1.Proctor's Reporting Guideline for Implementation Strategies: Interpretation, Application, and Challenges
Jiangyun CHEN ; Jinghan LIU ; Youping ZHUANG ; Xueying CHEN ; Siyuan LIU ; Xiaoshan CHEN ; Yeqing ZHAN ; Dongmei ZHONG ; Huadan HUANG ; Dong XU
Medical Journal of Peking Union Medical College Hospital 2026;17(1):263-273
The Proctor's reporting guideline for implementation strategies represents a landmark framework in the field of implementation science, aiming to address the issue of inconsistent reporting in implementation research by standardizing the naming, definition, and operationalization of implementation strategies, thereby enhancing the credibility and utility of research findings. This paper provides an in-depth interpretation of the core connotations of this reporting guideline and illustrates its application in developing interview outlines and specifying implementation strategies, using a brief smoking cessation intervention project as a case study. Through this reporting guideline, abstract recommendations for implementation are systematically transformed into clear, multidimensional operational guides, significantly improving the transparency of strategy connotations and the replicability of actual execution. Meanwhile, the case study highlights the flexibility of the guideline, which allows researchers to adapt the content and format of strategies based on local resources and cultural contexts, thus enhancing practical adaptability while maintaining scientific rigor. However, the application of Proctor's reporting guideline still faces challenges, primarily manifested in the potential confusion surrounding the constructs of temporality and dose in practice, as well as the challenges that the inherent flexibility of the guideline may pose to the assessment of fidelity and effectiveness. Despite these limitations, the reporting guideline remains a vital tool for implementation research; future efforts should focus on optimizing its application—through refining operational guidelines, standardizing flexible adaptations, and involving stakeholders—to better guide implementation studies and continuously promote high-quality development in the field.
2.Application of frontal electroencephalogram in assessment of anxiety, depression,and sleep quality in adolescents with emotional disorders
Ying YANG ; Haiting XU ; Weigang PAN ; Xinpeng XU ; Siyuan LIU ; Xiaohong LI
Sichuan Mental Health 2026;39(2):112-118
BackgroundEmotional disorders in adolescents have emerged as a prominent issue in recent years. Current mainstream clinical assessment approaches for such conditions predominantly rely on interviews and rating scales, which are limited by inherent drawbacks such as high subjectivity and recall bias. Accordingly, there exists an urgent clinical need for the development of objective, quantifiable auxiliary diagnostic tools.In previous studies, frontal electroencephalography (EEG) has demonstrated significant value in assessing depressive and anxiety. However, the lack of standardized quantitative metrics and intuitive visual analytical approaches has severely restricted clinical interpretability of EEG data and diminished patient engagement. To address these key limitations, the present study proposes an innovative analytical framework that converts frontal EEG signals into quantifiable visual metrics to enhance clinical comprehension and acceptance. ObjectiveTo explore the value of frontal EEG in assessing anxiety, depression, and sleep quality in adolescents with emotional disorders, with the aim of providing objective auxiliary tools for clinical diagnosis and assessment of adolescents with emotional disorders. MethodsThis cross-sectional study recruited 105 adolescents aged 12-18 years who visited the outpatient department of a specialized mental hospital in Beijing from April 2023 to April 2024. All participants met the diagnostic criteria for mood (affective) disorders or anxiety