The purpose of this study was to investigate the effect of aqueous extract of Corni Fructus on β-amyloid protein 25-35(Aβ_(25-35))-induced brain injury and neuroinflammation in Alzheimer's disease(AD) mice to provide an experimental basis for the treatment of AD by aqueous extract of Corni Fructus. Sixty C57BL/6J male mice were randomly divided into a sham group, a model group, a positive control group(huperizine A, 0.2 mg·kg~(-1)), a low-dose aqueous extract of Corni Fructus group(1.3 g·kg~(-1)), a medium-dose aqueous extract of Corni Fructus group(2.6 g·kg~(-1)), and a high-dose aqueous extract of Corni Fructus group(5.2 g·kg~(-1)). The AD model was induced by lateral ventricular injection of Aβ_(25-35) in mice except for those in the sham group, and AD model mice were treated with corresponding drugs by gavage for 24 days. The behavioral test was performed one week before animal dissection. Hematoxylin-eosin(HE) staining was performed to observe the morphology of neurons in the hippocampal region. Flow cytometry was used to detect the apoptosis level of primary hippocampal cells in mice. ELISA kits were used to detect the levels of β-amyloid protein 1-42(Aβ_(1-42)) and phosphorylated microtubule-associated protein Tau(p-Tau) in mouse brain tissues. Immunofluorescence and Western blot were used to detect the expression of related proteins in mouse brain tissues. MTT assay was used to detect the effect of compounds in aqueous extract of Corni Fructus on Aβ_(25-35)-induced N9 cell injury. Molecular docking was employed to analyze the interactions of caffeic acid, trans-p-hydroxy cinnamic acid, isolariciresinol-9'-O-β-D-glucopyranoside, esculetin, and(+)-lyoniresinol with β-amyloid precursor protein(APP), interleukin-6(IL-6), and tumor necrosis factor-α(TNF-α). Aqueous extract of Corni Fructus could improve the learning and memory abilities of Aβ_(25-35)-induced mice by increasing the duration of the autonomous activity, the rate of autonomous alternation, the preference coefficient, and the discrimination coefficient, and reduce Aβ_(25-35)-induced brain injury and neuroinflammation in mice by increasing the expression levels of interleukin-10(IL-10) and B-cell lymphoma-2(Bcl-2) in brain tissues, decreasing the expression levels of Aβ_(1-42), p-Tau, IL-6, TNF-α, cysteine aspartate-specific protease 3(caspase-3), cysteine aspartate-specific protease 9(caspase-9), and Bcl-2-associated X protein(Bax), and decreasing the number of activated glial cells in brain tissues. The results of cell experiments showed that esculetin and(+)-lyoniresinol could improve Aβ_(25-35)-induced N9 cell injury. Molecular docking results showed that caffeic acid, trans-p-hydroxy cinnamic acid, isolariciresinol-9'-O-β-D-glucopyranoside, esculetin, and(+)-lyoniresinol had good binding affinity with APP and weak binding affinity with IL-6 and TNF-α. Aqueous extract of Corni Fructus could ameliorate cognitive dysfunction and brain damage in Aβ_(25-35)-induced mice by reducing the number of apoptotic cells and activated glial cells in the brain and decreasing the expression level of inflammatory factors. Caffeic acid, trans-p-hydroxy cinnamic acid, isolariciresinol-9'-O-β-D-glucopyranoside, esculetin, and(+)-lyoniresinol may be the material basis for the anti-AD effect of aqueous extract of Corni Fructus.
Mice ; Male ; Animals ; Alzheimer Disease/drug therapy* ; Amyloid beta-Peptides/metabolism* ; Cornus/metabolism* ; Neuroinflammatory Diseases ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6 ; Aspartic Acid ; Cysteine/therapeutic use* ; Molecular Docking Simulation ; Mice, Inbred C57BL ; Brain Injuries ; Peptide Hydrolases ; Disease Models, Animal ; Mice, Transgenic

Citric Acid Cycle ; Citric Acid ; Malates/metabolism* ; Citrates/metabolism* ; Aspartic Acid/metabolism*

