CXCR4 antagonist AMD3100 reduced oxaliplatin resistance in colorectal cancer cells / 中国现代普通外科进展

Objective:colorectal cancer is one of the common malignant tumors in the gas-trointestinal tract.Oxaliplatin is the first-line drug for the treatment of advanced colorectal cancer,but drug resistance often occurs.The mechanism of CXCR4 in oxaliplatin resistance of colon cancer is not clear.This study intends to explore the mechanism of CXCR4 mediated oxaliplatin resistance and the potential therapeutic value of CXCR4 inhibitor AMD3100.Methods:oxaliplatin resistant strains HCT116 were constructed.The expression of CXCR4 and the phosphorylation level of PI3K-Akt signal pathway were detected by Q-PCR and Western blot.The phosphorylation level of PI3K-Akt signal pathway was detected by Q-PCR and Western blot.The effect of AMD3100 an-tagonizing CXCR4 or combined application of Akt inhibitor LY294002 on oxaliplatin resistance of drug-resistant cells was detected by CCK8.Results:CCK-8 was used to detect the proliferation activity of Oxaliplatin in HCT116 drug resistant group compared with control cells in the absence of drugs and at different concentrations.The results showed that there was no significant change in the activity of the resistant strains,while the control cells showed a significant decrease.Q-PCR and Western blot showed that the expression of CXCR4 and the phosphorylation level of PI3K-Akt increased significantly in the drug-resistant group(P＜0.05).After administration of CXCR4 inhibitor AMD3100,CXCR4 expression and PI3K-Akt phosphorylation decreased significantly(P＜0.05).AMD3100 enhanced the sensitivity of drug-resistant cell lines to oxaliplatin.The combination of AMD3100 and Akt inhibitor LY294002 can further enhance the sensitivity of drug-resistant cell lines to oxaliplatin.Conclusion:CXCR4 mediated activation of PI3K-Akt signaling pathway plays an important role in the resistance of colon cancer to oxaliplatin.AMD3100 may become a potential therapeutic drug against chemoresistance of colon cancer.

Esketamine induced apoptosis in human breast cancer cell line MDA-MB-468 and its mechanism / 中华内分泌外科杂志

Objective:To explore whether esketamine (ESK) can inhibit the proliferation and induce apoptosis of breast cancer cells, and explore the mechanism.Methods:CCK-8 assay was used to detect the inhibitory effect of ESK on the proliferation of breast cancer cells. Annexin V/PI staining was used to detect the morphological changes of cells; Apoptosis and reactive oxygen species were detected by flow cytometry. Western blot was used to detect apoptosis and pathway expression.Results:CCK-8 experiment results proved that ESK could inhibit the proliferation of breast cancer cells in a time-dependent manner. The survival rate of MDA-MB-468 cells treated with ESK at 20 μM was (35.47±2.61) %, which was statistically different from that treated with vinorelbine at the same concentration ( P<0.05). The IC50 value of ESK on MDA-MB-468 cells was (14.54±2.12) μM. After treatment with ESK, the mitochondrial membrane potential was significantly reduced. In the protein level, the expression of Cytochrome C, Bax and Caspase-3 was up-regulated, and the expression of Bcl-2 was down regulated, which induced the mitochondrial dependent apoptosis of MDA-MB-468 cells. ESK could up regulate the level of reactive oxygen species in MDA-MB-468 cells and regulate the expression of PI3K/AKT signaling pathway. Conclusions:ESK can inhibit the proliferation and migration of breast cancer cells and induce them to play a mitochondrial dependent apoptosis. Its mechanism is achieved by up regulating the level of ROS in breast cancer cells, thereby regulating the PI3K/AKT signaling pathway, which provides a theoretical basis for the development and utilization of Aln.

Effects of piperine on experimental colon carcinogenesis induced by DMH/DSS and its mechanism / 中国医师杂志

