Network pharmacology and molecular docking reveal the mechanism of resveratrol in oral squamous cell carcinoma treatment / 口腔疾病防治

Objective@#To explore the molecular mechanism of resveratrol (RES) in the treatment of oral squamous cell carcinoma (OSCC) through the use of biological information methods such as network pharmacology and molecular docking and to provide a theoretical reference for the clinical application of RES in the treatment of OSCC.@*Methods@#The Swiss Target Prediction(http://www.swisstargetprediction.ch), SEA (http://sea.bkslab.org)database, and Pharm mapper database(http://lilab-ecust.cn) were used to retrieve RES-related targets, and the DISGENET (www.disgenet.org), OMIM (https://omim.org) and GeneCards (https://www.genecards.org) databases were used to screen OSCC disease targets. The intersection of drugs and disease targets was determined, and Cytoscape 3.7.2 software was used to construct a "drug-diseasetarget pathway" network. The Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database was used to construct a target protein interaction network, and the DAVID database was used for enrichment analysis of key proteins. Finally, molecular docking validation of key proteins was performed using AutoDock and PyMOL. The enrichment analysis and molecular docking results were integrated to predict the possible molecular mechanisms of RES treatment in OSCC; western blot was used to determine the effect of resveratrol at different concentrations (50, 100) μmol/L on the expression of Src tyrosine kinase (SRC), epidermal growth factor receptor (EGFR), estrogen receptor gene 1 (ESR1), and phosphatidylinositol 3 kinase/protein kinase B (PI3K/AKT) signaling pathway proteins in OSCC HSC-3 cells.@*Results@#A total of 243 targets of RES drugs and 6 094 targets of OSCC were identified. A total of 116 potential common targets were obtained by intersecting drugs with disease targets. These potential targets mainly participate in biological processes such as in vivo protein self-phosphorylation, peptide tyrosine phosphorylation, transmembrane receptor protein tyrosine kinase signaling pathway, and positive regulation of RNA polymerase Ⅱ promoter transcription, and they interfere with the PI3K/AKT signaling pathway to exert anti-OSCC effects. The docking results of resveratrol with OSCC molecules indicated that key targets, such as EGFR, ESR1, and SRC, have good binding activity. The results of cell-based experiments showed that resveratrol inhibited the protein expression of SRC, EGFR, ESR1, p-PI3K, and p-AKT in HSC-3 cells in a dose-dependent manner.@*Conclusion@#RES can inhibit the expression of its targets EGFR, ESR1, SRC, p-PI3K, and p-AKT in OSCC cells.

Role of Eukaryotic Translation Elongation Factor 1 Family Members in the Tumorigenesis and Progression of Lung Adenocarcinoma / 中国医学科学院学报

Objective To investigate the role and mechanism of eukaryotic translation elongation factor 1(EEF1) family members (EEF1D,EEF1A1,and EEF1A2) in lung adenocarcinoma (LUAD) based on public databases.Methods We examined EEF1 member expression levels in human LUAD samples via The Cancer Genome Atlas in the UCSC Xena browser and the Clinical Proteomic Tumor Analysis Consortium.We analyzed the mRNA and protein levels of EEF1D,EEF1A1,and EEF1A2 and their correlations with pathological variables via the Mann-Whitney U test.The Kaplan-Meier curves were established to assess the prognostic values of EEF1D,EEF1A1,and EEF1A2.The single-sample gene set enrichment analysis algorithm was employed to explore the relationship between the expression levels of EEF1 members and tumor immune cell infiltration.Spearman and Pearson correlation analyses were performed to examine the relationship between the expression levels of EEF1 members and those of the genes in the phosphatidylinositol 3-kinase/protein kinase B signaling pathway.The immunohistochemical assay was employed to determine the expression levels of EEF1D,EEF1A1,and EEF1A2 in the LUAD tissue (n=75) and paracancer tissue (n=75) samples.Results The mRNA and protein levels of EEF1D,EEF1A1,and EEF1A2 showed significant differences between tumor and paracancer tissues (all P<0.001).The patients with high protein levels of EEF1A1 showed bad prognosis in terms of overall survival (P=0.039),and those with high protein levels of EEF1A2 showed good prognosis in terms of overall survival (P=0.012).The influence of the mRNA level of EEF1D on prognosis was associated with pathological characteristics.The expression levels of EEF1 members were significantly associated with the infiltration of various immune cells and the expression of key molecules in the phosphatidylinositol 3-kinase/protein kinase B signaling pathway.Conclusion EEF1D,EEF1A1,and EEF1A2 are associated with the progression of LUAD,serving as the candidate prognostic markers for LUAD.

