AIM To determine the contents of aspartic acid,glutamic acid,serine,glycine,threonine,citrulline,arginine,alanine,γ-amino-butyric acid,tyrosine,valine,phenlalanine,isoleucine,ornithine,leucine,lysine and proline in Gualoupi Injection and its intermediates,and to analyze their change laws.METHODS The OPA-FMOC online derivatization analysis was performed on a 45℃ thermostatic Waters XBridge C18 column(4.6 mm×100 mm,3.5 μm),with the mobile phase comprising of phosphate buffer solution-[methanol-acetonitrile-water(45 : 45 : 10)]flowing at 1 mL/min in a gradient elution manner,and the detection wavelengths were set at 262,338 nm.Principal component analysis and heatmap analysis were adopted in chemical pattern recognition for the corresponding intermediates in ten processes of six batches of samples.RESULTS Seventeen amino acids showed good linear relationships within their own ranges(R2>0.998 0),whose average recoveries were 83.4％-119.5％with the RSDs of 0.91％-7.94％.Different batches of samples in the same process were clustered,and the corresponding intermediates in different processed were clustered into three groups.Alcohol precipitation and cation exchange column demonstrated the biggest influences on amino acid composition.CONCLUSION This experiment can provide important references for the critical factors on quality control of Gualoupi Injection,thus ensure the stability and uniformity of final product.

A hydrophilic interaction chromatography tandem mass spectrometry method was developed for simultaneous quantification of 35 components in gualoupi injection. The analytes were separated with an ACQUITY XBridge Amide column using 20 mmol·L^-1 ammonium formate aqueous solution (pH 3.0) as mobile phase A and 20 mmol·L^-1 ammonium formate (pH 3.0)∶acetonitrile (1∶9) as mobile phase B for gradient elution. Mass spectrometry with dynamic multiple reaction monitoring and external standard method were used for quantitative analysis. A total of 35 components were determined in 10 batches of gualoupi injection. The results showed that the 35 compounds had a good linear relationship within their respective concentration ranges with the correlation coefficients (R² > 0.998 0), the recoveries ranged from 76.6% to 118.5%. The results showed that γ-aminobutyric acid, trigonelline, alanine, threonine, homoserine, citrulline, and leucine were abundant in gualoupi injection, while nicotinamide, methylsuccinic acid, cytosine and choline account for a low percentege. The present study provides an important reference for elucidation of the effective material basis and the improvement of quality standard of gualoupi injection.

Objective To investigate the Gram-positive coccus resistance in nationwide's tertiary hospitals and understand the trend of antimicrobial resistance.Methods All the clinical isolates were collected from 19 hospitals and the minimal inhibitory concentrations(MICs)were tested using agar/broth dilution method recommended.Results A total of 1 974 pathogenic Gram-positive coccus from 19 tertiary hospitals in 19 cities nationwide over the period from July 2021 to June 2022 were studied.Based on the MIC results,the prevalence of methicillin resistant Stapylococcus aureus(MRSA)and methicillin resistant Stapylococcus epidermidis(MRSE)were 36.4％and 79.9％respectively.No vancomycin insensitivity Staphylococcus was detected.Staphylococcus aureus were 100％susceptibility to linezolid and teicoplanin.Antibiotic resistance rate of Enterococcus faecalis and Enterococcus faecium to ampicillin were 3.1％and 92.9％.The detectation rate of vancomycin resistant Enterococcus(VRE)was 1.6％.Nonsusceptibility rate of Enterococcus faecalis to linezolid was 32.2％,two consecutive monitoring rises and nonsusceptibility rate of Enterococcus faecium(12.5％)was also significantly increased.The prevalence of penicillin non-susceptible Streptococcus pneumoniae(PNSSP)was 0.8％based on non-meningitis and parenteral administration criterion,decrease of nearly 30 percentage points from the previous surveillance.While for cases of oral penicillin,the rate was 71.8％,showing similar to last time.The results indicated that the number of strains with higher MIC value of penicillin(MIC ≥4 mg·L-1)decreased significantly.There were no significant differences of resistance rates of Stapylococcus aureus,Stapylococcus epidermidis,Enterococcus faecalis,Enterococcus faecium and Streptococcus pneumoniae among various groups such as different department,age,or specimen source.Conclusion VRE detection ratio stablized at a relatively low level.The number of Streptococcus pneumoniae with higher MIC value of penicillin decreased significantly compared with the previous monitoring.The increase of linezolidin-insensitive Enterococcus was noteworthy.

