1.Mechanism of Gegen Qinliantang in Regulating Microglia Polarization to Improve Diabetic Cognitive Impairment
Hui FENG ; Chunxiang ZHOU ; Tianyi REN ; Weiwei TAO ; Yun LING
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):1-10
ObjectiveTo explore the protective effect and underlying mechanism of Gegen Qinliantang on cognitive function in db/db mice with diabetic cognitive impairment (DCI). MethodsThirty-two 8-week-old male db/db mice were randomly assigned to the model group, dapagliflozin group (1.0 mg·kg-1·d-1), low-dose Gegen Qinliantang group (6.24 g·kg-1·d-1), and high-dose Gegen Qinliantang group (24.96 g·kg-1·d-1). Eight db/m mice served as the normal group. All mice were administered the corresponding treatment once daily by gavage for 10 consecutive weeks. Body weight and fasting blood glucose (FBG) were dynamically monitored. The Morris water maze test was used to evaluate cognitive function. Hematoxylin-eosin (HE) staining and Nissl staining were used to observe pathological changes in the hippocampus. Enzyme-linked immunosorbent assay (ELISA) was employed to measure the levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in hippocampal tissue. Immunofluorescence double staining was used to detect the co-expression of M1 microglial marker CD16/32 and ionized calcium-binding adapter molecule 1 (IBA1) in the hippocampus. Western blot analysis was performed to detect the protein expression of postsynaptic density protein 95 (PSD-95), synapsin (SYN), nuclear factor-κB p65 (NF-κB p65), and phosphorylated NF-κB p65 (p-NF-κB p65) in the hippocampus. ResultsCompared with the normal group, the model group showed significantly increased body weight and FBG levels (P<0.01), significantly prolonged escape latency and reduced platform crossings in the Morris water maze test (P<0.01), disordered arrangement of hippocampal neurons, nuclear pyknosis, increased neuronal necrosis, reduced Nissl bodies, decreased expression of synaptic proteins PSD-95 and SYN (P<0.01), increased CD16/32+ /IBA1+ positive rate, elevated levels of TNF-α and IL-1β, and an increased p-NF-κB p65/NF-κB p65 ratio (P<0.01). Compared with the model group, the dapagliflozin group exhibited significantly reduced FBG levels at weeks 5 and 10 (P<0.05, P<0.01) and increased body weight. The high-dose Gegen Qinliantang group showed significantly reduced FBG at week 10 (P<0.05). Escape latency was significantly reduced on days 3 and 5 of the water maze test in the dapagliflozin group and on day 5 in the high-dose Gegen Qinliantang group (P<0.05). Platform crossings were significantly increased in both the dapagliflozin group and the high-dose Gegen Qinliantang group (P<0.05). Hippocampal pathological damage was alleviated to varying degrees in the dapagliflozin group and the low- and high-dose Gegen Qinliantang groups, with significantly increased expression of PSD-95 and SYN (P<0.01). Further studies revealed that both low- and high-dose Gegen Qinliantang reduced hippocampal IL-1β levels and the CD16/32+/IBA1+ positive rate of microglia, while the high-dose group also significantly reduced hippocampal TNF-α levels and the p-NF-κB p65/NF-κB p65 (P<0.05, P<0.01). ConclusionGegen Qinliantang can improve hyperglycemia, cognitive dysfunction, and synaptic damage in DCI, inhibit M1 polarization of microglia and neuroinflammation, and its mechanism may be related to the inhibition of NF-κB activation.
