Objective: To study the suppressive effect of LRRC4 gene on human glioma U251 cells and further investigate its biological functions.
Methods: H&E, DNA and AgNORs stainings were performed on LRRC4-transfected U251 cells, mock-transfected U251 cells and non-transfected U251 cells, respectively. Quantitative analysis including cell morphometry, DNA content, DNA ploidy, silver stained argyrophilic nucleolar organizer regions (AgNORs) were investigated by image analysis. Flow cytometry was employed to determine the difference of cell cycle distribution and MTT staining was used to elucidate the activity of the LRRC4-transfected U251 cells.
Results: The morphological cell parameters such as area, perimeter and diameter, DNA content, chromosomal aneupoloidy, mean area of AgNORs particles and mean nucleus area of the LRRC4-transfected U251 cells were remarkably decreased compared to those of the mock-transfected and non-transfected U251 cells (P < 0.05, P < 0.01). Meanwhile, significant accumulation of cells in G(0)/G(1) phase but decrease of cells in S and G(2)/M phase, was observed in transfected U251 cells compared to those of the mock-transfected and non-transfected U251 cells (P < 0.05, P < 0.01). MTT staining showed that proliferation activity of both the mock- and non-trasfected U251 cells was significantly higher than that of the U251 cells transfected with LRRC4 gene (P < 0.01).
Conclusion: LRRC4 gene might be involved in tumor suppression by restraining DNA synthesis and the nucleoli organizer regions-associated proteins, keeping the cell cycles in phase G(0)/G(1) and reducing proliferation activity of the glioma cells. Morphometry combined with other techniques such as flow cytometry and MTT staining can well elucidate the biological function of novel genes.
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