OBJECTIVE To investigate the ameliorative effect and potential mechanism of imperatorin （IMP） on doxorubicin （DOX） resistance in breast cancer. METHODS The effects of maximum non-toxic concentration （100 μg/mL） of IMP combined with different concentrations of DOX （12.5， 25， 50， 75， 100 μg/mL） on the proliferation of MCF-7/DOX cells were determined by MTT method. MCF-7/DOX cells were divided into blank control group （1‰ dimethyl sulfoxide）， DOX group （50 μg/mL）， IMP+DOX group （100 μg/mL IMP+50 μg/mL DOX） and IMP group （100 μg/mL）. mRNA and protein expressions of multidrug resistance protein 1 （MDR1） and multidrug resistance-associated protein 1 in each group were measured. The relevant pathways and targets involved in the improvement of DOX resistance in breast cancer cells by IMP were screened and validated by using transcriptome sequencing technology， along with gene ontology （GO） enrichment analyses and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analyses. RESULTS Compared with DOX alone， the combination of IMP and DOX reduced the half inhibitory concentration of DOX on MCF-7/DOX cells from 81.965 μg/mL to 43.170 μg/mL， the reverse fold was 1.90， and the mRNA expression of MDR1 was significantly down-regulated （P＜0.05）. The results of GO enrichment analyses and KEGG pathway enrichment analyses indicated that the reversal of DOX resistance in breast cancer by IMP was mainly associated with the regulation of biological processes such as detoxification， multiple biological processes， and cell killing. The main pathway involved was the p53 signaling pathway， and the key targets mainly included constitutively photomorphogenic protein 1 （COP1）， cyclin E1 （CCNE1）， growth arrest and DNA damage-inducible protein 45A E-mail：tangxilan1983@163.com （GADD45A） and GADD45B. The results of the verification experiments showed that compared with DOX group， there was a trend of up-regulation of COP1 mRNA， and significant down- regulation of CCNE1， GADD45A， and GADD45B mRNA expression in IMP+DOX group （P＜0.05）. CONCLUSIONS The effect of IMP in ameliorating DOX resistance in breast cancer is related to its regulation of COP1， CCNE1， GADD45A and GADD45B targets in the p53 signaling pathway.

ADC189 is a novel drug of cap-dependent endonuclease inhibitor. In our study, its antiviral efficacy was evaluated in vitro and in vivo, and compared with baloxavir marboxil and oseltamivir. A first-in-human phase I study in healthy volunteers included single ascending dose (SAD) and food effect (FE) parts. In the preclinical study, ADC189 showed potent antiviral activity against various types of influenza viruses, including H1N1, H3N2, influenza B virus, and highly pathogenic avian influenza, comparable to baloxavir marboxil. Additionally, ADC189 exhibited much better antiviral efficacy than oseltamivir in H1N1 infected mice. In the phase I study, ADC189 was rapidly metabolized to ADC189-I07, and its exposure increased proportionally with the dose. The terminal elimination half-life (T_1/2) ranged from 76.69 to 98.28 hours. Of note, food had no effect on the concentration, clearance, and exposure of ADC189. It was well tolerated, with few treatment-emergent adverse events (TEAEs) reported and no serious adverse events (SAEs). ADC189 demonstrated excellent antiviral efficacy both in vitro and in vivo. It was safe, well-tolerated, and had favorable pharmacokinetic characteristics in healthy volunteers, supporting its potential for single oral dosing in clinical practice.