2.Effect of Zuoguiwan on Development of Skin Barrier in Neonatal Rat Model of Congenital Kidney Deficiency Based on Intercellular Connections
He YU ; Min XIAO ; Xiaocui JIANG ; Min ZHAO ; Yinjuan LYU ; Jian GONG ; Jigang CAO
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):11-18
ObjectiveTo study the effect of Zuoguiwan on the development of skin barrier in the neonatal rat model of congenital kidney deficiency and unveil the underlying mechanism. MethodsSixty rats were paired in a female-to-male ratio of 2∶1, and the pregnant rats were assigned into control, congenital kidney deficiency, and low- and high-dose (2 and 8 g·kg-1, respectively) Zuoguiwan groups. The pregnant rats in other groups except the control group were exposed to chronic unpredictable mild stress for the modeling of congenital kidney deficiency. The rats in the control group and congenital kidney deficiency group were administrated with normal saline by gavage, and those in Zuoguiwan groups with Zuoguiwan suspension by gavage from day 1 of pregnancy. The serum level of interleukin-6 (IL-6) in the neonatal rats on the day of birth was determined by enzyme-linked immunosorbent assay. The full-thickness skin of neonatal rats on the day of birth was removed from the same position on the back and stained with hematoxylin-eosin for observation of histopathological changes, measurement of skin thickness, and counting of hair follicles. Terminal deoxynucleotidyl transferase-mediated nick end labeling was used to detect the apoptosis of skin tissue cells. The expression of interleukin-6 receptor (IL-6R) and interleukin-17A (IL-17A) in the skin tissue was assessed by immunohistochemistry and Western blot, and the expression of occludin, connexin 43 (Cx43), and zonula occludens-1 (ZO-1) in the skin tissue was assessed by immunofluorescence and Western blot. ResultsCompared with those in the control group, the neonatal rats in the congenital kidney deficiency group showed a rise in the serum IL-6 level (P<0.01), decreases in stratum corneum thickness, skin thickness, and number of hair follicles (P<0.01), increases in the expression of IL-6R and IL-17A in the skin tissue (P<0.01) and the number of apoptotic cells (P<0.01), and decreases in the expression of occludin, Cx43, ZO-1 (P<0.05). Compared with the congenital kidney deficiency group, the low- and high-dose Zuoguiwan groups showed declines in serum IL-6 level (P<0.05). The low-dose group showed increased number of hair follicles (P<0.05), and the high-dose group presented thickened stratum corneum (P<0.01), increased number of hair follicles (P<0.01), and down-regulated expression of IL-6R and IL-17A in the skin tissue (P<0.05, P<0.01). Both Zuoguiwan groups showcased decreased number of apoptotic cells (P<0.05, P<0.01), and the high-dose group showed up-regulated expression of occludin, Cx43, and ZO-1 in the skin tissue (P<0.05, P<0.01). ConclusionZuoguiwan can reduce the levels of IL-6 in the serum and IL-6R and IL-17A in the skin tissue and improve the expression of intercellular junction proteins, thereby ameliorating the abnormal development of the skin barrier in the neonatal rat model of congenital kidney deficiency.
3.Visual Analysis of Literature Research on Compatibility of Bitter and Pungent Medicinals in Traditional Chinese Medicine
Wei SONG ; Lele YANG ; Zhongyi ZHANG ; Tao SHEN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):19-28
ObjectiveTo analyze the research situation, hotspots, and trend of the compatibility of bitter and pungent medicinals in traditional Chinese medicine by knowledge mapping and provide reference for the inheritance and clinical application of this theory. MethodsThe literature on the compatibility of bitter and pungent medicinals was retrieved from 6 databases, including China National Knowledge Infrastructure (CNKI), VIP, Wanfang Data, SinoMed, Web of Science, and PubMed. The literature was searched with the keywords of bitter medicinals, pungent medicinals, compatibility, Banxia Xiexin decoction, Zhuyu pills, and Zuojin pills. The search time ranged from January 1, 1984 to August 10, 2024. NoteExpress 3.7 was used for literature screening, and CiteSpace 6.1.R2 was used to visualize the number of publications, authors, institutional cooperation network, and the co-occurrence, clustering, time line, and burst of keywords. ResultsA total of 628 Chinese articles and 151 English articles related to the compatibility of bitter and pungent medicinals were included, and the annual number of publications rose with fluctuations. WEI Wei was the core author of Chinses articles and SHEN Tao and