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OBJECTIVE@#To investigate the molecular mechanism by which a novel naphthalene allyl trifluoromethyl benzocyclopentanone XX0335 inhibits the proliferation and induces apoptosis of lung cancer A549 cells.@*METHODS@#Lung cancer A549 cells were treated with 0.1% DMSO (control) or different concentrations (6.25, 12.5, and 25 μg/mL) of XX0335, and the changes in cell viability, cell cycle, proliferation and apoptosis were assessed with CCK-8 assay, EdU experiment, and flow cytometry. The effects of different concentrations of XX0335 on phosphorylation levels of proliferation-related proteins Akt, mTOR, Akt/mTOR and the expressions of cleaved PARP and cyclin D1 were determined using Western blotting. We also assessed the effect of XX0335 on tumor growth in a mouse model bearing A945 cell xenograft.@*RESULTS@#Treatment with XX0335 reduced the viability of A549 cells in a dose-dependent manner (P < 0.01) and significantly inhibited cell proliferation (P < 0.001). Flow cytometry showed that XX0335 treatment promoted apoptosis of the cells (P < 0.01) and caused an obvious increase of the number of G1-phase cells. Compared with DMSO, XX0335 significantly inhibited the phosphorylation of Akt and mTOR, increased the expression of cleaved PARP, and lowered the protein expression of cyclin D1. In the tumor-bearing mouse models, injection of XX0335 significantly decreased the tumor volume (P < 0.01).@*CONCLUSION@#XX0335 inhibits the proliferation, cycle and induces apoptosis of lung cancer A549 cells possibly by inhibiting the Akt/mTOR signal pathway.
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Animals , Humans , Mice , A549 Cells , Apoptosis , Cell Proliferation , Lung Neoplasms/metabolism , Naphthalenes/pharmacologyABSTRACT
Aim To explore mechanism of epigallocatechin-3-gallate (EGCG) on alleviation of hippocampal neuronal autophagy in APP/PSI transgenic mice. Methods 8-month old APP/PSI transgenic mice were randomly divided into three groups;model group (Tg), EGCG low dose group (Tg/EGCG-L), high dose group (Tg/EGCG-H). C57BL/6J mice were utilized as control. Learning and memory were detected by Morris water maze test. The hippocampal ULK1, P62, LC3 I I / LC3 I,mT0R and Aß M2 expressions were detected by Western blot, immunohistochemical staining and ELISA. Results Compared with NT mice, Tg mice showed a marked prolongation of the escape latency in MWM test (P <0. 05). Decreased ULK1 expression and increased P62, LC3 II/LC3 I and A ßM 2 were detected (P < 0. 05). EGCG-treated group showed marked improvement of all these abnormal changes (P < 0. 05). Conclusions EGCG treatment is able to improve cognitive function, which may be attributed to ameliorated autophagic networks dysfunction and reduced Aß plaques in the the hippocampi of APP/PS1 transgenic mice.
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Objective To investigate the effect of ginsenoside Rg3 on immune checkpoint PD-L1 in Lewis cells (LLC) and to explore the related mechanism. Methods The effects of ginsenoside Rg3 on the proliferation of LLC were observed by MTT assay and cell long-term dynamic monitoring. LLC cells treated with 20 ng/mL IFN-γ were used to construct experimental model with high expression of PD-L1 in vitro. The effect of ginsenoside Rg3 on expression of PD-L1 was detected by flow cytometry and immunofluorescence. The effect of ginsenoside Rg3 on the protein expression of PI3K/Akt/mTOR pathway was verified by Western blotting. Results Ginsenoside Rg3 at 16, 32, 64, and 128 μmol/L significantly inhibited the proliferation of LLC (P < 0.01) and reduced the expression of PD-L1 induced by IFN-γ (P < 0.05). Ginsenoside Rg3 at 32 and 64 μmol/L significantly decreased the protein expression of PI3K and mTOR (P < 0.01), and ginsenoside Rg3 at 16, 32, and 64 μmol/L significantly inhibited the phosphorylation of Akt proteins (P < 0.05). Conclusion Ginsenoside Rg3 significantly inhibited the expression of PD-L1 in LLC cells by inhibiting PI3K/Akt/mTOR pathway. Ginsenoside Rg3 blocked the tumor cells escaping immune response by PD-L1 over-expression, enhanced the immune response of T cells, and inhibited the growth of non-small cell lung cancer cells.
