Effect Of All-trans Retinoic Acid On The Differentiation, Invation And Metastasis In HepG2Hepatocellular Carcinoma Cell Line

ObjectiveBy far the most important kind of differentiation agent is All-trans-retinoicacid(ATRA), which has been successfully used in the leukemia cell differentiation.Current research showed that it also plays a role in some solid tumor. Nanog was closelyrelated to the tumors of genitourinary system and the differentiation of solid tumors.Furthermore, it expressed in the gastrointestinal tract and liver tumor too. We willinvestigate the effect of ATRA on the differentiation, invation and metastasis in HepG2hepatocellular carcinoma cell line, meanwhile, we will investigate the effect of ATRA oncancer stem cell differentiation gene Nanog, then to illustrate ATRA’s antitumor functionand its possible mechanism.MethodsThe proliferation of the HepG2was evaluated by MTT assay. Colony-formationassay in plate was used to test inhibition of colony-forming in HepG2by ATRA; ELISAwas used to analyse the changes of AFP after treatment with ATRA; the transcriptionlevels of Nanog and MMP9were assessed by reverse transcription-polymerase chainreaction(RT-PCR), and the protein level was assessed by western-blot; transwell andscratch test was used to detect the effects of ATRA on the invasion and migration inHepG2cells.Results1. ATRA suppresed the proliferation and anchoring-dependent growth inHepG2hepatocellular carcinoma cell lineMTT method showed that after treated with ATRA the proliferation of HepG2cells was suppressed in a dose and time-dependent; Colony-formation assay showed thatATRA also suppressed the anchoring-dependent growth of HepG2cells in adose-dependent manner.2. ATRA induced HepG2cells differentiationELISA showed that ATRA induced differentiation in HepG2cells, decreased thesecretion of AFP which represents hepatocellular malignancy.3. ATRA down-regulated MMP9’s expression in HepG2cellsRT-PCR and Western-blot indicated that after treated HepG2hepatocellularcarcinoma cell line with ATRA for24h, the mRNA and protein expression of MMP9was decreased in a dose-dependent manner.4. ATRA suppresed the invasion and metastasis in HepG2cellsThe scrach and transwell assay result suggested that after treated HepG2hepatocellular carcinoma cell line with ATRA for24h, the invasion and metastasis ofHepG2cells was prevented.5. ATRA down-regulated Nanog’s expression in HepG2cellsRT-PCR and Western-blot indicated that HepG2cells expressed nanog and ATRAcould down-regulate nanog’s mRNA and protein expression in a dose and time-dependent.Conclusion1.ATRA may induce HepG2cells differentiation, reduce the invasion and migrationcapabilities of HepG2.2.The mechanism of ATRA inducing HepG2cells differentiation may be associatedwith down-regulating the expression of cancer stem cell differentiation gene Nanog.