Objective To investigate the Gram-negative bacteria resistance in nationwide's tertiary hospitals and understand the trend of antimicrobial resistance.Method All the clinical isolates were collected from 19 hospitals and the minimal inhibitory concentrations(MICs)were tested using agar/broth dilution method recommended.Results A total of 4 066 pathogenic isolates from 19 tertiary hospitals in 19 cities nationwide over the period from July 2021 to June 2022 were studied.Based on the MIC results,Escherichia coli and Klebsiella pneumoniae showed extended spectrum β-lactamase(ESBLs)phenotype rates of 55.0％and 21.0％,respectively,ESBLs phenotype rate of Klebsiella pneumoniae keep going down.The ratios of carbapenems resistance Klebsiella pneumoniae increased by 5 percentage points compared with the previous monitoring.Carbapenems,moxalactam,sitafloxacin,β-lactam combination agents,fosfomycin trometamol,and amikacin displayed desirable antibacterial activity against Enterbacterales,susceptibal rates were above 75％.In addition,tigacycline,omacycline,colistin and fluoxefin maintained good antibacterial activity against their respective effective bacteria/species,and the bacterial sensitivity rates by more than 80％.Resistance rates of Pseudomonas aeruginosa and Acinetobacter baumannnii to imipenem were 26.3％and 72.1％and multidrug-resistant(MDR)detection rates were 41.1％and 77.3％,extensively drug-resistant(XDR)were 12.0％and 71.8％,respectively.Comparison of drug resistance rates from different wards,ages and specimen sources indicated that the proportion of resistance in Klebsiella pneumoniae and Acinetobacter baumannii isolated from intensive care unit(ICU)were significantly higher than non-ICU.Carbapenem resistance rates of Klebsiella pneumoniae isolated from ICU were more than 35％.Resistance rates of Haemophilus influenzae isolated in children to β-lactam,macrolide,clindamycin and ESBLs detection rate in Klebsiella pneumoniae isolated from children were more than those from adults and the old people,so bacterial resistance in children is an important problem in China.Conclusion ESBLs detection rate of Escherichia coli increased slightly after years of continuous decline.The proportion of carbapenem resistant Pseudomonas aeruginosa was stable,but the resistance rate of Klebsiella pneumoniae and Acinetobacter baumannii to carbapenems was still increased,which should be paid more attention.

Alveolar echinococcosis is a zoonotic parasitic disease caused by an infection with Echinococcosis multilocularis.The liver is the primary organ of alveolar echinococcosis.Alveolar echinococcosis is usually characterized by invasive growth and consequently iscalled"parasitic cancer."Resection of radical lesions is a preferred and effective treatment for hepatic alveolar echinococcosis.End-stage hepatic alveolar echinococcosis often occurs with parasiticcirrhosis,such as secondary biliary cirrhosis,congestive liver cirrhosis or Budd-Chiari syndrome.Few studies have examined hepatic multilocular echinococcosis leading to cirrhosis.This article reviews the aspects of hepatic alveolar echinococcosis involving the invasion of important blood vessels and bile ducts,thereby leading to secondary biliary cirrhosis and congestive liver cirrhosis caused by hepatic alveolar echinococcosis.

Animals ; Cell Proliferation ; Hepatic Stellate Cells ; Liver Cirrhosis/metabolism* ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; Rats