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Platycodin D(PD) has a significantly inhibitory effect on multiple malignant tumors, and can inhibit the proliferation of leukemia cells K562 and induce apoptosis. However, its effect in improving the sensitivity of drug-resistant cells to imatinib and their molecular mechanism remained unclear. To investigate the effect and mechanism of PD alone or combined with imatinib (IM) in inhibiting CML imatinib resistant cell line K562/R, the cell proliferation was examined by CCK8 assay to reveal the effect of PD on the inhibitory function of imatinib. Cell apoptosis was detected by Annexin V-FITC/PI double staining. Protein expressions of cleaved caspase-3, cleaved caspase-9, PARP, cleaved PARP, Bcr/abl, p-AKT and p-mTOR were detected by Western blot. The results showed that the inhibitory effect of PD combined with imatinib on the proliferation and apoptosis of K562/R cells was significantly higher than that of the control group and the single drug group. Protein expressions of cleaved caspase-3, cleaved caspase-9 and cleaved PARP were significantly up-regulated in the combination group, and protein expressions of PARP, Bcr/abl, p-AKT and p-mTOR were down-regulated. The results indicated that PD increased the sensitivity of drug-resistant cells to imatinib, and the inhibitory effect of PD combined with imatinib was significantly better than the single drug on cell proliferation, induction of apoptosis, inhibition of Bcr/abl protein and PI3K/AKT/mTOR signaling pathway.
Subject(s)
Humans , Antineoplastic Agents , Pharmacology , Apoptosis , Cell Proliferation , Drug Resistance, Neoplasm , Imatinib Mesylate , Pharmacology , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive , Drug Therapy , Pathology , Saponins , Pharmacology , Signal Transduction , Triterpenes , PharmacologyABSTRACT
Objective To investigate the mechanism of phenylethanoid glycosides of pedicularis muscicola Maxim ameliorating high altitude memory impairmentby activating mTOR signal pathway.Methods 60 clean male Wistar rats were randomly divided into normoxic control group, hypoxia group, PhGs low, medium and high dose groups(50, 200, 400 mg/kg by oral administration).Normoxic control and hypoxia groups were administered with sterile injection water for 7 days.On the fourth day of drug treatment, hypoxia and PhGs groups were exposed to a specially designed animal decompression chamber, which simulated 7 500 m high altitude environment.The expression levels of mTOR, P70S6K and 4E-BP1 mRNA in hippocampus were detected by SYBR Green real-time PCR.The expression levels of p-mTOR, p-P70S6k and p-4E-BP1 protein in hippocampus were detected by Western blot.Results For hypoxia group rats, mTORand P70S6k mRNA repression, p-mTOR and p-P70S6K protein repression were respectively decreased by 22.50%, 26.00%, 42.28% and 11.70%(P<0.05, P<0.01), 4E-BP1 mRNA repression and p-4E-BP1 protein repression were respectively increased by 41.28%, 111.86%(P<0.01) in comparison tonormoxic control group.Compared with hypoxia group,for PhGs low dose group rats, 4E-BP1 mRNA repression and p-4E-BP1 protein repression were respectively decreased by 77.33% and 82.4%(P<0.01), p-P70S6K protein repression was increased by 32.53%(P<0.01).For PhGs medium, high dose groups, mTOR and P70S6k mRNA repression,p-mTOR and p-P70S6K protein repression were respectively increased by 64.56%, 60.76%;14.86%, 20.27%;65.12%, 94.17% and 56.63%, 78.31%(P<0.01), 4E-BP1 mRNA repression and p-4E-BP1 protein repression were respectively decreased by 72.67%, 71.57% and 57.6%, 40%(P<0.01).Conclusion Phenylethanoid glycosides of Pedicularis muscicola Maxim can ameliorate high altitude-induced memory impairment.This protective mechanism may due to the activation of mTOR signal pathway.
