The Effect Of MALAT1in Cervical Cancer Cell Biology Phenotype

Background and objectiveIn recent years, the incidence of cervical cancer has a clear upward trend in certain areas and the patients were getting younger and younger. Studies have shown that the incidence of cervical cancer is closely related to persistent infection of high-risk human papillomavirus (human papillomaviruses, HPV). HPV infection is usually short and can be cleared by the body in women, therefore, there are some deficiencies in HPV test when identify persistent infection and temporary infection, so HPV is not a goog indicator for monitoring cervical cancer. Traditional method of surgery, radiotherapy and chemotherapy all exist certain limitations in cervical cancer therapy, so, it is necessary and urgent to study the mechanisms to find new target molecules that can be used to treat and monitor cervical cancer.lncRNA is a class of non-translating RNA, which could regulate gene expression in the epigenetic level, transcriptional level and post-transcriptional level and participated broadly in almost all physiological and pathological processes of body. We discovered an up-regulating lncRNA MALAT1in our previous space mutagenized cervical cancer cells. MALAT1is located on chromosome11q13with a length of8708bp. MALAT1is closely related to a variety of tumors and regulating their development, while the relationship of MALAT1and cervical cancer is still not reported in other laboratories. This study will detect the expression of MALAT1in cervical cancer cell lines and cervical epithelial cell samples by RT-PCR; establish a stable model of MALAT1silence cell lines using RNAi technology and stable MALAT1over-expressing cell line models using over-expression vector, then study the affection of MALAT1on cervical cancer cell biology phenotype through both positive and negative aspects; further explore MALAT1downstream molecules related to cell prolification.Method1. After collecting cervical cells and cervical epithelial cell samples, extracting RNA, using RT-PCR method to detect MALAT1expression, then analyzing the correlation of MALAT1expression and cervical cancer.2. Using MALAT1over-expression vector pEGFP-C1transfecting HaCat cell and SiHa cell, using MALAT1silencing vector pRNAT-U6.1/Neo transfecting CaSki cell, then positive stable cell lines were established by G418election and limiting dilution method. Cell proliferation assay, colony formation assay, wound healing assay and transwell assay were applied to detect MALAT1effect on cervical cancer cell biology phenotype.3. Cell cycle was analyzed after MALAT1change by flow cytometry. RT-PCR approach was used to explore cell cycle related molecules that related with MALAT1in regulating cell proliferation.Results1. MALAT1expressed highly in Hela, CaSki, SiHa, HCC94and cervical cervical epithelial lesion cells, but not in HaCat cells and normal cervical epithelial cells sample. It indicated that MALAT1was correlated to the development of cervical cancer.2. By double digestion and DNA sequencing identification, MALAT1over-expression vector pEGFP-C1/M1was successfully constructed. We successfully established stable MALAT1silence or over-expressing cell lines after transfecting corresponding vector; MALAT1silencing decreased cell growth, colony formation and cell metastatic ability; MALAT1over-expression didn’t impact cell growth but increased cell metastatic ability.3. After MALAT1silencing, G1phase cells were increased, and S phase cells were decreased. The expression of cyclinDl, cyclinE and CDK6were significantly reduced, CDK4did not change significantly, indicating that MALAT1affect cervical cancer cells proliferation by regulating G1/S phase.Conclusion1. MALAT1expressed highly in cervical cancer and was correlated with the development of cervical cancer. 2. MALAT1promotes cell proliferation and migration in cervical cancer cells.3. MALAT1regulates cell proliferation through change cell cycle related molecules.