| Cancer is one of the most important causes of death in the world,so scientists are trying their best to deal with it.We all know that the mutations of oncogenes and anti-oncogenes are the underlaying mechanism for tumorigenesis.The field of epigenetics has recently moved to the forefront of cancer studies.DNA methulation is the most common form of it.Recent work has revealed that the hypermethylation of CpG islands usually appear earlier than cell's hyperplasia,the global hypomethylation of genes happens within cancer organism.Methylation would not alter the sequence of DNA,but covalently add a methyl group to the 5-posisiton of cytosine,this can be reversed by demethylation drugs.Detection of changes in DNA methylation can be used for diagnosis and prognosis,and the CpG islands may become the treating targets.So searching the related genes has become the hot point of recent research.Homo sapiens LIM homeobox 6,LHX6,at 9q33.2 site of human chromosome, belongs to the LIM family.As a transcription factor,it can transfer signal between DNA and proteins.It is related to the teeth development and the interneurons migration from the ventral telecephalon to the neocortex.But we know little about how the gene is related to cancer.In MRH Estecio's studies,LHX6 is a sensitive methylation marker in head and neck carcinomas,2006.So we interested in LHX6,and expect it can be a methylation cancer marker in cancer cells.Based on these reports,we found that there are some CpG islands in promoter region,methylation of these CpG islands may impact on the expression of the gene.So we chose this area to study,useing Ms-PCR to analysis the methylation in A549,Hela,HepG2 and MCF-7 cell lines.And then we treated these cells with 5-Aza-CdR,a demethylation drug.Because the effect of 5-Aza-CdR on cancer cells is based on its concentration of it,we detect the effective concentration of 5-Aza-CdR on these cells.We observed the shape of the cells which has been treated with the best concentration of 5-Aza-CdR for 72 hours,and detect the proliferation of them by MTT assay.And analyse the proliferations of cells by MTT tests,using RT-PCR to detect the expression of LHX6,using MS-PCR to analyse the different levels of the gene's methylation.We found different levels of methylation in these four cell lines.A549 has a higher methylation level when compared with the others.We optimized 1×10-6mol/L as the best 5-Aza-CdR treatment concentration.Comparing with the control,the proliferation of these cells witch are treated by 5-Aza-CdR has been reduced.A549 cell's growth rate has reduced by 72.2%.Observing the shape through×100 microscope,the shape of the cells treated with 5-Aza-CdR has changed in different levels.RT-PCR results showed that the re-expression of LHX6 only can be found in 5-Aza-CdR treated A549 cells,but can not found in other cells,neither the control nor the samples.At the same sample of the A549 cells,we found the methylation level has been reduced.So there is relationship between re-expression of the gene LHX6 and the reduction of proliferation in A549 cells.In conclusion,we found A549 may have some methylation genes,which can be inhibited by 5-Aza-CdR significantly,and has relationship to the proliferation of the cells.LHX6 has CpG islands in its gene,and has hyper-methylation in A549 cells. De-methylation can re-expresse it,and this is related to the depressing of cell proliferation.So,LHX6 maybe one of the potential cancer marker in A549 cell line,and it maybe used for searching new methods for diagnosing and treating cancer.
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