disorders in the International Classification of Diseases, tenth edition (ICD-10). Frontal EEG signals were collected within a big data analytics-driven framework and further processed by EEG system to generate six quantitative cerebral function indices, namely brain load, tension and excitement, emotional stress, sleepiness index, cerebral vitality, and cerebral fatigue. In addition, validated standardized scales, including the Self-rating Anxiety Scale (SAS), the Self-rating Depression Scale (SDS), and the Pittsburgh Sleep Quality Index (PSQI), were administered for anxiety, depressive symptoms, and sleep quality, respectively. ResultsIn adolescent patients with emotional disorders, the SAS score exhibited significant positive correlations with brain load (rs=0.328, P<0.01), emotional stress (rs=0.341, P<0.01), and cerebral fatigue (rs=0.286, P<0.01). The SDS score was positively correlated with brain load (rs=0.275, P<0.01), emotional stress (rs=0.241, P<0.05), and cerebral fatigue (rs=0.311, P<0.01), while showing a significant negative correlation with cerebral vitality (rs=-0.212, P<0.05). Additionally, the PSQI total score demonstrated positive correlations with brain load (rs=0.340, P<0.01), emotional stress (rs=0.322, P<0.01), and cerebral fatigue (rs=0.229, P<0.05). ConclusionFrontal EEG-derived indices, including brain load, emotional stress, cerebral fatigue and cerebral vitality, may serve as objective markers for reflecting anxiety, depression, and sleep quality in adolescents with emotional disorders. [Funded by Beijing High level Innovation and Entrepreneurship Talent Support Program (number, 202504841041); Horizontal Joint Project]
3.Consistent Detection of Aquaporin-4 Antibodies:A Comparative Analysis Between Fixed and Live Cell-Based Assays
Jing WANG ; Linge WANG ; Xiaolin YANG ; Zhizhong LI ; Jinyu JIANG ; Qiao XU ; Siyuan HUANG ; Qing FU ; Yang YANG ; Rongrong ZHANG ; Lin YANG ; Ai CHEN ; Xiaopeng ZENG ; Ke XU ; Peng ZHENG ; Xinyue QIN ; Jinzhou FENG
Journal of Clinical Neurology 2026;22(2):212-220
Background:
and Purpose Live cell-based assays (LCBA) are increasingly used for serological antibody detection due to their ability to preserve antigen conformation, offering moderately higher sensitivity than fixed cell-based assays. However, the clinical necessity of prioritizing LCBA for the detection of aquaporin-4 immunoglobulin G (AQP4-IgG) in neuromyelitis optica spectrum disorder (NMOSD) remains unclear, especially when compared to its established role in diagnosing myelin oligodendrocyte glycoprotein antibody-associated disease.
Methods:
We compared the performance of live cell-based assays using immunofluorescence (LCBA-IF) and fixed cell-based assays using immunofluorescence (FCBA-IF) in detecting AQP4-IgG in 90 cases of NMOSD meeting 2015 International Panel for Neuromyelitis Optica Diagnosis criteria, alongside 40 controls. Additionally, we further investigated the relationship between AQP4-IgG titers as measured by LCBA-IF and FCBA-IF and clinical parameters in NMOSD patients.
Results:
Results showed 96.9% agreement between LCBA-IF and FCBA-IF (Cohen’s κ=0.935), with a strong Spearman correlation (0.977, p<0.001). Both methods demonstrated 100% specificity, with LCBA-IF showing slightly higher sensitivity compared to FCBA-IF. Within LCBAIF-tested groups, statistically significant differences in annualized relapse rates were observed across all pairwise comparisons (low-titer vs. moderate-titer, low-titer vs. high-titer, and moderate-titer vs. high-titer; all p<0.050). However, this association reached statistical significance in some FCBA-IF-tested groups.
Conclusions
Overall, there is a strong concordance between LCBA-IF and FCBA-IF in detecting AQP4-IgG, where LCBA-IF shows slightly higher sensitivity. Furthermore, there is a potential link between elevated AQP4-IgG titers and an increased risk of relapse, and this correlation may appear more pronounced when using LCBA-IF.
