OBJECTIVE To investigate the effects and mechanism of asperuloside （Asp） on the pyroptosis of intestinal epithelial cells in rats with ulcerative colitis （UC）. METHODS The male SD rats were randomly divided into Control group， model group （UC group）， ASP low-dose and high-dose groups [Asp-L， Asp-H groups， Asp 35， 70 mg/（kg·d）]， ASP high-dose group+AMPK inhibitor Compound C group [Asp-H+Compound C group， Asp 70 mg/（kg·d）+Compound C 0.2 mg/（kg·d）]， with 12 rats in each group. Except for Control group， the other groups were injected with 50% ethanol （0.25 mL）+5% 2，4， 6- trinitrobenzene sulfonic acid solution （2 mL/kg） into the intestinal cavity to construct UC model. After modeling， the rats in each drug group were given corresponding drug solution by gavage or （and） tail vein injection， once a day， for 14 consecutive days. After the last administration， the weight of rats in each group was measured， and the length of their colons was measured； disease activity index （DAI） score and colonic mucosal damage index （CMDI） score were performed， and the serum levels of inflammatory factors （interleukin-18， -1β， -6） were detected. The pathological changes of the colon tissue were observed. The expressions of pyroptosis-related proteins [caspase-1， gasdermin D （GSDMD）] in colon tissue， and pathway-related proteins such as adenosine monophosphate-activated protein kinase （AMPK）， thioredoxin-interacting protein （TXNIP）， NOD-like receptor protein 3 （NLRP3） and apoptosis-associated speck-like protein containing a CARD （ASC） were all detected. RESULTS Compared with Control group， the colon tissue structure of rats in UC group was damaged， with obvious infiltration of inflammatory cells and edema. Their body weight， colon length and phosphorylation level of AMPK protein were significantly reduced or shortened； DAI and CMDI scores， serum levels of inflammatory factors， and the protein expressions of caspase-1， GSDMD， TXNIP， NLRP3 and ASC in colon tissue were increased or upregulated significantly （P＜0.05）. Compared with UC group， the pathological damage of colon tissue in rats was relieved in Asp-L and Asp-H groups， and all quantitative indicators were significantly improved （P＜0.05）； the improvement effect of Asp-H group was more significant （P＜0.05）. Compound C could significantly reverse the improvement effect of high-dose of Asp on the above indicators in UC rats （P＜0.05）. CONCLUSIONS Asp can improve inflammatory damage in colon tissue and inhibit pyroptosis of intestinal epithelial cells in UC rats， which is associated with the activation of AMPK and inhibition of TXNIP/NLRP3 signaling pathway.

OBJECTIVE To investigate the effects and mechanism of asperuloside （Asp） on the pyroptosis of intestinal epithelial cells in rats with ulcerative colitis （UC）. METHODS The male SD rats were randomly divided into Control group， model group （UC group）， ASP low-dose and high-dose groups [Asp-L， Asp-H groups， Asp 35， 70 mg/（kg·d）]， ASP high-dose group+AMPK inhibitor Compound C group [Asp-H+Compound C group， Asp 70 mg/（kg·d）+Compound C 0.2 mg/（kg·d）]， with 12 rats in each group. Except for Control group， the other groups were injected with 50% ethanol （0.25 mL）+5% 2，4， 6- trinitrobenzene sulfonic acid solution （2 mL/kg） into the intestinal cavity to construct UC model. After modeling， the rats in each drug group were given corresponding drug solution by gavage or （and） tail vein injection， once a day， for 14 consecutive days. After the last administration， the weight of rats in each group was measured， and the length of their colons was measured； disease activity index （DAI） score and colonic mucosal damage index （CMDI） score were performed， and the serum levels of inflammatory factors （interleukin-18， -1β， -6） were detected. The pathological changes of the colon tissue were observed. The expressions of pyroptosis-related proteins [caspase-1， gasdermin D （GSDMD）] in colon tissue， and pathway-related proteins such as adenosine monophosphate-activated protein kinase （AMPK）， thioredoxin-interacting protein （TXNIP）， NOD-like receptor protein 3 （NLRP3） and apoptosis-associated speck-like protein containing a CARD （ASC） were all detected. RESULTS Compared with Control group， the colon tissue structure of rats in UC group was damaged， with obvious infiltration of inflammatory cells and edema. Their body weight， colon length and phosphorylation level of AMPK protein were significantly reduced or shortened； DAI and CMDI scores， serum levels of inflammatory factors， and the protein expressions of caspase-1， GSDMD， TXNIP， NLRP3 and ASC in colon tissue were increased or upregulated significantly （P＜0.05）. Compared with UC group， the pathological damage of colon tissue in rats was relieved in Asp-L and Asp-H groups， and all quantitative indicators were significantly improved （P＜0.05）； the improvement effect of Asp-H group was more significant （P＜0.05）. Compound C could significantly reverse the improvement effect of high-dose of Asp on the above indicators in UC rats （P＜0.05）. CONCLUSIONS Asp can improve inflammatory damage in colon tissue and inhibit pyroptosis of intestinal epithelial cells in UC rats， which is associated with the activation of AMPK and inhibition of TXNIP/NLRP3 signaling pathway.

