OBJECTIVES: To explore the mechanism of Shuangshu Decoction (SSD) for inhibiting growth of gastric cancer xenografts in nude mice. METHODS: Network pharmacology analysis was conducted to identify the common targets of SSD and gastric cancer cell ferroptosis, and bioinformatics analysis and molecular docking were used to validate the core targets. In the cell experiment, AGS cells were treated with SSD-medicated serum, Fer-1 (a ferroptosis inhibitor), or both, and the changes in cell viability, ferroptosis markers (ROS, Fe²⁺ and GSH), expressions of P53, SLC7A11 and GPX4, and mitochondrial morphology were examined. In a nude mouse model bearing gastric cancer xenografts, the effects of gavage with SSD, intraperitoneal injection of Fer-1, or their combination on tumor volume/weight, histopathology, and expressions of P53, SLC7A11 and GPX4 levels were evaluated. RESULTS: The active components in SSD (quercetin and wogonin) showed strong binding affinities to P53. In AGS cells, SSD treatment dose-dependently inhibited cell proliferation, increased ROS and Fe²⁺ levels, upregulated P53 expression, and downregulated the expressions of SLC7A11 and GPX4, but these effects were effectively attenuated by Fer-1 treatment. SSD also induced mitochondrial shrinkage and increased the membrane density, which were alleviated by Fer-1. In the tumor-bearing mouse models, gavage with SSD significantly reduced tumor size and weight, caused tumor cell necrosis, upregulated P53 and downregulated SLC7A11 and GPX4 expression in the tumor tissue, and these effects were obviously mitigated by Fer-1 treatment. CONCLUSIONS SSD inhibits gastric cancer growth in nude mice by inducing cell ferroptosis via the P53/SLC7A11/GPX4 axis.
Ferroptosis/drug effects* ; Animals ; Stomach Neoplasms/metabolism* ; Tumor Suppressor Protein p53/metabolism* ; Mice, Nude ; Phospholipid Hydroperoxide Glutathione Peroxidase ; Drugs, Chinese Herbal/pharmacology* ; Humans ; Amino Acid Transport System y+/metabolism* ; Mice ; Cell Line, Tumor ; Cell Proliferation/drug effects* ; Xenograft Model Antitumor Assays

In general, bioassay-guided fractionation and isolation of bioactive constituents from botanical materials frequently ended up with the reward of a single compound. However, botanical materials typically exert their therapeutic actions through multi-pathway effects due to the intrinsic complex nature of chemical constituents. In addition, the content of bioactive compounds in botanical materials is largely dependent on humidity, temperature, soil, especially geographical origins, from which rapid and accurate identification of plant materials is pressingly needed. These long-standing obstacles collectively impede the deep exploitation and application of these versatile natural sources. To address the challenges, a new paradigm integrating biogravimetric analyses and machine learning-driven origin classification (BAMLOC) was developed. The biogravimetric analyses are based on absolute qHNMR quantification and in vivo zebrafish model-assisted activity index calculation, by which bioactive substance groups jointly responsible for the bioactivities in all fractions are pinpointed before any isolation effort. To differentiate origin-different botanical materials varying in the content of bioactive substance groups, principal component analysis, linear discriminant analysis, and hierarchical cluster analysis in conjunction with supervised support vector machine are employed to classify and predict production areas based on the detection of volatile organic compounds by E-nose and GC-MS. Expanding BAMLOC to Codonopsis Radix enables the identification of polyacetylenes and pyrrolidine alkaloids as the bioactive substance group for immune restoration effect and accurately determines the origins of plants. This study advances the toolbox for the discovery of bioactive compounds from complex mixtures and lays a more definitive foundation for the in-depth utilization of botanical materials.

