1.Treatment Principles and Paradigm of Diabetic Microvascular Complications Responding Specifically to Traditional Chinese Medicine
Anzhu WANG ; Xing HANG ; Lili ZHANG ; Xiaorong ZHU ; Dantao PENG ; Ying FAN ; Min ZHANG ; Wenliang LYU ; Guoliang ZHANG ; Xiai WU ; Jia MI ; Jiaxing TIAN ; Wei ZHANG ; Han WANG ; Yuan XU ; .LI PINGPING ; Zhenyu WANG ; Ying ZHANG ; Dongmei SUN ; Yi HE ; Mei MO ; Xiaoxiao ZHANG ; Linhua ZHAO
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(5):272-279
To explore the advantages of traditional Chinese medicine (TCM) and integrative TCM-Western medicine approaches in the treatment of diabetic microvascular complications (DMC), refine key pathophysiological insights and treatment principles, and promote academic innovation and strategic research planning in the prevention and treatment of DMC. The 38th session of the Expert Salon on Diseases Responding Specifically to Traditional Chinese Medicine, hosted by the China Association of Chinese Medicine, was held in Beijing, 2024. Experts in TCM, Western medicine, and interdisciplinary fields convened to conduct a systematic discussion on the pathogenesis, diagnostic and treatment challenges, and mechanism research related to DMC, ultimately forming a consensus on key directions. Four major research recommendations were proposed. The first is addressing clinical bottlenecks in the prevention and control of DMC by optimizing TCM-based evidence evaluation systems. The second is refining TCM core pathogenesis across DMC stages and establishing corresponding "disease-pattern-time" framework. The third is innovating mechanism research strategies to facilitate a shift from holistic regulation to targeted intervention in TCM. The fourth is advancing interdisciplinary collaboration to enhance the role of TCM in new drug development, research prioritization, and guideline formulation. TCM and integrative approaches offer distinct advantages in managing DMC. With a focus on the diseases responding specifically to TCM, strengthening evidence-based support and mechanism interpretation and promoting the integration of clinical care and research innovation will provide strong momentum for the modernization of TCM and the advancement of national health strategies.
2.Expert consensus on neoadjuvant PD-1 inhibitors for locally advanced oral squamous cell carcinoma (2026)
LI Jinsong ; LIAO Guiqing ; LI Longjiang ; ZHANG Chenping ; SHANG Chenping ; ZHANG Jie ; ZHONG Laiping ; LIU Bing ; CHEN Gang ; WEI Jianhua ; JI Tong ; LI Chunjie ; LIN Lisong ; REN Guoxin ; LI Yi ; SHANG Wei ; HAN Bing ; JIANG Canhua ; ZHANG Sheng ; SONG Ming ; LIU Xuekui ; WANG Anxun ; LIU Shuguang ; CHEN Zhanhong ; WANG Youyuan ; LIN Zhaoyu ; LI Haigang ; DUAN Xiaohui ; YE Ling ; ZHENG Jun ; WANG Jun ; LV Xiaozhi ; ZHU Lijun ; CAO Haotian
Journal of Prevention and Treatment for Stomatological Diseases 2026;34(2):105-118
Oral squamous cell carcinoma (OSCC) is a common head and neck malignancy. Approximately 50% to 60% of patients with OSCC are diagnosed at a locally advanced stage (clinical staging III-IVa). Even with comprehensive and sequential treatment primarily based on surgery, the 5-year overall survival rate remains below 50%, and patients often suffer from postoperative functional impairments such as difficulties with speaking and swallowing. Programmed death receptor-1 (PD-1) inhibitors are increasingly used in the neoadjuvant treatment of locally advanced OSCC and have shown encouraging efficacy. However, clinical practice still faces key challenges, including the definition of indications, optimization of combination regimens, and standards for efficacy evaluation. Based on the latest research advances worldwide and the clinical experience of the expert group, this expert consensus systematically evaluates the application of PD-1 inhibitors in the neoadjuvant treatment of locally advanced OSCC, covering combination strategies, treatment cycles and surgical timing, efficacy assessment, use of biomarkers, management of special populations and immune related adverse events, principles for immunotherapy rechallenge, and function preservation strategies. After multiple rounds of panel discussion and through anonymous voting using the Delphi method, the following consensus statements have been formulated: 1) Neoadjuvant therapy with PD-1 inhibitors can be used preoperatively in patients with locally advanced OSCC. The preferred regimen is a PD-1 inhibitor combined with platinum based chemotherapy, administered for 2-3 cycles. 