1.Effect of Serum Containing Zhenwutang on Apoptosis of Myocardial Mast Cells and Mitochondrial Autophagy
Wei TANG ; Meiqun ZHENG ; Xiaolin WANG ; Zhiyong CHEN ; Chi CHE ; Zongqiong LU ; Jiashuai GUO ; Xiaomei ZOU ; Lili XU ; Lin LI
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(3):11-21
ObjectiveTo explore the effect of serum containing Zhenwutang on myocardial mast cell apoptosis induced by angiotensin Ⅱ (AngⅡ) and the mechanism of the correlation between apoptosis and mitochondrial autophagy. MethodsIn this experiment, AngⅡ and serum containing Zhenwutang with different concentrations were used to interfere with H9C2 cardiomyocytes for 24 h, and the survival rate of H9C2 cardiomyocytes was detected by cell counting kit-8 (CCK-8) to screen the optimal concentration for the experiment. Enzyme-linked immunosorbent assay (ELISA) was used to detect the content of B-type natriuretic peptide (BNP) in cell culture supernatant, and immunofluorescence was used to detect the cell surface area to verify the construction of the myocardial mast cell model. Subsequently, the experiment was divided into a blank group (20% blank serum), a model group (20% blank serum + 5×10-5 mol·L-1 AngⅡ), low-, medium-, and high-dose (5%, 10% and 20%) serum containing Zhenwutang groups, an autophagy inhibitor group (1×10-4 mol·L-1 3-MA), and autophagy inducer group (1×10-7 mol·L-1 rapamycin). The apoptosis level of H9C2 cells and the changes of mitochondrial membrane potential were detected by flow cytometry. The lysosomal probe (Lyso Tracker) and mitochondrial probe (Mito Tracker) co-localization was employed to detect autophagy. Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was used to detect Caspase-3, Caspase-9, B-cell lymphoma 2 (Bcl-2), Bcl-2-related X protein (Bax), and cytochrome C (Cyt C) in apoptosis-related pathways and the relative mRNA expression of ubiquitin ligase (Parkin), phosphatase and tensin homolog (PTEN)-induced kinase 1 (PINK1), and p62 protein in mitochondrial autophagy-related pathways. Western blot was used to detect cleaved Caspase-3, cleaved Caspase-9, Bax, Bcl-2, and Cyt C in apoptosis-related pathways, phosphorylated ubiquitin ligase (p-Parkin), phosphorylated PTEN-induced kinase 1 (p-PINK1), p62, and Bcl-2 homology domain protein Beclin1 in mitochondrial autophagy-related pathways, and the change of microtubule-associated protein 1 light chain 3 (LC3) Ⅱ/Ⅰ ratio. ResultsCCK-8 showed that when the concentration of AngⅡ was 5×10-5 mol·L-1, the cell activity was the lowest, and there was no cytotoxicity. At this concentration, the surface area of cardiomyocytes was significantly increased (P<0.01), and the content of BNP in the supernatant of culture medium was significantly increased (P<0.05). Therefore, AngⅡ with a concentration of 5×10-5 mol·L-1 was selected for the subsequent modeling of myocardial mast cells. Compared with the blank group, the model group and the autophagy inhibitor 3-MA group had a significantly increased apoptosis rate (P<0.01) and significantly decreased mitochondrial membrane potential (P<0.01). The results of immunofluorescence co-localization showed that compared with the blank group, the model group had a significantly decreased number of red and green fluorescence spots. The results of Real-time PCR showed that compared with that in the blank group, the relative mRNA expression of Bax, Caspase-3, Caspase-9, Cyt C, and p62 in the model group was significantly up-regulated (P<0.01), while the relative mRNA expression of Bcl-2, Parkin, and PINK1 was significantly down-regulated (P<0.01). In addition, the relative protein expression of Bax, cleaved Caspase-3, cleaved Caspase-9, Cyt C, and p62 was significantly up-regulated (P<0.01). The LC3Ⅱ/Ⅰ was significantly decreased, and the relative protein expression of Bcl-2, p-Parkin, p-PINK1, and Beclin1 was significantly down-regulated (P<0.01). Compared with the model group, the serum containing Zhenwutang groups and the autophagy inducer group