Objective:To analyze the expression of FAT1 gene in pancreatic adenocarcinoma and its relationship with clinicopathological features, prognosis, and immunotherapy for pancreatic adenocarcinoma.Methods:(1) Bioinformatics analysis: based on FAT1 mRNA expression and clinical data of 179 cases of pancreatic adenocarcinoma in the TCGA database, and FAT1 mRNA expression data of 328 cases of normal pancreatic tissues in the GTEx database. We analyzed the differences in FAT1 mRNA expression in pancreatic adenocarcinoma and normal pancreatic tissues and the relationship between FAT1 mRNA expression and the degree of differentiation, clinical stage, prognosis, immune cell infiltration, and immune checkpoint-associated genes in pancreatic adenocarcinoma. FAT1-related differentially expressed genes were analyzed by applying Limma 3.40.2 software package, and GO and KEGG enrichment analysis was performed on the differentially expressed genes. Immunohistochemical (IHC) of FAT1 in pancreatic adenocarcinoma and normal pancreatic tissues was analyzed by HPA database. (2) Validation of own tissue samples: tissue samples and clinical and prognostic data of 192 patients with pancreatic ductal adenocarcinoma admitted to Zhejiang Cancer Hospital from March 8, 2010 to September 30, 2020 were collected. IHC was performed on the tissue samples to verify the protein expression of FAT1 in pancreatic adenocarcinoma and its relationship with immune-related proteins, the degree of differentiation of pancreatic adenocarcinoma, clinical staging, and prognosis.Results:(1) Bioinformatics analysis: the FAT1 mRNA expression of 179 pancreatic adenocarcinoma tissues from the TCGA database was 5.55±1.04, which was higher than that of 328 normal pancreatic tissues with FAT1 mRNA from the GTEx database (2.95±0.53, P＜0.001). FAT1-specific IHC images showed that FAT1 expression was generally high in pancreatic adenocarcinoma tissues, and FAT1 expression shifted from the cell membrane to the cytoplasm. The FAT1 mRNA expression in the highly differentiated group (31 cases), the moderately differentiated group (96 cases), and the lowly differentiated group (52 cases) were 4.99±1.46, 5.51±0.80, and 5.68±1.08, the expression of pancreatic adenocarcinoma tissues were all higher than that of normal pancreatic tissues (all P＜0.001), and the FAT1 mRNA expression of the moderately differentiated group and the poorly differentiated group were all higher than that of the highly differentiated group (all P＜0.001). The median progression-free survival time (PFS) and median overall survival time (OS) of the 90 patients in the FAT1 mRNA low-expression group were 16.5 and 24 months, respectively, which were longer than those of the 89 patients in the FAT1 mRNA high-expression group (median PFS and OS were 13 and 18 months, respectively; P-values were 0.011 and 0.005, respectively). Multifactorial Cox regression analysis showed that FAT1 mRNA expression level was an independent influencing factor for OS in pancreatic adenocarcinoma patients ( HR=1.47, 95% CI: 1.09-1.99). Correlation analysis showed that FAT1 mRNA expression in pancreatic adenocarcinoma was positively correlated with B-cell infiltration, CD8+ T-cell infiltration, neutrophil infiltration, macrophage infiltration, and myeloid dendritic cell infiltration ( ρ=0.27, P＜0.001; ρ=0.28, P＜0.001; ρ=0.32, P＜0.001; ρ=0.21, P=0.004; ρ=0.32, P＜0.001), and also positively correlated with mRNA expression of CD274, HAVCR2, and PDCD1LG2 ( r=0.327, P＜0.001; r=0.231, P=0.002; r=0.258, P＜0.001). GO and KEGG enrichment analyses showed that FAT1 mRNA expression levels were associated with activation of the Wnt signaling pathway ( P=0.029), the PI3K/Akt pathway ( P<0.001), and other tumor microenvironment-related pathways. (2) Validation of own tissue samples: among 192 pancreatic adenocarcinoma tissues, FAT1 was highly expressed in 58 cases (30.21%), and the proportion of FAT1-expressing positive tumor cells was positively correlated with the combined positive score of PD-L1 and the number of CD3+ T-cells infiltration ( r=0.154, P=0.032; r=0.287, P＜0.001), and the protein expression of FAT1 had no correlation with the differentiation degree of pancreatic adenocarcinoma ( ρ=0.082, P=0.254). The median OS of 58 patients in the FAT1 high-expression group and 134 patients in the FAT1 low-expression group were 18.89 and 25.84 months, respectively, and the difference was not statistically significant (χ2=1.93, P=0.165). Conclusion:FAT1 gene is highly expressed in pancreatic adenocarcinoma tissues, may play an oncogenic role in pancreatic adenocarcinoma, may be an adverse influence on overall survival and progression-free survival of patients; FAT1 gene may be involved in multiple immune-related pathways and promote tumor immune escape.