Objective:To investigate the effects of piperine on experimental colon carcinogenesis induced by 1, 2-dimethylhydrazine (DMH)/sodium dextran sulfate (DSS) and its mechanism.Methods:36 mice were divided into control group, model group and piperine group, 12 mice in each group. The control group was given normal saline by gavage; The model group and piperine group were given 3.6 mg/(kg·d) of normal saline and piperine respectively after establishing the experimental colon cancer model induced by DMH/DSS. Tumor load and volume were observed. Hematoxylin and eosin (HE) staining was used to observe the histological change of colon in mice. RAS/PI3K/AKT related pathway protein expression was detected by Western blot.Results:The body weight gain, protein expression levels of cleaved poly-ADP ribose polymerase (PARP), cleaved caspase-3 in model group were significantly lower than those in the control group (all P<0.05). The protein expression levels of Bcl-2, Bax, pan-Ras, p-MEK, p-ERK, PI3K, p-AKT, NF-κBP65, c-Myc and cyclin D1 in model group were significantly higher than those in the control group (all P<0.05). The body weight gain, protein expression levels of cleaved PARP and cleaved caspase-3 in piperine group were significantly higher than those in model group (all P<0.05). The protein expression levels of Bcl-2, Bax, pan-Ras, p-MEK, p-ERK, PI3K, p-AKT, NF-κBP65, c-Myc and cyclin D1 in piperine group were significantly lower than those in model group (all P<0.05). Conclusions:Piperine can inhibit the occurrence of experimental colon cancer induced by DMH/DSS, which may involve multiple components of RAS/PI3K/AKT signal axis.

Protective Mechanism of Lavender Against Skin Photodamage Based on Network Pharmacology and Experimental Verification / 中国实验方剂学杂志

ObjectiveTo predict the active ingredients and mechanism of action of lavender in protecting skin photodamage based on network pharmacology and molecular docking technology，and further verify possible signal pathways via animal experiments. MethodThe active ingredients and potential targets of lavender were obtained by SwissTargetPrediction，PharmMapper， and literature. Skin photodamage-related targets were searched from GeneCards，Online Mendelian Inheritance in Man （OMIM），DrugBank and DisGeNET databases. After common targets of the two were screened out，STRING was adopted to analyze the protein-protein interaction （PPI） network，where topological analysis and core target screening were performed by CytoNCA plug-in of Cytoscape 3.8.2. Based on DAVID， gene ontology （GO） annotations and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis were carried out among the intersection targets， and the active ingredients of lavender and the signal pathway proteins were selected and verified via molecular docking with AutoDock vina 1.1.2. Finally， mouse photodamage model was established by UVB irradiating the bare skin of mouse back， and the skin condition was observed by naked eyes. Hematoxylin-eosin （HE） and picric acid-acid fuchsin staining （Van Gieson， VG） were used to observe the pathological changes of mouse skin tissues. Western blot was employed to detect the protein expression in mouse skin tissues to further validate the key signal pathways. ResultIn this study，6 active ingredients of lavender，526 potential targets，2 688 disease-related targets，and 258 intersection targets were screened out， and 16 core targets were obtained by PPI network. Additionally， 113 related signal pathways were obtained by KEGG pathway analysis，among which phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） pathway and nuclear transcription factor-κB （NF-κB） signal pathway might play a key role in skin photodamage protection by lavender. Molecular docking showed that the active ingredients and the signal pathway proteins were well docked. Animal experiments indicated that the total flavonoids of lavender improved the appearance and histopathological condition of mouse skin， reduced the relative expression levels of phosphorylated（p）-PI3K，p-Akt，and B cell lymphoma 2 （Bcl-2） proteins （P<0.05，P<0.01）， and increased relative expression level of Bcl-2-associated X protein（Bax） （P<0.05）. ConclusionLavender exerts synergistic effect in resisting skin photodamage，with the characteristics of multi-components，multi-targets，and multi-pathways， which provides a basis for subsequent in-depth research on the complex mechanism of lavender against skin photodamage.

Effects of long-chain noncoding RNA Linc00673 overexpression on proliferation and apoptosis of gastric cancer cell line MGC-803 / 中国应用生理学杂志

Objective: To investigate the effects of long-chain noncoding RNA Linc00673 overexpression on proliferation and apoptosis of gastric cancer cells and its mechanisms. Methods: The recombinant lentivirus expressing plasmid pLVX-Linc00673 and the control empty plasmid pLVX-NC were packaged and amplified in 293T cells, and the recombinant lentivirus was transfected into gastric cancer cell line MGC-803 to establish a cell line stably overexpressing Linc00673. The expression of Linc00673 gene was detected by real-time fluorescence quantitative PCR. The growth and proliferation of cells were observed by MTT assay and clone formation assay. Cell cycle and apoptosis were detected by flow cytometry. The expressions of cell cycle related regulatory genes were detected by qPCR. The expressions of key molecules in the PI3K/Akt signaling pathway and tumor proliferation related proteins were detected by Western blot. Results: The expressions of Linc00673 in gastric cancer cell line MGC-803, BGC-823 and AGS were significantly higher than that in normal gastric mucosa cell line GES-1 (P＜0.05). MGC-803 cell line with stable overexpression of LINC00673 was established, and the expression level of LincC00673 was 200 times higher than that of the control empty carrier group. Overexpression of Linc00673 promoted proliferation of MGC-803 cells (P＜0.05) and clone formation (P＜0.05), inhibited cell apoptosis and affected the G1→S phase progression of cell cycle (P＜0.01). Overexpression of Linc00673 could affect the expressions of cell cycle regulatory gene CCNG2, P19 and CDK1 in MGC-803. Western blot showed that Linc00673 overexpression not only promoted the expressions of the key molecule pAkt in PI3K / Akt signaling pathway and its downstream target NF-κ B and Bcl-2 protein, but also up regulated the expressions of tumor related factors β-catenin and EZH2 proteins. Conclusion: Overexpression of Linc00673 may promote proliferation and inhibit apoptosis of MGC-803 cells through PI3K/Akt signaling pathway.

