Abstract
Objective To investigate the role of leucine-rich repeats and immunoglobulin-like domains 1 (LRIG1) in U251 cells and its gene changes.Methods The stable cell line over-expressed LRIG1 (PEGFP-LRIG1) was established; PEGFP-N1-U251 cells and blank-U251 cells were used as controls.The expression changes ofLRIG1,GDNF and topoisomerase Ⅱ (TOPOⅡ) were detected by real time-PCR; Western blotting was employed to detect the LRIG1 protein expression; the growth curve of PEGFP-LRIG1-U251 cells and PEGFP-N1-U251 cells was drew from the 1st to 5th d,and the cell inhibition rate of temozolomide in these two groups was calculated.And then,siGDNF+siLRIG1 was performed to knock down the expressions of GDNF and LRIG1; besides that,siGDNF,siLRIG1 and/or blank control group were employed as controls to compare the proliferation index.Results The LRIG1 gene and protein expressions,GDNF and TOPOⅡ gene expressions were significantly different among the three groups (P＜0.05); the gene and protein expressions of LRIG1 were up-regulated and GDNF and TOPOⅡ gene expressions were down-regulated in the stable cell line LRIG1-U251 as compared with those in the PEGFP-N1-U251 cells and blank-U521 cells (P＜0.05).Cell counting in the stable cell line LRIG1-U251 was significantly lower than that in the PEGFP-N1-U251 cells on the 1-5 d of growth (P＜0.05).After temozolomide was added,cell inhabitation rate in the PEGFP-LRIG1-U251 group was obviously higher that that in the PEGFP-N1-U251 group (P＜0.05).SiGDNF+siLRIG1 could successfully knock down the expressions of LRIG1 and GDNF; the inhabitation rate of siLRIG1 was significantly higher than that of blank control group,and the that of siGDNF+siLRIG1 group was significantly lower than that in the siLRIGI group (P＜0.05).Conclusion LRIG1 can regulate proliferation of U251 cells by regulating GDNF.LRIG1 can inhibit GDNF and TOPOⅡ expressions and improve the chemosensitivity of U251 cells. Key words: Leucine-rich repeats and immunoglobulin-like domains 1; Glial cell line-derived neurotrophic factor; TopoisomeraseⅡ; U251 cell; Proliferation
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