2) During the efficacy evaluation of neoadjuvant therapy, radiographic assessment should follow the dual criteria of Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1 and immune RECIST (iRECIST). After surgery, systematic pathological evaluation of both the primary lesion and regional lymph nodes is required. For combination chemotherapy regimens, PD-L1 expression and combined positive score need not be used as mandatory inclusion or exclusion criteria. 3) For special populations such as the elderly (≥ 70 years), individuals with stable HIV viral load, and carriers of chronic HBV/HCV, PD-1 inhibitors may be used cautiously under the guidance of a multidisciplinary team (MDT), with close monitoring for adverse events. 4) For patients with a poor response to neoadjuvant therapy, continuation of the original treatment regimen is not recommended; the subsequent treatment plan should be adjusted promptly after MDT assessment. Organ transplant recipients and patients with active autoimmune diseases are not recommended to receive neoadjuvant PD-1 inhibitor therapy due to the high risk of immune related activation. Rechallenge is generally not advised for patients who have experienced high risk immune related adverse events such as immune mediated myocarditis, neurotoxicity, or pneumonitis. 5) For patients with a good pathological response, individualized de escalation surgery and function preservation strategies can be explored. This consensus aims to promote the standardized, safe, and precise application of neoadjuvant PD-1 inhibitor strategies in the management of locally advanced OSCC patients.
3.Analysis of HPV Infection Characteristics and Influencing Factors for Lesion Grade in Patients with Cervical Squamous Intraepithelial Lesion and Cervical Cancer
Jingjing HAN ; Lijie ZHANG ; Ruyu CAI ; Haili LI ; He WANG ; Le DANG ; Hongda CHEN ; Ming'e LI ; Lan ZHU
Medical Journal of Peking Union Medical College Hospital 2026;17(1):156-165
To summarize the distribution characteristics of human papillomavirus(HPV) infection types in patients with cervical squamous intraepithelial lesion(SIL) and cervical cancer(CC), and to explore the impact of HPV vaccination, HPV infection types, and general clinical data on different grades of cervical lesions. Clinical data of women attending the gynecological colposcopy clinic of Shenzhen People's Hospital from January 2020 to December 2023 were retrospectively collected. Patients with HPV genotyping records and histopathologically diagnosed SIL or CC were included and divided into three groups based on pathological results: low-grade squamous intraepithelial lesion(LSIL) group, high-grade squamous intraepithelial lesion(HSIL) group, and CC group. The distribution of high-risk HPV subtypes was analyzed among the three groups, and multivariate Logistic regression was used to identify influencing factors for high-grade cervical lesions. A total of 4162 patients were included, comprising 4057 cervical SIL patients(3317 LSIL and 740 HSIL) and 105 CC patients. The overall mean age was(39.9±11.2) years. The HPV infection rate was 95.1%(3959/4162), and 25.0%(1040/4162) of patients had received HPV vaccination. Among high-risk HPV infections, HPV 52, HPV 16, HPV 58, and HPV 18 were the most common subtypes. HPV 52 had the highest infection rate in the LSIL group(27.6%), while HPV 16 was the most prevalent in the HSIL group(45.3%) and CC group(64.9%). Multivariate Logistic regression analysis showed that HPV vaccination( HPV infection is common in patients with SIL and CC, but the distribution of high-risk HPV subtypes varies among different grades of cervical lesions. It is recommended to strengthen cervical cancer screening and monitoring of key high-risk HPV infections in older and multiparous women in Shenzhen, and to continue promoting HPV vaccination.