[摘 要] 目的：探究环状RNA hsa_circ_0081621对人喉鳞状细胞癌AMC-HN-8和TU177细胞恶性生物学行为的影响。方法：常规培养AMC-HN-8和TU177细胞，用转染试剂将si-NC、si-hsa_circ_0081621、空载体（vector）和hsa_circ_0081621过表达载体（hsa_circ_0081621-OE）转染至AMC-HN-8和TU177细胞，分别记作si-NC、si-hsa_circ_0081621、vector和hsa_circ_0081621-OE组。用CCK-8法、克隆形成实验、划痕愈合实验和Transwell小室实验分别检测敲减或过表达hsa_circ_0081621对AMC-HN-8和TU177细胞增殖、迁移和侵袭能力的影响。结果：在AMC-HN-8和TU177中成功地敲减或过表达了hsa_circ_0081621；敲减或过表达hsa_circ_0081621能显著抑制或促进AMC-HN-8和TU177细胞的增殖、迁移和侵袭能力（P < 0.01或P < 0.001或P < 0.000 1）。结论：hsa_circ_0081621可促进人喉鳞状细胞癌AMC-HN-8和TU177细胞的恶性生物学行为。

Emodin is a hydroxyanthraquinone compound that is widely distributed and has multiple pharmacological activities, including anti-diarrheal, anti-inflammatory, and liver-protective effects. Research indicates that emodin may be one of the main components responsible for inducing hepatotoxicity. However, studies on the mechanisms of liver injury are relatively limited, particularly those related to bile acids(BAs) metabolism. This study aims to systematically investigate the effects of different dosages of emodin on BAs metabolism, providing a basis for the safe clinical use of traditional Chinese medicine(TCM)containing emodin. First, this study evaluated the safety of repeated administration of different dosages of emodin over a 5-week period, with a particular focus on its impact on the liver. Next, the composition and content of BAs in serum and liver were analyzed. Subsequently, qRT-PCR was used to detect the mRNA expression of nuclear receptors and transporters related to BAs metabolism. The results showed that 1 g·kg~(-1) emodin induced hepatic damage, with bile duct hyperplasia as the primary pathological manifestation. It significantly increased the levels of various BAs in the serum and primary BAs(including taurine-conjugated and free BAs) in the liver. Additionally, it downregulated the mRNA expression of farnesoid X receptor(FXR), retinoid X receptor(RXR), and sodium taurocholate cotransporting polypeptide(NTCP), and upregulated the mRNA expression of cholesterol 7α-hydroxylase(CYP7A1) in the liver. Although 0.01 g·kg~(-1) and 0.03 g·kg~(-1) emodin did not induce obvious liver injury, they significantly increased the level of taurine-conjugated BAs in the liver, suggesting a potential interference with BAs homeostasis. In conclusion, 1 g·kg~(-1) emodin may promote the production of primary BAs in the liver by affecting the FXR-RXR-CYP7A1 pathway, inhibit NTCP expression, and reduce BA reabsorption in the liver, resulting in BA accumulation in the peripheral blood. This disruption of BA homeostasis leads to liver injury. Even doses of emodin close to the clinical dose can also have a certain effect on the homeostasis of BAs. Therefore, when using traditional Chinese medicine or formulas containing emodin in clinical practice, it is necessary to regularly monitor liver function indicators and closely monitor the risk of drug-induced liver injury.
Emodin/administration & dosage* ; Bile Acids and Salts/metabolism* ; Animals ; Male ; Liver/injuries* ; Chemical and Drug Induced Liver Injury/genetics* ; Drugs, Chinese Herbal/adverse effects* ; Humans ; Rats, Sprague-Dawley ; Mice ; Rats

