【摘要】：目的:探討海藻多糖(algal polysaccharides,AP)對肝癌細胞Hep3B的增殖和遷移的影響及其可能的作用機制,為治療肝癌提供新的思路。方法:(1)比較正常細胞與癌細胞中糖酵解(embden-meyerhof-parnas,EMP)途徑的表達差異,用紫外分光光度計比色法檢測EMP限速酶酶活力:己糖激酶(hexokinase,HK)、丙酮酸激酶(pyruvate kinase,PK),用乳酸測定試劑盒測定EMP產(chǎn)物:乳酸。(2)AP處理Hep3B后,檢測EMP表達水平變化。(3)AP處理細胞后,檢測肝癌細胞Hep3B的增殖和遷移能力及對上皮細胞-間充質轉化(EMT)的影響,方法包括MTT、q PCR和Transwell小室實驗。(4)MTT、Transwell和q PCR檢測HK抑制劑3-溴丙酮酸(3-bromopyruvate,3-BrPA)對肝癌Hep3B細胞的活力和遷移能力及EMT的影響。(5)Western blot和相關試劑盒檢測AP與3-BrPA分別處理細胞時,對EMP水平及Akt通路的影響。(6)AP聯(lián)合3-BrPA處理Hep B3后,檢測HK和Akt信號通路表達水平及細胞活力和遷移的變化。結果:(1)癌細胞(Hep3B、He La、SW480)EMP代謝水平均高于正常肝細胞(HL02),其中Hep3B差異最為顯著。(2)AP能抑制Hep B細胞EMP代謝水平,且抑制程度隨著AP濃度升高而升高,存在濃度依賴性。(3)AP可抑制肝癌細胞Hep3B的增殖和遷移,且抑制EMT的發(fā)生。AP濃度為200mg/ml,處理時間為48h時,生長抑制率達(55±2.8)%,細胞遷移數(shù)為對照組的(32±2.9)%;(4)3-BrPA可下調Hep3B細胞HK活性,并抑制細胞活力與遷移,且抑制EMT的發(fā)生。200μmol/L 3-BrPA作用細胞48h后,與對照組相比,細胞活性下降了(52±5.8)%(P0.001),細胞遷移數(shù)下降了(48±6.1)%(P0.01)。(5)AP和3-BrPA均下調EMP代謝水平,且抑制Akt信號通路。(6)AP與3-BrPA聯(lián)合處理Hep3B后,HK表達下降,顯著抑制Akt信號通路,細胞活性和遷移能力均較AP單用組顯著下降(P0.05)。結論:肝癌細胞Hep3B EMP代謝水平高于正常肝細胞,AP可下調Hep3B細胞EMP代謝水平,低EMP代謝水平可能通過下調EMT和Akt信號通路抑制Hep3B的增殖和遷移。當AP和3-BrPA聯(lián)合應用時,抑制肝癌遷移和增殖效果更明顯。
[Abstract]:Objective: to investigate the effect of seaweed polysaccharide (algal polysaccharides,AP) on the proliferation and migration of HCC cell line Hep3B and its possible mechanism, so as to provide a new idea for the treatment of HCC. Methods: (1) the expression difference of glycolysis (embden-meyerhof-parnas,EMP) pathway between normal cells and cancer cells was compared. The activities of EMP rate-limiting enzyme were detected by ultraviolet spectrophotometer: hexokinase (hexokinase,HK), pyruvate kinase (pyruvate kinase,PK), EMP product: lactic acid was measured by lactic acid assay kit. (2) after Hep3B treatment with AP, the expression level of EMP was detected. (3) after AP treatment, the expression level of EMP was detected. The proliferation and migration ability of hepatocellular carcinoma cell line Hep3B and the effect of MTT,Transwell and Q PCR on the proliferation and migration of HCC cells and the transformation of EMT into epithelial cells were detected by MTT,Transwell and Q PCR. (5) Western blot and related kits were used to detect the effect of AP and 3-BrPA on the activity and migration of HCC Hep3B cells, and the effect of HK inhibitor 3 bromopyruvate, 3 鈮，

本文編號：2519751