RKIP Low Expression In Gastric Cancer And Its Effect In Migration

Background&ObjectiveCurrently, gastric cancer is one of the most frequent cancers of the digestivesystem in our country and even around the world and has a very high morbidity andmortality.More than70%of gastric cancer occure in resource-poor developingcountries[1]. Since the early symptoms of gastric cancer is not obvious, when theclinical symptom appears, most of it has been in advanced stage, the treatment is poor.Therefore, the key factor in reducing the mortality of gastric cancer is early detection,early diagnosis and early treatment. Generally, the incidence of cancer ismultifactorial participation, multi-channel, multi-step process, is the result ofenvironmental factors and genetic factors working together. In recent years, the studyof epigenetic mechanisms in gastric cancer conformed that the inactivation of tumorsuppressor gene is closely related to the occurrence and development of gastriccarcinoma, this shows that the silencing of tumor suppressor genes is a major changein the molecular biology[2-5].In recent years, due to the discovery of RKIP play an important role in cellsignal transduction pathways, especially after the latest RKIP was found to inhibittumor cell metastasis[6], The research of RKIP has become a new research focus inthe field of tumor cell biology. RKIP is a multifunctional protein highly conserved,exists in in the human body widely, the human RKIP is located on the NO.12chromosome q24.22, contains4exons, the transcription of mRNA up to1434bp, encodes a187amino acid protein[7]. RKIP is a natural inhibitor of themitogen-activated protein kinase (Mitogen-activated protein kniase, MAPK) signaltransduction pathway. Yeung[8]firstly reported in Nature journal in1999that can becombined with Raf-1, thereby inhibiting Raf-1/MEK/ERK signaling pathway,therefore, it was renamed RKIP. In addition to inhibiting Raf-1/MEK/ERK RKIPsignaling pathway, RKIP is also involved in the G protein-coupled receptor signalingpathway[9]and NF-kB signaling pathway[10]regulation. Because the signaling pathwayis a major regulator of cell growth,differentiation,proliferation and apoptosis,therefore it is associated with tumor progression and metastasis, it has an importantrole in the development and prognosis of cancer, and more closely related to poorprognosis in cancer[11]. It was reported that RKIP can inhibit the metastasis of prostatecancer, breast cancer and melanoma cells,and when the expression of RKIPdownregulation or absent was related to metastasis and poor prognosis of prostatecancer, breast cancer and melanoma and other human malignancies[12,13]. However,the role of RKIP in gastric cancer (GC) metastasis has not been reported, so therelationship between the development of gastric cancer and RKIP still requires a lot ofscientific study to be further explored. Our aim is to study the relationship betweenRKIP and the occurrence,development of gastric cancer, as well as changes in theexpression levels of RKIP effect on GC cells in vitro invasion.Materials&Methods1.74cases of gastric cancer and its corresponding adjacent tissues (from foci greaterthan5cm) were taken surgically from December2011to February2013in ourhospital. All specimens from patients with informed consent, approved by the hospitalethics committee and reported. And no preoperative chemotherapy, radiotherapy andother biological treatments., and all patients signed informed consent as proof.Aftersurgical resection we wash away the blood of specimens with saline at4℃,thenspecimens are put into-80℃fridge to preserve．2. Human gastric cancer cell line SGC-7901provided by Basic Medical College ofZhengzhou University is moderately diffrentiated gastric adenocarcinoma cell. Theycome from a56-year-old woman whose cancer tissue of metastatic adenocarcinomalymph nodes. Cells culture at37°C,5%CO2saturated humidity conditions, withDMEM medium containing10%FBS.3. We use reverse transcription-polymerase chain reaction detection (RT-PCR) and Western blotting (Western Blot) to detect RKIP mRNA and the protein in the abovetissues． Plasmids that expressed sense and antisense RKIPcDNA byliposome transfection method and its corresponding empty vector were stablytransfected into SGC-7901cells,then the result of transfectiong was detected byWestern bloting. The effects of RKIP expression on in vitro cell invasion wereanalyzed in the transfected cells by Transwell cell migration assay.4. All the experimentaldata are used SPSS17.0statistical software for statisticalanalysis. The gray values of RKIP mRNA between gastric cancer tissues and adjacenttissues were compared with paired t-test. Using χ2test and Fisher exact probabilitiestest to analysis correlation between RKIP and clinical pathological factors. There werestatistically significant differences in P <0.05Results:1. The relative value of RKIP mRNA expression in gastric cancer tissues is0.12±0.02, while the relative value of RKIP mRNA expression in adjacent tissues is0.48±0.04, t=6.562,(P<0.01). The statistical analysis showed that the expressionlevel of RKIP mRNA in patients has a closely relationship with histologicdifferentiation, and TNM stage and lymph node metastasis, but has noting to do withgender, age, tumor size,(all P <0.05).2. The relative value of RKIP protein expression in gastric cancer tissues is0.08±0.02, while the relative value of RKIP mRNA expression in adjacent tissues is0.41±0.04, t=6.562,(P<0.01). The statistical analysis showed that the expression ofRKIP protein is related to histologic differentiation, lymph node metastasis and stagesof TNM in gastric cancer，regardless of gender, age, tumor size (all P <0.05).3. Transwell cell migration assay showed that upregulated the expression of RKIP inSGC-7901cells could decrease its in vitro cell invasion, whereas downregulated theexpression of RKIP in SGC-790cells could increase its in vitro cell invasion.Conclusions:1. The expression of RKIP mRNA was significantly down-regulated or loss ingastric cancer tissues.which was closely related to histologic differentiation, andTNM stage and lymph node metastasis, While the expression of RKIP mRNA wasobviously overexpression or normal in adjacent tissues.It points that the occurrence,development of gastric cancer may be associated with abnormalexpression of RKIP on gastric cancer.It reveals that RKIP may be a potentialtherapeutic target that could have a profound impaction prevention of metastasis ingastric cancer patients.2. Downregulated the exprssion of RKIP in SGC-7901could increase its in vitro cellinvasion, however upregulated the expression of RKIP in SGC-7901could decreaseits in vitro cell invasion. RKIP expression in gastric cancercells is negativly related toits in vitro cell invasion.