had significantly decreased apoptosis rate (P<0.01), and the decrease ratio of mitochondrial membrane potential is significantly lowered (P<0.01) in a dose-dependent manner. Additionally, both red and green fluorescence spots became more in these groups. In the 3-MA group, the number of red and green fluorescence spots decreased significantly. The relative mRNA expression of Bax, Caspase-3, Caspase-9, Cyt C, and p62 was significantly down-regulated (P<0.05, P<0.01), while that of Bcl-2, Parkin, and PINK1 was significantly up-regulated (P<0.01). In the serum containing Zhenwutang groups, the relative protein expression levels of Bax, cleaved Caspase-3, cleaved Caspase-9, Cyt C, and p62 were significantly down-regulated (P<0.05,P<0.01). The LC3Ⅱ/Ⅰ was significantly increased, and the relative protein expression levels of Bcl-2, p-Parkin, p-PINK1, and Beclin1 were significantly up-regulated (P<0.01). ConclusionThe serum containing Zhenwutang can reduce the apoptosis of myocardial mast cells and increase mitochondrial autophagy. This is related to the inhibition of intracellular Bax/Bcl-2/Caspase-3 apoptosis pathway and regulation of Parkin/PINK1 mitochondrial autophagy pathway.
2.Development and research of an AI-assisted decision-making platform in treatment of insomnia with acupuncture of Tongdu Yangxin acupoint prescription.
Chi WANG ; Chengyong LIU ; Xiaoqiu WANG ; Enqi LIU ; Juguang SUN ; Jin LU ; Min DING ; Wenzhong WU
Chinese Acupuncture & Moxibustion 2025;45(7):881-888
OBJECTIVE:
To construct and validate a predictive model for the therapeutic effect of acupuncture at Tongdu Yangxin prescription (acupoint prescription for promoting the circulation of the governor vessel and nourishing the heart) on insomnia, so as to develop an open-access interactive artificial intelligence (AI)-assisted decision-making platform.
METHODS:
Clinical data of 139 insomnia patients treated with Tongdu Yangxin acupuncture therapy were included. All the patients had received acupuncture at Baihui (GV20), Yintang (GV24+), bilateral Shenmen (HT7), and bilateral Sanyinjiao (SP6); and electric stimulation was attached to Baihui (GV20) and Yintang (GV24+), using a continuous wave and a frequency of 2 Hz. The treatment was delivered once every other day, 3 treatments a week, and for 2 consecutive weeks. Patients with Pittsburgh sleep quality index (PSQI) score reduction rate <50% were classified as the "no response group", and those with ≥50% were as the "response group". Outliers were addressed using the 1.5×IQR rule, and missing values were imputed via predictive mean matching. Key features were selected by intersecting the feature importance results from eXtreme Gradient Boosting (XGBoost) and random forest algorithms. After balancing class distribution using the Synthetic Minority Over-sampling Technique (SMOTE), 20% of the data was reserved as a validation set. The remained data underwent the stratified sampling iterations to generate 200 pairs of 3∶1 training-test sets, which was employed for training and internal validation of 8 machine learning algorithms. The optimal algorithm and data partitioning strategy were selected to construct the final model, followed by external validation. The best-performing model was deployed online via Streamlit to create an interactive AI platform.
RESULTS:
Key predictive features for model construction included insomnia duration, the total PSQI score, PSQI sleep efficiency subscore, the proportion of N1 and N2 sleep stages in total sleep duration, and the maximum pulse rate during sleep. The CatBoost-based model achieved an AUC of 0.92, the average precision of 0.77, and accuracy, average recall, and average F1-score of 0.75 on the test set. On the validation set, it attained an AUC of 0.84, with accuracy, average precision, average recall, and average F1-score all at 0.72, demonstrating robust predictive performance. An interactive AI platform was subsequently developed (https://tdyx-catboost.streamlit.app/).
