ObjectiveTo verify the therapeutic effect of Yinqi Sanhuang Jiedu decoction （YQSH） on carbon tetrachloride （CCl₄）-induced hepatic fibrosis in mice， and to explore whether its effect was related to the inhibition of neutrophil infiltration and the formation of neutrophil extracellular traps （NETs）. MethodsThe 36 C57BL/6J mice were randomly divided into control group， model group， positive drug silybin （SF） group （55 mg·kg^-1·d^-1）， YQSH-L group， YQSH-M group， and YQSH-H group （8.325， 16.65， 33.3 g·kg^-1·d^-1， respectively），n=6 in each group. Except for the control group， mice in all other groups were intraperitoneally injected with CCl₄ to induce hepatic fibrosis. After successful modeling， each drug administration group was given the corresponding drugs by gavage for eight weeks. Hematoxylin-eosin （HE） staining， Sirius red staining and Masson staining were used to observe the pathological changes of liver tissue. Liver elasticity was detected by a color Doppler ultrasound system. Immunohistochemistry and real-time fluorescent quantitative polymerase chain reaction （Real-time PCR） were performed to detect the protein expression and mRNA levels of C-X-C motif chemokine ligand 1 （CXCL1）， CXCL2 and CXCL5. Neutrophil levels were detected by flow cytometry. The expression of neutrophil elastase （NE） and myeloperoxidase （MPO） positive protein was observed by immunofluorescence. The contents of MPO， NE and CitH3 were detected by enzyme-linked immunosorbent assay （ELISA）. ResultsCompared with the control group， the liver of the model group showed obvious inflammatory cell infiltration and collagen deposition， and the liver elasticity， CXCL1， CXCL2， CXCL5 expression， neutrophil level， and MPO， NE and CitH3 levels were significantly increased （P<0.05， P<0.01）. Compared with the model group， inflammatory cell infiltration and collagen deposition in the liver tissue of mice were reduced after YQSH treatment. Moreover， the liver elasticity was reduced （P<0.01）. The protein expression （P<0.01） and mRNA level of CXCL1， CXCL2 and CXCL5 were decreased（P<0.05，P<0.01）. The neutrophil level was decreased （P<0.01）， the expression of MPO and NE positive protein was significantly decreased（P<0.05，P<0.01）， and the levels of MPO， NE and CitH3 were decreased （P<0.05， P<0.01）. ConclusionThe anti-hepatic fibrosis effect of YQSH may be related to its inhibition of chemokines （CXCL1， CXCL2， CXCL5）， reduction of neutrophil infiltration， and inhibition of NETs generation.

ObjectiveTo verify the therapeutic effect of Yinqi Sanhuang Jiedu decoction （YQSH） on carbon tetrachloride （CCl₄）-induced hepatic fibrosis in mice， and to explore whether its effect was related to the inhibition of neutrophil infiltration and the formation of neutrophil extracellular traps （NETs）. MethodsThe 36 C57BL/6J mice were randomly divided into control group， model group， positive drug silybin （SF） group （55 mg·kg^-1·d^-1）， YQSH-L group， YQSH-M group， and YQSH-H group （8.325， 16.65， 33.3 g·kg^-1·d^-1， respectively），n=6 in each group. Except for the control group， mice in all other groups were intraperitoneally injected with CCl₄ to induce hepatic fibrosis. After successful modeling， each drug administration group was given the corresponding drugs by gavage for eight weeks. Hematoxylin-eosin （HE） staining， Sirius red staining and Masson staining were used to observe the pathological changes of liver tissue. Liver elasticity was detected by a color Doppler ultrasound system. Immunohistochemistry and real-time fluorescent quantitative polymerase chain reaction （Real-time PCR） were performed to detect the protein expression and mRNA levels of C-X-C motif chemokine ligand 1 （CXCL1）， CXCL2 and CXCL5. Neutrophil levels were detected by flow cytometry. The expression of neutrophil elastase （NE） and myeloperoxidase （MPO） positive protein was observed by immunofluorescence. The contents of MPO， NE and CitH3 were detected by enzyme-linked immunosorbent assay （ELISA）. ResultsCompared with the control group， the liver of the model group showed obvious inflammatory cell infiltration and collagen deposition， and the liver elasticity， CXCL1， CXCL2， CXCL5 expression， neutrophil level， and MPO， NE and CitH3 levels were significantly increased （P<0.05， P<0.01）. Compared with the model group， inflammatory cell infiltration and collagen deposition in the liver tissue of mice were reduced after YQSH treatment. Moreover， the liver elasticity was reduced （P<0.01）. The protein expression （P<0.01） and mRNA level of CXCL1， CXCL2 and CXCL5 were decreased（P<0.05，P<0.01）. The neutrophil level was decreased （P<0.01）， the expression of MPO and NE positive protein was significantly decreased（P<0.05，P<0.01）， and the levels of MPO， NE and CitH3 were decreased （P<0.05， P<0.01）. ConclusionThe anti-hepatic fibrosis effect of YQSH may be related to its inhibition of chemokines （CXCL1， CXCL2， CXCL5）， reduction of neutrophil infiltration， and inhibition of NETs generation.

