ObjectiveTo investigate the effect of folic acid-modified crebanine polyethylene glycol-polylactic acid hydroxyacetic acid copolymer（PEG-PLGA） nanoparticles（FA-Cre@PEG-PLGA NPs， hereinafter referred to as NPs） combined with ultrasonic irradiation on subcutaneous tumor of liver cancer in Kunming（KM） mice. MethodsEighty-four healthy male KM mice were utilized to establish a subcutaneous tumor model of mouse hepatocellular carcinoma with H22 cells， then mice were randomly divided into model group， placebo group， hydroxycamptothecin group（8 mg∙kg^-1）， low， medium and high dose crebanine raw material groups（2， 2.5， 3 mg∙kg^-1， hereinafter referred to as the low， medium and high dose crebanine groups， respectively）， low， medium and high dose NPs groups（2， 2.5， 3 mg∙kg^-1）， and low， medium and high dose NPs combined with ultrasonic irradiation groups（2， 2.5， 3 mg∙kg^-1， hereinafter referred to as the low， medium and high dose combination groups， respectively）. The corresponding doses of drugs were administered via tail vein injection， the model group received no treatment， while the placebo group was injected with an equivalent amount of normal saline. Dosing was conducted for a total of 10 times on alternate days. The body mass of the mice was monitored， and parameters such as body mass change rate， thymus index， spleen index， tumor volume， tumor weight， relative tumor growth rate（T/C）， and tumor inhibition rate（TGI） were calculated. Pathological changes in liver and kidney tissues as well as the tumor were observed by hematoxylin-eosin（HE） staining. Additionally， the levels of aspartate aminotransferase（AST）， alanine aminotransferase（ALT）， blood urea nitrogen（BUN） and creatinine（CREA） in serum of mice were detected by biochemical method. Furthermore， the effect of ultrasound on the distribution of NPs in subcutaneous tumors of mouse hepatocellular carcinoma was observed by in vivo imaging technique. ResultsAmong different treatment methods， the combination of NPs and ultrasound irradiation had the best therapeutic effect. Compared with the model group， the body mass growth rates of mice in the medium and high combination groups decreased， while the thymus index and spleen index increased， but there was no statistically significant difference in serum AST， ALT， BUN and CREA levels， indicating that NPs combined with ultrasound irradiation had little effect on the normal physiological state of the body， oth groups had TGI>40% and T/C<60%， indicating a clear anti-tumor effect. Pathological analysis showed that compared with the NPs groups， the combination groups exhibited varying degrees of necrosis in tumor cells， accompanied by less damage to the liver and kidneys. In vivo imaging of small animals showed that compared with the high dose NPs group， the high dose combination group had stronger tumor targeting ability（P<0.01）. ConclusionNPs combined with ultrasonic irradiation can not only effectively targeted the drug to the tumor site， inhibit the subcutaneous tumor growth of mouse liver cancer， but also decrease damage to liver and kidney tissues.

Objective To explore the role of neogambogic acid in the characteristics of colorectal cancer stem cells (CRC-CSCs) through the Wnt/β-catenin signaling pathway. Methods The colorectal cells SW480 and HCT166 were divided into control group and neogambogic acid groups (1.5, 3, 6, and 12 μmol/L). The viability of CRC-CSCs was determined by MTT method, and spheroid and clone formation assays were used to assess the capacity of spheroid formation and self-renewal ability of the cells. The effects of neogambogic acid on the apoptosis and cell cycle of CRC-CSCs were evaluated by flow cytometry assays. Real-time quantitative PCR was used to detect the mRNA expression levels of relative markers (CD133, CD44, ALDH1, Oct4, and Nanog) of CRC-CSCs, and the protein expression levels of the self-renewal marker (PCNA), apoptosis markers (cleaved caspase-3 and cleaved caspase-9), and Wnt/β-catenin signaling pathway markers (p-GSK3β, GSK3β, β-catenin, and Wnt) were analyzed using Western blot. Results Compared with the control group, after neogambogic acid treatment, the viability of SW480 and HCT116 cells decreased (P<0.05), the spheroid forming ability and the clone numbers of CRC-CSCs decreased (P<0.001, P<0.01) but the cell apoptosis rate increased (P<0.01), and cell cycle was arrested in G₀/G₁ phase. Moreover, neogambogic acid downregulated the mRNA and protein expression of relative markers of CRC-CSCs (CD133, CD44, ALDH1, Oct4, and Nanog), PCNA, p-GSK3β, β-catenin, and Wnt (P<0.05) and upregulated the expression of cleaved caspase-3, cleaved caspase-9, and GSK3β (P<0.01). Conclusion Neogambogic can inhibit the stem cell properties of colorectal cells via inhibition of Wnt/β-catenin signaling pathway. As a result, neogambogic acid may be an attractive agent against colorectal cancer.

