Modulating NF-κB Pathway in Leukemia Cells

Human THP-1 (ATCC) and MOLM-14 (ATCC) cells were cultured in RPMI1640 medium supplemented with 10% heat-inactivated fetal bovine serum, 100 units/ml penicillin, 100 ug/ml streptomycin and 2 mM L-glutamine. Authenticity of the cells was confirmed by short tandem repeat (STR) analysis [44 (link)]. Cells were infected with lentiviral vectors expressing a MUC1 shRNA, NF-κB p65 shRNA or scrambled control shRNA vector (Sigma). Cells were selected and maintained in puromycin [40 (link)]. Cells were treated with the NF-κB pathway inhibitor BAY-11-7085 (Santa Cruz Biotechnology) or DMSO as a vehicle control.

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Tagde A., Rajabi H., Stroopinsky D., Gali R., Alam M., Bouillez A., Kharbanda S., Stone R., Avigan D, & Kufe D. (2016). MUC1-C induces DNA methyltransferase 1 and represses tumor suppressor genes in acute myeloid leukemia. Oncotarget, 7(26), 38974-38987.

Publication 2016

scrambled control shRNA vector BAY-11-7085

Bay 11 7085 Cell culture Cells Cultured medium Dmso Fetal bovine serum Glutamine Human Muc1 Nf κb inhibitor Nf κb p65 Penicillin Puromycin Short tandem repeat Shrna Streptomycin Vectors

Corresponding Organization : Harvard University

Other organizations : Beth Israel Deaconess Medical Center

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Variable analysis

independent variables

Lentiviral vectors expressing MUC1 shRNA
Lentiviral vectors expressing NF-κB p65 shRNA
Lentiviral vectors expressing scrambled control shRNA
NF-κB pathway inhibitor BAY-11-7085
DMSO as a vehicle control

dependent variables

Not explicitly mentioned

control variables

RPMI1640 medium supplemented with 10% heat-inactivated fetal bovine serum, 100 units/ml penicillin, 100 ug/ml streptomycin and 2 mM L-glutamine
Puromycin selection and maintenance

controls

Positive control: Scrambled control shRNA vector
Negative control: DMSO as a vehicle control

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