ObjectiveTo decipher the mechanism by which Zuoguiwan （ZGW） treat hyperthyroidism in rats with kidney-Yin deficiency based on the dopamine receptor D4 （DRD4）/nicotinamide adenine dinucleotide phosphate （NADPH） oxidase 4 （NOX4） signaling pathway. MethodsThe rat model of kidney-Yin deficiency was induced by unilateral intramuscular injection of dexamethasone （0.35 mg·kg^-1）. After successful modeling， the rats were randomized into model， methimazole （positive control， 5 mg·kg^-1）， low-， medium-， and high-dose （1.85， 3.70， 7.40 g·kg^-1， respectively） ZGW， and normal control groups. After 21 days of continuous gavage， the behavioral indexes and body weight changes of rats were evaluated. The pathological changes of the renal tissue were observed by hematoxylin-eosin staining. The serum levels of thyroid hormones ［triiodothyronine （T₃）， thyroxine （T₄）， thyroid-stimulating hormone （TSH）］， renal function indexes ［serum creatine （Scr） and blood urea nitrogen （BUN）］， energy metabolism markers ［cyclic adenosine monophosphate （cAMP） and cyclic guanosine monophosphate （cGMP）］， and oxidative stress-related factors ［superoxide dismutase （SOD）， malondialdehyde （MDA）， and NADPH）］ were measured by enzyme-linked immunosorbent assay （ELISA）. Western blot was employed to analyze the expression of DRD4， NOX4， mitochondrial respiratory chain complex proteins ［NADH：ubiquinone oxidoreductase subunit S4 （NDUFS4） and cytochrome C oxidase subunit 4 （COX4）］， and inflammation-related protein ［tumor necrosis factor-alpha （TNF-α）， interleukin-6 （IL-6）， p38 mitogen-activated protein kinase （MAPK）］ pathway in the renal tissue. ResultsCompared with the normal group， the model group showed mental malaise， body weight decreases （P<0.01）， inflammatory cell infiltration in the renal tissue， a few residual parotid glands in the thyroid， elevations in serum levels of T₃， T₄， Scr， BUN， cAMP， cAMP/cGMP， MDA， and NADPH （P<0.01）， down-regulation in protein levels of TSH， SOD， and DRD4 （P<0.05， P<0.01）， and up-regulation in expression of NOX4， p-p38 MAPK/p38 MAPK， and inflammatory factors （P<0.01）. Compared with the model group， ZGW increased the body weight （P<0.05， P<0.01）， reduced the infiltration of renal interstitial inflammatory cells， restored the thyroid structure and follicle size， lowered the serum levels of T₃， T₄， Scr， BUN， cAMP， cAMP/cGMP， MDA and NADPH （P<0.05， P<0.01）， up-regulated the expression of TSH， SOD and DRD4 （P<0.05， P<0.01）， and down-regulated the expression of NOX4， p-p38 MAPK/p38 MAPK， and inflammatory factors （P<0.05， P<0.01）. Moreover， high-dose ZGW outperformed methimazole （P<0.05）. ConclusionBy activating DRD4， ZGW can inhibit the expression of NOX4 mediated by the p38 MAPK pathway， reduce oxidative stress and inflammatory response， thereby ameliorating the pathological state of hyperthyroidism due to kidney-Yin deficiency. This study provides new molecular mechanism support for the clinical application of ZGW.

Objective To investigate the expression characteristics, prognostic value, and correlation with immune infiltration of vasoactive intestinal peptide receptor 2 (VIPR2) in esophageal adenocarcinoma (ESAD). Methods Transcriptome data of 80 ESAD and 10 normal tissues were retrieved from the TCGA database. Immune-related differential genes were identified by integrating the ImmPort database. Cox regression analysis was applied to evaluate the prognostic significance of VIPR2. GSVA and ssGSEA were employed to assess the association between VIPR2 and immune-cell infiltration. Functional enrichment was performed using GO, KEGG, and GSEA. Results VIPR2 expression was significantly downregulated in ESAD tissues (P<0.05). High VIPR2 expression was associated with prolonged overall survival (HR=0.246, P=0.047), serving as an independent prognostic factor. Fifteen immune-cell types, including CD8⁺ T cells, NK cells, and dendritic cells, showed significantly higher infiltration in the VIPR2-high group, whereas Th17-cell abundance was decreased (P<0.05). Enrichment analyses revealed that VIPR2 participated in multiple tumor-immune pathways and was positively correlated with PD-1 and CTLA-4 expression (P<0.01). Conclusion VIPR2 may serve as a potential biomarker in the prognostic assessment and could modulate the tumor-immune microenvironment in ESAD.