4.Role of exosomal miR-320c in gingerol-mediated defense against Staphylococcus aureus infection
Zhencai XING ; Mengxue XU ; Xiang KONG ; Jinghan SUN ; Zhen MA ; Yu GAO ; Siyuan DU ; Hong ZHENG ; Yakun LIU
Acta Universitatis Medicinalis Anhui 2026;61(5):795-802
ObjectiveTo investigate the inhibitory effect of gingerol, an active component of ginger, on Staphylococcus aureus (S. aureus) infection, and to preliminarily explore its mechanism related to extracellular vesicles (EVs). MethodsS.aureus infection models were established in human umbilical vein endothelial cell (HUVECs), Vero E6 cells, and C57BL/6 mice. Experimental groups included control, infection, and gingerol-treated groups. Bacterial load and VE-cadherin protein expression were detected using immunofluorescence and Western blot. EVs were isolated by size exclusion chromatography and characterized by transmission electron microscopy and nanoparticle tracking analysis. miRNA sequencing was performed on EVs. ResultsGingerol treatment significantly reduced the bacterial load in both in vitro and in vivo infection models and upregulated VE-cadherin expression. miRNA sequencing of EVs revealed that S. aureus infection upregulated the expression of hsa-miR-320c, while gingerol treatment reversed this abnormal expression. Bioinformatic analysis further predicted that the target genes of hsa-miR-320c were significantly enriched in cell junction-related pathways. ConclusionGingerol exhibits clear antibacterial and host-protective effects, by regulating hsa-miR-320c in EVs to maintain endothelial barrier integrity.
5.Quality Evaluation of Commercially Available Huoxiang Zhengqi Preparations
Yue GUO ; Siyuan LI ; Qingxia XU ; Sha CHEN ; Shuangyan LI ; Jipeng DI ; An LIU ; Yan LIU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(19):238-247
ObjectiveTo establish a comprehensive evaluation framework combining multi-component quantification with daily intake in response to the challenges posed by multiple coexisting dosage forms, single-faceted quality evaluation, and ambiguous clinical equivalence of Huoxiang Zhengqi preparations, and to compare the quality differences and superiority among eight commercially available dosage forms. MethodsA total of 75 batches of Huoxiang Zhengqi preparations from 25 manufacturers were investigated. Ultra-performance liquid chromatography-triple quadrupole-tandem mass spectrometry (UPLC-QqQ-MS/MS) coupled with gas chromatography-triple quadrupole-tandem mass spectrometry (GC-QqQ-MS/MS) was adopted to simultaneously quantify 20 core marker components. Normalization was performed based on equivalent raw herb concentrations to eliminate interferences from dosage forms, excipients, and specifications. Considering the specified clinical dosage, the actual daily intake level was calculated to comprehensively evaluate component retention, content uniformity, and potential differences in in vivo exposure among different dosage forms. ResultsThe quality of Huoxiang Zhengqi preparations differed greatly among dosage forms and manufacturers. On the basis of the recommended daily doses in the package inserts, the total daily intake of the 20 tested components ranged from 3.14 to 614.30 mg·d-1, while that of hesperidin, a quality-control marker, ranged from 0.03 to 40.65 mg·d-1. Atractylodin was detected only in tinctures and soft capsules, with concentrations ranging from (2.68±0.01) to (5.73±0.37) mg·L-1. The soft capsules from manufacturer M22 had the highest atractylodin level, while atractylodin was not detected in the other dosage forms. Nucleosides were most abundant in the oral liquids and tinctures (M4 to M13). The tincture from manufacturer M5 showed the highest nucleoside level of 29.12 mg·L-1, whereas the granules had the lowest nucleoside levels, with that of M23 being only 3.48 mg·L-1. Perillae Folium-related components varied greatly among batches, with an relative standard deviation (RSD) of 87.55%. The daily intake of hesperidin from all three granule products was below 3.6 mg·d-1, the reference value calculated for Huoxiang Zhengqi Shui according to the 2025 edition of the Chinese Pharmacopoeia. ConclusionThe levels of active components differed significantly among different dosage forms of Huoxiang Zhengqi preparations. Tinctures contained more types and higher overall levels of active components, while soft capsules were better at preserving volatile components. Granules and dripping pills had lower overall levels of active components. The comprehensive evaluation method established in this study can provide a basis for the quality control, process optimization, and the selection of high-quality products.
6.Sexual behaviors, HIV/AIDS knowledge awareness, and novel synthetic drug use among young students in Guangdong Province
Chinese Journal of School Health 2026;47(8):1102-1105
Objective:
To assess the status of sexual behaviors, HIV/AIDS knowledge awareness, and novel synthetic drug use among school attending young students in Guangdong Province, so as to provide evidence base for formulating targeted intervention strategies.
