Study On The Role Of Leucine-rich Glioma Inactivated Gene-1

ObjectiveTo investigate the loss of heterozygosity (LOH) of leucine-rich glioma inactivated gene-1(LGI1) in human glioma specimens and glioma cell lines as well as construct the plasmid of pcDNA3.1/LGI1 and research the relationship between the LGI1 gene and the glioma pathogenesis. Material and method1. The fresh specimens of tissue and blood of 25 the glioma patients were collected from the Neurosurgery Departments of Tianjin First Central Hospital and TianJin Huanhu Hospital. 1 fresh brain tissue specimen of human was also gained from decompression operation. Therefore, 5 glioma cell lines (C6, TJ905, A172, U251 and H4) were used in present study.2. Genomic DNA was extracted from all of the above specimens and glioma cell lines. Special primers were designed according to the reference. The PCR products were detected with the methods of electrophoresis and silver staining. If any LOH in banding patterns was found, the DNA sequence analyse was made directly. The expressions of LGI1 mRNA and protein in A172 and U251 gloma cell lines were detected with the methods of in situ hybridization and immunohistochemistry.3. pcDNA3.1/LGI1 plasmid was constructed and transfected into A172 glioma cell mediated by liposome. The cell proliferating activity and apoptosis were detected by methods of MTT and Annexin V-FITC. The expressions of LGI1 mRNA and protein in the positive clones were detected by RT-PCR and immunohistochemistry. ResultsThe any LOH was not found in 25 glioma specimens and 5 glioma cell lines. The expression of LGI1 gene was not found in A172 cell. pcDNA3.1/LGI1 vector can be effectivly transfected into A172 cells by liposome. After A172 cell was transfected by pcDNA3.1/LGI1, their proliferating activity was inhibited. The MTT OD value was reduced at 24 hour and 48 hour after the transfection (P<0.05, vsnormal control group). Early apoptic cells at 24 hour were much more than that at 48 hour after transfection. The results of RT-PCR and immunohistochemical detection showed that expressions of LGI1 mRNA and protein were positive significantly after transfection. ConclusionThe LOH of LGI1 gene is not possibly the main factor for the malignant progression of glioma. LGI1 may inhibit cell growth and proliferation.