Previously, we discovered that cells contain a 5-hydroxytryptamine (5-HT) degradation system (5DS), which includes 5-HT_2A receptor (5-HT_2AR), 5-HT synthase, and monoamine oxidase A (MAO-A). Among these, 5-HT_2AR has the ability to regulate the expression of 5-HT synthase and MAO-A, and activation of 5DS causes upregulation of these proteins at the same time, resulting in the production of reactive oxygen species (ROS) in the mitochondria. In this study, we investigated the relationship between interstitial pneumonia (IP) and 5DS activation, as well as the therapeutic effect of inhibiting 5DS on IP. Animal models of bleomycin (BLM)-induced IP in mice and radiation (Rad)-induced IP in rats were established, and the models were treated with the 5-HT_2AR antagonist sarpogrelate hydrochloride (SH), 5-HT synthesis inhibitor carbidopa (CDP), and their combination (SH∶CDP = 2∶1). The animal experiments were carried out in accordance with the regulations of the Animal Ethics Committee of China Pharmaceutical University. In the two IP models, immunohistochemistry staining and Western blot analysis showed that the expression of 5-HT synthase was significantly upregulated in all cells of lung tissue, while the expression of 5-HT_2AR and MAO-A was most significantly upregulated in the macrophages. Treatment with SH or CDP significantly reduced pulmonary interstitial thickening, alveolar atrophy with collapse, massive macrophage infiltration and interstitial fibrosis in the two IP models, as measured by HE and Masson staining, and a combination of both almost eliminated the lung tissue lesions. Moreover, treatment with the combination of SH and CDP almost completely eliminated increased ROS and malondialdehyde levels, decreased superoxide dismutase activity, increased tumor necrosis factor-α and interleukin-1β levels, and upregulated nuclear factor-κB phosphorylation and α‍-smooth muscle actin, matrix metalloproteinase-2, and collagen expression. SH and CDP worked together to create a synergistic effect. The findings suggested that the activation of 5DS, as evidenced by increased 5-HT synthesis in all cells of lung tissue and increased 5-HT synthesis and degradation in macrophages, is probably related to the occurrence of IP and that inhibition of 5DS can effectively treat IP.

OBJECTIVES: Ziyin Huatan Recipe (ZYHT), a traditional Chinese medicine comprised of Lilii Bulbus, Pinelliae Rhizoma, and Hedyotis Diffusa, has shown promise in treating gastric cancer (GC). However, its potential mechanism has not yet been clearly addressed. This study aimed to predict targets and molecular mechanisms of ZYHT in treating GC by network pharmacology analysis and to explore the role of ZYHT in GC both in vitro and in vivo. METHODS: Targets and molecular mechanisms of ZYHT were predicted via network pharmacology analysis. The effects of ZYHT on the expression of metastasis-associated targets were further validated by Western blot and quantitative real-time polymerase chain reaction. To explore the specific molecular mechanisms of the effects of ZYHT on migration and invasion, the runt-related transcription factor 3 (RUNX3) gene was knocked out by clustered regularly interspaced short palindromic repeats/Cas9, and lentiviral vectors were transfected into SGC-7901 cells. Then lung metastasis model of GC in nude mice was established to explore the anti-metastasis effect of ZYHT. Western blot and immunohistochemistry were used to explore the impact of ZYHT on the expression of metastasis-related proteins with or without RUNX3 gene. RESULTS: The network pharmacology analysis showed that ZYHT might inhibit focal adhesion, migration, invasion and metastasis of GC. ZYHT inhibited the proliferation, migration and invasion of GC cells in vitro via regulating the expression of metastasis-associated targets. Knocking out RUNX3 almost completely reversed the cell phenotypes (migration and invasion) and protein expression levels elicited by ZYHT. In vivo studies showed that ZYHT inhibited the metastasis of GC cells to the lung and prolonged the survival time of the nude mice. Knocking out RUNX3 partly reversed the metastasis of GC cells to the lung and the protein expression levels elicited by ZYHT. CONCLUSION ZYHT can effectively inhibit the invasion and migration of GC in vitro and in vivo, and its molecular mechanism may relate to the upregulation of RUNX3 expression.
Animals ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; China ; Gene Expression Regulation, Neoplastic ; Mice ; Mice, Nude ; Neoplasm Invasiveness ; Stomach Neoplasms/genetics*

OBJECTIVE: To explore the mechanism by which berberine inhibits ferroptosis of mouse hippocampal neuronal cells (HT22). METHODS: Cultured HT22 cells were pretreated with 30 or 60 μmol/L berberine for 2 h before exposure to 0.5 μmol/L erastin for 8 h, and the cell proliferation, intracellular ferric iron level, changes in intracellular reactive oxygen species (ROS) and cell apoptosis were detected using CCK-8, Fe²⁺ fluorescent probe, fluorescent dye (DAPI) and fluorescent probe (H2DCFH-DA). RT-qPCR and Western blotting were used to detect the mRNA and protein expressions of Nrf2, HO-1 and GPX4 in the cells. We further tested the effects of treatments with 2 μmol/L ML385 (a Nrf2 inhibitor), 60 μmol/L berberine and erastin in the cells to explore the protective mechanism of berberine against erastin-induced ferroptosis in the neuronal cells. RESULTS: Treatment with 0.5 μmol/L erastin significantly lowered the viability of HT22 cells (P < 0.05) and increased the production of ROS, cell apoptosis rate and ferric iron level (P < 0.05). Pretreatment with 30 and 60 μmol/L berberine both significantly increased the vitality of erastin-exposed cells (P < 0.05) and lowered the levels of intracellular ROS and ferric iron content (P < 0.05). RT-qPCR and Western blotting showed that berberine obviously promoted the expressions of Nrf2, HO-1 and GPX4 in the cells (P < 0.05), and treatment with ML385 significantly inhibited the Nrf2-HO-1/GPX4 pathway, increased intracellular ROS and ferric iron contents and mitigated the protective effect of berberine against erastin-induced ferroptosis (P < 0.05). CONCLUSION Berberine can inhibit erastin-induced ferroptosis in HT22 cells possibly by activating the Nrf2-HO-1/ GPX4 pathway.
Animals ; Berberine/pharmacology* ; Ferroptosis ; Fluorescent Dyes ; Hippocampus/metabolism* ; Iron/metabolism* ; Mice ; NF-E2-Related Factor 2/metabolism* ; Piperazines ; Reactive Oxygen Species/metabolism*