Background Atmospheric fine particulate matter (PM_2.5) can disrupt the metabolic homeostasis of the liver and accelerate the progression of liver diseases, but there are few studies on the effects of sub-chronic PM_2.5 exposure on the liver metabolome. Objectives To investigate the effects of sub-chronic exposure to concentrated PM_2.5 on hepatic metabolomics in mice by liquid chromatography-mass spectrometry (LC-MS), and to identify potentially affected metabolites and metabolic pathways. Methods Twelve male C57BL/6J (6 weeks old) mice were randomly divided into two groups: a concentrated PM_2.5 exposure group and a clean air exposure group. The mice were exposed to concentrated PM_2.5 using the "Shanghai Meteorological and Environmental Animal Exposure System" at Fudan University. The exposure duration was 8 h per day, 6 d per week, for a total of 8 weeks. The mice's liver tissues were collected 24 h after the completion of exposure. LC-MS was performed to assess changes in the hepatic metabolome. Orthogonal partial least squares discriminant analysis and t-test were employed to identify differentially regulated metabolites between the two groups under the conditions of variable important in projection (VIP)≥1.0 and P<0.05. Metabolic pathway enrichment analysis was performed using MetaboAnalyst 5.0 software and the Kyoto Encyclopedia of Genes and Genomes (KEGG). Results A total of 297 differentially regulated metabolites were identified between the concentrated PM_2.5 exposure group and the clean air group. Among these metabolites, 142 were upregulated and 155 were downregulated. A total of 38 metabolic pathways were altered, with 7 pathways showing significant perturbation (P<0.05). These pathways involved amino acid metabolism, glucose metabolism, nucleotide metabolism, as well as cofactor and vitamin metabolism. The 7 significant metabolic pathways were pantothenic acid and coenzyme A biosynthesis; purine metabolism; amino sugar and nucleotide sugar metabolism; arginine biosynthesis; alanine, aspartate and glutamate metabolism; aminoacyl-tRNA biosynthesis; and fructose and mannose metabolism. Conclusion The results from metabolomics analysis suggest that sub-chronic exposure to PM_2.5 may disrupt hepatic energy metabolism and induce oxidative stress damage. Aspartic acid, succinic acid, ornithine, fumaric acid, as well as purine and xanthine derivatives, were identified as potential early biomarkers of hepatic response to sub-chronic PM_2.5 exposure.

Objective:To analyze the expression of FAT1 gene in pancreatic adenocarcinoma and its relationship with clinicopathological features, prognosis, and immunotherapy for pancreatic adenocarcinoma.Methods:(1) Bioinformatics analysis: based on FAT1 mRNA expression and clinical data of 179 cases of pancreatic adenocarcinoma in the TCGA database, and FAT1 mRNA expression data of 328 cases of normal pancreatic tissues in the GTEx database. We analyzed the differences in FAT1 mRNA expression in pancreatic adenocarcinoma and normal pancreatic tissues and the relationship between FAT1 mRNA expression and the degree of differentiation, clinical stage, prognosis, immune cell infiltration, and immune checkpoint-associated genes in pancreatic adenocarcinoma. FAT1-related differentially expressed genes were analyzed by applying Limma 3.40.2 software package, and GO and KEGG enrichment analysis was performed on the differentially expressed genes. Immunohistochemical (IHC) of FAT1 in pancreatic adenocarcinoma and normal pancreatic tissues was analyzed by HPA database. (2) Validation of own tissue samples: tissue samples and clinical and prognostic