DAPT suppresses the proliferation of human glioma cell line SHG-44

OBJECTIVE@#To explore the suppressing effect of γ-secretase inhibitor DAPT on proliferation of human glioma cell line SHG-44 in vitro and its mechanism.@*METHODS@#The SHG-44 cell was treated by DAPT with different concentration. The proliferation of cells was detected by MTT assay; cell cycle and TSC of CD133(+) were determined by flow cytometry analysis technique; the key factor in Notch signaling pathway (Notch-1, Delta-1, Hes-1) was measured by reverse transcriptase-polymerase chain reaction and western blotting.@*RESULTS@#DAPT inhibited the growth and proliferation of SHG-44 cells significantly(P<0.05). And the inhibiting effect on SHG-44 cells produced by DAPT showed a dose-dependent manner. DAPT increased the rate of cells in G0/G1 phase of SHG-44 cells, while it decreased the rate of cells in S phase. TSC of CD133(+) was significantly reduced after DAPT treated SHG-44 cells. The expression of protein and mRNA of Notch-1, Delta-1 and Hes-1 were gradually downregulated with the increase of DAPT doses.@*CONCLUSIONS@#DAPT can downregulate these key factor in Notch signaling pathway, reduce the TSC of CD133+ and inhibit the proliferation of SHG-44 cells.

Establishment of SHG-44 cell line stably and highly expressing MCHR2 / 第二军医大学学报

Objective: To construct a eukaryotic expressing vector harboring human melanin-concentrating hormone receptor 2 (MCHR2) and to establish a SHG-44 cell line stably and highly expressing MCHR2. Methods: The full-length MCHR2 cDNA fragment was amplified from the human fetal brain cDNA library by PCR and was cloned into pcDNA3.1(+) to construct eukaryotic vector pcDNA3.1(+)/MCHR2; the latter was then transduced into SHG-44 cells by Lipofectamine™. After screening culture by G418, SHG-44 cells stably expressing MCHR2 were established. The transcription and expression of MCHR2 was identified by RT-PCR, Western blotting and immunofluorescence. Results: The full-length MCHR2 cDNA fragment was amplified and the eukaryotic expression vector pcDNA3.1(+)/MCHR2 was successfully constructed. The expression of MCHR2 was found positive by RT-PCR, Western blotting and immunofluorescence, indicating that the SHG-44 cell line stably and highly expressing MCHR2 was successfully established. Conclusion: The successful establishment of MCHR2-SHG-44 cell line provides a solid foundation for further study on MCHR2 function.