Objective: To explore the relationship between extracellular enzymes activity and virulence of Candida glabrata clinical isolates based on the infection model of Galleria mellonella larvae. Methods: Using experimental research methods, 71 strains of non-repetitive Candida glabrata were collected from Qinghai Provincial People's Hospital from June 2021 to January 2022. Bovine serum protein agar medium, egg yolk agar medium, sheep blood agar medium, Tween-80 agar medium and triglyceride agar medium were used to detect the aspartyl protease activity, phospholipase activity, hemolysis activity, esterase activity and lipase activity of Candida glabrata. Median lethal concentration (LC₅₀) was calculated by using 1.25×10⁸ CFU/ml,2.50×10⁸ CFU/ml,3.75×10⁸ CFU/ml,5.00×10⁸ CFU/ml suspension of Candida glabrata ATCC2001 to infect Galleria mellonella larvae. Histopathological and etiological analysis was performed to determine whether the infection model was successfully established. The clinical isolates of Candida glabrata were configured to infect Galleria mellonella larvae with LC₅₀ concentration to detect the pathogenicity of Galleria mellonella larvae.Spearman test or Pearson test were used to analyze the correlation between the extracellular enzyme activity of Candida glabrata clinical isolates and the pathogenicity of Galleria mellonella larvae. Results: 71 strains of Candida glabrata isolated clinically were detected to have low hemolytic activity after 2 days of culture. Aspartyl protease was detected after 4 days of culture, among which 7 strains (9.86%), 19 strains (26.76%) and 45 strains (63.38%) showed low, medium and high aspartyl protease activity. After 7 days of culture, 71 strains did not detect phospholipase, esterase and lipase activities. Candida glabrata on Galleria mellonella larvae of LC₅₀=2.5×10⁸ CFU/ml Fungal spore were found in the intestinal tissue pathological section of Galleria mellonella larvae in the experimental group, and Candida glabrata was identified by the microbial Mass Spectrometry after culture, while no fungi were found in the pathological section and culture of the control group. Spearman test shows that, there was a linear positive correlation between aspartyl protease activity and the survival rate of Galleria mellonella larvae (r = 0.73, P<0.01), the difference was statistically significant.Pearson test shows that, there was no significant linear relationship between hemolytic activity and survival rate of Galleria mellonella larvae (r = 0.16, P = 0.34), the difference was not statistically significant. Conclusion: The clinical isolates of Candida glabrata in this study had aspartyl protease activity and low hemolytic activity, but no phospholipase, esterase and lipase activity. The activity of aspartyl aspartyl protease of Candida glabrata was positively correlated with the pathogenicity of Galleria mellonella larvae.
Animals ; Sheep ; Larva/microbiology* ; Virulence ; Candida glabrata ; Agar ; Moths/microbiology* ; Esterases ; Aspartic Acid Proteases ; Lipase

This study investigated the potential mechanism of Cordyceps militaris(CM) against non-small cell lung cancer(NSCLC) based on serum untargeted metabolomics. Specifically, Balb/c nude mice were used to generate the human lung cancer A549 xenograft mouse model. The tumor volume, tumor weight, and tumor inhibition rate in mice in the model, cisplatin, Cordyceps(low-, medium-, and high-dose), and CM(low-, medium-, and high-dose) groups were compared to evaluate the influence of CM on lung cancer. Gas chromatography-mass spectrometry(GC-MS) was used for the analysis of mouse serum, SIMCA 13.0 for the compa-rison of metabolic profiles, and MetaboAnalyst 5.0 for the analysis of metabolic pathways. According to the pharmacodynamic data, the tumor volume and tumor weight of mice in high-dose CM group and cisplatin group decreased as compared with those in the model group(P<0.05 or P<0.01). The results of serum metabolomics showed that the metabolic profiles of the model group were significantly different from those of the high-dose CM group, and the content of endogenous metabolites was adjusted to different degrees. A total of 42 differential metabolites and 7 differential metabolic pathways were identified. In conclusion, CM could significantly inhibit the tumor growth of lung cancer xenograft mice. The mechanism is the likelihood that it influences the aminoacyl-tRNA biosynthesis, the metabolism of D-glutamine and D-glutamate, metabolism of alanine, aspartate, and glutamate, metabolism of glyoxylate and dicarboxylic acid, biosynthesis of phenylalanine, tyrosine, and tryptophan, arginine biosynthesis as well as nitrogen metabolism. This study elucidated the underlying mechanism of CM against NSCLC from the point of metabolites. The results would lay a foundation for the anticancer research and clinical application of CM.
Alanine/metabolism* ; Animals ; Arginine/metabolism* ; Aspartic Acid ; Carcinoma, Non-Small-Cell Lung/drug therapy* ; Cisplatin/pharmacology* ; Cordyceps ; Glutamic Acid ; Glutamine ; Glyoxylates/metabolism* ; Humans ; Lung Neoplasms/drug therapy* ; Metabolomics/methods* ; Mice ; Mice, Nude ; Nitrogen/metabolism* ; Phenylalanine/metabolism* ; RNA, Transfer/metabolism* ; Tryptophan/metabolism* ; Tyrosine/metabolism*