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OBJECTIVE Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disease with a high morbidity around 1/1000-1/400, characterized by progressive enlargement of fluid-filled cysts derived from renal tubular epithelial cells. Massive cysts gradually compress renal parenchyma destroying normal renal structures and compromising renal functions. Unfortunately, it will cause end-stage renal disease in most of the patients but without effective therapy now, who have to live on hemodialysis or kidney transplantation. Based on this present situation, it is of great significance to find early intervention to inhibit renal cyst development. The projective of this study was to investigate whether Ganoderma triterpenes (GT) can inhibit renal cyst development and study the related mechanism. METHODS and RESULTS First, we used MDCK cyst model, cultivated MDCK cells in vitro to form fluid-filled cysts surrounded by monolayer cells. GT inhibited MDCK cyst formation significantly, and inhibited cyst enlargement dose-dependently proving GT cyst inhibition in vitro. Then we used an embryonic kidney cyst model, wile-type mice kidneys were taken out on embryonic day 13.5 to form renal cysts stimulated with 8-Br-cAMP. GT inhibited embryonic kidney cyst development significantly in a dose-dependent and reversible manner proving GT cyst inhibition at organ level. Furthermore, we used two ADPKD mouse models with severe cystic kidney disease phenotypes. GT dramatically inhibited renal cyst development, decreased ADPKD mouse kidney volume and the cyst index inside proving GT cyst inhibition in vivo. By Western blot, we proved GT down-regulated Ras/MAPK signal pathway without detectable effect on mTOR signal pathway both in MDCK cells and two ADPKD mouse kidneys. CONCLUSION GT retard renal cyst development both in vitro and in vivo significantly. The related mechanisms were involved in GT promoting renal tubular epithelial cell differentiation, down-regulating intracellular cAMP level and Ras/MAPK signal pathway.
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AIM: To investigate the autophagy of breast cancer cells induced by baicalein and to explore its mechanism.METHODS: The effects of baicalein on the viability of MCF-7 cells and 4T1 cells were investigated by MTT assay, and the dosage of the drug was determined.The expression levels of microtubule-associated protein 1 light chain 3-II (LC3-II) and LC3-I in the MCF-7 cells and 4T1 cells treated with baicalein at doses of 25, 50 and 100 μmol/L, or combined with autophagy inhibitor 3-methyladenine (3-MA) were determined by Western blot.In order to confirm the role of baicalein in autophagy, the effect of 3-MA on the apoptosis of both MCF-7 cells and 4T1 cells induced by baicalein was analyzed by flow cytometry.The protein levels of p-mTOR, mTOR, p-AKT and AKT were examined by Western blot and the role of AKT-mTOR pathway in the induction of autophagy in breast cancer induced by baicalein was determined by the combination of activators.RESULTS: Baicalein at 50 μmol/L and above doses significantly inhibited the viability of breast cancer cells in a dose-and time-dependent manner.The expression of LC3-II/LC3-I in both MCF-7 cells and 4T1 cells was significantly enhanced after the action of baicalein, and the ratio of LC3-II/LC3-I was significantly decreased after 3-MA addition.The results of flow cytometry showed that, compared with baicalein group, the combination of baicalein and 3-MA promoted the levels of necrosis and apoptosis.Moreover, the protein levels of p-mTOR and p-AKT were significantly decreased and were rescued by EGF, while their total protein levels were not changed.CONCLUSION: Baicalein induces autophagy through AKT-mTOR pathway both in MCF-7 cells and 4T1 cells.
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The phosphoinositide-3 kinase (PI3K)/protein kinase B (PKB/AKT)/mammalian target of rapamycin (mTOR) pathway is asso-ciated with cell growth, proliferation, differentiation, apoptosis and metabolism. The abnormalities of this signal pathway are found in various malignant tumors. This pathway has also been investigated as an anti-tumor target. Recently, novel inhibitors have been stud-ied in clinical trials of lymphoma. This review summarizes the activation status of the PI3K/AKT/mTOR pathway and its use for targeted therapy of lymphoma.