BackgroundNegative symptoms of schizophrenia impose a substantial burden on society， and the current antipsychotic drugs have limited efficacy in treating these symptoms. Intermittent theta burst stimulation （iTBS）， as a novel neuromodulation technique， shows potential therapeutic promise. However， the evidence for its efficacy in treating negative symptoms of schizophrenia and its relationship with associated immuno-inflammatory changes remain to be further elucidated. ObjectiveTo investigate the efficacy of iTBS targeting the left dorsolateral prefrontal cortex （L-DLPFC） on persistent negative symptoms of schizophrenia， and to analyze its relationship with changes in peripheral blood inflammatory markers， so as to provide references for the clinical application of iTBS in treating persistent negative symptoms and to offer clues for further research into its potential biological mechanisms. MethodsA randomized， double-blind， sham-controlled clinical trial was conducted. A total of 60 patients with schizophrenia who were hospitalized at Xuzhou Oriental Peopple's Hospital from February 20， 2024 to May 10， 2025 were consecutively enrolled. All patients met the diagnostic criteria for schizophrenia according to the Diagnostic and Statistical Manual of Mental Disorders， fifth edition （DSM-5） and fulfilled the criteria for persistent negative symptoms. Participants were randomly assigned into the study group and the control group， with 30 patients in each group. The two groups respectively received 20 sessions of either active iTBS or sham stimulation over a 4-week period. Clinical symptoms were assessed using the Positive and Negative Syndrome Scale （PANSS） at the baseline， the end of treatment， and 3 months after treatment. Peripheral blood inflammatory markers were measured at the baseline and at the end of treatment， including white blood cell count， platelet count， neutrophil count， lymphocyte count， and monocyte count， the neutrophil-to-lymphocyte ratio （NLR）， monocyte-to-lymphocyte ratio （MLR）， and platelet-to-lymphocyte ratio （PLR） were also calculated. A linear mixed model was used to compare PANSS scores between the two groups. Spearman correlation analysis was performed to examine the association between reductions in inflammatory markers at the end of treatment and reductions in PANSS negative symptom subscale scores， with body mass index （BMI） and chlorpromazine-equivalent dose as covariates. Results①The time-group interaction effect for the PANSS negative symptom subscale score was statistically significant （F=5.410， P<0.01）. At the end of treatment and 3 months after treatment， the study group had significantly lower scores on the PANSS negative symptom subscale than the control group （P<0.01）. ② At the end of treatment， group differences in reduction scores for neutrophil count， NLR， and PLR were significant （Z=-2.262， -2.048， -2.070， P<0.05）. However， after FDR correction， the between-group differences in reductions in peripheral blood inflammatory markers were no longer statistically significant （P>0.05）. ③ After adjusting for BMI and chlorpromazine equivalents， the reduction in PANSS negative symptom subscale scores was positively correlated with the reduction in neutrophil count in the study group （r_s=0.382， P=0.045）. ConclusioniTBS targeting the L-DLPFC may be beneficial in improving persistent negative symptoms of schizophrenia， and its clinical efficacy may be associated with changes in neutrophil count. ［Funded by National Natural Science Foundation of China （number， 82371510）； Social Development Project of Jiangsu Province （number， BE2023668）. www.chictr.org.cn number， ChiCTR2500101060］

ObjectiveTo validate the therapeutic effect of Yinqi Sanhuang Jiedu decoction （YQSH） on a mouse model of carbon tetrachloride （CCl₄）-induced liver fibrosis and to investigate its correlations with the Piezo-type mechanosensitive ion channel component 1 （Piezo1）-Yes-associated protein （YAP） mechanical signaling axis and macrophage polarization. MethodsFifty-four C57BL/6J mice were randomized into a blank control group， a 4-week model group， a 6-week model group， a 8-week model group， a positive drug （silymarin， 55 mg·kg^-1·d^-1） group， and low-， medium-， and high-dose （8.325， 16.65， 33.3 g·kg^-1·d^-1， respectively） YQSH groups. Except the 6-week model group （n=12）， each of the other groups had 6 mice. Mice in other groups except the blank control group