Methods:
From September to December 2024, a convenience sampling method was employed to conduct an anonymous online survey among 41 119 students from 20 higher education institutions and vocational colleges in Guangdong Province. Chi square test was utilized for rate comparisons and univariate analysis, and a multivariate Logistic regression model was applied to analyze factors associated with novel synthetic drug use.
Results:
The self reported rate of sexual behavior was 5.2%. Among sexually active students, the rates of unprotected sexual intercourse, having temporary sexual partners, and having commercial sexual partners were 32.4%, 34.5%, and 26.3%, respectively. The awareness rate of HIV related knowledge was 75.7%. The overall use rate of novel psychoactive substances(NPS) was 1.0%, and among sexually active students it was 6.5%. Multivariate Logistic regression analysis showed that students with sexual experience ( OR =4.15) and those with HIV testing history ( OR =9.38) had higher risks of NPS use; among sexually active students, those with unprotected sexual intercourse ( OR =6.83) and those with commercial sexual partners ( OR = 4.68 ) had higher risks of NPS use (all P <0.05).
Conclusions
The HIV/AIDS knowledge level among surveyed students needs further improvement, and high risk sexual behaviors are cross linked with novel synthetic drug use. It is recommended to carry out integrated health education for all students and implement comprehensive targeted interventions for sexually active students.
7.Serial mediating roles of cognitive distortion and hopelessness in the associations of stressful life events with subclinical depressive and anxiety symptoms
Xiaolin SUN ; Xinpeng XU ; Weigang PAN ; Ying YANG ; Siyuan LIU ; Xiaohong LI
Sichuan Mental Health 2026;39(4):375-380
BackgroundStressful life events are associated with subclinical depressive and anxious symptoms, which may increase the risk of subsequent depressive and anxiety disorders. However, the mechanisms linking stressful life events to subclinical emotional symptoms remain unclear. ObjectiveTo explore the clinical characteristics of individuals with subclinical depression and anxiety following stressful events, and to analyze the mechanisms by which stressful events affect subclinical depressive and anxious symptoms, so as to provide a reference for the early identification and intervention of emotional symptoms in individuals under stress. MethodsFrom December 2024 to April 2025, 80 individuals with subclinical depression and/or anxiety (study group) and 80 healthy controls were recruited via the online official accounts of Beijing Anding Hospital, Capital Medical University and Beijing Huilongguan Hospital, Capital Medical University. Participants completed the Patients' Health Questionnaire Depression Scale-9 item (PHQ-9), the Generalized Anxiety Disorder Scale-7 item (GAD-7), the Life Event Scale (LES), the Self-rating Idea of Suicide Scale (SIOSS), and the Dysfunctional Attitude Scale (DAS). Spearman rank correlation analysis was used to examine the associations in the study group. Serial mediation analyses were conducted using model 6 of Process version 4.1, to test whether cognitive distortion and hopelessness mediated the associations of stressful life events with subclinical depressive and anxious symptoms. ResultsCompared with healthy controls, participants with subclinical depressive and anxious symptoms had higher scores on PHQ-9, GAD-7, LES, SIOSS hopelessness subscale, and DAS scores (Z/t=10.294, 9.197, 4.659, 6.918, 3.517, P<0.01). In the study group, GAD-7 score was positively correlated with LES, DAS, and SIOSS hopelessness subscale scores (rs=0.233, 0.465, 0.588, P<0.05); SIOSS hopelessness subscale score was positively correlated with DAS and LES scores (rs=0.393, 0.309, P<0.01) and cognitive distortion was positively correlated with stressful life events(rs=0.432, P<0.01). Stressful life events were directly associated with subclinical anxiety symptoms (β=0.040, 95% CI: 0.015–0.065, P<0.05), and showed a significant serial indirect association with subclinical anxiety symptoms (indirect effect=0.006, 95% CI: 0.001–0.017, P<0.01), accounting for 7.15% of the total effect. Stressful life associated with subclinical depressive symptoms (β=0.087, 95% CI: 0.060–0.115, P<0.01), however, the indirect effect through cognitive distortion and hopelessness were not statistically significant. ConclusionAfter experiencing stressful events, individuals with subclinical depression and anxiety exhibit more pronounced cognitive distortions and hopelessness. Stressful events can both directly affect subclinical anxiety symptoms and indirectly affect them through the serial mediating effects of cognitive distortion and hopelessness. Stressful events have a direct effect on subclinical depressive symptoms, whereas the mediating effects of cognitive distortion and hopelessness are no statistically significant. [Funded by the Training Program for the Development of High-level Public Health Technical Talents (number, Academic Leader-01-22); www.chictr.org.cn number, ChiCTR2400093516]