Humans ; Antiviral Agents/therapeutic use* ; Animals ; Male ; Adult ; Mice ; Female ; Endonucleases/antagonists & inhibitors* ; Influenza, Human/drug therapy* ; Young Adult ; Dibenzothiepins/pharmacology* ; Oseltamivir/pharmacology* ; Middle Aged ; Triazines/pharmacology* ; Thiepins/pharmacology* ; Influenza B virus/drug effects* ; Influenza A Virus, H1N1 Subtype/drug effects* ; Pyridines/pharmacology* ; Morpholines ; Pyridones

Objective : To investigate the effects of sinapine on lung tissue inflammation and mucus hypersecretion in asthmatic mice. Methods: Eight-week-old female C57BL/6J mice were randomly divided into Control group, ovalbumin(OVA) group, Sinapine group, and Sinapine+OVA group. The asthmatic mice model were established by intraperitoneal injection of OVA combined with aluminum hydroxide [Al(OH)3] suspension and OVA nasal stimulation. One hour before OVA nasal stimulation, the mice in Sinapine+OVA group and Sinapine group were intraperitoneally injected with sinapine solution, and the mice in OVA group and Control group were treated with the same dose of 0.9% sodium chloride solution. 24 hours after the last OVA stimulation, the inflammation of lung tissue of mice were observed by HE staining; the mucus secretion were evaluated by PAS staining; the mRNA expression levels of Interleukin-4(IL-4), Interleukin-5(IL-5), Interleukin-13(IL-13), tumor necrosis factor-alpha(TNF-α), Mucin 5ac(Muc5ac), and the mRNA of the key genes of Notch pathway such as Notch receptor 1(Notch1), Notch receptor 2(Notch2), Notch receptor 3(Notch3), and hes family bHLH transcription factor 1(Hes1) in lung tissues were detected by real-time fluorescent quantitative PCR(RT-qPCR); the expression levels of Notch1, Notch2, Notch3 and Hes1 proteins were determined by Western blot. Results : Compared with Control group, the inflammation score and PAS score of lung tissues of mice in OVA group increased(P<0.001); the mRNA expression levels of IL-4, IL-5, IL-13, TNF-α, and Muc5ac of mice in OVA group were enhanced(P<0.05); the mRNA and protein expression levels of Notch2, Notch3, and Hes1 of mice in OVA group significantly increased(P<0.001), while there was no significant difference in the mRNA and protein expression levels of Notch1. Compared with OVA group, the inflammation score and PAS score of lung tissues of mice in Sinapine+OVA group decreased(P<0.001); the mRNA expression levels of IL-4, IL-5, IL-13, TNF-α, and Muc5ac of mice in Sinapine+OVA group were reduced(P<0.05); the mRNA and protein expression levels of Notch2, Notch3, and Hes1 of mice in Sinapine+OVA group were downregulated(P<0.05), while there was no significant difference in the mRNA and protein expression levels of Notch1. Conclusion Sinapine can alleviate the lung tissue inflammation and mucus hypersecretion in asthmatic mice, and its mechanism may be related to the inhibition of Notch2/Notch3-Hes1 signal pathway.

ObjectiveTo investigate the possible mechanism of Huatan Sanjie Formula （化痰散结方， HSF） in treating Graves' disease （GD） from the perspective of thyroid angiogenesis. MethodsThirty-six BALB/c female mice were randomly divided into a normal control group （n=9） and a modeling group （n=27）. Mice in the modeling group were injected with 2.0×10⁹ PFU/ml of Ad-TSHR289 adenovirus into the tibialis anterior muscle to build GD model. Nine weeks after immunization， the successfully modeled mice were randomly divided into model group， methimazole （MMI） group and HSF group， with 9 mice in each group. The MMI group was given 5.2 mg/（kg·d） of methimazole tablets by gavage， while the HSF group was given HSF at a relative crude drug dosage of 7.02 g/（kg·d） by gavage. The normal control group and the model group were given 0.1 ml/10 g of pure water by gavage. All groups were administered intragastrically once a day for a total of 4 weeks. The levels of thyroxine （T4） and thyrotropin receptor autoantibodies （TRAb） in serum were detected by radioimmunoassay， while the pathological changes of the thyroid gland were assessed by HE staining. The vascular morphology of thyroid tissue was observed by CD34 immunohistochemical staining， and the microvessel density （MVD） was counted. The protein expression of vascular endothelial growth factor