CONCLUSION
This study successfully establishes and validates a CatBoost-based efficacy prediction model for Tongdu Yangxin acupuncture therapy in treatment of insomnia. The developed AI platform provides data-driven decision support for acupuncture-based insomnia management.
Humans
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Sleep Initiation and Maintenance Disorders/physiopathology*
;
Acupuncture Therapy
;
Male
;
Acupuncture Points
;
Female
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Middle Aged
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Adult
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Artificial Intelligence
;
Aged
;
Young Adult
3.Percutaneous coronary intervention vs . medical therapy in patients on dialysis with coronary artery disease in China.
Enmin XIE ; Yaxin WU ; Zixiang YE ; Yong HE ; Hesong ZENG ; Jianfang LUO ; Mulei CHEN ; Wenyue PANG ; Yanmin XU ; Chuanyu GAO ; Xiaogang GUO ; Lin CAI ; Qingwei JI ; Yining YANG ; Di WU ; Yiqiang YUAN ; Jing WAN ; Yuliang MA ; Jun ZHANG ; Zhimin DU ; Qing YANG ; Jinsong CHENG ; Chunhua DING ; Xiang MA ; Chunlin YIN ; Zeyuan FAN ; Qiang TANG ; Yue LI ; Lihua SUN ; Chengzhi LU ; Jufang CHI ; Zhuhua YAO ; Yanxiang GAO ; Changan YU ; Jingyi REN ; Jingang ZHENG
Chinese Medical Journal 2025;138(3):301-310
BACKGROUND:
The available evidence regarding the benefits of percutaneous coronary intervention (PCI) on patients receiving dialysis with coronary artery disease (CAD) is limited and inconsistent. This study aimed to evaluate the association between PCI and clinical outcomes as compared with medical therapy alone in patients undergoing dialysis with CAD in China.
METHODS:
This multicenter, retrospective study was conducted in 30 tertiary medical centers across 12 provinces in China from January 2015 to June 2021 to include patients on dialysis with CAD. The primary outcome was major adverse cardiovascular events (MACE), defined as a composite of cardiovascular death, non-fatal myocardial infarction, and non-fatal stroke. Secondary outcomes included all-cause death, the individual components of MACE, and Bleeding Academic Research Consortium criteria types 2, 3, or 5 bleeding. Multivariable Cox proportional hazard models were used to assess the association between PCI and outcomes. Inverse probability of treatment weighting (IPTW) and propensity score matching (PSM) were performed to account for potential between-group differences.
RESULTS:
Of the 1146 patients on dialysis with significant CAD, 821 (71.6%) underwent PCI. After a median follow-up of 23.0 months, PCI was associated with a 43.0% significantly lower risk for MACE (33.9% [ n = 278] vs . 43.7% [ n = 142]; adjusted hazards ratio 0.57, 95% confidence interval 0.45-0.71), along with a slightly increased risk for bleeding outcomes that did not reach statistical significance (11.1% vs . 8.3%; adjusted hazards ratio 1.31, 95% confidence interval, 0.82-2.11). Furthermore, PCI was associated with a significant reduction in all-cause and cardiovascular mortalities. Subgroup analysis did not modify the association of PCI with patient outcomes. These primary findings were consistent across IPTW, PSM, and competing risk analyses.
CONCLUSION
This study indicated that PCI in patients on dialysis with CAD was significantly associated with lower MACE and mortality when comparing with those with medical therapy alone, albeit with a slightly increased risk for bleeding events that did not reach statistical significance.
