.008-2.373, P=0.046) were independent risk factors for recurrence after MWA in patients with BCLC stages 0 to A HCC. Univariate analysis, GAR≥0.9 was not associated with postoperative recurrence ( HR=1.315, 95% CI: 0.869-1.989, P=0.195). Conclusion:Preoperative GAR (≥0.9) is an independent risk factor affecting overall survival in patients with BCLC stages 0 to A HCC after MWA.
8.Acupuncture Treatment Strategies for Crohn's Disease Based on the Principle of "Shaoyang as the Pivot"
Chunhui BAO ; Jin HUANG ; Xinyi ZHU ; Zhou HAO ; Luyi WU ; Huirong LIU ; Huangan WU
Journal of Traditional Chinese Medicine 2025;66(10):1017-1022
The shaoyang meridian is an important pivot between the internal organs and meridians system, with the functions of regulating qi and blood, balancing yin and yang, and coordinating the ascending and descending movement of qi. Dysfunction of the shaoyang pivot can lead to spleen and kidney deficiency, impaired liver and gallbladder qi regulation, and stagnation of qi and blood. It is believed that the onset and progression of Crohn's disease are closely related to shaoyang pivot dysfunction, with the core pathogenesis characterized by shaoyang disharmony, spleen deficiency, dampness retention, and blood stasis. Based on this understanding, the treatment principle centers on harmonizing the shaoyang pivot, supplemented by methods such as warming and nourishing the spleen and stomach, tonifying shaoyang, and soothing the liver and benefiting the gallbladder. Acupuncture is employed to target key acupoints along the shaoyang meridian to restore its regulatory functions, improve spleen and stomach transformation and transportation, facilitate liver and gallbladder qi flow, and promote the circulation of qi and blood. This provides a practical therapeutic approach for acupuncture-based treatment of Crohn's disease.
9.Expert recommendations for diagnosis and treatment routes of severe infections in elderly people based on immune function evaluation
Lina ZHANG ; Chunhui LI ; Zhihong ZUO ; Zhanwen WANG ; Fulai YUAN ; Chuan-chang LI ; Qiong CHEN ; Wei LIU ; Anhua WU ; Zhaoxin QIAN
Chinese Journal of Infection Control 2025;24(8):1027-1032
The aging trend is intensifying currently,but there is still a lack of standardized diagnosis and treat-ment schemes for severe infections in elderly people.This paper focuses on the recommendations for immune-related clinical diagnosis and treatment routes as well as the idea of risk stratified diagnosis and treatment for elderly peo-ple,aiming to effectively prevent infectious diseases in elderly people and perform stratified management through systematic and scientific means of immune function monitoring and regulation,so as to enhance the standardized level of diagnosis and treatment as well as clinical treatment effect of infection in elderly people.
10.Molecular mechanisms and diagnostic value of seminal plasma exosomal miR-26a-5p targeting PTEN in idiopathic teratozoospermia
Chunhui LIU ; Ruipeng WU ; Zhiqiang WANG ; Wensheng SHAN ; Shaojun LI
The Journal of Practical Medicine 2025;41(20):3256-3266
Objective This study aims to elucidate the functional mechanism through which seminal plasma exosomal miR-26a-5p and its target genes contribute to idiopathic teratozoospermia,with emphasis on their regulatory pathways in sperm morphogenesis defects,thereby establishing a theoretical foundation for novel diagnostic markers and targeted therapies.Methods A case-control study design was adopted,enrolling 154 male subjects(84 in the teratozoospermia group[TZS]and 70 in the normal control group[NC]).Seminal plasma exosomes were isolated to screen differentially expressed miRNAs,followed by prediction and analysis of target genes and signaling pathways.Dual-luciferase reporter gene assays were employed to validate the direct binding of miR-26a-5p to PTEN.Quantitative real-time PCR and Western blot were used to measure miRNA and protein expression levels.Spearman correlation analysis evaluated relationships between miR-26a-5p,PTEN,and sperm morphological parameters,while ROC curve analysis assessed diagnostic efficacy.Results Seminal plasma exosomal miRNA sequencing identified 11 differentially expressed miRNAs,with miR-26a-5p significantly upregulated in the TZS group(P<0.05)and negatively correlated with the percentage of normal sperm morphology in TZS(r=-0.762,P<0.05).PTEN was confirmed as a direct target of miR-26a-5p,with its expression significantly reduced in the TZS group(P<0.05)and positively correlated with normal sperm morphology(r=0.821,P<0.05).A negative correlation was observed between miR-26a-5p and PTEN(r=-0.878,P<0.05).Dual-luciferase assays demonstrated that miR-26a-5p specifically inhibited PTEN 3'UTR activity(45%reduction in luciferase activity,P<0.05).Functional enrichment analysis revealed that the miR-26a-5p-PTEN axis exacerbates oxidative stress,DNA damage,and abnormal spermatocyte division by regulating the PI3K/AKT/mTOR,p53,Wnt/β-catenin,and NF-κB pathways.ROC analysis showed diagnostic efficacy for teratozoospermia with AUCROC values of 0.785(sensitivity 78.9%,specificity 71.3%)for miR-26a-5p and 0.754(sensitivity 73.3%,specificity 75.1%)for PTEN.Conclusions miR-26a-5p suppresses PTEN through targeted inhibition,activating PI3K/AKT/mTOR signaling pathways while impairing DNA repair functions.This cascade leads to accumulated oxidative stress and sperm morphological abnormalities.The combined pattern of elevated seminal plasma exosomal miR-26a-5p expression and reduced PTEN levels demonstrates diagnostic potential for idiopathic teratozoospermia,with its molecular mechanism providing a theoretical foundation for biomarker development.Targeted silencing of miR-26a-5p,either alone or in combination with AKT/mTOR inhibitors and antioxidant therapy,may represent a novel strategy for improving sperm morphology.

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