A （VEGFA） and vascular endothelial growth factor receptor 2 （VEGFR2） in thyroid was detected by Western-blot. ResultsCompared to those in the normal control group， the thyroid volume of the mice in the model group significantly increased with excessive congestion， and the pathology showed significant thyroid follicular hyperplasia， columnar and proliferated epithelial cells， and enlarged follicle size； serum T4 and TRAb significantly increased， as well as the count of thyroid MVD， and the protein expressions of thyroid VEGFA and VEGFR2 （P＜0.01）. Compared to those in the model group， the thyroid glands of the mice in the MMI group and the HSF group were significantly reduced， and the congestion was improved； pathology showed that thyroid follicular hyperplasia and epithelial cell proliferation were reduced， with smooth edges of the follicles and the significantly reduced inward protrusion； serum T4 and TRAb significantly decreased， as well as the thyroid MVD， thyroid VEGFA and VEGFR2 protein expressions （P＜0.05 or P＜0.01）. There was no significant difference in all indicators between the MMI group and the HSF group （P＞0.05）. ConclusionHSF may inhibit thyroid angiogenesis by down-regulating thyroid VEGFA/VEGFR2 signaling pathway， thereby improving goitre and hyperfunction in GD mice.

Objective To observe the effects of Daizong Prescription on glycogen metabolism in adipose tissue of obese mice;To explore its regulatory mechanism in activating browning in the white adipose tissue.Methods A obesity model was established by feeding high-fat diet to C57BL/6J mice.The obese mice were divided into model group,metformin group(0.15 g/kg),and Daizong Prescription low-(0.20 g/kg)and high-dosage(0.40 g/kg)groups.Mice fed a standard diet were set as the normal group,with 12 mice in each group.Each medication group was given corresponding drugs by gavage for 6 consecutive weeks.Body mass and fasting blood glucose were monitored,serum triglycerides(TG),total cholesterol(TC),high-density lipoprotein cholesterol(HDL-C),and low-density lipoprotein cholesterol(LDL-C)contents were measured.Brown adipose tissue from the interscapular region and white adipose tissue from the inguinal,perirenal and epididymal region were collected,the adipose tissue mass was measured,and the body fat coefficient was calculated.HE staining was performed to observe morphological changes in adipose tissue,PAS staining was used to observe glycogen distribution in adipose tissue,immunohistochemistry staining was performed to detect the expressions of Gys2,Ppp1r3c,and GSK-3β in inguinal white adipose tissue.Results Compared with the normal group,the body mass and fasting blood glucose in different time points of the model group significant increase(P<0.05,P<0.01),and serum TC and HDL-C contents significantly increased(P<0.01);the mass and body fat coefficient of white adipose tissue in inguinal,perirenal,and epididymis significantly increased(P<0.01),the cells in white adipose tissue in inguinal were hypertrophic and appeared as large vacuoles,with less glycogen accumulation,the expressions of Gys2 and Ppp1r3c significantly decreased(P<0.01).Compared with the model group,the mice in Daizong Prescription high-dosage group showed a significant decrease in body mass and fasting blood glucose at 4 and 6 weeks of administration(P<0.05,P<0.01),and the contents of serum TG,TC,HDL-C,and LDL-C were significantly decreased(P<0.01);the mass and body fat coefficient in white adipose tissue of perirenal and epididymal significantly decreased(P<0.05,P<0.01),and the mass of inguinal white adipose tissue significantly decreased(P<0.05),multiple irregularly shaped small vacuoles could be seen in inguinal white adipose tissue,accompanied by nuclear aggregation and increased glycogen accumulation,the expressions of Gys2 and Ppp1r3c significantly increased(P<0.01).There was no significant difference in the expression of GSK-3β inguinal white adipose tissue of mice among the groups.Conclusion Daizong Prescription can increase the activity of Gys2 by upregulating the expression of Ppp1r3c,promote glycogen synthesis,induce browning of adipose tissue,increase fat heat production,and improve obesity and related disorders of glycolipid metabolism.