β-arrestin-2 inhibits autophagy by up-regulating PI3K/Akt signal to alleviate liver ischemia-reperfusion injury in mice / 器官移植

Objective To verify whether β-arrestin-2 inhibits autophagy by up-regulating PI3K/Akt signal to protect the liver from ischemia-reperfusion injury (IRI) in mice. Methods Twelve β-arrestin-2 knockout (KO) and twelve wild-type (WT) C57BL/6 mice were randomly divided into the KO+sham group, KO+IRI group, WT+sham group and WT+IRI group, six mice in each group. The mouse models with 70% liver IRI were established or sham operation was performed. Relevant experiments were carried out at 6 h after liver reperfusion or operation. The expression levels of apoptosis signal protein cleaved Caspase-3, proliferation signal protein Ki-67 and the PI3K/Akt signal protein p-Akt were detected by immunohistochemical staining. Results Immunohistochemical staining demonstrated that compared with the corresponding sham group, the positive cell count for cleaved Caspase-3, Ki-67 and p-Akt in liver tissues of mice was significantly increased in the KO+IRI and WT+IRI groups (all P < 0.01). Compared with the WT+IRI group, the positive cell count for cleaved Caspase-3 in liver tissues of mice was significantly increased, whereas the positive cell count forKi-67 and p-Akt was significantly decreased in the KO+IRI group (both P < 0.05). Conclusions β-arrestin-2 can mitigate the liver cell apoptosis and promote the repair of injury after IRI in mice. Moreover, β-arrestin-2 inhibits autophagy by up-regulating the PI3K/Akt signal to alleviate liver IRI in mice.

Exploration on mechanism of anti-coronavirus of Sophora flavescens through multi-pathway / 中草药

The coronavirus disease 2019 (COVID-19), originated in Wuhan city, Hubei Province, China in December 2019, and then quickly spread to most provinces and regions in China and even spread to many countries abroad. COVID-19 is characterized by wide epidemic, strong infectivity, rapid onset and critical condition. In the face of this epidemic, all parts of the country quickly set off a peak in the fight against COVID-19, but no effective drug for COVID-19 has been developed in the short term. Recently, many hospitals have combined traditional Chinese medicine with western medicine in treatment, and the clinical effect is remarkable, which proves the antiviral effect of traditional Chinese medicine. A large number of pharmacological and clinical studies have proved that the Chinese materia medica S. flavescens has significant antiviral effect. In this paper, the mechanism of anti-coronavirus effect of S. flavescens is expounded from multiple pathways, such as type I interferon, NF-κB signal pathway, ERK signal pathway, PI3K/Akt signal pathway and matrine alkaloids, etc. It is intended to provide reference for clinical treatment of coronavirus infection pneumonia and research and development of related drugs of S. flavescens.

UC-MSCs affects apoptosis and proliferation of human lung adenocarcinoma A549 cells via PI3K/AKT signaling pathway / 中国肿瘤生物治疗杂志

