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Salidroside (SAL), a major bioactive compound of Rhodiola crenulata, has significant anti-hypoxia effect, however, its underlying molecular mechanism has not been elucidated. In order to explore the protective mechanism of SAL, the lactate dehydrogenase (LDH), reactive oxygen species (ROS), superoxide dismutase (SOD) and hypoxia-induced factor 1α (HIF-1α) were measured to establish the PC12 cell hypoxic model. Cell staining and cell viability analyses were performed to evaluate the protective effects of SAL. The metabolomics and bioinformatics methods were used to explore the protective effects of salidroside under hypoxia condition. The metabolite-protein interaction networks were further established and the protein expression level was examined by Western blotting. The results showed that 59 endogenous metabolites changed and the expression of the hub proteins of CK2, p-PTEN/PTEN, PI3K, p-Akt/Akt, NF-κB p65 and Bcl-2 were increased, suggesting that SAL could increase the expression of CK2, which induced the phosphorylation and inactivation of PTEN, reduced the inhibitory effect on PI3K signaling pathways and activated the PI3K/Akt/NF-κB survival signaling pathway. Our study provided an important insight to reveal the protective molecular mechanism of SAL as a novel drug candidate.
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Objective To investigate the effect of Wulong Xiaozheng Pill (WXP) on the migration and invasion of human gastric cancer cell-line BGC-823 and its mechanism. Methods WXP, IGF-1, and LY294002 were added on BGC-823 cells. Then the inhibitory effect of WXP was detected by MTT assay. Transwell assay was performed to determine the migration and invasion capacity of on BGC-823 cells. Expressions of VEGF, MMP-2, and MMP-9 were detected by ELISA, while the expressions of related proteins and mRNA in PI3K/NF-κB signaling pathway were detected by Western blotting and RT-PCR. Results WXP can inhibit the proliferation, adhesion, invasion, and migration of BGC-823 cells. In addition, WXP inhibited the expression of VEGF, MMP-2 and MMP-9 protein in BGC-823 cells. WXP significantly inhibited the expression of p-PI3K, p-Akt, p-IKK-a, and p-NF-κB p65Ser 276 proteins, PI3K, Akt, IKKa, and NF-κB mRNA, which showed a time-dependent and dose-dependent manner. Conclusion WXP inhibit the capacity, migration and invasion of BGC-823 cells by blocking PI3K/NF-κB signaling pathway.
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Objective:To evaluate the effects of dexamethasone on systemic lupus erythematosus complicated with cognitive dysfunction.Methods:Ten wild type mice and 20 MRL/lpr mice were applied for the research.MRL/lpr mice were randomly assigned to a MRL/lpr group and a MRL/lpr + dexamethasone (1.5 mg/kg) group.Interleukin-6 (IL-6),IL-1β,and tumor necrosis factor alpha (TNF-α) in serum and hippocampus were detected.The protein phosphorylation levels of phosphoinositide 3-kinase (P-PI3K),protein kinase B (P-Akt),NF-kappa-B inhibitor alpha (P-IκBa) and nuclear transcription factor kappa-B p65 (P-NF-κB p65) were detected by Western blot,the level of P-NF-κB p65 also was detected by immunohistochemistry.Results:Treatment with dexamethasone (1.5 mg/kg) alleviated the cognitive dysfunction and decreased the levels of IL-6,IL-1 β and TNF-α in serum and hippocampus,and reduced the levels of P-PI3K,P-Akt,P-IκBa and P-NF-κB p65 in hippocampus in MRL/lpr mice.Conclusion:Dexamethasone may play a protective role in the cognitive function by decreasing the levels of TNF-α and IL-1 β in the hippocampus of MRL/lpr lupus mice.
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Objective To study the effect and mechanism of Water extract from Jiangtang Decoction (WEJTD) on diabetes mellitus and diabetic nephropathy (DN) in spontaneous type 2 diabetes mellitus model KK-Ay mice.Methods Totally 50 KK-Ay mice were randomly divided into five groups:model group,metformin (positive drug,250 mg/kg) group,WEJTD low,medium and high dose (2,4,and 8 g/kg) group,with 10 C57BL/6J mice as normal group.The relative drugs were ig administered once a day for 12 weeks,and mice in control group and model group were perfused with distilled water of equal volume.After 12 weeks' oral administration,mice were put into metabolism cages,and the food-intake,water-intake and urine volume were calculated and collected.Blood were collected to detect the concentration of IL-6,ICAM-1 and TNF-α.Then mice were executed,and HE staining and PASM staining were used to check the effect of WEJTD on kidney.Western blotting and qRT-PCR were used to detect the concentration of PI3K,Akt,NF-κB,IL-6,ICAM-1 and TNF-α in kidney.Results WEJTD can alleviate the symptoms of diabetes,such as food ration,polydipsia and polyuria (P ＜ 0.05,0.01,and 0.001);Relief the pathological changes of kidney and significantly decreased glycogen deposition (P ＜ 0.001),down-regulate the increase of IL-6,ICAM-1 and TNF-α in serum and kidney (P ＜ 0.05,0.01 and 0.001),up-regulate the phosphorylation of PI3K and Akt (P ＜ 0.05,0.01,and 0.001),and inhibit the phosphorylation of NF-κB (P ＜ 0.001).Conclusion WEJTD had positive effects on kidney morphology of KK-Ay,and the underlying mechanism might be related to the regulation of PI3K-Akt and NF-κB-mediated inflammation.