Objective:To discuss the effect of Jiegeng Yuanshen Tang(JGYST)on airway tissue inflammation and mucus secretion in the mice with allergic asthma,and to clarify the related mechanism.Methods:Forty male C57BL/J mice were randomly divided into control group,JGYST group,ovalbumin(OVA)group,and OVA + JGYST group.The mice in OVA group and OVA +JGYST group were sensitized with 50 μg OVA via intraperitoneal injection twice weekly,followed by 20 μg OVA nasal drops daily for 7 d to induce asthma;the mice in OVA +JGYST group were gavaged with 200 μL JGYST 1 h before each OVA challenge,and the administration lasted for 7 d;the mice in control group were given equivalent dose of PBS via intraperitoneal injection,nasal drops,and gavage;the mice in JGYST group were given the same dose of PBS for intraperitoneal and nasal administration and gavaged with the same dose of JGYST.The pathomorphology of lung tissue of the mice in various groups was observed by HE staining and periodic acid-Schiff(PAS)staining,and the inflammation and PAS scores were calculated;flow cytometry method was used to detect the numbers of eosinophils,neutrophils,helper T lymphocyte 1(Th1)cells,helper T lymphocyte 2(Th2)cells,and dendritic cells(DCs),as well as the percentage of mature DCs and level of reactive oxygen species(ROS)in lung tissue of the mice in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of interleukin-4(IL-4),interleukin-10(IL-10),and tumor necrosis factor-α(TNF-α)mRNA in lung tissue of the mice in various groups.Results:The HE and PAS staining results showed that the mice in control group had intact airway and alveolar structure,without infiltration of inflammatory cells or mucus secretion;compared with control group,there was a large number of infiltrating inflammatory cells in airway tissue of the mice in OVA group,and the inflammation and PAS scores were increased(P<0.01);compared with OVA group,the infiltration of inflammatory cells in airway tissue of the mice in JGYST group and OVA + JGYST group was decreased,and the inflammation and PAS scores were significantly decreased(P<0.01).The flow cytometry results showed that compared with control group,the numbers of eosinophils,Th2 cells,and DCs in lung tissue of the mice in OVA group were increased(P<0.05 or P<0.01),and the percentage of mature DCs and level of ROS were significantly increased(P<0.01);compared with OVA group,the numbers of eosinophils,Th2 cells,and DCs in lung tissue of the mice in JGYST group and OVA + JGYST group were decreased(P<0.01),and the percentage of mature DCs and level of ROS were significantly decreased(P<0.01).The RT-qPCR results showed that compared with control group,the expression levels of IL-4,IL-10,and TNF-α mRNA in lung tissue of the mice in OVA group were increased(P<0.01);compared with OVA group,the expression levels of IL-4 and TNF-α mRNA in lung tissue of the mice in JGYST group and OVA + JGYST group were decreased(P<0.01),while the expression level of IL-10 mRNA was increased(P<0.01).Conclusion:JGYST can alleviate the airway tissue inflammation and mucus secretion in the mice with allergic asthma,and its mechanism may be related to reducing the number of Th2 cells and DCs,decreasing the ROS level and expression level of proinflammatory cytokine,and increasing the expression level of anti-inflammatory cytokine.