Humans
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Percutaneous Coronary Intervention/methods*
;
Male
;
Female
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Coronary Artery Disease/drug therapy*
;
Retrospective Studies
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Renal Dialysis/methods*
;
Middle Aged
;
Aged
;
China
;
Proportional Hazards Models
;
Treatment Outcome
4.Jianpi Qinghua Formula improves metabolic-associated fatty liver disease by modulating PGC1α/PPARα/CPT1A pathway.
Yan-Yan XIAO ; Xu HAN ; Qing-Guang CHEN ; Jun-Fei XU ; Chi CHEN ; Fan GONG ; Hao LU
China Journal of Chinese Materia Medica 2025;50(9):2505-2514
Based on the regulation of mitochondrial fatty acid β-oxidation through the PGC1α/PPARα/CPT1A pathway, this study investigated the effect of Jianpi Qinghua Formula on the mitochondrial fatty acid β-oxidation pathway in the livers of mice with metabolic-associated fatty liver disease(MAFLD) induced by a high-fat diet. MAFLD mice were fed a high-fat diet to establish the model, and after successful modeling, the mice were divided into the model group, the Jianpi Qinghua Formula group, and the metformin group, with an additional control group. Each group was treated with the corresponding drug or an equivalent volume of saline via gavage. Body mass and food intake were measured regularly during the experiment. At the end of the experiment, blood lipid levels and liver function-related indices were measured, liver pathological changes were observed, and protein expression levels of PGC1α, PPARα, PPARγ, and CPT1A were detected by Western blot. The results showed that, with no difference in food intake, compared to the model group, the body mass of the Jianpi Qinghua Formula group and the metformin group was reduced, liver weight and liver index decreased, and levels of cholesterol, triglycerides, and low-density lipoprotein cholesterol(LDL-C) were lowered. Additionally, a decrease in alanine aminotransferase(ALT) and aspartate aminotransferase(AST) was observed. Hematoxylin and eosin(HE) staining revealed reduced pathological damage to hepatocytes, while oil red O staining showed improvement in fatty infiltration. The liver disease activity score decreased, and transmission electron microscopy revealed improvement in mitochondrial swelling and restoration of internal cristae. Western blot analysis indicated that Jianpi Qinghua Formula significantly increased the expression of PGC1α, PPARα, and CPT1A proteins in the liver and reduced the expression of PPARγ. These results suggest that the Jianpi Qinghua Formula improves mitochondrial function, promotes fatty acid oxidation, and alleviates the pathological changes of MAFLD. In conclusion, Jianpi Qinghua Formula can improve MAFLD by mediating mitochondrial fatty acid β-oxidation through the PGC1α/PPARα/CPT1A pathway.
Animals
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PPAR alpha/genetics*
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Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha/genetics*
;
Drugs, Chinese Herbal/administration & dosage*
;
Mice
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Carnitine O-Palmitoyltransferase/genetics*
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Male
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Liver/metabolism*
;
Fatty Liver/genetics*
;
Humans
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Mice, Inbred C57BL
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Diet, High-Fat/adverse effects*
5.Research Advances in the Construction and Application of Intestinal Organoids.
Qing Xue MENG ; Hong Yang YI ; Peng WANG ; Shan LIU ; Wei Quan LIANG ; Cui Shan CHI ; Chen Yu MAO ; Wei Zheng LIANG ; Jun XUE ; Hong Zhou LU
Biomedical and Environmental Sciences 2025;38(2):230-247
The structure of intestinal tissue is complex. In vitro simulation of intestinal structure and function is important for studying intestinal development and diseases. Recently, organoids have been successfully constructed and they have come to play an important role in biomedical research. Organoids are miniaturized three-dimensional (3D) organs, derived from stem cells, which mimic the structure, cell types, and physiological functions of an organ, making them robust models for biomedical research. Intestinal organoids are 3D micro-organs derived from intestinal stem cells or pluripotent stem cells that can successfully simulate the complex structure and function of the intestine, thereby providing a valuable platform for intestinal development and disease research. In this article, we review the latest progress in the construction and application of intestinal organoids.