data of 192 patients with pancreatic ductal adenocarcinoma admitted to Zhejiang Cancer Hospital from March 8, 2010 to September 30, 2020 were collected. IHC was performed on the tissue samples to verify the protein expression of FAT1 in pancreatic adenocarcinoma and its relationship with immune-related proteins, the degree of differentiation of pancreatic adenocarcinoma, clinical staging, and prognosis.Results:(1) Bioinformatics analysis: the FAT1 mRNA expression of 179 pancreatic adenocarcinoma tissues from the TCGA database was 5.55±1.04, which was higher than that of 328 normal pancreatic tissues with FAT1 mRNA from the GTEx database (2.95±0.53, P＜0.001). FAT1-specific IHC images showed that FAT1 expression was generally high in pancreatic adenocarcinoma tissues, and FAT1 expression shifted from the cell membrane to the cytoplasm. The FAT1 mRNA expression in the highly differentiated group (31 cases), the moderately differentiated group (96 cases), and the lowly differentiated group (52 cases) were 4.99±1.46, 5.51±0.80, and 5.68±1.08, the expression of pancreatic adenocarcinoma tissues were all higher than that of normal pancreatic tissues (all P＜0.001), and the FAT1 mRNA expression of the moderately differentiated group and the poorly differentiated group were all higher than that of the highly differentiated group (all P＜0.001). The median progression-free survival time (PFS) and median overall survival time (OS) of the 90 patients in the FAT1 mRNA low-expression group were 16.5 and 24 months, respectively, which were longer than those of the 89 patients in the FAT1 mRNA high-expression group (median PFS and OS were 13 and 18 months, respectively; P-values were 0.011 and 0.005, respectively). Multifactorial Cox regression analysis showed that FAT1 mRNA expression level was an independent influencing factor for OS in pancreatic adenocarcinoma patients ( HR=1.47, 95% CI: 1.09-1.99). Correlation analysis showed that FAT1 mRNA expression in pancreatic adenocarcinoma was positively correlated with B-cell infiltration, CD8+ T-cell infiltration, neutrophil infiltration, macrophage infiltration, and myeloid dendritic cell infiltration ( ρ=0.27, P＜0.001; ρ=0.28, P＜0.001; ρ=0.32, P＜0.001; ρ=0.21, P=0.004; ρ=0.32, P＜0.001), and also positively correlated with mRNA expression of CD274, HAVCR2, and PDCD1LG2 ( r=0.327, P＜0.001; r=0.231, P=0.002; r=0.258, P＜0.001). GO and KEGG enrichment analyses showed that FAT1 mRNA expression levels were associated with activation of the Wnt signaling pathway ( P=0.029), the PI3K/Akt pathway ( P<0.001), and other tumor microenvironment-related pathways. (2) Validation of own tissue samples: among 192 pancreatic adenocarcinoma tissues, FAT1 was highly expressed in 58 cases (30.21%), and the proportion of FAT1-expressing positive tumor cells was positively correlated with the combined positive score of PD-L1 and the number of CD3+ T-cells infiltration ( r=0.154, P=0.032; r=0.287, P＜0.001), and the protein expression of FAT1 had no correlation with the differentiation degree of pancreatic adenocarcinoma ( ρ=0.082, P=0.254). The median OS of 58 patients in the FAT1 high-expression group and 134 patients in the FAT1 low-expression group were 18.89 and 25.84 months, respectively, and the difference was not statistically significant (χ2=1.93, P=0.165). Conclusion:FAT1 gene is highly expressed in pancreatic adenocarcinoma tissues, may play an oncogenic role in pancreatic adenocarcinoma, may be an adverse influence on overall survival and progression-free survival of patients; FAT1 gene may be involved in multiple immune-related pathways and promote tumor immune escape.