ZHAO Yanling were the core authors of English articles. China Academy of Chinese Medical Sciences formed a close cooperative network with many research institutions at home and abroad. The high-frequency keywords included pungent dispersing and bitter descending, Coptidis Rhizoma, Euodiae Fructus, Banxia Xiexin decoction, Zuojin pills, compatibility, spleen and stomach diseases, and gut microbiota, which mainly involved common prescriptions, diseases responding specifically to traditional Chinese medicine, mechanism research and so on. The keyword bursts concentrated in famous doctors' experience, network Meta-analysis, inflammatory bowel disease, diabetes mellitus, intestinal microbiota, etc. The compatibility of bitter medicinals and pungent medicinals was widely used in clinical practice. Based on pungent dispersing and bitter descending, the middle energizer was regulated for treating spleen and stomach diseases and inflammatory bowel disease. Bitter and pungent medicinals descending the turbid were used to treat diabetes mellitus and hyperlipidemia by lowering blood glucose and lipid levels. The mild bitter and slightly pungent medicinals were used as supplements to treat depression and COVID-19 by regulating Qi movement. ConclusionIn the past 40 years, the number of publications on the compatibility of bitter and pungent medicinals has been increasing, and the research team has begun to take form. Clinically, the compatibility of bitter and pungent medicinals is the main treatment for spleen and stomach diseases and glycolipid metabolic diseases. At present, the modern scientific essence of the compatibility of bitter and pungent medicinals is mainly explained based on famous doctors' experience inheritance, network meta-analysis, and intestinal flora.
4.Clinical Efficacy of Zhuyuwan in Treatment of Hyperlipidemia with Syndrome of Phlegm Turbidity and Obstruction
Lele YANG ; Danmei LUO ; Jiao CHEN ; Xiaobo ZHANG ; Wei SONG ; Wenyu ZHU ; Xin ZHOU ; Xueping LI ; Tao SHEN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):29-37
ObjectiveTo observe the clinical efficacy and safety of Zhuyuwan in the treatment of hyperlipidemia. MethodsIn this study, hyperlipidemia patients treated in the Hospital of Chengdu University of Traditional Chinese Medicine (TCM) from September 2022 to December 2023 were randomly assigned into a control group and an observation group. Finally, 162 valid cases were included, encompassing 74 cases in the control group and 88 cases in the observation group. The control group was treated with atorvastatin calcium tablets, and the observation group with atorvastatin calcium tablets + Zhuyuwan extract granules. Both groups were treated for 8 weeks. The efficacy in terms of blood lipid level recovery, blood lipid levels, TCM syndrome distribution, efficacy in terms of TCM syndrome, and TCM symptom scores were compared between the two groups as well as between before and after treatment. Liver and kidney functions were monitored for safety assessment. ResultsIn terms of blood lipid level recovery, the total response rate in the observation group was 86.36% (76/88) and that in the control group was 86.49% (64/74), with no statistically significant difference between the two groups. After treatment, both groups showed declines in levels of triglyceride (TG), total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) (P<0.05) and elevations in the level of high-density lipoprotein cholesterol (HDL-C) (P<0.05). Moreover, the observation group outperformed the control group in recovering the levels of TG, LDL-C, and HDL-C (P<0.05, P<0.01). In terms of TCM syndrome, hyperlipidemia was mostly caused by phlegm turbidity and obstruction. The total response rate in terms of TCM syndrome in the observation group was 87.30% (55/63), which was higher than that (63.46%, 33/52) in the control group (χ2=9.102, P<0.01). After treatment, the scores of total TCM symptoms, primary symptoms, and secondary symptoms decreased in both groups (P<0.05), and the observation group had lower scores than the control group (P<0.01). The observation group was superior to the control group in alleviating obesity, chest tightness, and low food intake (P<0.05). In terms of safety, the level of aminotransferase was slightly elevated in the control group, and no obvious adverse reaction was observed in the observation group, with no statistical significance in the incidence of adverse reactions. ConclusionZhuyuwan combined with atorvastatin can not only recover blood lipid levels and alleviate TCM symptoms but also reduce the occurrence of adverse reactions.