In this study, lipopolysaccharide(LPS), ovalbumin(OVA), and compound 48/80(C48/80) were administered to establish non-infectious pneumonia models under simulated clinical conditions, and the correlation between their pathological characteristics and traditional Chinese medicine(TCM) syndromes was compared, providing the basis for the selection of appropriate animal models for TCM efficacy evaluation. An acute pneumonia model was established by nasal instillation of LPS combined with intraperitoneal injection for intensive stimulation. Three doses of OVA mixed with aluminum hydroxide adjuvant were injected intraperitoneally on days one, three, and five and OVA was administered via endotracheal drip for excitation on days 14-18 to establish an OVA-induced allergic pneumonia model. A single intravenous injection of three doses of C48/80 was adopted to establish a C48/80-induced pneumonia model. By detecting the changes in peripheral blood leukocyte classification, lung tissue and plasma cytokines, immunoglobulins(Ig), histamine levels, and arachidonic acid metabolites, the multi-dimensional analysis was carried out based on pathological evaluation. The results showed that the three models could cause pulmonary edema, increased wet weight in the lung, and obvious exudative inflammation in lung tissue pathology, especially for LPS. A number of pyrogenic cytokines, inclading interleukin(IL)-6, interferon(IFN)-γ, IL-1β, and IL-4 were significantly elevated in the LPS pneumonia model. Significantly increased levels of prostacyclin analogs such as prostaglandin E2(PGE2) and PGD2, which cause increased vascular permeability, and neutrophils in peripheral blood were significantly elevated. The model could partly reflect the clinical characteristics of phlegm heat accumulating in the lung or dampness toxin obstructing the lung. The OVA model showed that the sensitization mediators IgE and leukotriene E4(LTE4) were increased, and the anti-inflammatory prostacyclin 6-keto-PGF2α was decreased. Immune cells(lymphocytes and monocytes) were decreased, and inflammatory cells(neutrophils and basophils) were increased, reflecting the characteristics of "deficiency", "phlegm", or "dampness". Lymphocytes, monocytes, and basophils were significantly increased in the C48/80 model. The phenotype of the model was that the content of histamine, a large number of prostacyclins(6-keto-PGE1, PGF2α, 15-keto-PGF2α, 6-keto-PGF1α, 13,14-D-15-keto-PGE2, PGD2, PGE2, and PGH2), LTE4, and 5-hydroxyeicosatetraenoic acid(5S-HETE) was significantly increased, and these indicators were associated with vascular expansion and increased vascular permeability. The pyrogenic inflammatory cytokines were not increased. The C48/80 model reflected the characteristics of cold and damp accumulation. In the study, three non-infectious pneumonia models were constructed. The LPS model exhibited neutrophil infiltration and elevated inflammatory factors, which was suitable for the efficacy study of TCM for clearing heat, detoxifying, removing dampness, and eliminating phlegm. The OVA model, which took allergic inflammation as an index, was suitable for the efficacy study of Yiqi Gubiao formulas. The C48/80 model exhibited increased vasoactive substances(histamine, PGs, and LTE4), which was suitable for the efficacy study and evaluation of TCM for warming the lung, dispersing cold, drying dampness, and resolving phlegm. The study provides a theoretical basis for model selection for the efficacy evaluation of TCM in the treatment of pneumonia.
Animals ; Disease Models, Animal ; Mice ; Pneumonia/genetics* ; Medicine, Chinese Traditional ; Male ; Humans ; Cytokines/immunology* ; Female ; Lipopolysaccharides/adverse effects* ; Lung/drug effects* ; Drugs, Chinese Herbal ; Ovalbumin ; Mice, Inbred BALB C

Panax ginseng as a perennial herb of Araliaceae, exhibits pharmacological effects such as central nervous system stimulation, anti-tumor properties, and cardiovascular and cerebrovascular protection. The B3 gene family plays a crucial role in growth and development, antioxidant activity, stress resistance, and secondary metabolism regulation of plants and has been extensively studied in various plants. However, the identification and analysis of the B3 gene family in P. ginseng have not been reported. In this study, a total of 145 B3 genes(PgB3s) with complete open reading frames(ORF) were identified from P. ginseng and classified into five subfamilies based on domain types. Through correlation analysis with ginsenoside content, SNP/InDels analysis, and interaction analysis with key enzyme genes, 15 PgB3 transcripts were found to be significantly correlated with ginsenoside content and exhibited a close interaction network with key enzyme genes involved in ginsenoside biosynthesis, which indicated that these genes may participate in the regulation of ginsenoside biosynthesis. Additionally, this study found that PgB3 genes exhibited induced expression in response to methyl jasmonate(MeJA) stress, which aligned with the presence of abundant stress response elements in their promoters, confirming the important role of the B3 gene family in P. ginseng in stress resistance. The results of this study revealed the potential functions of PgB3 genes in ginsenoside biosynthesis and stress response, providing a significant theoretical basis for further research on the functions of PgB3 genes and their regulatory mechanisms.
Panax/metabolism* ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism* ; Ginsenosides/biosynthesis* ; Multigene Family ; Phylogeny