Objective : To study the effect of lipopolysaccharide ( LPS) -induced immune system activation on syn- aptic structure and DISC1. Methods : 40 rats were divided into 4 groups，which were PBS group，PBS + MK-801 group，LPS group and LPS + MK-801 group．The rats in LPS group and LPS + MK-801 group were administrated with LPS solution (0. 2 mg / kg，i．p．BIW) for 6 times in total， the rats in PBS group and PBS + MK801 group were administrated with PBS．At the third time when the rats were administrated with LPS solution，the rats in PBS + MK-801 group and LPS + MK-801 group were administrated with MK-801 solution (0. 5 mg / kg，i．p．BIW) for a total of 4 times，the rats in PBS group and LPS group were adminis- trated with saline．The head shaking and the open field tests were then evaluated in the rats．Immediately，the rats were sacrificed，and their brain were saved for testing. RT-qPCR , Golgi staining，immunohistochemistry and ELISA kits were used to detect DISC1 gene expression，dendritic spine status，microglia activation and inflammatory cytokine content in cortex，respectively. gene expression in the cortex of rats with schizophrenia． Results : Compared with the PBS group，the expression of DISC1 gene of the rats in the PBS + MK-801 group increased (P ＜0. 01) ，the score of head shaking increased (P ＜0. 01) ，and the distance of the open field increased (P ＜0. 000 1) ; As well as the LPS + MK-801 group，compared with the LPS group，the expression of DISC1 gene increased (P＜0. 000 1) ，the head shaking score and the distance of the open field increased (P＜0. 000 1) . In addition，compared with the PBS + MK-801 group，the expression of DISC1 gene， the shaking head score and the open field distance of the LPS + MK-801 group further improved (P ＜0. 000 1) . Furthermore，compared with PBS group and PBS + MK-801 group，the content of iba1 in the cortex of LPS group and LPS + MK-801 group significantly increased (P ＜0. 05 ) ，the content of inflammatory cytokines significantly increased (P＜0. 05) ，and the length and density of dendritic spines significantly decreased (P＜0. 05) ． Conclusion The activation of the immune system induced by LPS can contribute to the susceptibility of schizophrenia model， and the possible mechanism is that it cooperates with MK-801 to increase the expression of DISC1 gene，which ulti- mately leads to abnormal mental behavior in rats.

For diabetes patients, especially elderly diabetes patients, insulin is widely used as an important treatment to control hyperglycemia.However, since a high percentage of elderly diabetes patients use high doses of insulin, it is common to incur issues such as increased insulin resistance and inappropriate treatment.Exogenous insulin-induced autoantibody production is associated with severe insulin resistance and refractory hyperglycemia and hypoglycemia and is referred to as exogenous insulin antibody syndrome.With hypoglycemia, high insulin levels may be inconsistent with C-peptide levels.Increasing the dose of insulin to control blood glucose may be counterproductive.It is necessary to quickly and accurately make a diagnosis and make personalized adjustments to the glucose-lowering regimen to avoid serious consequences.

As a highly malignant gastrointestinal tumor, pancreatic cancer is highly invasive and metastatic, which leads to the low overall survival rate of patients with pancreatic cancer. Studies have shown that long non-coding RNA (lncRNA) is involved in the development, progression, invasion, and metastasis of pancreatic cancer through epigenetic, transcriptional or post-transcriptional regulation. Dysregulated expression of lncRNA is observed in pancreatic cancer and induces epithelial mesenchymal transition (EMT) through specific regulatory mechanisms, thereby causing the changes in the biological behavior of tumor cells. This article reviews the mechanisms of lncRNA in promoting EMT, regulating tumor biological function as competing endogenous RNA, and affecting the development, invasion, and metastasis of pancreatic cancer via multiple pathways by regulating the ferroptosis, autophagy, and exosome of tumor cells, in order to provide a theoretical basis and new targets for the early diagnosis and treatment of pancreatic cancer.

Pancreatic cancer stroma plays a critical role in tumor progression, invasion, metastasis and resistance.Targeting tumor cell alone could not meet the demand for prolonging patients'' survival.Growing studies have laid emphasis on developing combined regimens between targeting pancreatic cancer stroma and chemotherapy, radiotherapy and immunotherapy.We are faced with some new opportunities in spite of the great challenges brought to the research and development of targeting drugs owing to the complicated stroma components, crosstalking signal pathways and abnormal angiogenesis of pancreatic cancer.In this article, recent advances in therapeutic strategies of targeting pancreatic cancer stroma are reviewed and analyzed from the aspects of extracellular matrix (ECM), cancer associated fibroblasts (CAFs) and vessels, in the hope of providing some novel ideas for targeting therapy against pancreatic cancer.