5.Zhuyuwan in Treatment of Hyperlipidemia Complicated with Carotid Atherosclerosis: A Randomized Controlled Trial
Wei SONG ; Lele YANG ; Zhongyi ZHANG ; Tao SHEN
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):38-45
ObjectiveTo observe the efficacy and safety of Zhuyuwan in the treatment of hyperlipidemia (syndrome of phlegm turbidity and obstruction) complicated with carotid atherosclerosis. MethodsA total of 120 patients with hyperlipidemia (syndrome of phlegm turbidity and obstruction) complicated with carotid atherosclerosis were randomly allocated into a treatment group and a control group, with 60 patients in each group. The control group orally took rosuvastatin calcium tablets, 10 mg each time, once a day. The treatment group took Zhuyuwan granules, 1 dose a day in 3 times. Both groups received diet and exercise guidance and were treated for 24 consecutive weeks. The traditional Chinese medicine (TCM) symptom scores were recorded before and after treatment. The levels of blood lipids [total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C)], carotid color ultrasound [carotid intima-media thickness (IMT), plaque number, maximum plaque area, and maximum plaque thickness], vascular intimal repair-related factors [nitric oxide (NO), endothelin-1 (ET-1), soluble intercellular adhesion molecule 1 (sICAM-1), and thromboxane B2 (TXB2)], and oxidative stress-related indicators [superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px)] were measured before and after treatment. The occurrence of adverse reactions was observed during treatment, and the clinical efficacy of the two groups was compared after treatment. ResultsThe trial was completed and 112 patients (57 in the treatment group and 55 in the control group) were included in the statistical analysis, and the general information of the two groups was comparable. The total response rate in the treatment group was 87.72% (50/57), which was higher than that (74.55%, 41/55) in the control group (χ2=4.823, P<0.05). After treatment, the scores of main TCM symptoms such as body obesity, chest tightness, and limb numbness, the scores of secondary symptoms such as palpitation, insomnia, tastelessness in the mouth, reduced appetite, tongue and pulse manifestation scores, and the total TCM symptom score decreased in both groups (P<0.05, P<0.01). Moreover, the treatment group had lower scores of main symptoms such as body obesity, head heaviness, chest tightness, nausea, salivation, and limb numbness, scores of secondary symptoms such as tastelessness in the mouth and reduced appetite, and total TCM symptom score than the control group (P<0.05, P<0.01). After treatment, both groups showed declines in TC, TG, LDL-C, IMT, plaque number, maximum plaque area, maximum plaque thickness, ET-1, sICAM-1, and TXB2 (P<0.05, P<0.01) and elevations in levels of HDL-C, NO, and SOD (P<0.05, P<0.01). The treatment group had lower TC, TG, LDL-C, IMT, plaque number, maximum plaque area, maximum plaque thickness, ET-1, sICAM-1, and TXB2 (P<0.01) and higher levels of HDL-C, NO, SOD, and GSH-Px (P<0.05, P<0.01) than the control group. During treatment, adverse reactions occurred in 3 cases (5.26%, 1 case of dizziness, 1 case of acid reflux, and 1 case of constipation) in the treatment group and 4 cases (7.27%, 2 cases of upper respiratory tract infection, 1 case of abdominal pain, and 1 case of low back pain) in the control group. ConclusionZhuyuwan can effectively reduce blood lipid levels, inhibit the formation of carotid atherosclerotic plaque, and alleviate the syndrome of phlegm turbidity and obstruction by promoting vascular intima repair and improving the antioxidant function.
