ABSTRACT
Objective To investigate the in vitro cytotoxic effects of targeted silencing of long non-coding RNA colorectal cancer associated transcripts 2(CCAT2)by siRNA on multiple myeloma cell RPMI 8226 and U266.Methods Two strands of siRNA targeting at CCAT2,including siCCAT2-1 and siCCAT2-2,were transfected into RPMI 8226 and U266 cells at logarith-mic growth phase by liposome.The siRNA with the best silencing efficiency was screened for the subsequent experiments as ex-periment group by real-time fluorescence quantitative PCR(qPCR).The cells transfected with scramble sequence served as nega-tive control group and cells without transfection as blank control group.The cell proliferation rates were measured by cell count-ing kit-8.The apoptotic rates were measured by Annexin Ⅴ-FITC/PI double staining via flow cytometry.The mRNA levels of apoptosis related genes were detected by qPCR method.The numbers of invasive and metastatic cells were detected by Tran-swell method.Results The CCAT2 levels of multiple myeloma cell lines were reduced after transfection with siCCAT 2-1 and siCCAT2-2 in comparison with cells of negative control group and blank control group(P<0.05).The inhibition rates of RPMI 8226 and U266 cells were higher by transfection with siCCAT 2-2 than transfection with siCCAT2-1(P<0.05),and siCCAT2-2 was chosen to verify the cytology function.The inhibitory rates of RPMI 8226 and U266 cells in the experimental group was higher than those in the control group and the negative control group in a time dependent manner(P<0.05).Compared with RPMI 8226 cells of other two groups,the apoptotic rates and mRNA levels of Bax and Bad were increased but number of trans-membrane cells and mRNA level of Bcl-2 were decreased in experimental group(P<0.05).There was no significant difference between the control group and the negative control group on the above indices(P>0.05).Conclusion In multiple myeloma cell lines,siRNA targeting CCAT2 expression can inhibit the proliferation,invasion and metastasis and induce apoptosis of CCAT2, which may be valuable in the prevention and treatment of multiple myeloma.
ABSTRACT
Objective To investigate the role and possible mechanism of combination use of chloroquine (CQ) with either dexamethasone (DEX) or radiation on multiple myeloma (MM) cell line U266 .Methods Cell viability of U266 treated with CQ alone ,or CQ combined with either DEX or radiation was measured by cell counting kit-8 (cck8) .CalcuSyn method was used to assess effect of drugs interaction .Cell viability and apoptosis of U266 pre-treated with CQ were also measured by cck8 and flow cytometry after radiation .Expression of B-cellymphoma-2 (Bcl-2) in U266 cells treated by CQ combined with DEX or radiation was determined by Western blot analysis .Results Either CQ or DEX displayed a dose dependent cell proliferation in-hibitory effect on U266 cells .Cytotoxic effect of DEX (125 μmol/L) on U266 cells was enhanced and expression of Bcl-2 pro-tein in U266 cells was decreased by combining with CQ (3 .9 μmol/L) .U266 cells were sensitized to radiation and cell death was induced by CQ (1 .0 μmol/L) .Conclusion CQ could sensitize cytotoxic effect of DEX or radiation on U 266 cells ,and the former was possibly related to down-regulation of Bcl-2 protein .