OBJECTIVE: To elucidate the mechanism of Lizhong Decoction (LZD) in treating dextran sodium sulfate (DSS)-induced colitis in mice based on metabonomics. METHODS: Thirty-six mice were randomly divided into 6 groups, including normal, model, low- (1.365 g/kg), medium- (4.095 g/kg) and high dose (12.285 g/kg) LZD and salazosulfadimidine (SASP) groups, 6 mice in each group. Colitis model mice were induced by DSS admistration for 7 days, and treated with low, medium and high dose LZD extract and positive drug SASP. Metabolic comparison of DSS-induced colitis and normal mice was investigated by using ultra-performance liquid chromatography coupled to quadrupole time-of-flight mass (UPLC-Q-TOF/MS) combined with Metabolynx™ software. RESULTS: The metabolic profiles of plasma and urine in colitis mice were distinctly ameliorated after LZD treatment (P<0.05). Potential biomarkers (9 in serum and 4 in urine) were screened and tentatively identified. The endogenous metabolites were mainly involved in primary bile acid, sphingolipid, linoleic acid, arachidonic acid, amino acids (alanine, aspartate, and glutamate), butanoate and glycerophospholipid metabolism in plasma, and terpenoid backbone biosynthesis, glycerophospholipid and tryptophan metabolism in urine. After LZD treatment, these markers notably restored to normal levels. CONCLUSIONS The study revealed the underlying mechanism of LZD on amelioration of ulcerative colitis based on metabonomics, which laid a foundation for further exploring the pathological and physiological mechanism, early diagnosis, and corresponding drug development of colitis.
Mice ; Animals ; Colitis, Ulcerative/drug therapy* ; Tryptophan/adverse effects* ; Aspartic Acid ; Dextrans/adverse effects* ; Drugs, Chinese Herbal/adverse effects* ; Colitis/drug therapy* ; Biomarkers/metabolism* ; Amino Acids/adverse effects* ; Glycerophospholipids/therapeutic use* ; Sphingolipids/adverse effects* ; Bile Acids and Salts/adverse effects* ; Glutamates/adverse effects* ; Alanine/adverse effects* ; Arachidonic Acids/adverse effects* ; Linoleic Acids/adverse effects* ; Terpenes

OBJECTIVE: To clarify the functional effects of differential expression of ring finger and tryptophan-aspartic acid 2 (RFWD2) on dendritic development and formation of dendritic spines in cerebral cortex neurons of mice. METHODS: Immunofluorescent staining was used to identify the location and global expression profile of RFWD2 in mouse brain and determine the co-localization of RFWD2 with the synaptic proteins in the cortical neurons. We also examined the effects of RFWD2 over-expression (RFWD2-Myc) and RFWD2 knockdown (RFWD2-shRNA) on dendritic development, dendritic spine formation and synaptic function in cultured cortical neurons. RESULTS: RFWD2 is highly expressed in the cerebral cortex and hippocampus of mice, and its expression level was positively correlated with the development of cerebral cortex neurons and dendrites. RFWD2 expression was detected on the presynaptic membrane and postsynaptic membrane of the neurons, and its expression levels were positively correlated with the length, number of branches and complexity of the dendrites. In cultured cortical neurons, RFWD2 overexpression significantly lowered the expressions of the synaptic proteins synaptophysin (P < 0.01) and postsynapic density protein 95 (P < 0.01), while RFWD2 knockdown significantly increased their expressions (both P < 0.05). Compared with the control and RFWD2-overexpressing cells, the neurons with RFWD2 knockdown showed significantly reduced number of dendritic spines (both P < 0.05). CONCLUSION RFWD2 can regulate the expression of the synaptic proteins, the development of the dendrites, the formation of the dendritic spines and synaptic function in mouse cerebral cortex neurons through ubiquitination of Pea3 family members and c-Jun, which may serve as potential treatment targets for neurological diseases.
Animals ; Aspartic Acid/metabolism* ; Cerebral Cortex ; Dendritic Spines/metabolism* ; Mice ; Neurons/metabolism* ; Synapses ; Tryptophan/metabolism*