8.Molecular Mechanism of Astragali Radix and Hedyotis diffusa Regulating Sp1-miR-582-3p-p27 Axis to Inhibit Lung Adenocarcinoma Proliferation
Haipeng SUN ; Lufan XU ; Xue LIU ; Siyuan LIU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(4):116-124
ObjectiveTo investigate the effects of excessive activation of the Specific protein 1 (Sp1)-MicroRNA-582-3p (miR-582-3p)-cyclin-dependent kinase inhibitor (p27) axis on the cell cycle and proliferation of A549 lung adenocarcinoma cells at the cellular level, thereby investigating the possible mechanisms by which Astragali Radix and Hedyotis diffusa(A-H) regulate the axis to inhibit A549 cells proliferation. MethodsLentiviral plasmid transfection technology was used to obtain four stable A549 transgenic cell lines: shRNA-Sp1+LV-miR-582-3p mimic, shRNA-Sp1+LV-mNC, shRNA-NC+LV-miR-582-3p mimic, and shRNA-NC+LV-mNC. Real-time PCR was used to detect the effect of Sp1 on miR-582-3p expression in A549 lung adenocarcinoma cells. CCK-8 and colony formation assay were used to detect the effect of Sp1 on cell proliferation through miR-582-3p, while flow cytometry was used to detect the effect of Sp1 on cell cycle via miR-582-3p. Western blot was used to detect the effect of Sp1 on cyclin in A549 cells through miR-582-3p. CCK-8 was employed to detect the effects of different concentrations (5%, 10%, 20%, 40%, 80%) of A-H on A549 cell viability, and the optimal concentration was selected for subsequent mechanism exploration. Stable transgenic strains oe-Sp1 and control oe-Vector were constructed using the above-mentioned transfection techniques. Real-time PCR was then used to investigate the effect of A-H on miR-582-3p expression in A549 cells, while Western blot was employed to detect the effect of A-H on Sp1 and p27 expression in A549 lung adenocarcinoma cells. ResultsCompared with the shRNA-NC+LV-mNC group, the expression of miR-582-3p was significantly inhibited in the shRNA-Sp1+LV-mNC group (P<0.01). Compared with the shRNA-Sp1+LV-mNC group, the shRNA-Sp1+LV-miR-582-3p mimic group significantly reversed miR-582-3p expression (P<0.01), indicating that Sp1 has a significant regulatory effect on miR-582-3p. Compared with the shRNA-Sp1+LV-mNC group, the shRNA-Sp1+LV-miR-582-3p mimic group significantly reversed cell proliferation (P<0.01), suggesting that miR-582-3p can partially reverse the proliferative effect of Sp1 on A549 lung adenocarcinoma. Compared with the shRNA-Sp1+LV-mNC group, the shRNA-Sp1+LV-miR-582-3p mimic group showed a significant decrease in G0/G1 proportions, while the G2/M and S phase proportions increased significantly (P<0.01), indicating that miR-582-3p can partially reverse the effect of Sp1 on the A549 cell cycle. Compared with the shRNA-Sp1+LV-mNC group, the shRNA-Sp1+LV-miR-582-3p mimic group significantly reversed the expression of various proteins with the decrease of p27 and the increase of Cyclin D1, CDK4, Cyclin E, CDK2, p-Rb, and E2F3 (P<0.01), indicating that miR-582-3p can partially reverse the effect of Sp1 on the cell cycle of A549 lung adenocarcinoma. Compared with the blank group, the serum containing A-H significantly inhibited A549 cell viability in a concentration-dependent manner. Therefore, 10% A-H was selected for intervention of subsequent mechanism experiments. Compared with A-H+oe-Vector, A-H+oe-Sp1 significantly reversed the downregulation of Sp1, miR-582-3p, and the upregulation of p27 (P<0.01). This suggests that Sp1 can partially reverse the effects of A-H on miR-582-3p and p27 levels in A549 lung adenocarcinoma cells. ConclusionOveractivation of the Sp1-miR-582-3p-p27 axis significantly promotes the proliferation of A549 lung adenocarcinoma cells, and the potential mechanism of the inhibitory effect of A-H on the proliferation of A549 cells may be related to its inhibition on the overactivation of the Sp1-miR-582-3p-p27 axis.