6.Mechanisms of Zhuyuwan in Treating both Intrahepatic Cholestasis and Ulcerative Colitis Based on Homotherapy for Heteropathy
Jun HAN ; Yueqiang WEN ; Zongying XU ; Dan LUO ; Li ZHOU ; Xueyi LI ; Yufan DAI ; Lele YANG ; Tao SHEN ; Han YU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):46-53
ObjectiveThe theory of homotherapy for heteropathy is one of the classical rules in traditional Chinese medicine. Taking this theory as a breakthrough point, this study employed gas chromatography-mass spectrometry (GC-MS) to elucidate the mechanism underlying the therapeutic effects of Zhuyuwan on both intrahepatic cholestasis (IC) and ulcerative colitis (UC) from the viewpoint of serum metabolic homeostasis. MethodsThe rat models of α-naphthylisothiocyanate (ANIT)-induced cholestasis and 2,4,6-trinitro-benzenesulfonic acid (TNBS)-induced UC were treated with low (0.6 g·kg-1) and high (1.2 g·kg-1) doses of Zhuyuwan by gavage. In the experiment regarding IC, 24 Sprague-Dawley (SD) rats were randomly assigned into four groups: normal, ANIT model, low-dose Zhuyuwan, and high-dose Zhuyuwan. In the experiment regarding UC, 24 SD rats were randomly allocated into four groups: normal, TNBS model, low-dose Zhuyuwan, and high-dose Zhuyuwan. Firstly, the two disease models and the intervention effects of Zhuyuwan on the two diseases were evaluated based on serum levels of biochemical indicators [alanine aminotransferase (ALT), aspartate transaminase (AST), γ-glutamyltranspeptidase (γ-GT), and total bile acid (TBA)], colon damage score, colon weight index, disease activity index, and histopathological changes in rats. Secondly, the rat serum samples were analyzed by gas chromatography-mass spectrometry (GC-MS) to screen the common core pathways of the two disease models, and the expression of core genes in the pathways was determined by Real-time PCR, on the basis of which the biological mechanism of the treatment of the two disease models by Zhuyuwan was ultimately elucidated. ResultsThe results of the experiment regarding IC showed that the ANIT model group had higher ALT, AST, γ-GT, and TBA levels than the normal group (P<0.01). Compared with the ANIT model group, the low-dose Zhuyuwan group showed declined ALT and TBA levels (P<0.01) and the high-dose Zhuyuwan group showed lowered ALT, TBA, AST, and γ-GT levels (P<0.01). The results of the experiment regarding UC showed that compared with the normal group, the TNBS model group presented increases in the colonic damage score, colon weight index, and disease activity index (P<0.01). Compared with the TNBS model group, the low-dose Zhuyuwan group showcased declines in colon weight index (P<0.01) and disease activity index (P<0.05), and the high-dose Zhuyuwan group showed reductions in the colon damage score, colon weight index, and disease activity index (P<0.01). GC-MS metabolomics analysis combined with qRT-PCR demonstrated that Zhuyuwan had a similar inverse regulatory effect on arginine metabolism disruption in the above two disease models. ConclusionZhuyuwan exhibited definite therapeutic effects on both IC and UC, and the regulation of arginine biosynthesis pathway is the core mechanism for the treatment of both diseases by Zhuyuwan.
