Figure 4.

Cytokines secretion by irradiated and nonirradiated SV-BR-1-GM cells. (A) SV-BR-1-GM cells from 4- and 24-h-old clinical formulations were seeded in 6-well plates and cultured in full medium. Culture supernatants (SNs) were harvested after 1 day and 3 days of culturing then assessed for cytokine release. Note the substantially reduced levels of granulocyte–macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-6 for the 24 h compared to the 4 h old formulation. Values shown are arithmetic means from triplicate wells ± SDs, expressed as pg cytokine/1 million viable cells (at time of seeding)/24 h. For KITLG, one of the 24 h, 3d wells was excluded as the obtained cytokine levels were substantially higher than those of the other two wells. For IL-8, ELISAs for the 4 and 24 h samples were conducted on different days. (B) Cytokine levels in the Lactated Ringer’s solution (LRS) fractions of the formulations from (A). For IL-10, data are only shown for the 24 h sample. Values shown are arithmetic means from technical duplicates, expressed as pg cytokine/1 million total cells. (C) Nonirradiated human mammary epithelial cells (HMECs) and SV-BR-1-GM cells were cultured in parallel then assessed for cytokine release 24 h after replacing the culture medium. Note that the IL-6 and IL-8 levels from the nonirradiated SV-BR-1-GM cells shown here are substantially lower than those from the irradiated SV-BR-1-GM cells shown in (A). Values shown are arithmetic means from triplicate wells ± SDs, expressed as pg cytokine/1 million viable cells/24 h, whereby cell viability was determined on the day of medium change (initiation of cytokine accumulation) from cells cultured in parallel wells.