4.Epidemiological characteristics and disease burden of liver cancer in Guangdong Province
Ying ZHANG ; Yixuan CHEN ; Rong CAO ; Yue GAO ; Yutong HAN ; Ye WANG ; Ruilin MENG ; Xueyan ZHENG ; Yu LIAO ; Zhuanping ZENG
Journal of Public Health and Preventive Medicine 2026;37(1):68-72
Objective To analyze the epidemiological characteristics and disease burden of liver cancer in Guangdong Province in 2020, and to provide a scientific foundation for the development of regionalized prevention and control strategies for liver cancer. Methods According to the cancer registry data of Guangdong Province, the incidence, mortality and age-standardized rate by Chinese standard population in 2020 were calculated to analyze the epidemiological characteristics of liver cancer. The disability adjusted life years (DALYs), year of life loss (YLL), year of lived with disability (YLD), and cause-eliminated life expectancy were used to assess the disease burden of liver cancer. Results In 2020, the crude incidence rate and the age-standardized incidence rate of liver cancer in Guangdong Province were 27.79/100 000 and 20.84/100 000,respectively, and the crude mortality rate and the age-standardized mortality rate of liver cancer were 25.49/100,000 and 17.64/100 000, respectively. The total DALY and DALY rate of liver cancer in Guangdong Province were 515 311 person-years and 513.83/100 000, respectively. After eliminating the causes of death from liver cancer, the life expectancy in Guangdong Province increased from 84.60 years to 84.99 years. All indicators consistently demonstrated that the burden of liver cancer was higher in males than that in females, and the burden of liver cancer was higher in rural areas than that in urban areas. Conclusion Liver cancer in Guangdong Province exhibits a high incidence, mortality and disease burden level in 2020. There are obvious differences of gender, age and region in cancer burden. It is necessary to strengthen liver cancer screening and diagnosis and treatment in men, the elderly and those in rural areas to reduce the burden of liver cancer gradually in Guangdong Province.
5.QingNangTCM: a parameter-efficient fine-tuning large language model for traditional Chinese medicine
Xuming TONG ; Liyan LIU ; Yanhong YUAN ; Xiaozheng DING ; Huiru JIA ; Xu YANG ; Sio Kei IM ; Mini Han WANG ; Zhang XIONH ; Yapeng WANG
Digital Chinese Medicine 2026;9(1):1-12
Objective:
To develop QingNangTCM, a specialized large language model (LLM) tailored for expert-level traditional Chinese medicine (TCM) question-answering and clinical reasoning, addressing the scarcity of domain-specific corpora and specialized alignment.
Methods:
We constructed QnTCM_Dataset, a corpus of 100 000 entries, by integrating data from ShenNong_TCM_Dataset and SymMap v2.0, and synthesizing additional samples via retrieval-augmented generation (RAG) and persona-driven generation. The dataset comprehensively covers diagnostic inquiries, prescriptions, and herbal knowledge. Utilizing P-Tuning v2, we fine-tuned the GLM-4-9B-Chat backbone to develop QingNangTCM. A multi-dimensional evaluation framework, assessing accuracy, coverage, consistency, safety, professionalism, and fluency, was established using metrics such as bilingual evaluation understudy (BLEU), recall-oriented understudy for gisting evaluation (ROUGE), metric for evaluation of translation with explicit ordering (METEOR), and LLM-as-a-Judge with expert review. Qualitative analysis was conducted across four simulated clinical scenarios: symptom analysis, disease treatment, herb inquiry, and failure cases. Baseline models included GLM-4-9B-Chat, DeepSeek-V2, HuatuoGPT-II (7B), and GLM-4-9B-Chat (freeze-tuning).
Results:
QingNangTCM achieved the highest scores in BLEU-1/2/3/4 (0.425/0.298/0.137/0.064), ROUGE-1/2 (0.368/0.157), and METEOR (0.218), demonstrating a balanced and superior normalized performance profile of 0.900 across the dimensions of accuracy, coverage, and consistency. Although its ROUGE-L score (0.299) was lower than that of HuatuoGPT-II (7B) (0.351), it significantly outperformed domain-specific models in expert-validated win rates for professionalism (86%) and safety (73%). Qualitative analysis confirmed that the model strictly adheres to the “symptom-syndrome-pathogenesis-treatment” reasoning chain, though occasional misclassifications and hallucinations persisted when dealing with rare medicinal materials and uncommon syndromes.