Organoids/cytology*
;
Intestines/physiology*
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Humans
;
Animals
;
Pluripotent Stem Cells
6.Biological principles of "food and medicine homologous"
Jin-wen DING ; Xiang-yin CHI ; Yu ZHANG ; Lu-lu WANG ; Jian-dong JIANG ; Yuan LIN
Acta Pharmaceutica Sinica 2024;59(6):1509-1518
With the rapid society development and broad recognition of "Healthy China", the demands for good life and health are increasing. Accordingly, the concept of "food and medicine homologous" have been attractive. The concept of "food and medicine homologous" has a long history in China, and is an essence of various ideas in traditional Chinese medicine, such as diet therapy, medicated diet, regimen and preventive treatment of disease, representing an important field in health science. Many studies have found that the active ingredients of "food and medicine homologous" substances are multiple types, multiple mechanisms and multiple targets, exerting their biological effects after oral administration and chemical or metabolic transformation. In this review, the chemical basis and biological principles of various "food and medicine homologous" substances were summarized as compounds, biological macromolecules and intestinal flora. By focusing on the intestinal flora, we discussed the detailed biological principles of several classic "food and medicine homologous" substances. The scientific significance of "food and medicine homologous" concept were also discussed. This review explores the concept of "food and medicine homologous" from the perspective of modern medicine, in order to provide insights for future drug development and human health.
7.Advances in High-throughput Protein Structural Bioinformatics
Progress in Biochemistry and Biophysics 2024;51(9):1989-1999
This review provides a comprehensive summary of the latest advancements in high-throughput protein structural bioinformatics, a field that has undergone a revolutionary transformation with the advent of deep learning-based protein structure prediction systems like AlphaFold2. These systems have significantly increased the accuracy, speed, and scale of protein structure prediction, resulting in an exponential growth in the number of protein structures available for analysis. Notably, the AlphaFold Protein Structure Database (AFDB) has amassed over 214 million protein structures, surpassing the PDB’s 50-year cumulative data by over 1 000-fold within several months. Big data is driving the comprehensive upgrade of protein structural bioinformatics. This review focuses on three main areas: structure data management, tool development, and structure data mining. In the realm of structure data management, the review spotlights the optimization strategy of AlphaFold-like systems, which significantly reduces the resource requirements for protein folding, enabling more researchers to make custom structure predictions and further enlarging the data scale. The resulting “data explosion” has exerted increased pressure on storage and bandwidth, prompting the development of cutting-edge tools such as Foldcomp, PDC, and ProteStAr for compressing PDB files. Moreover, the review underscores the critical role of public repositories like ModelArchive and PDB-Dev in archiving and sharing third-party AlphaFold models. It also highlights the utilization of independent services like MineProt and 3D-Beacons to create more interactive and accessible data portals. In terms of tool development, the review spotlights recent breakthroughs in structure alignment algorithms, represented by Foldseek, which enable ultra-fast searching of large protein structure databases. It also covers tools for functional annotation of proteins based on their structures, including AlphaFill for ligand annotation, DeepFRI for Gene Ontology (GO) annotation, TT3D for protein-protein interaction (PPI) prediction, among others. It is proposed that 3Di sequences born concurrently with Foldseek can enhance many sequence-based deep learning models developed in the pre-AlphaFold era, enabling them to be applied to structure-based function prediction. The challenges on traditional molecular docking methods in the high-throughput era are mentioned at last, in a gesture to arouse the attention of researchers. Finally, the review explores the burgeoning field of structure data mining. Whole proteome structuring has become feasible in recent years, and scientists are processing large structure datasets from an omics viewpoint, continuously identifying analyzable elements and optimizing methodologies, as well as utilizing newly developed tools to push the boundaries. Notable examples include the identification of new protein families, the development of protein structure clustering, and the integration of AlphaFold with conventional experimental techniques to solve large structures. These advancements are paving the way for a deeper understanding of protein structure and function and have the potential to unlock new discoveries in the life sciences. However, the review also acknowledges the challenges and limitations that persist in the field, including the lack of diversity in high-throughput software for protein structural bioinformatics and the existing bottleneck in rapidly predicting protein complex structures. Overall, structural bioinformatics is expected to play an even more crucial role in the life sciences with the development of high-throughput methodology.