OBJECTIVE To investigate the targeting effect of folic acid-modified crebanine nanoparticles （FA-Cre@PEG- PLGA NPs， hereinafter referred to as “NPs”） combined with ultrasound irradiation on M109 cells in vitro and in vivo after administration， and explore the anti-tumor mechanism. METHODS CCK-8 assay was used to detect the inhibitory effect of NPs combined with ultrasound irradiation on the proliferation of M109 cells， and the best ultrasound time was selected. Using human lung cancer A549 cells as a control， the targeting of NPs combined with ultrasound irradiation to M109 cells was evaluated by free folic acid blocking assay and cell uptake assay. The effects of NPs combined with ultrasound irradiation on the migration， invasion， apoptosis， cell cycle and reactive oxygen species （ROS） levels of M109 cells were detected by cell scratch test， Transwell chamber test and flow cytometry at 1 h after 958401536@qq.com administration； the changes of mitochondrial membrane potential （MMP） were observed by fluorescence inverted microscope. A mouse subcutaneous tumor model of M109 cells was constructed， and the in vivo tumor targeting of NPs combined with ultrasound irradiation was investigated by small animal in vivo imaging technology. RESULTS NPs combined with ultrasound irradiation could significantly inhibit the proliferation of M109 cells， and the optimal ultrasound time was 1 h after administration. The free folic acid could antagonize the inhibitory effect of NPs on the proliferation of M109 cells， and combined with ultrasound irradiation could partially reverse this antagonism. Compared with A549 cells， the uptake rate of NPs in M109 cells was significantly higher （P＜0.01）， and ultrasound irradiation could promote cellular uptake. NPs combined with ultrasound irradiation could inhibit the migration and invasion of M109 cells and block the cell cycle in the G0/G1 and G2/M phases. Compared with control group， the apoptosis rate of M109 cells and ROS level were increased significantly （P＜0.01）， while the MMP decreased significantly （P＜0.01） in the different concentration （100， 200， 300 μg/mL） groups of M109 cells. Compared with the mice in non-ultrasound group， the fluorescence intensity and tumor-targeting index of the tumor site in the 0 h ultrasound group were significantly enhanced （P＜0.05 or P＜0.01）. CONCLUSIONS NPs combined with ultrasound irradiation have a strong targeting effect on M109 cells in vitro and in vivo， the anti-tumor mechanism includes inhibiting cell migration and invasion， blocking cell cycle， and inducing apoptosis.