Conclusion
Combining domain-specific corpus construction with parameter-efficient prompt tuning enhances the reasoning behavior and domain adaptation of LLMs for TCM-related tasks. This work provides a technical framework for the digital organization and intelligent utilization of TCM knowledge, with potential value for supporting diagnostic reasoning and medical education.
6.Mechanisms of Qizhujianwei Granules in Blocking Malignant Progression of Gastric Intraepithelial Neoplasia
Yuling YU ; Yanmin WANG ; Siqi WANG ; Yateng SUN ; Yunhe WANG ; Yonghuang YAN ; Xinyu YANG ; Siqi HAN ; Yuhong SONG ; Yuhan WANG ; Cai ZHANG ; Zeqi SU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):143-151
ObjectiveTo investigate the effects of Qizhujianwei granules (QZJW) on abnormal proliferation and malignant transformation of gastric mucosal cells in rats with gastric intraepithelial neoplasia (GIN) and to explore the related mechanisms. MethodsA total of 80 SPF male Wistar rats were used. A GIN rat model was established using a four-factor comprehensive method consisting of methylnitronitrosoguanidine (MNNG), ranitidine, irregular feeding patterns, and sodium salicylate. Except for the normal group, after successful modeling, the rats were randomly divided according to body weight into a model group, a Moluodan group (0.55 g·kg-1), and a QZJW group (7.34 g·kg-1), with 12 rats in each group. All groups were treated for 8 weeks. The general characteristics of the rats and morphological changes of the gastric mucosa were observed. Histopathological changes of the gastric mucosa were examined by hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum levels of pepsinogenⅠ (PGⅠ), pepsinogenⅡ (PGⅡ), and gastrin (G-17), as well as the expression level of transforming growth factor-β1 (TGF-β1) in gastric mucosal tissue, and the PGⅠ/PGⅡ ratio was calculated. Immunohistochemistry (IHC) was used to detect the localization and expression levels of proliferating cell nuclear antigen (Ki-67) and Vimentin in gastric mucosal tissue. Western blot analysis was used to determine the protein expression levels of Wnt family member 3A (Wnt3a), β-catenin, CyclinD1, proto-oncogene Cmyc, transforming growth factor-β receptor Ⅰ (TGFβRⅠ), intracellular signaling transducers Smad2/3, phosphorylated (p)-Smad2/3, twist family transcription factor (Twist1), and Vimentin in gastric mucosal tissue. ResultsCompared with the normal group, the model group showed characteristic changes including dim eyes, pale ears and claws, dark-red tongue, and reduced luster of the tail. The gastric mucosa appeared pale, with surface congestion and erosion. The gastric mucosal glands were disordered, the nuclear-to-cytoplasmic ratio increased, and local tumor cells were observed. Serum PGⅠ and PGⅡ levels and the PGⅠ/PGⅡ ratio were significantly decreased (P<0.01), while the level of G-17 was significantly increased (P<0.01). The protein expression levels of Ki-67, Wnt3a, β-catenin, CyclinD1, Cmyc, TGF-β1, TGFβRⅠ, Smad2/3, Twist1, and Vimentin in gastric mucosal tissue were significantly increased (P<0.05, P<0.01), whereas the ratio of p-Smad2/3 to Smad2/3 was significantly decreased (P<0.05). Compared with the model group, the general characteristics and gastric mucosal conditions of rats in the Moluodan group and the QZJW group were improved. HE staining showed that QZJW could effectively block the malignant progression of GIN. Serum PGⅠ and PGⅡ levels and the PGⅠ/PGⅡ ratio were significantly increased (P<0.05, P<0.01), while the level of G-17 was significantly decreased (P<0.01). The protein expression levels of Ki-67, Wnt3a, β-catenin, CyclinD1, Cmyc, TGF-β1, TGFβRⅠ, Smad2/3, Twist1, and Vimentin in gastric mucosal tissue were significantly decreased (P<0.05, P<0.01). ConclusionQZJW have a therapeutic effect on rats with GIN. The mechanism may involve inhibition of the Wnt/β-catenin signaling pathway to regulate the cell cycle and suppress abnormal cell proliferation. Meanwhile, it may inhibit epithelial-mesenchymal transition by suppressing the TGF-β1/Smad/Twist1 signaling pathway, thereby blocking the malignant progression of GIN.