8.A clinical randomized controlled study on the psycho-cardiological therapy for patients with coronary atherosclerosis disease
Lijun ZHANG ; Yunpeng CHI ; Dongfang HE ; Guo LI ; Nan LU ; Yanwei LI ; Sen WANG ; Meiyan LIU
Chinese Journal of Cardiology 2024;52(9):1051-1057
Objective:To explore the prognosis efficacy of psycho-cardiological therapy and management on patients with coronary atherosclerosis disease (CAD).Methods:This was a clinical randomized controlled study. This study included inpatients with CAD at the cardiology department in Beijing Anzhen Hospital, Capital Medical University from August 2021 to January 2024. The patients enrolled in this study were asked for basic information, and received measurements for depression, anxiety, sleep quality and living quality by the scales of Patient Health Questionnaire-9 (PHQ-9), General Anxiety Disorder scale-7 (GAD-7), Athens Insomnia Scale (AIS), EuroQol 5-dimension 5-level (EQ-5D-5L) respectively. The patients were randomly grouped into a regular treatment group and a psycho-cardiological treatment group which included WeChat management or antidepressant/antianxiety medical therapy according to the situation. After the patients discharging from hospital for 2, 4, 12, 24, 48, 72, and 96 weeks, professional cardiovascular doctors would follow up by telephone, WeChat, and outpatient department, including scales (2-48 weeks), and cardiac events (2-96 weeks). Kaplan-Meier survival curve and multivariate Cox proportional hazards model were used for analyzing the association between psycho-cardiological treatment and cardiac events.Results:This study recruited a total of 552 patients with CAD, aged 61.0(54.0, 67.0) years, and 379 (68.7%) were male. There were 279(50.5%) in the regular treatment group and 273(49.5%) in the psycho-cardiological treatment group. After treatment for 4, 12 and 48 weeks, the PHQ-9 score in psycho-cardiological was significantly lower than the regular treatment group; After treatment for 12 weeks, the EQ-5D-5L effective value in psycho-cardiological group was higher than the regular treatment group; After treatment for 2, 4, 12, 24 and 48 weeks, the EQ-5D-5L VAS score in psycho-cardiological group was higher than the regular therapy group (all P<0.05). The Kaplan-Meier survival curve showed that, during the different follow-up periods, the rate of cardiac events in psycho-cardiological treatment group was lower than regular treatment group (log-rank P<0.001). The multivariate Cox proportional hazards model adjusted the factor of age, the psycho-cardiological treatment contributed to reducing the cardiac events rate by 80.3% ( HR=0.197, 95% CI: 0.067-0.582, P=0.003). Conclusion:Psycho-cardiological treatment is beneficial for improving psychological stress, living quality, and reducing cardiac events, and helps to improve prognosis and psycho-cardiological rehabilitation in CAD patients.
