Effect Of The Rab27B On Hepatocellular Carcinoma Growth And Metastasis

Primary liver cancer is the fifth most frequently diagnosed cancer worldwide butthe third most frequent cause of cancer death.An estimated748,300new liver cancercases and695,900cancer deaths occurred worldwide in2008。Eighty-five percent ofcases occur in developing countries, half of these cases and deaths were estimated tooccur in China，and it is the fastest growing cause of cancer death in the United States.Among primary liver cancers, hepatocellular carcinoma (HCC) represents the majorhistological subtype, accounting for70%to85%of the total liver cancer burdenworldwide.Risk factors for HCC include hepatitis B, hepatitis C, alcohol ingestion, andcirrhosis from any cause. Many high risk patients are diagnostic for HCC even late fortreatment. Among various prognostic indicators of hepatocellular carcinoma diseaserecurrence and metastasis are key factors affecting the outcome of primary HCC therapy.Identification of specific and sensitive biomarkers for metastatic potential and theirregulatory genes in HCC is very significant for both early clinical detection and validprophylactic therapy.Tumors are ecosystems characterized by an intense communication between cancercells and stromal cells. These heterotypic interactions are a prerequisite for invasivegrowth and the final stage of multi-step tumor progression—metastasis. Communicationin the tumor ecosystem involves direct cell-cell interactions, but also the release ofsingle molecules (growth factors, cytokines, chemokines, proteases, matrix molecules)and membrane vesicles that enclose soluble factors within a lipid bilayer. The RABs,are the largest subfamily of the Ras superfamily of small GTPases is widely conservedin metazoan with more than60genes in the human genome, that regulate various types of intracellular membrane traffic in all eukaryotesStudies have shown that，Rab27B is expressed in breast cancer, stomach cancerand other tumor cells of epithelial origin, and related with breast cancer metastasis.Early study found that the expression of Rab27B was increased in HCC cells with highmetastatic ability, However，there has been no report about the relation and mechanismsunderlying Rab27B and HCC biological behavior researchTo systematically explore the expression,localization and function of Rab27B genein human human hepatoma cell lines. We determine the localization of Rab27A andRab27B in MHCC97H cells used by immunofluorescence assay and the Laser-ConfocalMicroscopy (CLSM) firstly,then western blot were used to quantitatively examine thedifferential expression of Rab27B protein in seven kinds of hepatocellular carcinomacells.In order to determine the correlation between Rab27B with HCC occurrence,development and metastasis, we screened stable knockdown or overexpressed Rab27Bcell line and the overexpress/decrease effect was identified by RT-PCR and westernblotting analysis compared with the negative control transfected cells, theexpression of Rab27B was obviously suppressed in MHCC97H cells transfected withRab27B siRNA at protein levels (P<0.05), and in MHCC97L cell overexpressionRab27B at protein levels (P<0.05)The ability of cell proliferation, adhesion, migration and invasion weresignificantly decreased in shRab27B cell compared with the negative control transfectedcells, while increased the endogenous expression of Rab27B and promote themigration of MHCC97L cell.The result indicating that the Rab27B is associated withHCC metastasis in vitro.To sum up, by using RNAi and gene over-expression technology,we establishedthe stable Rab27B knockdown MHCC97H cell,and overexpression Rab27B MHCC97Lcell. This research confirmed the Rab27B involved in HCC metastasis and will help study the effect of Rab27B on the regulatory pathway of cell phenotype as well as themolecular mechanism of HCC.