7.Construction of A Conceptual Framework for the Integration of Traditional Chinese and Western Medicine in Evolutionary Syndrome Differentiation and Treatment Across Full-cycle of Parkinson's Disease
Yu WANG ; Jianing MEI ; Hongping ZHAO ; Yunzhe TANG ; Zijun WEI ; Qinliang TAO ; Xueyi HAN ; Jiyuan HU ; Yunyun ZHANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):270-279
Parkinson's disease (PD) is a complex neurodegenerative disease involving multiple systems and neurotransmitters. Due to the high clinical heterogeneity of PD,it is urgent to establish a comprehensive and long-term traditional Chinese medicine (TCM) management model. In this paper,the conceptual framework of full-cycle management of PD is preliminarily constructed:based on the evolution of the pathophysiological mechanisms of protein deposition and neurotransmitter disorder in PD,the three-stage syndrome characteristics of the prodromal stage (predominant healthy Qi with subtle pathogenic factors),the early clinical stage (declining healthy Qi with growing pathogenic factors) and the middle and late stages (overwhelming pathogenic factors with deficient healthy Qi) are longitudinally described. Through the syndrome differentiation of visceral manifestations,the etiology and pathogenesis of PD motor and non-motor symptoms were comprehensively analyzed,while the matching treatment methods and prescriptions were inferred,and the modular scheme of the combining main symptoms,accompanying symptoms and secondary symptoms was proposed. The conceptual gap of TCM regarding motor complications ('variable syndrome') and PD-related hyperpyrexia syndrome ('critical syndrome') was explained. This framework reflects the characteristics of combination of disease and syndrome and overall constant motion,and provides new theories and research ideas for individualized and whole-process management of PD in TCM.
8.Mechanisms of Qizhujianwei Granules in Blocking Malignant Progression of Gastric Intraepithelial Neoplasia
Yuling YU ; Yanmin WANG ; Siqi WANG ; Yateng SUN ; Yunhe WANG ; Yonghuang YAN ; Xinyu YANG ; Siqi HAN ; Yuhong SONG ; Yuhan WANG ; Cai ZHANG ; Zeqi SU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):143-151
ObjectiveTo investigate the effects of Qizhujianwei granules (QZJW) on abnormal proliferation and malignant transformation of gastric mucosal cells in rats with gastric intraepithelial neoplasia (GIN) and to explore the related mechanisms. MethodsA total of 80 SPF male Wistar rats were used. A GIN rat model was established using a four-factor comprehensive method consisting of methylnitronitrosoguanidine (MNNG), ranitidine, irregular feeding patterns, and sodium salicylate. Except for the normal group, after successful modeling, the rats were randomly divided according to body weight into a model group, a Moluodan group (0.55 g·kg-1), and a QZJW group (7.34 g·kg-1), with 12 rats in each group. All groups were treated for 8 weeks. The general characteristics of the rats and morphological changes of the gastric mucosa were observed. Histopathological changes of the gastric mucosa were examined by hematoxylin-eosin (HE) staining. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum levels of pepsinogenⅠ (PGⅠ), pepsinogenⅡ (PGⅡ), and gastrin (G-17), as well as the expression level of transforming growth factor-β1 (TGF-β1) in gastric mucosal tissue, and the PGⅠ/PGⅡ ratio was calculated. Immunohistochemistry (IHC) was used to detect the localization and expression levels of proliferating cell nuclear antigen (Ki-67) and Vimentin in gastric mucosal tissue. Western blot analysis was used to determine the protein expression levels of Wnt family member 3A (Wnt3a), β-catenin, CyclinD1, proto-oncogene Cmyc, transforming growth factor-β receptor Ⅰ (TGFβRⅠ), intracellular