9.Modified Wenshen Yixin Formula (温肾益心方加减) for Coronary Heart Disease Complicated with Hypothyroidism of Spleen-kidney Yang Deficiency:A Prospective Real-world Study of 51 Cases
Aolin LI ; Zhipeng YAN ; Lu LIAN ; Qianqian ZHANG ; Chi ZHANG ; Boyu ZHU ; Lei WEI ; Zhihan YANG ; Junping ZHANG
Journal of Traditional Chinese Medicine 2024;65(20):2116-2125
ObjectiveTo observe the clinical efficacy and relative mechanism of the Modified Wenshen Yixin Formula (温肾益心方加减, MWYF) as an auxiliary treatment of coronary heart disease (CHD) complicated with hypothyroidism of spleen-kidney yang deficiency. MethodsA total of 135 CHD patients complicated with hypothyroidism and spleen-kidney yang deficiency were included and divided into control group (67 cases) and experimental group (68 cases) according to the patients' wishes of herbal medicine administration. The control group was given conventional western medicine, while the treatment group was additionally given MWYF, 1 dose per day; both groups were treated for 8 weeks. The traditional Chinese medicine (TCM) syndrome scores, angina scores, SF-36 scores, thyroid function indicators including thyroid stimulating hormone (TSH), thyroxine (T4) and triiodothyronine (T3), as well as serum cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP), monocyte chemoattractant ligand 2 (CCL2), and tumor necrosis factor-related activator protein (CD40L) levels before and after treatment were compared between the two groups. The dosage and reduction and discontinuation rate of thyroid hormone preparations after treatment were compared between the two groups. The effectiveness regarding TCM syndrome and angina pectoris was evaluated, and the safety was assessed. ResultsBias was adjusted by matching on propensity score, and 102 cases were finally included in the statistical analysis, with 51 cases in each group. The total effective rate regarding TCM syndrome [94.12% (48/51) versus 64.71% (33/51)], the total effective rate regarding angina pectoris [80.39% (41/51) versus 62.75% (32/51)], and the reduction and discontinuation rate of thyroid hormone preparation [39.21% (20/51) versus 5.88% (3/51)] were significantly higher in the experimental group than those in the control group (P<0.05 or P<0.01). After treatment, the total TCM syndrome score, individual scores of major symptoms , the major symptoms score, the secondary symptoms score, angina pectoris score, and TSH level were significantly reduced (P<0.01), while all dimensions of SF-36 scores, T4, T3, and cAMP levels significantly increased in both groups (P<0.05 or P<0.01). The dosage of thyroid hormone preparations and the levels of cGMP, CCL2, and CD40L in the experimental group significantly decreased after treatment (P<0.01). When compared between the two groups after treatment, the total TCM syndrome score, the major symptoms score, the scores of individual major symptom (chest tightness, chest pain, fear of cold, cold limbs, waist and kness soreness and weakness), the secondary symptoms score, angina pectoris score, TSH, cGMP, CCL2, and CD40L levels of the experimental group were significantly lower than those of the control group (P<0.05 or P<0.01), while all dimension scores of SF-36, T4, T3, and cAMP levels were significantly higher (P<0.01). A total of three adverse events occurred during treatment, none of which were judged to be related to the interventions of this study. ConclusionMWYF can significantly ameliorate the TCM syndrome, angina pectoris, quality of life and thyroid function in CHD patients complicated with hypothyroidism and spleen-kidney yang deficiency, and can promote the reduction and disconti-nuation of thyroid hormone preparations. The mechanism may be related to the regulation of cAMP/cGMP balance, the regulation of hypothalamic-pituitary-thyroid metabolic axis and the reduction of immune inflammation.