@#Objective: To investigate the effects of umbilical cord-derived mesenchymal stem cells (UC-MSCs) on apoptosis and proliferation of human lung adenocarcinoma A549 cells via PI3K/AKT signaling pathway, and to explore the mechanism. Methods: UCMSCs were isolated from human umbilical cord tissues by enzyme digestion method and cultured in vitro. The immunophenotypes of the obtained MSCs were identified by flow cytometry. The culture supernatant of UC-MSCs was collected to establish an indirect in vitro co-culture system of UC-MSCs conditioned medium and lung adenocarcinomaA549 cell line. Proliferation ofA549 cells was detected by CCK-8 assay; apoptosis ofA549 cells was determined byAnnexin V/PI double staining, and cell cycle distribution of tumor cells was determined by PI staining. The transcription levels of apoptosis and proliferation associated downstream genes in the PI3K/AKT pathway, such as CyclinD1, BAX and Bcl-2, were detected by quantitative polymerase chain reaction (qPCR). Moreover, Wb was utilized to detect the expression levels of PI3K/AKT pathway-related proteins. Results: The culture flask was filled with fibroblast-like cells, arranged in parallel, with spiral growth after three weeks of isolation and culture of human umbilical cord tissues. The flow cytometry results revealed that the MSC markers CD73, CD90 and CD105, but not CD45 and HLA-DR, were expressed on obtained cells.After indirect in vitro co-culture of UC-MSCs conditioned medium and lung adenocarcinoma A549 cells, the proliferation rate of A549 cells was significantly decreased; the apoptosis rate was significantly increased, and the cell cycle was obviously arrested at the G1 phase as compared with the control group (all P<0.01). The transcription levels of PI3K/AKT signaling pathway-related factors, CyclinD1 and Bcl-2 were down-regulated, and the transcription level of BAX was up-regulated (all P<0.01). The total AKT was not changed, but p-AKT protein expression was decreased in a dose-dependent manner inA549 cells cultured in UC-MSCs conditioned medium (P<0.01). Conclusion: UC-MSCs can affect the proliferation and the apoptosis of A549 cells, and arrest cells in G1 phase. The main mechanism is that UC-MSCs can inhibit the PI3K/AKT signaling pathway in A549 cells, providing an experimental basis for exploring the safety and effectiveness of clinical application of UC-MSCs.

Clinical study on transcription and protein expression of tumor necrosis factor α mRNA in peripheral blood and activation of signal path PI3K/AKT in patients with chronic heart failure / 实用医学杂志

Objective The study was to investigate the activation of tumor necrosis factor α (TNF-α) mRNA transcription and protein expression in peripheral blood and activation of signal path PI3K/AKT in patients with chronic heart failure. Methods From February 2015 to April 2018, 244 patients with heart failure in the cardiovascular department of our hospital were selected as heart failure group, while 244 healthy cases were enrolled as the control group at the same time. The peripheral blood samples of two groups were collected. We detected the transcription and protein expression of TNF-α mRNA and the activation of PI3K, AKT in peripheral blood. The left ventricular ejection fraction (LVEF) were measured in two groups. The correlations between influencing factors and LVEF were analyzed. Results The levels of PI3K, AKT in the heart failure group were higher than those in the control group. The differences were statistically significant respectively (P < 0.05). The mRNA relative content and protein content of TNF-α in peripheral blood mononuclear cells of heart failure group were higher compared with those of control group (P < 0.05). The LVEF of heart failure group was significantly lower than that of the control group (34.50 ± 6.33) ％ versus (55.60 ± 2.49) ％, P < 0.001). Among 244 patients with heart failure, Spearman correlation analysis showed that there were significant positive correlations between TNF-a mRNA and protein expression levels and the levels of PI3K, AKT respectively (P < 0.05). Multiple factors unconditional Logistic regression analysis showed that the TNF-α mRNA, protein expression and PI3K, AKT levels in peripheral blood were independent risk factors for LVEF (P < 0.05). Conclusion The expression levels of PI3K, AKT and TNF-α are all significantly increased in chronic heart failure patients, which could participate in the occurrence and development of heart failure.

Shikonin suppresses invasion and migration of non-small cell lung cancer cells A549 / 中国免疫学杂志

Objective: Lung cancer is the most commonly diagnosed cancers and the leading cause of cancer death worldwide. This study aims to explore the effect and mechanism of shikonin on invasion and migration of non-small cell lung cancer cells A549. Methods: Cell viability was detected by CCK-8. Transwell assay was performed to analyze the invasion of A549. Wound healing assay was used to measure the migration of A549. The expression of vascular endothelial growth factor ( VEGF), matrix metalloprotein 14 ( MMP-14), fibronectin ( FN), Vimentin, PI3 K, p-AKT and p53 was tested by Western blot. Results: Cell viability of A549 was reduced by shikonin. Compared with control group, the invasion of A549 in shikonin group ( 20, 50, 100 mmol/L) was decreased as well as the migration ( P＜0. 05). The expression of VEGF, MMP-14, Fn and Vimentin in shikonin group ( 20, 50, 100 mmol/L) was lower than control group ( P＜0. 05). Compared with A549 group, the expression of PI3 K and p-AKT in IGF-1 group was increased with alleviated expression of p53 ( P＜0. 05). Compared with IGF-1 group, the expression of PI3 K and p-AKT in shikonin group ( 20, 50, 100 mmol/L) declined with enhancive expression of p53 ( P ＜ 0. 05). Conclusion: Shikonin suppresses invasion and migration of NSCLC cells A549 through inhibition of PI3 K/AKT signal pathway.