MEX3A promotes proliferation and migration of colorectal cancer cells via PI3K/Akt signaling pathway / 解剖学报

Objective To investigate the expression of MEX3A in colorectal cancer (CRC), and to explore the effect and mechanism of MEX3A on the proliferation and migration of colorectal cancer cells. Methods Totally 327 cases of data(41 normal tissues and 286 tumor tissues) were obtained from TCGA database, and 104 cases of clinical samples (77 cases tissues and 27 paracancerous tissues) were collected for immunohistochemistry, then analysed the differences in MEX3A expression between CRC tissues and normal tissues. Western blotting and immunofluorescence staining were used to evaluate the differential expression of MEX3A in CRC cell lines. CL187 cells were selected as the follow-up research vector. Small interfering RNA of MEX3A(siMEX3A) was transfected into CL187 cells to inhibit the expression of MEX3A. The proliferation and migration of CL187 cells were measured by MTT, colony formation assay and Transwell assay. The expression of PI3K, p-PI3K, Akt and p-Akt were detected by Western blotting. Results TCGA database, immunohistochemistry and Western blotting analysis showed that MEX3A was highly expressed in colorectal cancer. The result of immunofluorescence staining showed that MEX3A was concentrated in the cytoplasm and the nucleus. In MTT, colony formation assay and Transwell assay, the proliferation and migration ability of CL187 cells in siMEX3A group decreased significantly than those in control group (P<0.05). Western blotting result showed that the expression of p-PI3K and p-Akt in siMEX3A group down-regulated significantly (P<0.05), and the inhibition of proliferation and migration ability of CL187 cells induced by siMEX3A group could be reversed by 740 Y-P via activating the PI3K/Akt signaling pathway. Conclusion MEX3A is highly expressed in colorectal cancer and promotes the proliferation and migration of CRC cells via PI3K/Akt signaling pathway.

Mechanism of insulin-like growth factor 2 regulating human ovarian granulosa cell proliferation through the phosphatidylinositol 3-kinase/ protein kinase B signaling pathway / 解剖学报

Objective To explore the effect and mechanism of insulin-like growth factor 2 ( IGF2 ) on the proliferation of human ovarian granulosa cells ( KGN ). Methods KGN cells cultured in vitro and treated with different concentrations of IGF2 were divided into control group and IGF2 group (25 μg/L, 50 μg/L, 100 μg/L), and then cells were divided into control group, 100 μg/L IGF2 group, LY294002 group, and IGF2 +LY294002 group after intervened the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway with LY294002. MTS and 5-ethynyl-2'- deoxyuridine (EdU) method was used to detect the effect of IGF2 on KGN cell proliferation, and enzyme linked immunosorbent assay was used to detect the contents of estrogen and progesterone in cell culture supernatant. The expressions of insulin like growth factor 1 receptor (IGF1R), protein kinase B (Akt), phosphorylated protein kinase B(p-Akt) and CYP19A1 protein in each group were detected by Western blotting. Results With the concentration gradient of IGF2, the proliferation rate of KGN cells and the secretion of estrogen and progesterone gradually increased. The cell proliferation rate and hormone level in the group treated with lOOfig/L IGF2 were the highest (P<0.01), while the PI3K/Akt signaling pathway was inhibited, and the cell proliferation rate and hormone secretion decreased significantly (P<0.01). The protein expression levels of IGF1R, p-Akt and CYP19A1 in different concentration groups increased significantly (P<0.05). While the expression of the above proteins were affected by intervened the PI3K/Akt signaling pathway. Compared with the control group, the protein expression of IGF1R and p-Akt increased significantly in IGF2 group and IGF2 +LY294002 group(P<0.01), CYP19A1 increased significantly in IGF2 group(P<0.01), the protein expression of p-Akt and CYP19A1 decreased significantly in LY294002 group (P<0.05), there was no significant difference in the protein expression of IGF 1R. Compared with the IGF2 group, the protein expression of p-Akt and CYP19A1 decreased in IGF2 +LY294002 group (P<0.01), there was no statistically significant difference in the protein expression of IGF1R, and the expression levels of IGF1R, p-Akt and CYP19A1 were significantly reduced in LY294002 group (P<0.01). Conclusion IGF2 may promote the proliferation and secretion of human ovarian granulosa cells through the PI3K/Akt signaling pathway mediated by IGF1R.

