ABSTRACT
The effects of RNAi-mediated gene silencing of LRIG1 on proliferation and invasion of the human glioma cell line U251-MG and the possible mechanisms were explored in this study.The plasmids pGenesi12-LRIG1-shRNA1 and pGenesi12-LRIGl-shRNA2 were transfected into U251-MG glioma cells respectively by using Lipofectamine 2000 and the transfected cells in which the LRIGI expression was stably suppressed were selected by G418.The cells transfected with negative shRNA served as control.The expression levels of LRIG1 mRNA and protein were measured by qRT-PCR and Western blotting,respectively.The cell cycle was analyzed by flow cytometry.The results showed that LRIG1 mRNA expression was reduced by 70％ and 58％ and LRIG1 protein expression by 58％ and 26％ in U251-MG cells transfected with pGenesil2-LRIGl-shRNAl and pGenesil2-LRIG1-shRNA2 relative to the negative shRNA-transfected U251-MG cells.The proliferative capacity of the LRIG1 specific siRNA-transfected cells was stronger than that of control cells.Cell cycle analysis showed that silencing LRIG 1 significantly increased the percentage of S phase cells and the proliferation index (P＜0.01).Moreover,silencing LRIG1 could promote the invasion of U251-MG cells (P＜0.05).These findings suggested that LRIG1-targeting siRNA can exert a dramatically inhibitory effect on RNA transcription and protein expression of LRIG1,and LRIG1down-regulation could promote the proliferation of U251-MG cells,arrest U251-MG cells in S phase,and enhance the invasion of U251-MG cells.
ABSTRACT
The Leucine-rich repeats and immunoglobulin-like domains-2 (LRIG2) gene expression in pituitary adenoma and its correlation with tumor invasiveness were studied.The expression of LRIG2 mRNA and protein in human pituitary adenoma obtained surgically was detected by RT-PCR (39 cases)and immunohistochemical staining (30 cases).It was found that LRIG2 was mostly localized at the nucleus of the pituitary adenoma cells.Its expression was significantly higher in the invasive cases than in the non-invasive cases.LRIG2 protein was positive in 14 cases out of 21 cases of invasive adenoma,but only 2 cases were positive in 9 cases of non-invasive adenoma.The positive expression rate of LRIG2 mRNA was 91.3％ in invasive cases (total 23 cases) and 62.5％ in non-invasive cases (total 16 cases),respectively.LRIG2 gene is overexpressed in invasive pituitary adenoma.It may play an important role in pituitary adenoma invasiveness and further studies are necessary to elucidate the mechanism under this phenomenon.
ABSTRACT
This study examined the effects of over-expression of leucine-rich repeats and immunoglobulin-like domains 3 (LRIG3) on the cell cycle and survival of human glioma cell line U87 and U251 and explored the possible mechanisms.The LRIG3 gene was transduced into U87 and U251 cells respectively by using lentivirus and the transduced cells were selected by puromycin.The changes in LRIG3 mRNA and protein levels were measured by RT-PCR and Western blotting.The apoptosis rate was detected by Annexin V-FITC/PI double labeling and the cell cycle was flow cytometrically analyzed.Compared with control cells,LRIG3 mRNA expression in U251 and U87 cells transduced with pLVX-DsRed-LRIG3-Monomer-N1 were increased by 77.6％ and 129.7％,and LRIG3 protein expression was raised by 141.3％ and 322.7％,respectively.Cell cycle analysis showed that LRIG3 over-expression increased the percentage of cells at G0/G1 phase (P＜0.01).Over-expressed LRIG3 could significantly promote the apoptosis of U87 and U251 cells (P＜0.05).These findings suggest that the over-expression of LRIG3 could arrest the cell cycle in G0/G1 phase,and promote apoptosis of U87 and U251 cells.
ABSTRACT
The effects of RNAi-mediated gene silencing of LRIG3 expression on cell cycle and survival of human glioma cell line GLI 5 and the possible mechanisms were explored.The plasmids pGenesil2-LRIG3-shRNA l and pGenesil2-LRIG3-shRNA2 were transfected into GL 15 glioma cells respectively by using Metafectine,and the transfected cells that stably suppressed LRIG3 expression were selected by G418.The control cells were transfected with negative control shRNA.The changes in LRIG3 mRNA and protein levels were measured by RT-PCR and Western blot.The apoptosis rate and cell cycle were analyzed by flow cytometry.As compared with the negative shRNA-transfected GL15 cells,LRIG3 mRNA expression in GLI5 cells transfected with pGenesil2-LRIG3-shRNAl and pGenesil2-LRIG3-shRNA2 was silenced by 52.4%,63.8%,and LRIG3 protein expression was re-duced by 50.9% and 67.4% respectively.The LRIG3-specific siRNA transfected cells had higher proliferation rate than control cells.Cell cycle analysis showed that silencing LRIG3 increased the percentage of G2/M phase cells and the proliferation index significantly (P<0.01).Silencing LRIG3 could inhibit the apoptosis of GLl5 cells (P<0.05).These findings suggest that the siRNA targeting LRIG3 gene shows a dramatic inhibitory effect on RNA transcription and protein expression,then promoting the proliferation of GLI5 cells,arresting GLI5 cells in G2/M phase,and suppressing apoptosis ofGL15 cells.