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OBJECTIVE To investigate the effects of Setaria italica extract on improving insomnia model mice and to explore its potential mechanisms. METHODS The mice were randomly assigned into blank group， model group， positive control group （diazepam， 2.6 mg/kg）， and S. italica extract low-dose， medium-dose and high-dose groups （1.2， 2.4， 4.8 g/kg）， with 10 mice in each group. Except for the blank group， all other groups received intraperitoneal injection of para-chlorophenylalanine （PCPA） to establish the insomnia model. After modeling， the blank group and model group were given a constant volume of normal saline intragastrically， and administration groups were given relevant medicine intragastrically， with a volume of 0.01 mL/g， once a day， for 7 consecutive days. After the administration， the open-field test was conducted to observe the praxiological changes of mice， and to determine the levels of 5-hydroxytryptamine （5-HT） and 5-hydroxyindoleacetic acid （5-HTAA） in the hippocampal tissue， as well as the contents of 5-HT， brain-derived neurotrophic factor （BDNF）， interleukin-2 （IL-2）， IL-6， B-cell lymphoma-2 （Bcl- 2）， and Bcl-2-associated X protein （Bax） in the serum. The expression of phosphoinositide 3-kinase/protein kinase B/nuclear factor- κB （PI3K/Akt/NF-κB） signaling pathway related protein was determined in the hippocampus of mice. RESULTS Compared with the model group， the total exercise time of mice in S. italica extract high-dose group was significantly prolonged， but the total rest time was significantly shortened （P＜0.01）； the number of standing times and modification times were significantly reduced （P＜ 0.01）. The contents of 5-HT， BDNF， and Bcl-2 in serum， and Bcl-2/Bax were significantly increased， while the contents of IL-2， IL-6， and Bax were significantly reduced （P＜0.05 or P＜ 0.01）. The content of 5-HTAA in the hippocampal tissue and 202104010910029）；the phosphorylation levels of PI3K and Akt proteins were increased significantly， while the phosphorylation level of NF-κB p65 protein was decreased significantly （P＜0.05）.CONCLUSIONS High-dose of S. italica extract demonstrates significant therapeutic effects on insomnia in mice， and the mechanism of which may be associated with the regulation of PI3K/Akt/NF-κB signaling pathway.
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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 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.
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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.