Effects of EI injection on Brain Energy and PI3K/AKT Signaling Pathway in IBO Basal Nucleus Microinjected with Dementia Model Rats / 世界科学技术-中医药现代化

Objective: To investigate the effects of EI injection on learning and memory ability and brain energy of two-way Meynert basal injection of Ibotenic acid (IBO) dementia model rats. Methods: A rat model of dementia wasestablished by bilateral meynert basal injection of IBO. After 8 weeks of EI injection, Morris water maze was used todetect the learning and memory ability of rats. Congo red staining was used to observe the deposition of Aβ plaque inhippocampal CA1 and cortical areas of rats. The changes of ATP, ADP and AMP in brain tissue of each group weredetermined by HPLC. The content of insulin in rat brain tissue was detected by ELISA kit. The expression of key proteinin PI3K/AKT signaling pathway was detected by Western blot. Results: Compared with the sham operation group, theescape latency of the model group was significantly prolonged, the number of entering the platform, the time andpercentage of crossing the platform quadrant decreased significantly (P ＜ 0.05); Aβ plaque deposition was observed inthe hippocampus and cortex; ATP/AMP ratio and insulin content were significantly decreased (P ＜ 0.05); brain tissue PI3 K and AKT protein were low expression (P＞ 0.05) . After intervention with EI injection, the escape latency of themodel rats was significantly shortened, the number of entering the platform and the time of crossing the platform quadrantincreased significantly (P ＜ 0.05); the hippocampus and cortex red staining was alleviated; the brain tissue ATP/AMPratio and insulin content increased significantly (P ＜ 0.05) . Conclusion: EI injection can improve the learning andmemory function of IBO-induced dementia model rats, which is related to the improvement of brain energy.

Chlamydia trachomatis pORF5 protein inhibits apoptosis through activating PI3K-Akt pathway / 中国免疫学杂志

Objective:To study the relationship between apoptosis and the pORF5 protein of Chlamydia trachomatis,and further to explore its molecular mechanisms,which could lay a foundation for chlamydial pathogenic mechanisms.Methods:pGEX-6p/pORF5 recombinant expression vector was transformed to XL1-blue E.Coli to express GST-pORF5 fusion protein,and GST-pORF5 fusion protein was purified with Glutathione Sepharose 4B Beads,and cleaved to get pORF5 protein without GST tag by PreScission protease.The pORF5 protein was used to stimulate HeLa cells at different concentrations,then Western blot was used to evaluate the ex-pression of Bax,Bcl-2 and phosphorylation of PI3K/Akt at different time points,Hoechst staining and Flow cytometry were applied to measure the apoptosis of HeLa cells.Before treated with pORF5 protein for 24 h,HeLa cells were pretreated with PI3K inhibitor LY294002 for 1 h,the expression of Bax,Bcl-2 and the phosphorylation of Akt were evaluated by Western blot,apoptosis rates were also determined.Results:The pORF5 protein changed the expression of Bax and Bcl-2 in dose-and time-dependent manners,pORF5 increased the expression of Bcl-2 and decreased the expression of Bax at the concentration of 10 μg/ml,and there was obvious change at concentration of 15 μg/ml for 24 h.The apoptosis rates of pORF5 treated group were reduced by 27.3% and 8.4% respectively when compared with TNF-α treated group(P<0.01) and untreated group (P<0.05).Akt was phosphorylated after stimulated with pORF5 protein for 15 min,and reached its peak at 30 min.PI3K/Akt inhibitor led to the decrease of the expression of Bcl-2 and phosphorylation of Akt and increase of the expression of Bax,furthermore,PI3K/Akt inhibitor reversed pORF5-mediated anti-apoptosis, the apoptosis rate in LY294002 treated group was increased by 13.0%,when compared with the control group(P<0.01).Conclusion:pORF5 protein could inhibit apoptosis through activating PI3K/Akt signal pathway by induction of Bcl-2 and suppression of Bax.