7.Exploration of Zhuyuwan in Treatment of Atherosclerosis from Perspective of Lipid Transport Disorder
Wei SONG ; Zhongyi ZHANG ; Hairong QIU ; Mei ZHAO ; Zubing ZHOU ; Tao SHEN ; Yong ZHANG
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):54-61
This article discusses the key pathogenesis of atherosclerosis (AS) based on the physiological characteristics and pathological changes of lipids and introduces the therapeutic effect of Zhuyuwan on AS, aiming to provide a theoretical basis for the treatment of cardiovascular diseases from the spleen. As essential substances, lipids have the same essence but different forms. They circulate throughout the body with body fluids under the action of Yang Qi to nourish the nutrient Qi and support the defensive Qi. Lipid metabolism disorder often leads to the obstruction of Qi movement, the accumulation of dampness and turbidity, and the generation of phlegm and blood stasis. It has been proven that the formation of vulnerable plaques in AS is attributed to the interaction of three pathogenic factors: deficiency of healthy Qi, phlegm-turbidity, and collateral stasis. Their pathological essence is closely related to abnormal lipid metabolism. As lipids constitute the thick and dense components of body fluids, their impaired dispersion may lead to phlegm-turbidity and blood stasis, the pathological process of which is predominantly ascribed to the dysfunction of the spleen in distributing essence. Therefore, AS is rooted in spleen-stomach disorder, manifests as plaques formed by pathological product accumulation in vessels, with lipid transport disorder as its core pathogenesis. Specifically speaking, the dysfunction of spleen in transportation with accumulation of dampness-turbidity marks the initial stage, and blood turbidity and coagulation and phlegm-nodules accumulating in vessels represent the intermediate phase. Cold accumulation and stagnated heat transforming into toxins represent the terminal stage. Zhuyuwan, first recorded in Taiping Holy Prescriptions for Universal Relief, contains equal proportions of Coptidis Rhizoma and Evodiae Fructus. Coptidis Rhizoma, bitter and cold, exerts descending and purging actions to assist stomach Qi in lowering turbidity. Evodiae Fructus, pungent-bitter and hot, disperses obstruction and promotes free flow to support spleen Qi in ascending the clear. The compatibility of Coptidis Rhizoma and Evodiae Fructus ascends the clear and descends the turbid to harmonize Yin and Yang, assisting the spleen in distributing essence and resolving lipid accumulation to reduce lipid levels. In terms of the therapeutic mechanism, Zhuyuwan modulates lipid metabolism by correcting immune-inflammation network imbalance, improving gut microbiota composition and metabolism, and enhancing reverse cholesterol transport. By analyzing the pathological characteristics of lipid transport disorder in AS, this study delves into the intrinsic connections between cardiovascular disease and lipid transport disorder, giving novel insights into the prevention and treatment of AS.
8.Shaoyaotang Containing Serum Mediates Fas/FasL Pathway to Inhibit Lipopolysaccharide Induced Inflammation and Apoptosis of Caco-2 Cells
Yuting YANG ; Dongsheng WU ; Hui CAO ; Yu ZHANG ; Nianjia XIE ; Bo ZOU ; Daguang CHEN ; Erle LIU ; Yi LU ; Zhaowen LYU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):62-69
ObjectiveTo investigate the effects of different concentrations of Shaoyaotang-containing serum on lipopolysaccharide (LPS)-induced inflammation of human colorectal adenocarcinoma (Caco-2) cells by inhibiting apoptosis via activating the tumor necrosis factor (TNF) receptor superfamily member 6 (Fas)/Fas ligand (FasL) pathway. MethodsCaco-2 cells were allocated into blank, model (LPS, 10 mg·L-1), Shaoyaotang-containing serum (5%, 10%, 15%, 20%), and Fas inhibitor (KR-33493, 20 mmol·L-1) groups. Except the blank group, the other groups were stimulated with 10 mg·L-1 LPS for 24 h for the modeling of inflammation. After successful modeling, the blank, Fas inhibitor, and model groups were treated with blank serum, and the Shaoyaotang-containing serum groups were treated with the serum samples at corresponding concentrations for 24 h. The Fas inhibitor group was subjected to KR-33493 pretreatment for 1 h. Cell proliferation and viability were examined by the cell-counting kit-8 (CCK-8) method. The levels of interleukin (IL)-6, IL-1β, and TNF-α were measured by enzyme-linked immunosorbent assay. Apoptosis was detected by flow cytometry. The protein and mRNA levels of Fas, FasL, cysteinyl aspartate-specific proteinase (Caspase)-3, Caspase-9, B-cell