Objective: To explore the effect of hyperbaric oxygen preconditioning with different frequency on the survival rate of flap and ischemia-reperfusion injury in rats after transplantation, and to explore the best preconditioning conditions to improve the survival rate of rat flaps after transplantation. Methods: Thirty-six Sprague Dawley rats were randomly divided into four groups according to the random number table method, 9 groups in each group.Four groups of rats were pretreated with hyperbaric oxygen pretreatment for 0, 2, 4, and 6 days before the operation, control group, pretreatment 2 d group, pretreatment 4 d group, and pretreatment 6 d group. Taking the midline of the back of the rat as the axis, an ultra-long random flap with a pedicle at the tail end and about 1 cm from the superior iliac spine was designed and cut to a size of 10.0 cm×2.5 cm. The survival of the flaps in each group was observed and the final survival area and survival rate of the flaps were measured on the 7th day after surgery. On the 7th day after operation, the tissue was taken at a distance of 5 cm from the pedicle, and the histopathology was observed; The content of superoxide dismutase (SOD) and malondialdehyde (MDA) in flap tissue was detected by immunohistochemistry, and the expression rate of positive cells in each group was calculated. Immunofluorescence was used to detect the expression of interleukin-6 (IL-6) in the flap tissue. Results: On the 7th day after the operation, the survival area and survival rate of the transplanted flaps in the hyperbaric oxygen pretreatment group were significantly higher than those in the control group (P<0.05), and the pretreatment 4 d and 6 d groups were significantly higher than the pretreated 2 d group (P<0.05), but there was no significant difference between the pretreated 4 d group and the 6 d group (P=0.095). Pathological observation on the 7th day after operation showed that there was some necrosis in the control group, the vascular cells in the pretreated 2 d group had more vascular structures, and more neovascularization was observed in the pretreated 4 d group. The inflammatory cells were the least in the 6 d pretreatment group, and the neovascularization was the same as the pretreatment 4 d group. The absorbance A value of SOD in the control group was 0.009 7±0.000 3, and the positive expression rate was 20%, which was significantly lower than that in the hyperbaric oxygen pretreatment group. The difference was statistically significant (P<0.05). However, the absorbance A value of MDA in the control group was 0.055 1±0.003 0, and the positive expression rate was 55%, which was significantly higher than that in the hyperbaric oxygen pretreatment group. The difference was statistically significant (P<0.05). Among them, the SOD absorbance A value of the pretreated 2 d group was 0.023 8±0.003 0, and the positive expression rate was 30%, which was lower than the pretreatment 4 d group (absorbance A value 0.046 9±0.003 0, positive expression rate 35%) and 6 d group (absorbance A value 0.047 2±0.003 6, positive expression rate 40%), The MDA absorbance A value of the pretreated 2 d group was 0.037 2±0.003 2, and the positive expression rate was 30%, which was higher than the pretreatment 4 d group (absorbance A value 0.014 7±0.002 4, positive expression rate 5%) and 6 d group (absorbance A value 0.017 0±0.001 8, positive expression rate 10%), the difference was statistically significant (P<0.05). However, there was no significant difference in the expression of SOD and MDA between the pretreated 4 d group and the pretreated 6 d group (P>0.05). The expression of IL-6 in the hyperbaric oxygen pretreatment group was significantly lower than that in the control group (absorbance A value 44.937 0±0.594 0), the difference was statistically significant (P<0.05). The absorbance A value in the pretreated 2 d group was 41.698 0±0.724 0, which was significantly higher than the pretreatment 4 d group (absorbance A value 34.049 0±0.323 0) and 6 d group (absorbance A value 33.524 0±0.639 0). The difference was statistically significant (P<0.05). There was no significant difference in the expression of the pretreated 4 d group compared with the 6 d group (P=0.068). Conclusions Hyperbaric oxygen preconditioning can significantly promote the survival of rat flaps after transplantation. Preoperative hyperbaric oxygen preconditioning once daily for 4 consecutive days, can enhance the tolerance of flap tissue to ischemia and anoxia and reduce tissue ischemia-reperfusion injury.