OBJECTIVE: To explore the role of 5-hydroxytryptamine (5-HT) in type 2 diabetes mellitus (T2DM)-related hepatic inflammation and fibrosis. METHODS: Male C57BL/6J mice were used to establish T2DM model by high-fat diet feeding combined with intraperitoneal injection of streptozotocin. Then, the mice with hyperglycemia were still fed with high-fat diet for nine weeks, and treated with or without 5-HT_2A receptor (5-HT_2AR) antagonist sarpogrelate hydrochloride (SH) and 5-HT synthesis inhibitor carbidopa (CDP) (alone or in combination). To observe the role of 5-HT in the myofibroblastization of hepa-tic stellate cells (HSCs), human HSCs LX-2 were exposed to high glucose, and were treated with or without SH, CDP or monoamine oxidase A (MAO-A) inhibitor clorgiline (CGL). Hematoxylin & eosin and Masson staining were used to detect the pathological lesions of liver tissue section, immunohistochemistry and Western blot were used to analyze protein expression, biochemical indicators were measured by ELISA or enzyme kits, and levels of intracellular reactive oxygen species (ROS) were detected by fluorescent probe. RESULTS: There were up-regulated expressions of 5-HT_2AR, 5-HT synthases and MAO-A, and elevated levels of 5-HT in the liver of the T2DM mice. In addition to reduction of the hepatic 5-HT levels and MAO-A expression, treatment with SH and CDP could effectively ameliorate liver lesions in the T2DM mice, both of which could ameliorate hepatic injury and steatosis, significantly inhibit the increase of hepatic ROS (H₂O₂) levels to alleviate oxidative stress, and markedly suppress the production of transforming growth factor β1 (TGF-β1) and the development of inflammation and fibrosis in liver. More importantly, there was a synergistic effect between SH and CDP. Studies on LX-2 cells showed that high glucose could induce up-regulation of 5-HT_2AR, 5-HT synthases and MAO-A expression, increase intracellular 5-HT level, increase the production of ROS, and lead to myofibroblastization of LX-2, resulting in the increase of TGF-β1 synthesis and production of inflammatory and fibrosis factors. The effects of high glucose could be significantly inhibited by 5-HT_2AR antagonist SH or be markedly abolished by mitochondrial 5-HT degradation inhibitor CGL. In addition, SH significantly suppressed the up-regulation of 5-HT synthases and MAO-A induced by high glucose in LX-2. CONCLUSION Hyperglycemia-induced myofibroblastization and TGF-β1 production of HSCs, which leads to hepatic inflammation and fibrosis in T2DM mice, is probably due to the up-regulation of 5-HT_2AR expression and increase of 5-HT synthesis and degradation, resulting in the increase of ROS production in mitochondria. Among them, 5-HT_2AR is involved in the regulation of 5-HT synthases and MAO-A expression.
Male ; Mice ; Humans ; Animals ; Hepatic Stellate Cells/pathology* ; Transforming Growth Factor beta1/pharmacology* ; Serotonin/metabolism* ; Reactive Oxygen Species/metabolism* ; Diabetes Mellitus, Type 2/complications* ; Hydrogen Peroxide/metabolism* ; Mice, Inbred C57BL ; Liver Cirrhosis/etiology* ; Hyperglycemia/pathology* ; Monoamine Oxidase/metabolism* ; Inflammation ; Glucose/metabolism* ; Cytidine Diphosphate/pharmacology*