Objective:To discuss the effect of Aspergillus fumigatus(Af)on DNA damage and interleukin(IL)-33 expression in the human bronchial epithelial cells,and to clarify its related mechanism.Methods:Different concentrations(1,5,and 10 mg·L-1)of Af were used to stimulate the bronchial epithelial BEAS-2B cells to select the appropriate stimulation concentration.When the BEAS-2B cells were treated with N-acetylcysteine(NAC)and Af,the cells were divided into control group,Af group,NAC group,and Af+NAC group.When the BEAS-2B cells were treated with DNA double-strand break repair inhibitor NU7441 and Af,the cells were divided into control group,Af group,NU7441 group,and Af+NU7441 group.The comet assay was used to detect the percentages of comet tail DNA of cells in various groups;immunofluorescence method was used to detect the expression levels of DNA damage-related protein phosphorylated H2AX(yH2AX)in the cells in various groups;2,7-dichlorofluorescein diacetate(DCFH-DA)fluorescence probe was used to detect the levels of reactive oxygen species(ROS)in the cells in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of interleukih-33(IL-33),thymic stromal lymphopoietin(TSLP),and interleukih-25(IL-25)mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of phosphorylated nuclear factor κB(p-NF-κB),phosphorylated ataxia telangiectasia mutated(p-ATM),and γH2AX proteins in the cells in various groups.Results:Compared with control group,the percentage of comet tail DNA and the expression level of γH2AX in the cells in 1 mg·L-1 Af group showed no significant difference(P＞0.05),while the percentage of comet tail DNA and the expression level of γH2AX in the cells in 5 mg·L-1 Af group were significantly increased(P＜0.01);compared with 5 mg·L-1 Af group,the percentage of comet tail DNA and the expression level of γH2AX in the cells in 10 mg·L-1 Af group were significantly increased(P＜0.01).Compared with control group,the ROS levels in the bronchial epithelial cells in 1 mg·L-1 Af group was significantly increased(P＜0.05);compared with 1 mg·L-1 Af group,the ROS level in the cells in 5 mg·L-1 Af group was significantly increased(P＜0.01);compared with 5 mg·L-1 Af group,the ROS level in the cells in 10 mg·L-1 Af group was significantly increased(P＜0.05).After treatment of NAC,compared with Af group,the percentage of comet tail DNA(P＜0.01),the expression level of γH2AX(P＜0.05),and the ROS level(P＜0.01)in the cells in Af+NAC group were significantly decreased;after treatment of NU7441,compared with Af group,the percentage of comet tail DNA and the expression level of yH2AX in the cells in Af+NU7441 group were significantly increased(P＜0.01).The RT-qPCR results showed that after treatment of NAC,compared with control group,the expression level of IL-33 mRNA in the cells in Af group was significantly increased(P＜0.05);compared with Af group,the expression level of IL-33 mRNA in the cells in Af+NAC group was significantly decreased(P＜0.05);after treatment of NU7441,compared with Af group,the expression level of IL-33 mRNA in the cells in Af+NU7441 group was significantly increased(P＜0.05).The Western blotting results showed that after treatment of NAC,compared with control group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af group were significantly increased(P＜0.05);after treatment of NU7441,compared with Af group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af+NAC group were significantly decreased(P＜0.05);After treat ment of NU7441,compared with Af group,the expression levels of p-NF-κB,p-ATM,and γH2AX proteins in the cells in Af+NU7441 group were significantly increased(P＜0.05).Conclusion:Af promotes the IL-33 expression in the human bronchial epithelial cells by causing DNA damage,and its mechanism may be related to the activation of ATM/NF-κB signaling pathway.