signaling transducers Smad2/3, phosphorylated (p)-Smad2/3, twist family transcription factor (Twist1), and Vimentin in gastric mucosal tissue. ResultsCompared with the normal group, the model group showed characteristic changes including dim eyes, pale ears and claws, dark-red tongue, and reduced luster of the tail. The gastric mucosa appeared pale, with surface congestion and erosion. The gastric mucosal glands were disordered, the nuclear-to-cytoplasmic ratio increased, and local tumor cells were observed. Serum PGⅠ and PGⅡ levels and the PGⅠ/PGⅡ ratio were significantly decreased (P<0.01), while the level of G-17 was significantly increased (P<0.01). The protein expression levels of Ki-67, Wnt3a, β-catenin, CyclinD1, Cmyc, TGF-β1, TGFβRⅠ, Smad2/3, Twist1, and Vimentin in gastric mucosal tissue were significantly increased (P<0.05, P<0.01), whereas the ratio of p-Smad2/3 to Smad2/3 was significantly decreased (P<0.05). Compared with the model group, the general characteristics and gastric mucosal conditions of rats in the Moluodan group and the QZJW group were improved. HE staining showed that QZJW could effectively block the malignant progression of GIN. Serum PGⅠ and PGⅡ levels and the PGⅠ/PGⅡ ratio were significantly increased (P<0.05, P<0.01), while the level of G-17 was significantly decreased (P<0.01). The protein expression levels of Ki-67, Wnt3a, β-catenin, CyclinD1, Cmyc, TGF-β1, TGFβRⅠ, Smad2/3, Twist1, and Vimentin in gastric mucosal tissue were significantly decreased (P<0.05, P<0.01). ConclusionQZJW have a therapeutic effect on rats with GIN. The mechanism may involve inhibition of the Wnt/β-catenin signaling pathway to regulate the cell cycle and suppress abnormal cell proliferation. Meanwhile, it may inhibit epithelial-mesenchymal transition by suppressing the TGF-β1/Smad/Twist1 signaling pathway, thereby blocking the malignant progression of GIN.
9.Construction of A Conceptual Framework for the Integration of Traditional Chinese and Western Medicine in Evolutionary Syndrome Differentiation and Treatment Across Full-cycle of Parkinson's Disease
Yu WANG ; Jianing MEI ; Hongping ZHAO ; Yunzhe TANG ; Zijun WEI ; Qinliang TAO ; Xueyi HAN ; Jiyuan HU ; Yunyun ZHANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(10):270-279
Parkinson's disease (PD) is a complex neurodegenerative disease involving multiple systems and neurotransmitters. Due to the high clinical heterogeneity of PD,it is urgent to establish a comprehensive and long-term traditional Chinese medicine (TCM) management model. In this paper,the conceptual framework of full-cycle management of PD is preliminarily constructed:based on the evolution of the pathophysiological mechanisms of protein deposition and neurotransmitter disorder in PD,the three-stage syndrome characteristics of the prodromal stage (predominant healthy Qi with subtle pathogenic factors),the early clinical stage (declining healthy Qi with growing pathogenic factors) and the middle and late stages (overwhelming pathogenic factors with deficient healthy Qi) are longitudinally described. Through the syndrome differentiation of visceral manifestations,the etiology and pathogenesis of PD motor and non-motor symptoms were comprehensively analyzed,while the matching treatment methods and prescriptions were inferred,and the modular scheme of the combining main symptoms,accompanying symptoms and secondary symptoms was proposed. The conceptual gap of TCM regarding motor complications ('variable syndrome') and PD-related hyperpyrexia syndrome ('critical syndrome') was explained. This framework reflects the characteristics of combination of disease and syndrome and overall constant motion,and provides new theories and research ideas for individualized and whole-process management of PD in TCM.