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ABSTRACT:Objective To explore the expressions of Toll-like receptor4/NF-κB and PI3K/AKT/NF-κB signa-ling pathways in rat ulcerative colitis (UC)induced by the combined enema of trinitrobenzene sulphonic acid and ethano and the interventional effect of electroacupuncture on them.Methods Totally 240 male Wistar rats were randomly divided into 6 groups:normal control group,model control group,electroacupuncture group,TLR4mAb group,LY294002 group,and TLR4mAb combined with LY294002 (T&L)group.The combined enema of trinitro-benzene sulphonic acid (TNB)and ethanol was intrarectally administered for 4 weeks to induce UC.At the same time of modeling ,Zusanli point was electro-acupunctured in electroacupuncture group while intraperitoneal injec-tion of TLR4mAb and LY294002 was given respectively to the corresponding group.Each rat was treated with the above-mentioned TLR4mAb injection and LY294002 injection in T&L group for 4 weeks.The disease activity index (DAI)of all the rats was evaluated daily.The rats were killed after 4 weeks.The colonic mucosa damage index (CMDI)and tissue damage index (TDI)were evaluated by a pathologic grading system.The expressions of P-Akt and active NF-κB protein in the colon mucosa were determined by Western blotting.TLR4 mRNA,PI3K mRNA, AKT mRNA,NF-κB mRNA,TNF-αmRNA and IL-1βmRNA expressions were measured with RT-PCR.Results Compared with those in normal control group,TLR4 mRNA,PI3K mRNA,P-AKT,active NF-κB,TNF-αmRNA and IL-1βmRNA expressions as well as DAI,CMDI and TDI were all increased obviously in model control group (P <0.01).Compared with those in model control group,TLR4mRNA expression was decreased obviously in TLR4mRNA group (P <0.01),the expressions of PI3KmRNA and P-AKT were decreased obviously in LY294002 group (P <0.01 ).Not only TLR4mRNA expression but also PI3KmRNA and P-AKT expressions were decreased significantly in electroacupuncture group and T&L group (P <0.01 ).Corresponding to the above-mentioned chan-ges,active NF-κB,TNF-αmRNA and IL-1βmRNA expressions as well as DAI,CMDI and TDI were decreased obvi-ously in all the treated groups compared with those in model control group (P <0.05 or P <0.01),but the six inde-xes were better in electroacupuncture group and T&L group than in TLR4mAb group and LY294002 group (P <0.05).There were obvious positive correlations of active NF-κB with TNF-αmRNA and IL-1β mRNA expressions (r 1 =0.579,P <0.05;r 2 =0.561,P <0.05).Conclusion Electroacupuncture can significantly decrease NF-κB activity and TNF-αmRNA and IL-1β mRNA expressions in UC rats,thus alleviating the severity of UC,which is closely correlated to its blocking both TLR4/NF-κB and PI3K/AKT/NF-κB signaling pathways.
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Objective: To investigate the effects of chemokine (C-X-C motif) ligand 8 (CXCL8) gene silencing on cell proliferation and invasion of human colon cancer cells, and to investigate its relationship with phosphatidylinositol 3-kinase/protein kinase B/nuclear factor-kappa B (PI3K/Akt/NF-κB) signaling pathway. Methods: The expression levels of CXCL8 mRNA and protein in four human colon cancer HT-29, WiDr, CaCo-2 and CoLo320 cells were detected by reverse transcription-PCR and Western blotting, respectively. The four colon cancer cells were transfected with the small interfering RNA targeting CXCL8 gene (CXCL8 siRNA) by LipofectAMINE 2000, then the expression of CXCL8 protein was detected by Western blotting. The proliferation and invasion abilities of colon cancer cells after CXCL8 gene silencing were detected by WST-1 method and Transwell invasion assay, respectively. The phosphorylation levels of PI3K, Akt and NF-κB proteins in HT-29 cells with CXCL8 gene silencing were detected by Western blotting. Results: CXCL8 mRNA and protein were highly expressed in four colon cancer cell lines. After CXCL8 siRNAs were transfected into the colon cancer cells, the expression of CXCL8 protein was significantly inhibited (all P < 0.01), while the proliferation and invasion abilities of colon cancer cells were significantly decreased (all P < 0.01). CXCL8 gene silencing resulted in blockage of PI3K, Akt and NF-κB protein phosphorylation induced by CXCL8 in colon cancer HT-29 cells (all P < 0.01). Conclusion: CXCL8 gene silencing significantly inhibits the proliferation and invasion of colon carcinoma cells. It may be related to down-regulation of protein activity in PI3K/Akt/NF-κB signaling pathway.