Effects of silencing NOTCH1 gene by eukaryotic vector in multiple myeloma RPMI 8226 cell line

Abstract

Abstract:

AIM: To discuss the impact of NOTCH1 shRNA on cell proliferation and apoptosis in multiple myeloma RPMI 8226 cell line. METHODS: Eukaryotic expression vector for human NOTCH1 gene (NOTCH1 shRNA) was synthesized, and eukaryotic expression vector was transfected into RPMI 8226 cells by LipofectamineTM2000. The influence of cell growth by NOTCH1 shRNA was determined by MTT. Cell apoptosis and cell-cycle distribution were measured by Flow cytometry. Western blot was used for the detection of NOTCH1, P21 and P27. ELISA was used for detecting IL-6 secretion affected by NOTCH1 shRNA.RESULTS:NOTCH1 shRNA inhibited cell growth through blocking cell cycle in G0/G1 phase. The NOTCH1 shRNA group's cell growth rate was (49.09±2.31)%, Neg-shRNA group was (91.73±2.67)%, and blank group was (98.10±0.41)%, (P<0.05). The cell phase in G0/G1 with NOTCH1 shRNA group, Neg-shRNA group and blank group was (66.23±3.71)%, (42.27±3.65)%, (40.70±0.92)% respectively (P<0.05). The S phase was (31.03±1.49)%, (56.53±1.76)%, (51.83±1.45)% , respectively (P<0.05). NOTCH1 shRNA induces cell apoptosis in RPMI 8226 cells. The cell apoptotic rate was (46.67±1.31)% in NOTCH1 shRNA group, (1.37±0.63)% in Neg-shRNA group and (0.85±0.43)% in Blank group (P<0.05), respectively. The NOTCH1 protein was reduced and the expression of P27 and P21 protein were increased by transfected NOTCH1 shRNA. NOTCH1 shRNA downregulated IL-6 secretion. OD value of IL-6 secretion was 28.18±3.50, 167.08±7.97 and 185.86±5.61, respectively through three groups (P<0.05). CONCLUSION: NOTCH1 shRNA inhibits cell growth and induces cell apoptosis RPMI 8226 cell line, which may be a new treatment target in multiple myeloma.

Key words: NOTCH1, RNA interference, multiple myeloma, RPMI 8226 cell

CLC Number:

R965.2

HUANG Xiaolu, MA Xudong, LU Quanyi. Effects of silencing NOTCH1 gene by eukaryotic vector in multiple myeloma RPMI 8226 cell line[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2017, 22(9): 978-983.

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