ObjectiveTo explore the quality differences between steamed products and raw products of Citri Reticulatae Pericarpium（CRP）. MethodThe color of steamed products and raw products of CRP was determined from the perspective of appearance by electronic eye technique， and the quality differences between them was objectively characterized by the luminous value（L^*）， yellow-blue value（b^*）， red-green value（a^*） and total chromatic value（E^*_ab）. Based on this， ultra-high performance liquid chromatography（UPLC） was used to establish a fingerprint evaluation method with the mobile phase of acetonitrile（A）-0.1% formic acid aqueous solution（B） for gradient elution（0-5 min， 5%A； 5-30 min， 5%-20%A； 30-60 min， 20%-52%A）， detection wavelength at 270 nm， flow rate of 0.3 mL·min^-1 and column temperature of 30 ℃. The quality differences between steamed products and raw products of CRP were compared from the perspective of chemical composition， and correlation analysis was used to reveal the correlation between the difference in appearance color and the difference in internal chemical composition. ResultAfter being steamed， L^*， b^* and E^*_ab of CRP showed an overall decreasing trend， indicating that the color of the steamed products darkened and deepened from yellow to blue but still tended to be yellow， while a^* showed an overall increasing trend， indicating that the color of the steamed products tended to red. A total of 24 peaks were identified in the fingerprint profiles of raw products and steamed products of CRP， and 13 of the main peaks were identified. The precision， stability and repeatability studies showed that compared with the reference peak （peak 14， hesperidin）， the relative standard deviations（RSDs） of the relative peak area and relative retention time of the remaining peaks were<3.0%.The results of chemometric statistical analysis showed that there were some differences between raw products and steamed products of CRP， and 7 main differential components were identified， among which 5-hydroxymaltol（peak 1） and 5-hydroxymethylfurfural（peak 2） were the characteristic components of steamed products. The correlation analysis results showed that， in addition to the above two characteristic components， four components of peak 4， peak 10 （vicenin-2）， peak 23 （tangeretin） and peak 24 （5-demethylnobiletin） also correlated significantly with the color change （E^*_ab） of the samples （P<0.05， P<0.01）. ConclusionBefore and after steaming， not only the chemical composition changes， but also the color. Comparing the characteristic peaks of chemical composition difference and color difference before and after steaming of CRP， it is found that 5-hydroxymaltol， 5-hydroxymethylfurfural and peak 4 are common characteristic difference components， which can provide a reference for establishing the characteristic quality control method of steamed products， and quickly evaluating the quality difference between raw products and steamed products of CRP.

Brain/diagnostic imaging* ; Head

Objective:To investigate the effect of ultrasound monitoring of inferior vena cava collapse index (IVC-CI) guiding fluid replacement on circulation in elderly patients during induction of general anesthesia.Methods:A total of 71 elderly patients who underwent elective surgery under general anesthesia and tracheal intubation at Hunan Provincial People′s Hospital from April 2021 to September 2022 were randomly divided into control group (35 cases) and observation group (36 cases) using a random number table method. Before anesthesia, both groups of patients underwent IVC ultrasound examination and calculated the IVC-CI value. For patients with IVC-CI≥40%, the observation group was given 8 ml/kg of crystal solution before anesthesia induction, while the control group was not treated. The incidence of hypotension, the use of vasoactive drugs, and the total infusion volume from anesthesia induction to skin incision were recorded in two groups. Mean arterial blood pressure (MBP), heart rate (HR), oxygen saturation (SpO 2), cardiac index (CI), and cardiac volume variability (SVV) before anesthesia (T 0), 5 min after induction (T 1), 1 min after tracheal intubation (T 2), 5 min after tracheal intubation (T 3), 10 min after tracheal intubation (T 4), and 1 min before skin incision (T 5) were recorded and compared between the two groups. Results:The incidence of hypotension (27.8% vs 60.0%) and utilization rate of vasoactive drugs (25.0% vs 48.6%) in the observation group were lower than those in the control group, and the total infusion volume during anesthesia induction was higher than that in the control group, with statistical significance (all P<0.05). SVV, CI and MBP at T 1, T 3, T 4 and T 5 were significantly different from those at T 0 in the control group ( F=3.85, 14.66, 3.96, all P<0.05). SVV, CI and MBP at T 1, T 3, T 4 and T 5 in the observation group were significantly different from those at T 0 ( F=3.51, 13.20, 4.35, all P<0.05). There was no significant difference in SVV, CI, MBP, HR and SpO 2 between 2 groups (all P>0.05). Conclusions:For the elderly patients with preoperative IVC-CI≥40%, pre-filling with 8ml/kg crystal solution before anesthesia induction can significantly reduce the incidence of hypotension and the utilization rate of vasoactive drugs in the elderly patients during anesthesia induction.