ObjectiveTo identify the prototypical components and metabolites absorbed into blood and cerebrospinal fluid of Schisandrae Chinensis Fructus（SCF） based on sequential metabolism combined with liquid chromatography-mass spectrometry. MethodBlood and cerebrospinal fluid samples of integrated metabolism， intestinal metabolism and hepatic metabolism were collected from male SD rats after gavage and in situ intestinal perfusion administration， and ultra-performance liquid chromatography-quadrupole/electrostatic field orbitrap high-resolution mass spectrometry（UPLC Q-Exactive Orbitrap MS） was used to analyze and compare the differences in the spectra of SCF extract， blank plasma， administered plasma， blank cerebrospinal fluid and administered cerebrospinal fluid with ACQUITY UPLC BEH Shield RP18 column（2.1 mm×100 mm， 1.7 µm）， the mobile phase was acetonitrile（A）-0.1% formic acid aqueous solution（B） for gradient elution（0-7 min， 95%B； 7-12 min， 95%-35%B； 12-17 min， 35%-15%B； 17-20 min， 15%-12%B； 20-22 min， 12%-5%B； 22-23 min， 5%B； 23-25 min， 5%-95%B； 25-28 min， 95%B）. And heated electrospray ionization（HESI） was used with positive and negative ion modes， the scanning range was m/z 100-1 500. The prototypical constituents and their metabolites absorbed into blood and cerebrospinal fluid of SCF were identified according to the retention time， characteristic fragments， molecular formulae and the information of reference substances. ResultA total of 42 chemical components were identified in the extract of SCF， including lignans， flavonoids， amino acids， tannins， and others， of which lignans were the main ones. A total of 27 prototypical components and 14 metabolites were identified in plasma samples from different sites. A total of 15 prototypical components and 9 metabolites were identified in cerebrospinal fluid. The main metabolic reactions involved in the formation of metabolites were mainly demethylation， methylation， demethoxylation and hydroxylation. ConclusionThrough the systematic identification of the prototypical components and metabolites of SCF in rats， it provides data support for further better exploring the material basis of SCF in the treatment of central nervous system diseases.

Objective To evalute the drug resistance characteristics of tuberculosis(TB)patients of all ages in Guangdong Province during 2014-2020,and provide prevention and treatment strategies of tuberculosis.Method We used 39,048 clinical isolates of Mycobacterium tuberculosis(MTB)belonging to patients with confirmed TB from 2014 to 2020,from 32 TB drug-resistant surveillance sites in Guangdong Province,and we retrospectively analyzed the laboratories data of patients with drug-resistant TB,and grouped patients by age and region,to explore the trend of drug-resistance of MTB clinical isolates,the trend and incidence differences of multi-resistant TB(including monodrug-resistant TB(MR-TB),polydrug-resistant TB(PDR-TB),multidrug-resistant TB(MDR-TB)and exten-sively drug-resistant TB(XDR-TB)),and resistance characteristics of MTB clinical isolates to drugs in focus(rifam-picin and ofloxacin).Result The differences in the resistance rates of MTB clinical isolates to nine antituberculosis drugs among patients at 32 TB drug resistance surveillance sites in Guangdong Province from 2014 to 2020 were not statistically significant(P>0.05).The rates of MR-TB,PDR-TB,MDR-TB,XDR-TB,and total resistance isolates of MTB clinical isolates were 14.46%,5.16%,5.16%,4.58%,and 1.29%,respectively.he pediatric group had a higher MR rate(15.4%)than the adult and geriatric groups,while the adult and geriatric groups had higher MDR rates(5.0%and 5.0%,respectively).The geriatric group also had a higher XDR rate(2.1%),with statistically significant differences(P<0.001).The rates of MR-TB(14.8%),PDR-TB(5.3%),MDR-TB(4.7%),XDR-TB(1.4%),ofloxacin resistance(11.33%)and rifampicin resistance(6.92%)of MTB clinical isolates were higher in patients from the Pearl River Delta than in other regions of Guangdong Province,with statistically significant differ-ences(P<0.001).Conclusion According to the data from the surveillance sites,the epidemiological trend of drug-resistant TB in Guangdong Province is leveling off during the period 2014-2020.However,the incidence of drug-resistant TB is higher in specific populations(e.g.children and the elderly),and the incidence of drug-resistant TB and the rate of drug resistance to drugs in focus are higher in the Pearl River Delta than in other regions of Guang-dong Province,necessitating further investigation and the development of novel prevention and control strategies.