Integration of network and experimental pharmacology to decipher the antidiabetic action of Duranta repens L / 中西医结合学报

OBJECTIVE@#Duranta repens is reported to contain a wide array of secondary metabolites, including α-amylase and α-glucosidase inhibitors, and - has potent antioxidant activity. The present study evaluated the network pharmacology of D. repens (whole plant) with targets related to diabetes mellitus and assessed its outcome by evaluating the effects of the hydroalcoholic extract of D. repens in streptozotocin-nicotinamide-induced diabetes mellitus in rats.@*METHODS@#Phytoconstituents of D. repens were retrieved from an open-source database and published literature, and their targets were predicted for diabetes mellitus using BindingDB and the therapeutic target database. Protein-protein interaction was predicted using STRING, and pathways involved in diabetes mellitus were identified using the Kyoto Encyclopedia of Genes and Genomes pathway browser. Druglikeness, ADMET profile (absorption, distribution, metabolism, excretion and toxicity) and cytotoxicity of compounds modulating proteins involved in diabetes were predicted using MolSoft, admetSAR2.0 and CLC-Pred, respectively. The interaction network among phytoconstituents, proteins and pathways was constructed using Cytoscape, and the docking study was performed using AutoDock4.0. The hydroalcoholic extract of D. repens was evaluated using streptozotocin-nicotinamide-induced diabetes mellitus animal model for 28 d, followed by an oral glucose tolerance test. At the end of the study, biochemical parameters like glycogen content, hepatic enzymes, antioxidant biomarkers and lipid profiles were quantified. Further, the liver and pancreas were collected for a histopathology study.@*RESULTS@#Thirty-six different secondary metabolites from D. repens were identified to regulate thirty-one targets involved in diabetes mellitus, in which protein-tyrosine phosphatase 1B (PTP1B) was primarily targeted. Enrichment analysis of modulated proteins identified 12 different pathways in diabetic pathogenesis in which the phosphatidylinositol 3-kinase-protein kinase B (PI3K-Akt) signaling pathway was chiefly regulated. The docking study found that durantanin I possessed the highest binding affinity (-8.9 kcal/mol) with PTP1B. Similarly, ADMET profiling showed that the majority of bioactive constituents from D. repens had higher human intestinal absorptivity and minimal cytotoxicity to normal cell lines, than tumor cell lines. Further, an in vivo animal study reflected the efficacy of the hydroalcoholic extract of D. repens to lower the elevated blood glucose level by stimulating insulin secretion, maintaining pancreatic β cell mass, regulating glycolysis/gluconeogenesis and enhancing the glucose uptake in skeletal muscles.@*CONCLUSION@#The present study reflected the probable network interaction of bioactive constituents from D. repens, their targets and modulated pathways, which identified the prime regulation of the PI3K-Akt signaling pathway and PTP1B protein. Modulation of PTP1B protein and PI3K-Akt signaling pathway could contribute to enhancing glucose uptake, insulin production and glycolysis and decreasing gluconeogenesis in diabetes, which was evaluated via the experimental study.