9.Effects of wogonin on joint inflammation in collagen-induced arthritis rats via the endoplasmic reticulum stress pathway
Yuru WANG ; Siyuan LI ; Ye XU ; Yumeng ZHANG ; Yang LIU ; Huiqin HAO
Chinese Journal of Tissue Engineering Research 2025;29(5):1026-1035
BACKGROUND:Rheumatoid arthritis is an inflammatory disease.Many studies have shown that wogonin has a good anti-inflammatory effect on rheumatoid arthritis,but its exact efficacy and specific mechanism of action remain to be clarified. OBJECTIVE:To investigate the mechanism of wogonin ameliorating joint inflammation by regulating endoplasmic reticulum stress pathway in rats with collagen-induced arthritis. METHODS:(1)At the animal level:Female Wistar rats were divided into healthy control group,arthritis model group and wogonin treatment group.Rat models of arthritis in the latter two groups were established by subcutaneous injection of bovine type Ⅱ collagen and adjuvant.In the wogonin group,wogonin was given by gavage for 28 consecutive days after modeling.During this period,the rats in each group were weighed,and arthritis score and ankle swelling were measured every 7 days.After the experiment,the pathological changes of the joint were observed,the mRNA and protein levels of endoplasmic reticulum stress pathway GRP78 and CHOP were detected by qRT-PCR,western blot,and immunohistochemistry.(2)At the cellular level,cell counting kit-8 was used to detect the cytotoxic effect of wogonin on fibroblast-like synoviocytes from rats with collagen-induced arthritis.The fibroblast-like synoviocytes induced by thapsigargin were treated with different concentrations of wogonin.The levels of interleukin-1β and tumor necrosis factor-α in the cell supernatant were detected by ELISA,and the intracellular reactive oxygen species in each group were determined by DCFH-DA probe method.The mRNA and protein levels of GRP78,IRE1α,XBP1s and CHOP were detected by qRT-PCR and western blot,respectively. RESULTS AND CONCLUSION:Compared with the healthy control group,arthritis index score and ankle swelling degree in the arthritis model group were increased(P<0.01),synovial hyperplasia,inflammatory cell infiltration,cartilage destruction and bone erosion were observed in pathological sections,and the mRNA and protein expressions of GRP78 and CHOP in the ankle were significantly increased(P<0.01),which were mainly located in synovial tissue and articular surface.Compared with the arthritis model group,the arthritis index score and ankle swelling degree in the wogonin treatment group were decreased(P<0.05),synovial hyperplasia and the number of inflammatory cells were decreased,cartilage destruction and bone erosion were alleviated,the mRNA and protein expression levels of GRP78 and CHOP in the ankle were decreased(P<0.05),particularly in synovial tissue and on the articular surface.There was no significant difference in body mass among the three groups(P>0.05).In the cell experiment,200 μmol/L wogonin significantly reduced the survival rate of fibroblast-like synoviocytes(P<0.01).Compared with the blank control group,the levels of interleukin-1β,tumor necrosis factor-α,content of reactive oxygen species,and mRNA and protein expression of GRP78,IRE1α,XBP1s,and CHOP in the thapsigargin group were significantly increased(P<0.05);compared with the thapsigargin group,50 and 100 μmol/L wogonin significantly reduced the levels of interleukin-1β and tumor necrosis factor-α in the cell supernatant(P<0.05,P<0.01),and 100 μmol/L wogonin significantly reduced the content of reactive oxygen species(P<0.01)and down-regulated the mRNA and protein expression levels of GRP78,IRE1α,XBP1s and CHOP(all P<0.05).These results suggest that wogonin can effectively alleviate joint inflammatory responses in rats with collagen-induced arthritis,and the endoplasmic reticulum stress pathway may be the key target of its intervention.