Effects and mechanism of resveratrol on apoptosis of vascular smooth muscle cell in diabetic rats / 中国免疫学杂志

Objective:To explore effects and mechanism of resveratrol on apoptosis of vascular smooth muscle cell in diabetic rats. Methods: The mRNA level of monocyte chemoattractant protein 1 (MCP1),macrophage migration inhibitory factor (MIF) and in-terleukin 18 (IL-18) was detected by quantitative real-time reverse transcription PCR (qRT-PCR). Myocardial fibrosis was analyzed by hematoxylin-eosin (HE) staining and Masson staining. Apoptosis was tested by flow cytometry. The expression of Cleaved caspase-3, Cleaved caspase-9,B cell lymphoma 2 (Bcl-2),Bcl-2-associated X protein (Bax),PI3K,p-PI3K,AKT and p-AKT was measured by Western blot. Results: The mRNA levels of MCP1,MIF and IL-18 in STZ-induced diabetic were reduced by resveratrol (P＜0. 05). The aggravation of myocardial fibrosis in STZ-induced diabetic rats was ameliorated by resveratrol. Apoptosis of vascular smooth muscle cell in STZ-induced diabetic rat group was higher than control group (P＜0. 05 ) . Compared with STZ-induced diabetic rat group, apoptosis of vascular smooth muscle cell in STZ+ resveratrol group was decreased (P＜0. 05). What′s more,the expression of Cleaved caspase-3,Cleaved caspase-9 and Bax in STZ-induced diabetic rats were inhibited by resveratrol(P＜0. 05). And the expression of Bcl-2 in STZ-induced diabetic rats was elevated by resveratrol(P＜0. 05). Compared with control group,the rate of p-PI3K/PI3K and p-AKT/AKT in STZ-induced diabetic rat group was decreased (P＜0. 05). The rate of p-PI3K/PI3K and p-AKT/AKT in STZ+ resveratrol group was higher than STZ-induced diabetic rat group ( P＜0. 05). Conclusion: Resveratrol represses apoptosis of vascular smooth muscle cell in STZ-induced diabetic rats through activating of PI3K/AKT signal pathway.

Analysis and validation of PI3K/AKT signaling pathway associated with overall survival time of patients with HER2-positive breast cancer / 解放军医学杂志

Objective To analyze and validate the key molecular targets correlated with the overall survival of patients with HER2-positive breast cancer.Methods First,the survival time and transcriptome data of patients with HER2-positive breast cancer in stage Ⅰ / Ⅱ and Ⅲ/Ⅳ were downloaded from the TCGA database.The significantly differential genes between overall survival ＜2 years and ＞8.5 years in stage Ⅰ / Ⅱ were picked out by edgeR package,and the pathways were enriched by KEGG.Similarly,the differential genes between overall survival ＜2 years and ＞7 years in stage Ⅲ/Ⅳ were analyzed.Furthermore,KEGG pathway analysis was performed using the differential genes overlapped by stage Ⅰ /Ⅱ and Ⅲ/Ⅳ.Second,the relationships between the expression levels of key node genes and other genes in enriched pathway and the overall survival of patients with HER2-positive breast cancer were validated by KMplot database.Last,the correlation between the activity of pathway enriched in KEGG and the resistance to anti-HER2 treatment was validated in HER2-positive breast cancer cell line BT474.Results In patients with stage Ⅰ / Ⅱ HER2-positive breast cancer whose overall survival was ＜2 years,PI3K/AKT was the 9th signaling pathway enriched by up-regulated differential genes.In patients with stage Ⅲ/Ⅳ whose overall survival was ＜2 years,PI3K/AKT was the 2nd signaling pathway enriched by up-regulated differential genes.Furthermore,PI3K/AKT was the first signal pathway enriched by the overlapping upregulated genes of patients in stage Ⅰ / Ⅱ and Ⅲ / Ⅳ whose overall survival was ＜2 years.Patients with high expression of PI3K and AKT (key node genes) or CFAP221 and COL4A6 (other genes) of PI3K/AKT pathway had shorter overall survival than those with low expression.PI3K inhibitors could enhance the growth inhibitory effect of HER2 small molecule inhibitor on HER2-positive breast cancer cell line BT474.Conclusions The overexpression of PI3K/AKT pathway is associated with the shorter overall survival in HER2-positive breast cancer patients,and associated with anti-HER2 resistance in HER2-positive breast cancer cell line.

