【摘要】：目的:探討海藻多糖(algal polysaccharides,AP)對(duì)肝癌細(xì)胞Hep3B的增殖和遷移的影響及其可能的作用機(jī)制,為治療肝癌提供新的思路。方法:(1)比較正常細(xì)胞與癌細(xì)胞中糖酵解(embden-meyerhof-parnas,EMP)途徑的表達(dá)差異,用紫外分光光度計(jì)比色法檢測(cè)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(cè)定試劑盒測(cè)定EMP產(chǎn)物:乳酸。(2)AP處理Hep3B后,檢測(cè)EMP表達(dá)水平變化。(3)AP處理細(xì)胞后,檢測(cè)肝癌細(xì)胞Hep3B的增殖和遷移能力及對(duì)上皮細(xì)胞-間充質(zhì)轉(zhuǎn)化(EMT)的影響,方法包括MTT、q PCR和Transwell小室實(shí)驗(yàn)。(4)MTT、Transwell和q PCR檢測(cè)HK抑制劑3-溴丙酮酸(3-bromopyruvate,3-BrPA)對(duì)肝癌Hep3B細(xì)胞的活力和遷移能力及EMT的影響。(5)Western blot和相關(guān)試劑盒檢測(cè)AP與3-BrPA分別處理細(xì)胞時(shí),對(duì)EMP水平及Akt通路的影響。(6)AP聯(lián)合3-BrPA處理Hep B3后,檢測(cè)HK和Akt信號(hào)通路表達(dá)水平及細(xì)胞活力和遷移的變化。結(jié)果:(1)癌細(xì)胞(Hep3B、He La、SW480)EMP代謝水平均高于正常肝細(xì)胞(HL02),其中Hep3B差異最為顯著。(2)AP能抑制Hep B細(xì)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(xì)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(shí)間為48h時(shí),生長(zhǎng)抑制率達(dá)(55±2.8)%,細(xì)胞遷移數(shù)為對(duì)照組的(32±2.9)%;(4)3-BrPA可下調(diào)Hep3B細(xì)胞HK活性,并抑制細(xì)胞活力與遷移,且抑制EMT的發(fā)生。200μmol/L 3-BrPA作用細(xì)胞48h后,與對(duì)照組相比,細(xì)胞活性下降了(52±5.8)%(P0.001),細(xì)胞遷移數(shù)下降了(48±6.1)%(P0.01)。(5)AP和3-BrPA均下調(diào)EMP代謝水平,且抑制Akt信號(hào)通路。(6)AP與3-BrPA聯(lián)合處理Hep3B后,HK表達(dá)下降,顯著抑制Akt信號(hào)通路,細(xì)胞活性和遷移能力均較AP單用組顯著下降(P0.05)。結(jié)論:肝癌細(xì)胞Hep3B EMP代謝水平高于正常肝細(xì)胞,AP可下調(diào)Hep3B細(xì)胞EMP代謝水平,低EMP代謝水平可能通過下調(diào)EMT和Akt信號(hào)通路抑制Hep3B的增殖和遷移。當(dāng)AP和3-BrPA聯(lián)合應(yīng)用時(shí),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(hào)：2519751