10.Study on the protection of gingival epithelial barrier by interleukin-22 through regulating microbiota and E-cadherin expression
Chi ZHANG ; Lu ZHANG ; Junxian REN ; Jinyu LI ; Lingping TAN ; Li GAO ; Chuanjiang ZHAO
Chinese Journal of Stomatology 2024;59(7):653-662
Objective:To investigate the regulatory effect and mechanism of interleukin-22 (IL-22) on the gingival epithelial barrier in the context of periodontal inflammation.Methods:IL-22 knockout (IL-22 KO) mice were constructed, and periodontitis mice models were established through oral gavage with polymicrobial inoculation. DNAs were extracted from the oral plaques of IL-22 KO periodontitis mice group ( n=7) and their wild-type littermates periodontitis group ( n=7) to establish a periodontitis-related oral microbiota database"PD-RiskMicroDB", determining the relationship between changes in oral microbiota and microbial function in two groups using 16S rRNA sequencing results. Gingival epithelial cells (GEC) were cultured by modified trypsinization method, and were stimulated with 100 μg/L IL-22, Porphyromonas gingivalis (Pg) (multiplicity of infection:100), separately or together for 3 and 12 hours. The experimental groups were as follows: control group (no stimulation), IL-22 group, Pg group and Pg+IL-22 group. The expression of barrier protein E-cadherin in each group at 3 h was detected by immunofluorescence, real-time fluorescence quantitative PCR (RT-qPCR) and Western blotting. Fluorescein isothiocyanate-dextran-mediated epithelial cell permeability experiment was conducted to clarify the changes in permeability of GEC in each group at 3 and 12 h. The mRNA expressions of E-cadherin in the gingival epithelium of wild-type littermates periodontitis group and IL-22 KO periodontitis group were detected by RT-qPCR. Fifteen C57BL/6 wild-type mice were randomly divided into control group ( n=5), periodontitis group ( n=5) and periodontitis+IL-22 treatment group ( n=5). RT-qPCR and immunohistochemistry (IHC) staining were used to detect the expression level of E-cadherin in the gingival epithelium of each group. Results:16S rRNA sequencing results showed that the composition of oral microbiota changed in IL-22 KO periodontitis group, of which the abundance of bacterial genera related to periodontal tissue invasion was significantly increased (linear discriminant analysis score: 2.22, P=0.009), compared with wild-type littermates periodontitis group. In vitro cell experiments showed that after Pg infection for 3 hours, the cell connections of GEC in Pg group were interrupted, and the fluorescence intensity of E-cadherin was reduced in Pg group compared with the control group. Meanwhile, the mRNA and protein expression levels of E-cadherin (mRNA: 0.69±0.12; protein: 0.60±0.12) were downregulated compared with the control group [mRNA: 1.00±0.00 ( P=0.043); protein: 1.04±0.08 ( P=0.003)], respectively. The fluorescence intensity of E-cadherin in the Pg+IL-22 group was enhanced compared with Pg group, and expression levels of E-cadherin mRNA (1.16±0.10) and protein (0.98±0.07) in Pg+IL-22 group showed a significant increase compared with Pg group [mRNA: 0.69±0.12 ( P=0.005); protein: 0.60±0.12 ( P=0.007)]. The result of epithelial permeability test showed that there was no statistical difference in epithelial permeability among control group, Pg group, IL-22 group and Pg+IL-22 group with treatment for 3 hours ( F=0.20, P=0.893). While when the treatment time turned to be 12 hours, the epithelial barrier permeability showed a significant increase in Pg group (1.39±0.15) compared with control group (1.00±0.00, P=0.027), and a decrease in Pg+IL-22 group (1.02±0.18) compared with Pg group (1.39±0.15, P=0.034). In vivo, the mRNA expression of E-cadherin in the gingival epithelium of IL-22 KO periodontitis group decreased significantly (0.32±0.21) compared with wild-type littermates periodontitis group (1.01±0.01) ( t=5.70, P=0.005). Moreover, RT-qPCR and IHC staining results showed that the mRNA expression level of E-cadherin (0.40±0.07) and absorbance value of E-cadherin positive expression (0.02±0.00) in gingival epithelial tissue of periodontitis group were both significantly down-regulated compared with control group [mRNA: 1.00±0.00 ( P=0.005); absorbance value of E-cadherin positive expression: 0.04±0.01 ( P=0.006)]. Meanwhile, the mRNA expression level of E-cadherin (1.06±0.24) and the absorbance value of E-cadherin positive expression (0.03±0.01) were both observed increase in periodontitis+IL-22 treatment group compared with periodontitis group ( P=0.003, P=0.039). Conclusions:IL-22 may exert a protective effect on the gingival epithelial barrier in an inflammatory environment by regulating the invasiveness of oral microbiota and the expression of host barrier protein.

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