received intraperitoneal injections of 10% CCl₄ twice weekly for the modeling of liver fibrosis. Drug interventions began one week after the initial modeling through gavage， and the blank control and model groups received 0.2 mL of normal saline via gavage. The histopathological changes and collagen deposition in the liver were observed via hematoxylin-eosin （HE）， Masson's trichrome， and Sirius red staining. Serum activities of alanine aminotransferase （ALT） and aspartate aminotransferase （AST）， as well as serum levels of total protein （TP）， albumin （ALB）， total bilirubin （TBIL）， hyaluronic acid （HA）， laminin （LN）， procollagen type Ⅲ （PCⅢ）， and collagen type Ⅳ （Ⅳ-C）， were measured. The levels of tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β） in the liver tissue were determined by enzyme-linked immunosorbent assay （ELISA）. The protein and mRNA levels of Piezo1 and YAP1 in the liver tissue were determined by immunohistochemistry （IHC） and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， respectively. Co-localization of Piezo1 with YAP1， and YAP1 with inducible nitric oxide synthase （iNOS） was observed by the immunofluorescence （IF） assay. The proportions of M1-type （F4/80⁺CD80⁺） and M2-type （F4/80⁺CD206⁺） macrophages in the liver tissue were examined by flow cytometry. Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analyses and gene set enrichment analysis （GSEA） were performed through transcriptomic sequencing. ResultsCompared with the blank control group， the model group exhibited gradually worsened liver fibrosis at the 4^th， 6^th， and 8^th weeks. The 6- and 8-week model groups showcased inflammatory cell infiltration and collagen deposition in the liver （P<0.01） and upregulated protein levels of both Piezo1 and YAP1 （P<0.01）. Compared with the model groups， treatment with YQSH improved the liver function （P<0.05， P<0.01）， alleviated liver fibrosis （P<0.05， P<0.01）， and reduced intrahepatic inflammatory cell infiltration and collagen deposition （P<0.01）. Furthermore， the treatment downregulated the protein and mRNA levels of Piezo1 and YAP1 （P<0.05， P<0.01）， lowered the levels of inflammatory factors TNF-α and IL-1β （P<0.05， P<0.01）， and decreased the ratio of CD80 （M1-type）/CD206 （M2-type） macrophage proteins （P<0.01）. The IF assay showed co-localization of YAP1 with Piezo1 and iNOS. Compared with the model groups， YQSH treatment downregulated the expression of Piezo1， YAP1， and iNOS （P<0.05， P<0.01）. Transcriptomic analysis suggested that the anti-liver fibrosis effect of YQSH may be related to the Hippo signaling pathway （P<0.05）. ConclusionThe anti-liver fibrosis effect of YQSH may be related to its inhibition of the Piezo1-YAP signaling axis and regulation of macrophage polarization.

ObjectiveTo validate the therapeutic effect of Yinqi Sanhuang Jiedu decoction （YQSH） on a mouse model of carbon tetrachloride （CCl₄）-induced liver fibrosis and to investigate its correlations with the Piezo-type mechanosensitive ion channel component 1 （Piezo1）-Yes-associated protein （YAP） mechanical signaling axis and macrophage polarization. MethodsFifty-four C57BL/6J mice were randomized into a blank control group， a 4-week model group， a 6-week model group， a 8-week model group， a positive drug （silymarin， 55 mg·kg^-1·d^-1） group， and low-， medium-， and high-dose （8.325， 16.65， 33.3 g·kg^-1·d^-1， respectively） YQSH groups. Except the 6-week model group （n=12）， each of the other groups had 6 mice. Mice in other groups except the blank control group received intraperitoneal injections of 10% CCl₄ twice weekly for the modeling of liver fibrosis. Drug interventions began one week after the initial modeling through gavage， and the blank control and model groups received 0.2 mL of normal saline via gavage. The histopathological changes and collagen deposition in the liver were observed via hematoxylin-eosin （HE）， Masson's trichrome， and Sirius red staining. Serum activities of alanine aminotransferase （ALT） and aspartate aminotransferase （AST）， as well as serum levels of total protein （TP）， albumin （ALB）， total bilirubin （TBIL）， hyaluronic acid （HA）， laminin （LN）， procollagen type Ⅲ （PCⅢ）， and collagen type Ⅳ （Ⅳ-C）， were measured. The levels of tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β） in the liver tissue were determined by enzyme-linked immunosorbent assay （ELISA）. The protein and mRNA levels of Piezo1 and YAP1 in the liver tissue were determined by immunohistochemistry （IHC） and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）， respectively. Co-localization of Piezo1 with YAP1， and YAP1 with inducible nitric oxide synthase （iNOS） was observed by the immunofluorescence （IF） assay. The proportions of M1-type （F4/80⁺CD80⁺） and M2-type （F4/80⁺CD206⁺） macrophages in the liver tissue were examined by flow cytometry. Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analyses and gene set enrichment analysis （GSEA） were performed through transcriptomic sequencing. ResultsCompared with the blank control group， the model group exhibited gradually worsened liver fibrosis at the 4^th， 6^th， and 8^th weeks. The 6- and 8-week model groups showcased inflammatory cell infiltration and collagen deposition in the liver （P<0.01） and upregulated protein levels of both Piezo1 and YAP1 （P<0.01）. Compared with the model groups， treatment with YQSH improved the liver function （P<0.05， P<0.01）， alleviated liver fibrosis （P<0.05， P<0.01）， and reduced intrahepatic inflammatory cell infiltration and collagen deposition （P<0.01）. Furthermore， the treatment downregulated the protein and mRNA levels of Piezo1 and YAP1 （P<0.05， P<0.01）， lowered the levels of inflammatory factors TNF-α and IL-1β （P<0.05， P<0.01）， and decreased the ratio of CD80 （M1-type）/CD206 （M2-type） macrophage proteins （P<0.01）. The IF assay showed co-localization of YAP1 with Piezo1 and iNOS. Compared with the model groups， YQSH treatment downregulated the expression of Piezo1， YAP1， and iNOS （P<0.05， P<0.01）. Transcriptomic analysis suggested that the anti-liver fibrosis effect of YQSH may be related to the Hippo signaling pathway （P<0.05）. ConclusionThe anti-liver fibrosis effect of YQSH may be related to its inhibition of the Piezo1-YAP signaling axis and regulation of macrophage polarization.

OBJECTIVE: To evaluate the protective effects of gentiopicroside (GPS) against reactive oxygen species (ROS)-induced NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome activation in endothelial cells, aiming to reduce atherosclerosis. METHODS: Eight-week-old male ApoE-deficient mice were randomly divided into 2 groups (n=10 per group): the vehicle group and the GPS treatment group. Both groups were fed a high-fat diet for 16 weeks. GPS (40 mg/kg per day) was administered by oral gavage to the GPS group, while the vehicle group received an equivalent volume of the vehicle solution. At the end of the treatment, blood and aortic tissues were collected for assessments of atherosclerosis, lipid profiles, oxidative stress, and molecular expressions related to NLRP3 inflammasome activation, ROS production, and apoptosis. Additionally, in vitro experiments on human aortic endothelial cells treated with oxidized low-density lipoprotein (ox-LDL) were conducted to evaluate the effects of GPS on NLRP3 inflammasome activation, pyroptosis, apoptosis, and ROS production, specifically examining the role of the sirtuin 1 (SIRT1)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. SIRT1 and Nrf2 inhibitors were used to confirm the pathway's role. RESULTS: GPS treatment significantly reduced atherosclerotic lesions in the en face aorta (P<0.01), as well as in the thoracic and abdominal aortic regions, and markedly decreased sinus lesions within the aortic root (P<0.05 or P<0.01). Additionally, GPS reduced oxidative stress markers and proinflammatory cytokines, including interleukin (IL)-1 β and IL-18, in lesion areas (P<0.05, P<0.01). In vitro, GPS inhibited ox-LDL-induced NLRP3 activation, as evidenced by reduced NLRP3 (P<0.01), apoptosis-associated speck-like protein containing a CARD, cleaved-caspase-1, and cleaved-gasdermin D expressions (all P<0.01). GPS also decreased ROS production, apoptosis, and pyroptosis, with the beneficial effects being significantly reversed by SIRT1 or Nrf2 inhibitors. CONCLUSION GPS exerts an antiatherogenic effect by inhibiting ROS-dependent NLRP3 inflammasome activation via the SIRT1/Nrf2 pathway.
NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Reactive Oxygen Species/metabolism* ; Iridoid Glucosides/therapeutic use* ; NF-E2-Related Factor 2/metabolism* ; Animals ; Atherosclerosis/metabolism* ; Inflammasomes/drug effects* ; Male ; Sirtuin 1/metabolism* ; Signal Transduction/drug effects* ; Humans ; Endothelial Cells/pathology* ; Mice ; Oxidative Stress/drug effects* ; Apoptosis/drug effects* ; Lipoproteins, LDL ; Mice, Inbred C57BL