OBJECTIVE: To explore the amino acid metabolomics characteristics of myeloid-derived suppressor cells (MDSCs) in mice with sepsis induced by the cecal ligation and puncture (CLP). METHODS: The sepsis mouse model was prepared by CLP, and the mice were randomly divided into a sham operation group (sham group, n = 10) and a CLP model group (n = 10). On the 7th day after the operation, 5 mice were randomly selected from the surviving mice in each group, and the bone marrow MDSCs of the mice were isolated. Bone marrow MDSCs were separated to measure the oxygen consumption rate (OCR) by using Agilent Seahorse XF technology and to detect the contents of intracellular amino acids and oligopeptides through ultra-performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) technology. Different metabolites and potential biomarkers were analyzed by univariate statistical analysis and multivariate statistical analysis. The major metabolic pathways were enriched using the small molecular pathway database (SMPDB). RESULTS: The proportion of MDSCs in the bone marrow of CLP group mice (75.53% ± 6.02%) was significantly greater than that of the sham group (43.15%± 7.42%, t = 7.582, P < 0.001), and the basal respiratory rate [(50.03±1.20) pmol/min], maximum respiration rate [(78.07±2.57) pmol/min] and adenosine triphosphate (ATP) production [(25.30±1.21) pmol/min] of MDSCs in the bone marrow of CLP group mice were significantly greater than the basal respiration rate [(34.53±0.96) pmol/min, (t = 17.41, P < 0.001)], maximum respiration rate [(42.57±1.87) pmol/min, (t = 19.33, P < 0.001)], and ATP production [(12.63±0.96) pmol/min, (t = 14.18, P < 0.001)] of sham group. Leucine, threonine, glycine, etc. were potential biomarkers of septic MDSCs (all P < 0.05). The increased amino acids were mainly enriched in metabolic pathways, such as malate-aspartate shuttle, ammonia recovery, alanine metabolism, glutathione metabolism, phenylalanine and tyrosine metabolism, urea cycle, glycine and serine metabolism, β-alanine metabolism, glutamate metabolism, arginine and proline metabolism. CONCLUSION The enhanced mitochondrial oxidative phosphorylation, malate-aspartate shuttle and alanine metabolism in MDSCs of CLP mice may provide raw materials for mitochondrial aerobic respiration, thereby promoting the immunosuppressive function of MDSCs. Blocking the above metabolic pathways may reduce the risk of secondary infection in sepsis and improve the prognosis.
Adenosine Triphosphate/metabolism* ; Alanine/metabolism* ; Animals ; Aspartic Acid/metabolism* ; Biomarkers/metabolism* ; Chromatography, Liquid ; Glycine/metabolism* ; Malates/metabolism* ; Mice ; Myeloid-Derived Suppressor Cells/metabolism* ; Sepsis/complications* ; Tandem Mass Spectrometry