Background Exposures to environmental pollution and specific occupational hazards exacerbate pulmonary fibrosis which has a complex pathogenesis and lacks effective therapeutic drugs. The extract from Dracocephalum moldavica L. can alleviate pulmonary fibrosis through anti-inflammatory and anti-pyroptosis pathways, but its mechanism of prevention and treatment for pulmonary fibrosis remains unclear. Objective To elucidate the targets and potential mechanism underlying the anti-pulmonary fibrosis efficacy of Dracocephalum moldavica L. extract by employing an amalgamation of network pharmacology and empirical verification. Methods The chemical composition of the extract of Dracocephalum moldavica L. was retrieved with the help of China National Knowledge Infrastructure (CNKI) and Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). The disease targets related to pulmonary fibrosis were inquired using Gene Cards and DisGeNET. A protein-protein interaction (PPI) was constructed using the Search Tool for the Retrieval of Interacting Genes (STRING) database and Cytoscape software. The predicted potential targets were analyzed by the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses through the Database for Annotation, Visualization and Integrated Discovery (DAVID) and validated by molecular docking. Thirty-two rats were randomly divided into a control group, a model group, a low-dose group of Dracocephalum moldavica L. extract (100 mg·kg⁻¹), and a high-dose group of Dracocephalum moldavica L. extract (400 mg·kg⁻¹), with eight rats in each group. A rat model of pulmonary fibrosis was constructed using bleomycin (5 mg·kg⁻¹) intratracheal instillation, and an equal volume of saline was instilled into the control group. After modelling, 400 and 100 mg·kg⁻¹ of Dracocephalum moldavica L. extract were given the high-dose and low-dose groups by gavage, and an equal volume of saline was given by gavage to the control group and the model group, once per day, for consecutive 28 d. The animals were then neutralized, and lung tissues were collected. Structural changes in rat lung tissue were evaluated by observing stained pathological sections. Western blot (WB) was used to detect fibrosis-related proteins type I collagen (Col-I), α-smooth muscle actin (α-SMA), phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT) in lung tissues. Real-time fluorescence quantitative PCR (RT-qPCR) was used to detect α-SMA and Col-I mRNA levels in lung tissue. Enzyme-linked immunosorbent assay (ELISA) was used to detect tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in rats. Results A total of 378 key chemical components of the Dracocephalum moldavica L. extract and 1611 lung fibrosis-related targets were identified. Among them, 574 potential targets of Dracocephalum moldavica L. extract acting on lung fibrosis were obtained. The key targets determined by Degree value were albumin (ALB), tumour necrosis factor (TNF), AKT1, etc. The results of KEGG analysis suggested that the potential targets of Dracocephalum moldavica L. extract against pulmonary fibrosis mainly involved the PI3K-AKT, HIF-1 signaling pathway, TNF signaling pathway, and MAPK signaling pathway. The molecular docking results showed that the binding energy between the active components (quercetin, apigenin, aesculetin, quercitrin) of Dracocephalum moldavica L. extract and the core targets of pulmonary fibrosis (TNF, IL-6, ALB, AKT1) were all <-29.288 kJ·mol⁻¹, indicating good binding ability. The animal validation results showed that compared with the control group, the rats in the model group had disrupted alveolar structure, obvious inflammatory cell infiltration, positive blue-striped collagen fibre deposition, and increased Col-I and α-SMA protein expression and transcription levels (P<0.001), p-PI3K and p-AKT expression levels (P<0.001), and levels of inflammatory factors TNF-α, IL-6, and IL-1β (P<0.001). Compared with the model group, the high- and low-dose groups of Dracocephalum moldavica L. extract alleviated the progression of pulmonary fibrosis, reduced inflammatory cell infiltration, and attenuated collagen fibre deposition in rats, with a decrease in the protein expression and transcription levels of Col-I and α-SMA (P<0.01), the expression levels of p-PI3K and p-AKT (P<0.001), and the levels of inflammatory factors IL-6 and TNF-α (P<0.05, P<0.001). Conclusion Dracocephalum moldavica L. extract manifests anti-pulmonary fibrotic properties through the modulation of the PI3K-AKT signaling cascade and the suppression of inflammatory reactions.