10.Mechanisms of Huanglian Jiedutang and Its Major Active Constituents in Inhibiting LPS-induced M1 Polarisation of BV2 Microglia
Haojia ZHANG ; Kai WANG ; Kunjing LIU ; Xin LAN ; Zijin SUN ; Chunyu WANG ; Wenyuan MA ; Wei SHAO ; Jinhua HAN ; Liyang DONG ; Changxiang LI ; Xueqian WANG ; Youxiang CUI ; Fafeng CHENG ; Qingguo WANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(11):44-55
ObjectiveTo investigate whether Huanglian Jiedutang (HLJD) and its major active constituents (geniposide, baicalin, and berberine) can inhibit the inflammatory response of BV2 cells under lipopolysaccharide (LPS) stimulation via the high-mobility group protein B1 (HMGB1)/Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway, and to explore differences in therapeutic efficacy among the three monomers, their combined formula, and HLJD under equal content ratios. MethodsBV2 microglial cells were used as the primary experimental model. Cell viability was assessed using the cell counting kit-8 (CCK-8) method to examine the effects of different concentrations of dimethyl sulfoxide (DMSO, 0.8%, 0.4%, 0.2%, 0.1%, and 0.05%) on cell viability. IncuCyte was employed to monitor the growth of cells under different concentrations of HLJD (200, 100, 50, 25, 12.5, 6.25 mg·L-1). Nitric oxide (NO) assay was used to screen the optimal HLJD concentration. High-performance liquid chromatography (HPLC) determined the content of geniposide, baicalin, and berberine in HLJD, and experimental groups were subsequently established according to the relative proportions of these constituents. CCK-8 assay evaluated cell viability under different treatments. Enzyme-linked immunosorbent assay (ELISA) measured levels of inflammatory factors (TNF-α, IL-1β, IL-6, IL-10) in the supernatant. Flow cytometry assessed the effects of treatments on M1-type polarization of BV2 cells. Western blot determined the expression levels of HMGB1, TLR4, and NF-κB-related proteins. ResultsCompared with the blank group, DMSO at concentrations ≤0.2% did not affect cell viability within 48 h. BV2 cell growth plateaued at 24 h after treatment with 200 mg·L-1 HLJD. Under stimulation with 2 mg·L-1 LPS, this concentration of HLJD effectively reduced NO release, and 6 h pre-treatment had a stronger inhibitory effect on NO than direct administration. HPLC results showed that 1 mg of HLJD freeze-dried powder contained approximately 24 μg of geniposide, 15 μg of baicalin, and 30 μg of berberine. Based on these ratios, experimental groups were blank, LPS (2 mg·L-1), HLJD (200 mg·L-1), monomer combination, geniposide (4.8 mg·L-1), baicalin (3 mg·L-1), and berberine (6 mg·L-1). The monomer combination group consisted of all three active constituents dissolved together. LPS and HLJD or its active constituents did not affect cell viability compared with the blank group. LPS significantly increased TNF-α, IL-1β, IL-6, and IL-10 in the supernatant (P<0.01). HLJD and its active constituents significantly reduced pro-inflammatory factors TNF-α, IL-1β, and IL-6 (P<0.05, P<0.01) while upregulating anti-inflammatory IL-10 (P<0.01), with the monomer combination showing the strongest effect (P<0.05, P<0.01). Compared with the blank group, LPS significantly increased the proportion of CD80⁺CD86⁺ (M1-type) BV2 cells (P<0.01). HLJD and its constituents partially inhibited M1 polarization (P<0.05, P<0.01), with the monomer combination exhibiting the most pronounced effect (P<0.05, P<0.01). Compared with the blank group, LPS upregulated HMGB1, TLR4, and NF-κB-related proteins (P<0.01), whereas HLJD and its active constituents significantly reduced their expression (P<0.05, P<0.01), with the monomer combination having the strongest regulatory effect (P<0.05, P<0.01). ConclusionHLJD and its major active constituents (geniposide, baicalin, berberine) can inhibit LPS-induced inflammatory responses in BV2 cells. The combination of the three active constituents demonstrates the most potent anti-inflammatory effect, significantly attenuating M1-type polarization of BV2 cells via the HMGB1/TLR4/NF-κB signaling pathway.


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