In this paper,metabolomics and network pharmacology were used to investigate the bioactive components of Harrisonia perforata and their possible mechanisms of action. Metabolites in the flowers,fruits,branches,leaves and stalks of H. perforata were analyzed by ultra-high performance liquid chromatography-quadrupole-time-of-flight mass spectrometry. Meanwhile,multiple statistical analysis methods including principal component analysis( PCA) and orthogonal partial least squares discriminant analysis( OPLS-DA)were applied to screen and identify differential compounds. With metabolomics method,9 differential compounds were preliminarily identified from leaves and other non-traditional medicinal parts. Subsequently,these compounds were explored by using network pharmacology. With gastrointestinal absorption and drug-likeness as limiting conditions,they were imported into the Swiss ADME,from which 7 compounds with potential medicinal activity were obtained. Then,their targets were predicted by PharmMapper,with Human Protein Targets Only and Normalized Fit Score>0. 9 set as limiting conditions,and 60 standardized potential targets were identified with Uniprot. KEGG( Kyoto encyclopedia of genes and genomes) pathway data was obtained using metascape and the " potential active ingredients-target-pathway" network was constructed with Cytoscape 3. 7. 2. The enrichment analysis of KEGG demonstrated that the 60 targets were enriched in 78 signaling pathways( min overlap: 3,P value cutoff: 0. 01,min enrichment: 1. 5),many of which are related to anti-bacteria,anti-inflammation and anti-virus,such as IL-17 signaling pathway,RIG-I-like receptor signaling pathway and NOD-like receptor signaling pathway. Finally,depending on the clinical activity of H. perforata,the relevant signaling pathways were analyzed through experimental data and literature. Dehydroconiferyl alcohol was reported to have the anti-inflammatory effect and perforamone D to possess the antimycobacterial activity. The KEGG pathway enrichment analysis showed that dehydroconiferyl alcohol could act on the Alzheimer's disease( AD) signaling pathway by targeting CDK5 R1 and BACE1. ACh E inhibitor is the most promising drug to treat AD,while dehydroconiferyl alcohol has been proved to inhibit ACh E according to literature. The experimental results revealed that the extract of leaves of H. perforata can effectively inhibit the growth of Staphylococcus aureus. These are consistent with the enrichment analysis results of KEGG. This study explored the bioactive components and pharmacodynamics of the leaves of the H. perforata,laying a theoretical foundation for its in-depth development and rational application.
Amyloid Precursor Protein Secretases ; Aspartic Acid Endopeptidases ; Drugs, Chinese Herbal/pharmacology* ; Humans ; Metabolomics ; Simaroubaceae

OBJECTIVES: To explore the metabolite characteristics in medial prefrontal cortex (mPFC) by METHODS: A total of 46 patients with the first-episode schizophrenia (FES), 49 people with clinical high risk (CHR), 61 people with genetic high risk (GHR), and 58 healthy controls (HC) were enrolled. The levels of N-acetylaspartylglutamate+N-acetylaspartate (tNAA), choline-containing compounds (Cho) and myo-inositol (MI), glutamate+glutamine (Glx) in medial prefrontal cortex were measured by single-voxel RESULTS: There were significant differences in Glx, tNAA, and MI concentrations among 4 groups (all CONCLUSIONS The decreased levels of MI and Glx in the FES patients suggest that there may be glial functional damage and glutamatergic transmitter dysfunction in the early stage of the disease. The compensatory increase of metabolites may be a protective factor for schizophrenia in the genetic individuals.
Aspartic Acid ; Glutamic Acid ; Glutamine ; Humans ; Magnetic Resonance Imaging ; Magnetic Resonance Spectroscopy ; Proton Magnetic Resonance Spectroscopy ; Schizophrenia

Physalin B (PB), one of the major active steroidal constituents of Solanaceae Physalis plants, has a wide variety of biological activities. We found that PB significantly down-regulated β-amyloid (Aβ) secretion in N2a/APPsw cells. However, the underlying mechanisms are not well understood. In the current study, we investigated the changes in key enzymes involved in β-amyloid precursor protein (APP) metabolism and other APP metabolites by treating N2a/APPsw cells with PB at different concentrations. The results indicated that PB reduced Aβ secretion, which was caused by down-regulation of β-secretase (BACE1) expression, as indicated at both the protein and mRNA levels. Further research revealed that PB regulated BACE1 expression by inducing the activation of forkhead box O1 (FoxO1) and inhibiting the phosphorylation of signal transducer and activator of transcription 3 (STAT3). In addition, the effect of PB on BACE1 expression and Aβ secretion was reversed by treatment with FoxO1 siRNA and STAT3 antagonist S3I-201. In conclusion, these data demonstrated that PB can effectively down-regulate the expression of BACE1 to reduce Aβsecretion by activating the expression of FoxO1 and inhibiting the phosphorylation of STAT3.
Alzheimer Disease ; Amyloid Precursor Protein Secretases/metabolism* ; Amyloid beta-Peptides/metabolism* ; Aspartic Acid Endopeptidases/metabolism* ; Down-Regulation ; Forkhead Box Protein O1/genetics* ; Humans ; Phosphorylation ; STAT3 Transcription Factor/metabolism* ; Secosteroids