c-Jun and p53 overexpression do not activate DKK1 transcription in myeloma cells, and overexpression of DKK1 does not affect myeloma cell apoptosis and drug response. (A) c-Jun or p53 proteins were overexpressed in JJN3 and OCI-MY5 cell lines using a lentiviral system. Western blot analysis of the nuclear extracts confirmed c-Jun or p53 up-regulation, as compared to each control transfected with empty vector. DKK1 expression, evaluated in the same samples by real-time PCR, was not activated. (B) JJN3 and OCI-MY5 cell lines were transfected with DKK1 overexpression vector using a lentiviral system. DKK1 protein was confirmed by Western blot analysis of cytosolic fractions (normalized to β-tubulin internal control). JJN3 and OCI-MY5 cell lines transfected with DKK1 or empty vector were treated with thalidomide or lenalidomide; after 48 hours of treatment, the percent of apoptotic cells was determined using flow cytometry (Table 1).