Effects of Aripiprazole on PC12 Cell Injury Induced by Aβ25-35 and Its Mechanism / 中国药房

OBJECTIVE:To study the effects of aripiprazole on PC12 cell injury induced by amyloid β-protein(Aβ25-35)and its mechanism. METHODS:PC12 cells were randomized into normal control group,model group (20 μmol/L Aβ25-35),aripiprazole low-concentration,medium-concentration and high-concentration groups(5,10,20 μmol/L aripiprazole+20 μmol/L Aβ25-35). These groups were cultured with culture medium containing relevant medicine for 48 h,with 6 wells in each group. The viability(optical density value)of PC12 cell was measured by MTT assay,and PC12 cell apoptosis was measured by Hoechst staining. The activi-ties of Caspase-3 and Caspase-9 were determined by spectrophotometry. The protein expression of Bcl-2,Bax and PI3K and the phosphorylation of Akt were assayed by Western blot assay. RESULTS:Compared with normal control group,optical density value of model group was decreased while apoptotic rate was increased;the activities of Caspase-3 and Caspase-9,and the protein expres-sion of Bax were increased;the protein expression of Bcl-2 and PI3K,the phosphorylation of Akt were decreased(P<0.01). Com-pared with model group,optical density value of aripiprazole low-concentration,medium-concentration and high-concentration groups were increased,while apoptotic rate and the activities of Caspase-3 and Caspase-9 were decreased;the protein expression of Bcl-2 and PI3K and the phosphorylation of Akt were enhanced;while the protein expression of Bax were decreased in aripiprazole medium-concentration and high-concentration groups(P<0.05 or P<0.01). CONCLUSIONS:Aripiprazole can suppress cell apop-tosis of PC12 cell induced by Aβ25-35,which is related to activating PI3K/Akt signal pathway.

Effect of Xiaotan Jieyu decoction on PI3K/Akt pathway of MCF-10AT cells and its mechanism / 第二军医大学学报

Objective To study the effect of Xiaotan Jieyu (XTJY) Decoction on PI3K/Akt signal pathway of MCF-10AT cells of breast precancerous lesion and the ralated mechanism. Methods The MCF-10AT cells were randomly divided into following groups: XTJY group, PI3K/Akt inhibitor LY294002 (LY) group, XTJY+LY group and control (CON) group. After corresponding drug intervention, CCK-8 method was used to observe the MCF-10AT cell proliferation and growth inhibition at 24 h and 48 h, flow cytometry was used to detect the changes of cell cycle at 48 h, and the expression changes of PTEN, PI3K and Akt protein were detected by Western blotting analysis at 48 h. Results The proliferation of MCF-10AT cells were significantly inhibited at 24 h and 48 h in XTJY, LY and XTJY+LY groups, and the inhibitory rate of MCF-10AT cells in XTJY+LY group was significantly higher than those in XTJY and LY groups (P<0.05). Compared with CON group, the percentages of G0/G1 phase cells in XTJY, LY and XTJY+LY groups were significantly increased at 48 h (P<0.05), while the percentages of S phase and G2/M phase cells were significantly decreased (P<0.05); besides, there were significant differences between XTJY+LY group and the other two groups (P<0.05). Compared with CON group, the expressions of PI3K and p-AKT protein in XTJY, LY and XTJY+LY groups were significantly decreased at 48 h (P<0.05), and the expression of PTEN protein was significantly increased (P<0.05). Conclusion XTJY prescription may exert inhibitory and apoptotic effect on MCF-10AT cells through inhibiting the PI3K/Akt signal pathway.

Study of mechanisms of hesperidin onanti-lung cancer effect based on microfluidic chip technology / 中国药理学通报

Aim To investigate the effect of hesperidin on human lung cancer cell A549 and the possible mechanism.Methods The cell apoptosis and necrosis of A549 treated with hesperidin were measured by the Hoechst 33342/PI fluorescent dye based on microfluidic chip technology.Cell cycle and apoptosis rate were evaluated by flow cytometry(FCM).The expressions of the related genes were detected through the real-time fluorescent quantitative PCR technology(RT-PCR) including VEGF, PI3K and PTEN.The protein expressions of Bcl-2, Cyclin B1, PI3K, Akt and PTEN were detected by Western blot after hesperidin intervention.Results The proliferation of A549 cells was significantly inhibited by hesperidin in a dose-dependent manner.FCM results showed that hesperidin could not only influence the G0/G1 phase and S phase, but also promote the apoptosis of lung cancer cells.Meanwhile, the apoptosis and necrosis rate was increased from(6.7±0.6)% to(27.9±1.1)% compared with that of control group(P<0.05).From the level of molecular, the gene expressions of VEGF and PI3K were decreased, while the PTEN was increased after hesperidin stimulation.Western blot results showed that the expression of protein Bcl-2, Cyclin B1 and Akt were decreased, which all showed close relationship with cell apoptosis, cell cycle and PI3K-Akt signaling pathway.The expression of PI3K was increased, but the change of PTEN was not statistically significant compared with that of control group.Conclusion Hesperidin induces lung cancer cell apoptosis through PI3K-Akt signaling pathway, which blocks cancer cell division and destroys the balance of related protein expression.