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OBJECTIVE@#To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism.@*METHODS@#The effect of bortezomib on the viability of HeLa cell was measured by MTT assay. The effect of bortezomib on cell migration and invasion was measured by Transwell assay and invasion experiment respectively. The activation of Akt/mTOR signaling pathway and expression level of MMP2, MMP9 were assayed by western blot.@*RESULTS@#MTT assay indicated bortezomib (2.5 μM, 5 μM, 10 μM) could inhibit HeLa cell viability, and the inhibitory rate was highest at 48 h. Transwell assay and invasion experiment results showed that bortezomib inhibited HeLa cell migration and invasion. Western blotting assays presented bortezomib could suppress the phosphorylation of Akt and mTOR, and down-regulate the expression of MMP2 and MMP9.@*CONCLUSIONS@#These results suggested bortezomib could inhibit migration and invasion in cervical carcinoma HeLa cell, which might be related to Akt/mTOR signal pathway.
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Objective: To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism. Methods:The effect of bortezomib on the viability of HeLa cell was measured by MTT assay. The effect of bortezomib on cell migration and invasion was measured by Transwell assay and invasion experiment respectively. The activation of Akt/mTOR signaling pathway and expression level of MMP2, MMP9 were assayed by western blot. Results:MTT assay indicated bortezomib (2.5μM, 5μM, 10μM) could inhibit HeLa cell viability, and the inhibitory rate was highest at 48 h. Transwell assay and invasion experiment results showed that bortezomib inhibited HeLa cell migration and invasion. Western blotting assays presented bortezomib could suppress the phosphorylation of Akt and mTOR, and down-regulate the expression of MMP2 and MMP9. Conclusions:These results suggested bortezomib could inhibit migration and invasion in cervical carcinoma HeLa cell, which might be related to Akt/mTOR signal pathway.
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Objective: To explore the effect of bortezomib on migration and invasion of cervical carcinoma HeLa cell and specific molecular mechanism. Methods: The effect of bortezomib on the viability of HeLa cell was measured by MTT assay. The effect of bortezomib on cell migration and invasion was measured by Transwell assay and invasion experiment respectively. The activation of Akt/mTOR signaling pathway and expression level of MMP2, MMP9 were assayed by western blot. Results: MTT assay indicated bortezomib (2.5μM, 5μM, 10μM) could inhibit HeLa cell viability, and the inhibitory rate was highest at 48h. Transwell assay and invasion experiment results showed that bortezomib inhibited HeLa cell migration and invasion. Western blotting assays presented bortezomib could suppress the phosphorylation of Akt and mTOR, and down-regulate the expression of MMP2 and MMP9. Conclusions: These results suggested bortezomib could inhibit migration and invasion in cervical carcinoma HeLa cell, which might be related to Akt/mTOR signal pathway.
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Protein kinase B( Akt)is an intermediate signal molecule in PI3K-Akt-mTOR signaling pathway which plays an important role in development and incidence of colorectal cancer when activated by phosphorylation. As target of drugs,Akt has become a focus in the treatment of colorectal cancer. Clinical trials research proves that many kinds of Akt inhibitors have good antitumor activity. In recent years,the Akt inhibi-tors are more and more be taken seriously in colorectal cancer treatment.
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Phosphoinositide-3 kinase (PI3K)/protein kinase B (PKB/AKT)/mammalian target of rapamycin (mTOR) pathway is associated with cell proliferation, survival and migration. It plays an important role in the process of tumorgenesis. As a kind of antitumor target, numerous researches have focused on this pathway in recent years. This review summarizes the advances in regulation, protein expression and targeted therapy of PI3K-AKT/mTOR pathway for gastric cancer, and predicts the hot spots of this pathway in gastric cancer in the future studies. Copyright © 2014 by TUMOR.