10.Material basis and action mechanism of drug-containing serum of Modified Erxian Pill inhibiting macrophage pyroptosis
Siyuan LI ; Yuru WANG ; Ye XU ; Di GUO ; Nan NAN ; Yang LIU ; Jie ZHAO ; Huiqin HAO
Chinese Journal of Tissue Engineering Research 2025;29(19):4029-4037
BACKGROUND:Our previous study found that Modified Erxian Pill could alleviate inflammation in collagen-induced arthritis rats,but its mechanism needs to be further verified. OBJECTIVE:To analyze the components absorbed in the blood of Modified Erxian Pill,and observe the effect of the drug-containing serum of Modified Erxian Pill on pyroptosis of J774A.1 macrophages. METHODS:(1)Analysis of components absorbed in the blood of Modified Erxian Pill:Ultra-high performance liquid chromatography-high resolution mass spectrometry was used to detect and identify Modified Erxian Pill and its components absorbed in the blood.(2)Effect of the drug-containing serum of Modified Erxian Pill on pyroptosis of J774A.1 macrophages:Molecular docking technology was used to initially verify the sesquiterpenoids and NLRP3 in components absorbed in the blood of Modified Erxian Pill.J774A.1 macrophages were randomly divided into blank control group,lipopolysaccharide+adenosine triphosphate group,and lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill with low(2.5%),medium(5%),and high(10%)dose groups.The release of lactate dehydrogenase in the cell supernatant of each group was detected according to the kit instructions.The levels of interleukin-1β and interleukin-18 in cell supernatant were detected in each group by ELISA.The cell membrane damage was detected by Hoechst/PI staining.The expression levels of NLRP3,Caspase-1,GSDMD,and GSDMD-N protein in the cells of each group were detected by western blot assay. RESULTS AND CONCLUSION:(1)A total of 32 active components of Modified Erxian Pill were identified,and 21 components entered the blood.The main components into blood included a variety of sesquiterpenoids.(2)Molecular docking results showed that 3-O-Acetyl-13-deoxyphomenone,Incensol oxide,Atractylenolide III,Rupestonic acid,and 3,7-Dihydroxy-9,11-eremophiladien-8-one had good binding activity with NLRP3.(3)Compared with the blank control group,lactate dehydrogenase activity and the expression levels of interleukin-1β and interleukin-18 were significantly increased in cell supernatant of lipopolysaccharide+adenosine triphosphate group(P<0.001).Hoechst/PI staining showed that the number of PI-positive cells was significantly increased.After the intervention of lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill group,all of them showed different degrees of reduction.(4)Compared with the blank control group,NLRP3,Caspase-1,GSDMD,and GSDMD-N protein expression levels were significantly increased in the lipopolysaccharide+adenosine triphosphate group(P<0.05).Compared with lipopolysaccharide+adenosine triphosphate group,the protein expressions of NLRP3,Caspase-1,GSDMD,and GSDMD-N were significantly decreased in the lipopolysaccharide+adenosine triphosphate+Modified Erxian Pill group(P<0.05),and had a certain dose dependence.These findings verify that the drug-containing serum of Modified Erxian Pill may inhibit the pyroptosis of J774A.1 macrophages by regulating the NLRP3/Caspase-1/GSDMD pathway.


Result Analysis
Print
Save
E-mail