Effects of phosphatidylinositol-3 kinase-protein kinase B signaling pathway on iodine-intake ability of lactating rats / 中华地方病学杂志

Objective To observe the variation of phosphatidylinositol-3 kinase (PI3K),protein kinase B (AKT),sodium iodide symporter (NIS) mRNA and protein expression in rat mammary tissues and serum insulin growth factor Ⅰ (IGF-1) under different iodine nutrition levels,and to study the role of PI3K-AKT signaling pathway in the process of mammary gland intaking iodine during lactation period.Methods Totally 130 Wistar rats (100 female rats,30 male rats) were randomly divided into five groups with 20 female rats in each group:①control group (NI):was feed with normal diet and iodine content 50 μg/L in deionized water;②low iodine group 1 (LI1 group):was feed with low iodine diet and deionized water;③low iodine group 2 (LI2):was feed with low iodine diet and iodine content 5 μg/L in deionized water;④high iodine group 1 (HI1 group):was feed with normal diet and iodine content 3 000 μg/L in deionized water;⑤high iodine group 2 (HI2):was feed with normal diet and iodine content 10 000 μg/L in deionized water.After feeding for 3 months,females were mated with male rats,then male rats were taken out and every female rat was feed individually.Urinary iodine level of rats in lactation period 10 days after giving birth was tested.Blood and mammary tissue samples of rats in lactation period were taken after killing them.Enzyme linked immunosorbent assay (ELISA) was used to detect serum IGF-1 level,real-time fluorescence quantification PCR to detect the mRNA expression of mammary gland PI3K,AKT and NIS,Western blotting to detect mammary gland PI3K,total AKT,phosphorylation AKT (p-AKT) and NIS protein expression.Results The medians urinary iodine of lactation period rats in LI1 and LI2 (3.16,6.36 μg/L) were significantly lower than that in NI group (162.59 μg/L),and were significantly higher in HI1 and HI2 (2 356.27,11 507.29 μg/L) than that in NI group.The differences were statistically significant (all P ＜ 0.01).Compared with control group [(8.84 ± 2.12) μg/L],the content of serum IGF-1 increased significantly in lactation period rats in LI1 and LI2 groups [(13.30 ± 2.37) and (10.90 ± 1.92) μg/L,all P＜ 0.01].The real-time fluorescence quantification PCR detection results indicated that the differences were statistically significant by comparing NIS,AKT,PI3K mRNA expression of the mammary tissues of lactation period rats in the five groups (F=14.916,36.477,14.994,all P＜ 0.01).Among them,NIS mRNA expression quantities in LI1 and LI2 groups (0.75 ± 0.40,0.89 ± 0.51) were significantly higher than that in NI group (0.53 ± 0.31),and significantly lower in HI2 group (0.30 ± 0.24) than that in NI group (P ＜ 0.05 or ＜ 0.01).AKT mRNA expression quantities in LI1 and LI2 groups (0.90 ± 0.19,0.64 ± 0.22) were significantly higher than that in NI group (0.43 ± 0.22),and significantly lower in HI2 group (0.29 ± 0.15) than that in NI group (P ＜ 0.05 or ＜ 0.01).PI3K mRNA expression quantity in LI1 group (0.85 ± 0.42) was significantly higher than that in NI group (0.50 ± 0.24),and significantly lower in HI2 group (0.28 ± 0.10) than that in NI group (all P ＜ 0.01).Western blot detection results indicated that the differences were statistically significant by comparing mammary gland NIS protein expression of lactation period rats in the five groups (F=4.510,P＜ 0.01).Among them,LI1 group (1.67 ± 0.97) was significantly higher than NI group (0.87 ± 0.43,P ＜ 0.05).The differences were statistically significant by comparing the p-AKT protein expression among groups (F =3.528,P ＜ 0.05).Among them,HI2 group (1.10 ± 0.30) was significantly higher than NI group (0.75 ± 0.23,P ＜0.05).The differences were not statistically significant by comparing total AKT and PI3K protein expression among groups (F =0.558,1.319,all P ＞ 0.05).Conclusion The inhibitory effect of PI3K-AKT signaling pathways on NIS in the mammary gland was weaker than the effect of iodine intake.But the expression of functional p-AKT was gradually increased with the increment of iodine intake,which had been presented inhibit effect on NIS expression in lactating mammary gland.