lymphoma 2 (Bcl-2), and Bcl-2-associated X protein (Bax) were determined by Western blot and Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR), respectively. ResultsCompared with the blank group, the model group presented a decrease in cell survival rate (P<0.01). Compared with that in the model group, the cell survival rate showed no significant change in the 5% Shaoyaotang-containing serum group but increased in the 10%, 15%, and 20% Shaoyaotang-containing serum groups (P<0.01). Since there was no statistical difference between the 5% Shaoyaotang-containing serum group and the model group, 10%, 15%, and 20% Shaoyaotang-containing sera were selected for the follow-up study. Compared with the blank group, the model group showed risen levels of IL-6, IL-1β, and TNF-α (P<0.01), an increased apoptosis rate (P<0.01), up-regulated protein and mRNA levels of Fas, FasL, Caspase-3, Caspase-9, and Bax (P<0.01), and down-regulated protein and mRNA levels of Bcl-2 (P<0.01). Compared with the model group, the Fas inhibitor group and the 10%, 15%, and 20% Shaoyaotang-containing serum groups showed declined levels of IL-6, IL-1β, and TNF-α (P<0.01), decreased apoptosis rates (P<0.01), down-regulated protein and mRNA levels of Fas, FasL, Caspase-3, Caspase-9, and Bax (P<0.05, P<0.01), and up-regulated protein and mRNA levels of Bcl-2 (P<0.05, P<0.01). In addition, the 15% and 20% Shaoyaotang-containing serum groups had lower levels of IL-6, IL-1β, and TNF-α (P<0.05, P<0.01), lower apoptosis rates (P<0.05, P<0.01), lower protein and mRNA levels of Fas, FasL, Caspase-3, Caspase-9, and Bax (P<0.05, P<0.01), and higher protein and mRNA levels of Bcl-2 (P<0.05, P<0.01) than the 10% Shaoyaotang-containing serum group. ConclusionThe Shaoyaotang-containing serum can reduce the content of inflammatory factors in Caco-2 cells, down-regulate the protein and mRNA levels of Fas, FasL, Caspase-3, Caspase-9, and Bax, and up-regulate the protein and mRNA levels of Bcl-2 under the intervention of LPS by regulating the Fas/FasL pathway and inhibiting the apoptosis of intestinal epithelial cells in ulcerative colitis.
9.Shaoyaotang Alleviates Damage of Tight Junction Proteins in Caco-2 Cell Model of Inflammation by Regulating RhoA/ROCK Pathway
Nianjia XIE ; Dongsheng WU ; Hui CAO ; Yu ZHANG ; Yuting YANG ; Bo ZOU ; Da ZHAO ; Yi LU ; Mingsheng WU
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):70-77
ObjectiveTo investigate the protective effect and mechanism of Shaoyaotang (SYD) on the lipopolysaccharide (LPS)-induced damage of tight junction proteins in the human colorectal adenocarcinoma (Caco-2) cell model of inflammation via the Ras homolog gene family member A (RhoA)/Rho-associated coiled-coil forming protein kinase (ROCK) pathway. MethodsCaco-2 cells were grouped as follows: Blank, model (LPS, 10 mg·L-1), SYD-containing serum (10%, 15%, and 20%), and inhibitor (Fasudil, 25 μmol·L-1). After 24 hours of intervention, the cell viability in each group was examined by the cell-counting kit 8 (CCK-8) method. Enzyme-linked immunosorbent assay was employed to determine the levels of endothelin-1 (ET-1), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) and Western blot were employed to determine the mRNA and protein levels, respectively, of RhoA, ROCK2, claudin-5, and zonula occludens-1 (ZO-1) in cells of each group. ResultsCompared with the blank group, the model group showcased a marked reduction in the cell viability (P<0.01), elevations in the levels of ET-1, TNF-α, IL-1β, and IL-6 (P<0.01), declines in both mRNA and protein levels of ZO-1 and claudin-5 (P<0.01), and rises in mRNA and protein levels of RhoA and ROCK2 (P<0.01). Compared with the model group, the Shaoyaotang-containing serum (10%, 15%, and 20%) groups had enhanced cell viability (P<0.01), lowered levels of ET-1, TNF-α, IL-1β, and IL-6 (P<0.01), up-regulated mRNA and protein levels of ZO-1 and claudin-5 (P<0.05, P<0.01), and down-regulated mRNA and protein levels of RhoA and ROCK2 (P<0.01). Moreover, the inhibitor group and the 15% and 20% Shaoyaotang-containing serum groups had lower levels of ET-1, TNF-α, IL-1β, and IL-6 (P<0.05, P<0.01), higher mRNA and protein levels of ZO-1 and claudin-5 (P<0.05, P<0.01), and lower mRNA and protein levels of RhoA and ROCK2 (P<0.05, P<0.01) than the 10% Shaoyaotang-containing serum group. ConclusionThe Shaoyaotang-containing serum can lower the levels of LPS-induced increases in levels of inflammatory cytokines and endothelin to ameliorate the damage of tight junction proteins of the Caco-2 cell model of inflammation by regulating the expression of proteins in the RhoA/ROCK pathway.