Bladder cancer （BCa） is the most common malignant tumor of the urinary system， and its incidence is increasing year by year. At present， for all patients with resectable non-metastatic muscle-invasive BCa， radical cystectomy + bilateral pelvic lymph node dissection is strongly recommended， but they still face the risk of recurrence， metastasis and death. In recent years， the proportion of patients with advanced and metastatic BCa is increasing among patients with newly diagnosed BCa. Although current treatment models are diverse， they often struggle to achieve significant efficacy due to their low effectiveness and adverse effects， resulting in low survival rates for patients with advanced and metastatic BCa. Therefore， the treatment of BCa still faces great challenges， and there is an urgent need to discover an effective new antitumor drug. With the improvement of medical standards， traditional Chinese medicine has shown great advantages in the treatment of BCa. Traditional Chinese medicine is mild and easy to accept， and can inhibit tumor progression through a multi-pathway， multi-way and multi-target manner， so as to exert its anticancer effect. Taraxaci Herba is a medicinal and food homologous plant， which has many biological activities， such as antibacterial， anti-inflammatory， anti-oxidation， anti-tumor， protecting liver and gallbladder， reducing blood sugar and enhancing immunity， and it has shown a clear anticancer effect in breast cancer， liver cancer， gastric cancer， tongue cancer and lung cancer. By reviewing previous studies worldwide， this article summarizes the mechanism of Taraxaci Herba extract in inducing autophagy and apoptosis， inhibiting cell migration and invasion， regulating cell cycle and proliferation， regulating cell metabolism， inhibiting tumor angiogenesis， combining the effects of chemotherapeutic drugs， and regulating the transduction of related signal pathways. On this basis， this study systematically elaborates on the potential mechanism of Taraxaci Herba against BCa， in order to provide a theoretical basis for the research and treatment of BCa.

Objective To investigate the expression and significance of cyclin dependent kinase inhibitor 3(CDKN3)in human papillomavirus type 16(HPV16)-positive cervical cancer.Methods CDKN3 expression in pan-cancer was retrieved and downloaded from the Gene Expression Profiling Interactive Analysis(GEPIA)platform,and the expression levels of CDKN3 between normal cervical tissues(13 samples)and cervical cancer tissues(306 samples)were compared.Subsequently,GSE39001 data of HPV16-positive cervical cancer was sourced and downloaded from the Gene Expression Omnibus(GEO)database,and the expression levels of CDKN3 mRNA in HPV16-positive cervical cancer tissues(43 samples)and normal cervical tissues(12 samples)were compared.Immunohistochemical method was used to detect the expression of CDKN3 in 12 ca-ses of HPV16-positive cervical cancer,12 cases of HPV16-positive cervical precancerous lesions,10 cases of HPV16-positive chronic cervicitis and 7 cases of HPV-negative normal cervical samples collected from the Af-filiated Hospital of Guizhou Medical University.SiHa(HPV16-positive),HeLa(HPV18-positive)and HCC94(HPV-negative)cervical cancer cell lines were selected,and their CDKN3 expression were detected by West-ern blot.Results The GEPIA platform analysis showed that CDKN3 was highly expressed in pan-cancer,and the expression level of CDKN3 in cervical cancer tissue was significantly higher than that in normal cervical tissue(P<0.05).The GEO dataset reflected a significantly increased CDKN3 mRNA expression level in HPV16-positive cervical cancer compared to normal cervical tissue(P<0.001).Immunohistochemical verifi-cation showed that the positive expression rates of CDKN3 in HPV16-positive cervical cancer,HPV16-positive cervical precancerous lesion,HPV16-positive chronic cervicitis and HPV-negative normal cervical tissues were 91.7%,58.3%,0 and 0,respectively.Western blot analysis of cervical cancer cells showed that the expression level of CDKN3 in SiHa(HPV16-positive)cells was significantly higher than that in HeLa(HPV18-positive)and HCC94(HPV-negative)cells(P<0.05).Conclusion CDKN3 is a new oncogene of HPV16-positive cer-vical cancer,which may be used as a marker of cervical precancerous lesions and cervical cancer screening,and may provide a theoretical basis for subsequent mechanism research and targeted therapy.

