The Expression Of Ndrg2 Gene In HHCC And The Preliminary Studies On Its Function

The Ndrg2 gene(N-myc downstream regulated gene 2 )was first discovered and cloned from normal human whole brain cDNA library in 1999. This gene locates on human chromosome 14qll. 2, including 16 exons and 15 introns, and its full mRNA is 2, 024bp in length, encoding the 357 amino acid , 40KD Ndrg2 protein. Ndrgl was the first member found of Ndrg gene family, which was isolated in N-myc mutant mouse embryos and found be repressed by N-myc and c-myc. Human Ndrg gene family consists of 4 members: Ndrgl, Ndrg2, NdrgS, Ndrg4, which has high similarity in nuclear acid or amino acid sequence. However, their expression levels in spatio-temporal distribution and periods of cells and tissues development are distinct.The knowledge about the function of the Ndrg gene family camemainly from the studies on Ndrgl. Now, many intracellular and extracellular factors have been found which could regulate the expression of Ndrgl. Homocystein, tunicamycin, p53, retinoic acid, oxidative and stress condition such as hypoxia, nickel and other reductive agents, et al, can upregulate Ndrgl expression, while the androgen, proto-oncogene N-myc and c-myc can repress its expression. Inferring from all the references obtained, the functions of Ndrggene family may be ' involved in cellular differentiation events, maintaining the balance of cell redox potential, and counteracting cell malignant transformation and so on.Our preliminary studies showed that the mRNA expression patterns of Ndrg2 geue was dominant in the white matter, gray matter and neuron of brain, salivary gland, skeletal muscle, and lower in bone marrow, testis, peripherial blood and placenta, however did not express in leukocyte, colorectal cancer and some tumor cell line such as Hela S3, leukemia cell, Burkitt's lymphoma, adenocarcinoma SW-480, all above suggested that Ndrg2 gene had the relationship with tumor, might be a new suppressor gene candidate. So, to study its function and regulation has important theoretical meaning and foreseeing clinical value, but up to now .there have been no reports about the precise cellular and molecular function of Ndrg2 gene. Our research is aimed to observe the expression of Ndrg2 gene in HHCC and explore its preliminary function.The major findings of the present work are as follows: 1. Semiquantitative RT-PCR analysis indicated that Ndrg2 gene expression in HHCC was lower than that in paracancerous normaltissue, which suggested that Ndrg2 gene might be involved in the carcinogenesis and development of HHCC, but there was no relationship between the expression of Ndrg2 gene and the Edmondson grades and the DNA multiploid.2. Immunohistochemistry indicated that the Ndrg2 protein located in cytoplasm.3. The sense pIRES2-EGFP-ndrg2 vector and antisense pIRES2-EGFP-anti-ndrg2 vector was successfully constructed, and the HepG2 cell, which expressed Ndrg2 gene negatively, was sifted as the experimental group.4. Transient transfection of Ndrg2 gene induced G1 phase arrest and apoptosis in HepG2 cells after 48h, while there was no obvious changes appearing in QZG cells after transient gene tranfection with antisense Ndrg2 gene.5. The EGFP-ndrg2-HepG2 cells were obtained after stable gene transfecttion .Its growth, proliferation, cloning-forming rate and carcinogenic ability were lower than that of HepG2 cells. These suggested that Ndrg2 gene could arrest the growth of tumor cells, and mainly acted in the transformed cells.These findings about the different expression of Ndrg2 gene in HHCC and paracancerous normal tissue, the function of inhibiting the growth of HepG2 cells suggest that Ndrg2 gene play an role in HHCC development.