ObjectiveTo screen and evaluate the antidepressant compounds of Curcumae Rhizoma， and explore its mechanism of regulating the nuclear factor erythroid 2-related factor 2（Nrf2）/glutathione（GSH） peroxidase 4（GPX4）/GSH pathway from an antioxidant perspective. MethodsThe antioxidant activities in vitro of 11 characteristic components from Curcumae Rhizoma， including curcumol， curgerenone， curdione， curzerene， curcumenol， curcumenone， dehydrocurdione， isocurcumenol， furanodienone， furanodiene and zederone， were detected using 1，1-diphenyl-2-picrylhydrazyl（DPPH） and 2，2'-azinobis-（3-ethylbenzothiazoline-6-sulphonic acid） diammonium salt（ABTS） radical scavenging assays. The depression in Drosophila melanogaster was induced by chronic unpredictable mild stress（CUMS）， and W1118 wild-type male D. melanogaster were randomly divided into blank group， model group， curcumol group， curgerenone group， curdione group， curzerene group， curcumenol group，curcumenone group， dehydrocurdione group， isocurcumenol group， furanodienone group， furanodiene group， zederone group and fluoxetine group（10 μmol·L^-1）. The treatment groups received a dose of 0.1 g·L^-1 of 11 characteristic components from Curcumae Rhizoma， while the blank and model groups were administered equivalent volumes of solvent. The sucrose preference test， climbing test and forced swimming test were used to evaluate the behavioral indicators of depression in D. melanogaster. Liquid chromatography-mass spectrometry（LC-MS） was used to detect the levels of 5-hydroxytryptamine（5-HT） and dopamine（DA） in the brain of D. melanogaster， and the entropy weight method was used to comprehensively evaluate neurobehavioral and neurotransmitter indicators， resulting in the identification of the antidepressant active components of Curcumae Rhizoma. In addition， a mouse depression model was established by CUMS， and C57BL/6J mice were randomly divided into blank group， model group， low and high dose groups of curzerene（0.5， 1 mg·kg^-1）， and fluoxetine group（10 mg·kg^-1） to confirm the antidepressant effect of the optimal active ingredient by behavioral analysis. Flow cytometry was used to detect the content of reactive oxygen species（ROS） in the hippocampus of mice from each group. Enzyme-linked immunosorbent assay was used to detect the contents of adenosine triphosphate（ATP）， superoxide dismutase（SOD）， catalase（CAT） and GSH. Transmission electron microscope（TEM） was used to observe the effect of curzerene on the ultrastructure of mitochondria in hippocampal tissue. Western blot was performed to determine the level of Nrf2 protein， and Nrf2 inhibitor（ML385） was used to verify the relationship between the antidepressant effect of curzerene and regulation of Nrf2. Real time fluorescence quantitative polymerase chain reaction（Real-time PCR） was employed to detect the effect of curzerene on the mRNA expression level of GPX. ResultsIn vitro antioxidant experiments showed that curzerene and curgerenone exhibited the most significant ability to scavenge free radicals， and comprehensive evaluation results of entropy weight method indicated that curzerene stood out as the most promising active component. Compared with the blank group， the model group exhibited a significant decrease in sucrose preference coefficient and the number of times entering the open field center（P<0.01）， as well as a significant increase in immobility time in the forced swimming and tail suspension tests（P<0.01）， and the ROS content in hippocampus significantly elevated（P<0.01）， while the ATP content significantly reduced（P<0.01）. In the hippocampal neurons of the model group， mitochondrial cristae were disordered， with vacuolation of the inner membrane and severe damage. Nrf2 protein expression level in the model group was significantly decreased（P<0.05）， and the antioxidant enzymes SOD， CAT and GSH contents were also significantly reduced（P<0.05， P<0.01）， and the gene expression levels of GPX1， GPX4 and GPX7 were significantly decreased（P<0.01）. Compared with the model group， the high-dose group of curzerene showed a significant increase in the sucrose preference coefficient and the number of times entering the open field center（P<0.05）， as well as a significant decrease in immobility time in the forced swimming and tail suspension tests（P<0.05， P<0.01）. The ROS content in the hippocampus of the high-dose group of curzerene was significantly reduced（P<0.01）， while the ATP content was significantly increased（P<0.05）. The neuronal mitochondrial damage in the hippocampus of the high-dose group of curzerene was alleviated， and the expression level of Nrf2 protein was significantly increased（P<0.05）. The Nrf2 inhibitor ML385 reversed the improvement of curzerene on depressive behaviors in CUMS mice. The GSH content in the hippocampal neurons of the high-dose group of curzerene was significantly increased（P<0.01）， while there were no significant differences in SOD and CAT contents. The expression level of GPX4 gene in the hippocampal neurons of the high-dose group of curzerene was significantly increased（P<0.05）， while there were no significant differences in other GPX genes. ConclusionCurzerene is the best component with antidepressant activity in Curcumae Rhizoma. It may improve mitochondrial dysfunction to exert its antidepressant effect by regulating Nrf2 and its downstream GPX4/GSH pathway rather than CAT or SOD pathways.

Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.