Objective To analyze the clinical outcomes of early invasive fungal disease(IFD)in patients after allogenetic hematopoietic stem cell transplantation(allo-HCST)with metagenomic next-generation sequencing(mNGS).Methods A retrospective analysis was conducted on patients undergoing allo-HCST in our Bone Marrow Transplantation Center between July 2021 and October 2022.These patients experienced one of the following conditions within 100 d after transplantation:① Patients with persistent fever and negative blood culture after empiric antimicrobial therapy for 72 h or longer;② Hyperpyrexia of unknown origin occurred again after effective anti-infection in the past;③ Symptoms in lower respiratory tract associated with lung lesions on CT scan,and empiric anti-infective therapy was ineffective.Peripheral blood or bronchoscopic alveolar lavage fluid were tested with mNGS,and overall survival(OS)and non-relapse mortality(NRM)were analyzed.Results There were 60 patients enrolled in this study.For the peripheral blood samples of 47 cases and bronchoalveolar lavage fluid samples of 13 cases,mNGS found that 19 cases were negative to pathogens,30 cases were non-fungal positive,and 11 case were fungal positive,including 3 cases of aspergillus,5 cases of mucor,2 cases of Candida tropicalis,and 1 case of Trichosporon asahii.Of the 11 patients with fungal positive,8 achieved complete remission after antifungal therapy according to the mNGS results.The 1-year OS and NRM of the 60 patients were 70.0％(95％CI:64.1％～75.9％)and 20.0％(95％CI:11.9％～32.5％),respectively,while those of the fungal infection patients were 54.5％(95％CI:49.5％～69.5％)and 36.4％(95％ CI:15.5％～70.3％),respectively.No significant differences were seen in 1-year OS(P=0.487)and 1-year NRM(P=0.358)among the negative,fungal infection and non-fungal infection patients,neither OS(P=0.238)and NRM(P=0.154)between the fungal infection and the non-fungal infection patients.Conclusion mNGS can rapidly diagnose the early IFD after allo-HSCT,which is helpful for timely and effective treatment and improves the prognosis of patients.

Objective Based on the theory of"lung governs the skin and hair"in"Yellow Emperor's Inner Classic",this paper analyzes the medication rules of whitening and freckle-removing.The aim of this study is to provide reference for the clinical practice of traditional Chinese medicine(TCM)theory and the medication in TCM cosmetics.Methods"Chinese Medical Classics"was used to search the records of whitening and freckle-related drugs.The frequency,nature,flavor,meridian tropism and compatibility laws of TCM for whitening and freckle-removal were analyzed by statistics and association rules.The network pharmacology research was used to analyze the whitening and freckle-removing effects and mechanisms of high-frequency drugs.Then,the potential active ingredients were analyzed.The whitening and anti-freckle effect was verified through cytotoxicity experiments and melanin content detection.Results A total of 171 external prescriptions were selected in eligible articles,including 261 Chinese medicinals,most of which were pungent and belong to the lung meridian.The most frequently used Chinese medicinals was"Erbai Yixin"(EBYX,Angelicae Dahuricae Radix,Typhonii Rhizoma,Asari Radix et Rhizoma).Network pharmacological analysis showed that the core targets of EBYX for whitening and removing freckles are TP53,EGFR,ALB,etc.,which are mainly involved in oxygen perception and response,skin immune regulation,skin cell growth,differentiation,stress,inflammatory response,and other biological processes.Based on the results of molecular docking,biological analysis proved that the active ingredients of EBYX are chrysophanol,gallic acid and caffeic acid,which have inhibitory effects on the proliferation of melanoma cells and melanin production.Conclusion Most of the ancient prescriptions for whitening and removing freckles are pungent and belong to the lung meridian,which embodies the theory of"lung governs the skin and hair".The high-frequency drug EBYX may play a role by regulating skin redox,immunity and inflammation.The active ingredients of EBYX have an inhibitory effect on melanin formation.This study enriches the scientific connotation of TCM whitening and freckle-removing prescriptions based on the theory of"lung governs the skin and hair",realizes interdisciplinary integration and provides support for the modernization of TCM.