Manumycin-induced apoptosis in U937 human myeloid leukemia cells via blocking PI3K/Akt signaling pathway / 实用医学杂志

Objective To explore the effects of manumycin on U937 cells and its mechanisms.Methods Apoptosis was detected by flow cytometry using annexin V and propidium iodide (PI) after treatment with manumycin (2 μmol/L) for different time.The reactive oxygen species (ROS) was detected by flow cytometry using Reactive Oxygen Species Assay Kit.The expressions of PI3K,P-Akt and Akt were detected by Western blotting.Results Compared with the 0 h treatment,manumycin treatment resulted in apoptosis (P ＜ 0.05) and ROS generation (P ＜ 0.05) gradually in U937 cell lines.Furthermore,manumycin also inhibited the activation of phosphatidylinositol-3 kinase (PI3K)/Akt pathway.Moreover,NAC could decrease manumycin-induced ROS generation and inhibit the effects of manumycin on the PI3K/Akt pathway and protect U937 cells from apoptosis induced by manumycin.Conclusion Manumycin induces apoptosis in U937 human leukemia cells through the regulation of the PI3K/Akt pathway and ROS production plays a critical role in the progress.

Mechanism Study on Gymnema Sylvestre Total Saponins Alleviating Insulin Resistance via Regulation of PI3κ/AKT Signal Transduction Pathway in Adipose Tissues / 世界科学技术-中医药现代化

This study was aimed to observe the effects of gymnema sylvestre total saponins (GA) on insulin resistance (IR) in adipose tissues of KKay mice,in order to discuss the action mechanism from the PI3K/AKT signal transduction pathway.A total of 27 KKay mice were randomly divided into the model group (DM),Pioglitazone group (BG) and GA group.Nine normal C57BL/6J mice were used as the normal control group (NC).Intragastric administration of drugs was given for eight weeks.The bodyweight and food intake of all mice were tested each week.After the experiment was completed,fasting plasma insulin (Fins) and fasting plasma glucose (FPG) were detected and the insulin sensitive index (ISI) was calculated.Expressions of PDK-1,AKT,P-AKT (Ser473),P-AKT (Thr308) in adipose tissues of epididymis in mice were detected.And expressions of PI3K-p85 mRNA,PTEN mRNA,APN mRNA were also measured.The results showed that compared with DM group,bodyweight of BG and GA groups were decreased (P＜0.05,P＜0.05);FPG and Fins level of BG and GA groups were decreased (P＜0.05,P＜0.05,P＜0.05,P＜0.05),ISI increased (P＜0.05,P＜0.05);APN mRNA increased in BG and GA group (P＜0.01,P＜0.01);PI3K-p85 mRNA increased in GA group (P＜0.05);P-AKT (Thr308) protein expression increased in BG and GA group (P＜0.05,P＜0.01);P-AKT (Ser473) protein expression increased in GA group (P＜0.05);the phosphorylation of AKT (Thr 308) was enhanced in BG and GA group (/9＜0.01,P＜ 0.01);the phosphorylation of AKT (Ser473) was enhanced in GA group (P＜0.01,P＜0.01).PDK-1 protein expression was decreased in BG and GA group (P＜0.05);PTEN mRNA decreased in BG and GA group (P＜0.05,P＜0.01).It was concluded that GA can ameliorate IR by sensitizing PI3κ/AKT signal pathway in adipose tissues of KKAy mice.

Icaritin promotes the apoptosis of leukemia K562 cells / 白血病·淋巴瘤

Objective To investigate the effect and mechanism of the icaritin on the human cronic myeloid leukemia K562 cells. Methods The K562 cells at logarithmic growth phase were divided into the control group and the icaritin group. The cells in the control group were normally treated and the cells in the icaritin group were incubated with 8 μmol/L icaritin. Methyl thiazolyl tetrazolium (MTT) method and flow cytometry were used to examine the proliferation and apoptotic changes in the two groups after incubation for 72 h, respectively. Gene expression of p85 and Akt were detected by RT-PCR. The protein changes of p85, Akt, p-p85, p-Akt cleavage-caspase-3 and caspase-3 were detected by Western blot. Results Compared with the control group, the proliferation rate of K562 cells in the icaritin group was significantly decreased (P 0.05). Conclusion Icaritin could induce the proliferation and promote the apoptosis of K562 cell, and its mechanism may be achieved through activating the PI3K-Akt signal transduction pathway.