10.Shaoyaotang Restores Th17/Treg Cell Balance by Regulating Glucose Metabolism Reprogramming in Treatment of Ulcerative Colitis
Yiwen WANG ; Yiling XIA ; Erle LIU ; Shaijin JIANG ; Bo ZOU ; Dongsheng WU ; Youwei XIAO ; Hui CAO
Chinese Journal of Experimental Traditional Medical Formulae 2025;31(13):78-85
ObjectiveTo investigate the effect of Shaoyaotang on T helper cell 17/regulatory T lymphocyte(Th17/Treg) cell balance in ulcerative colitis and decipher the intervention mechanism based on glucose metabolism reprogramming. MethodsThe mouse model of ulcerative colitis was established by the dextran sulfate sodium (DSS) method. Forty-eight C57BL/6 mice were randomly allocated into normal, model, Western drug control (mesalazine, 0.39 g·kg-1·d-1), Shaoyaotang (15.54 g·kg-1·d-1), inhibitor (2-deoxy-D-glucose, 2-DG, 100 mg·kg-1·d-1), and inhibitor (2-DG, 100 mg·kg-1·d-1) + Shaoyaotang (15.54 g·kg-1·d-1) groups. Mice were administrated with the corresponding drugs by gavage for 7 days. The general conditions and the colon injury degree were observed 24 h after the last administration. The expression of interleukin (IL)-10 and IL-17 in the colon tissue was detected by immunohistochemical staining. Western blot and Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) were performed to determine the protein and mRNA levels, respectively, of hypoxia-inducing factor-1α (HIF-1α), lactate dehydrogenase (LDHA), and hexokinase 2 (HK2) in the colon tissue. Th17/Treg cell differentiation was detected by flow cytometry. Enzyme-linked immunosorbent assay was employed to measure the levels of lactic acid and glucose in the colon tissue and IL-10, IL-17, and IL-6 in the serum. ResultsCompared with the normal group, the model group showed decreases in body weight and disease activity index (DAI) (P<0.05), elevations in levels of HIF-1α, LDHA, HK2, IL-17, IL-6, Th17 cells, lactic acid, and glucose in the colon tissue (P<0.05), and declines in the levels of of IL-10 and Treg cells (P<0.05). Compared with the model group, the drug administration groups showed increases in body weight and DAI (P<0.05), declines in levels of HIF-1α, LDHA, HK2, IL-17, IL-6, Th17 cells, lactic acid, and glucose in the colon tissue (P<0.05), and rises in levels of IL-10 and Treg cells (P<0.05). Shaoyaotang+2-DG group had the most obvious effect. ConclusionShaoyaotang can relieve diarrhea and bloody stool in mice with ulcerative colitis by restoring the Th17/Treg cell balance via regulation of glucose metabolism reprogramming, thus playing a role in the treatment of ulcerative colitis.

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