Synergistic effects of nimesulide and 5-fluorouracil on tumor growth and apoptosis in the implanted hepatoma in mice

Abstract

Aim: To compare the effect of nimesulide or/and 5-fluorouracil (5-FU) on tumor growth inhibition and apoptosis in mice with the implanted hepatoma and to observe their possible interactions.

Methods: The inhibitory effects on tumor growth was evaluated by inhibition rate. Apoptosis was assessed by the ultrastructural, flow cytometry features and the DNA ladder demonstrated by agarose gel electrophoresis. PGE(2) level was determined by radioimmunoassay. Expression levels of c-jun, c-fos and p53 were evaluated by western blotting.

Results: Nimesulide or 5-FU alone inhibited the growth of hepatoma, while a synergistic effect was observed for a combined use of both. More pronounced morphologic changes for tumor cell apoptosis and the DNA ladder were found for the latter treatment. Expression levels of c-jun and p53 were found to be elevated for the tumors from mice treated with nimesulide and 5-FU comparing to those with either of them, but a reduced PGE(2) level was observed only for the treatment with nimesulide. No change was detected on c-fos expression.

Conclusion: Nimesulide and 5-FU appear to have synergistic effects for the growth inhibition and apoptosis induction. Both were found to be overexpressed in p53 and c-jun proteins, rather than that of c-fos, associations with the resulted apoptosis.

Figure 1

Electro micrographs of nimesulide plus 5-FU treated mice hepatoma. A, control; B, nimesulide plus 5-FU.

Figure 2

DNA ladder pattern of hepatoma tissues as demon-strated by agarose gel 1.5% electrophoresis. M, DNA markers; lanes 0-3, control, nimesulide 20 mg/kg, 5-FU 20 mg/kg, nimesulide 20 mg/kg + 5-FU 10 mg/kg.

Figure 3

Data of flow cytometry of mouse hepatoma without any treatment as a control (A), or following 21 d treatment with nimesulide 20 mg/kg (B), 5-FU 20 mg/kg (C) and nimesulide 20 mg/kg + 5-FU 10 mg/kg (D). The sub-G1 peak to the left of the G1 peak represents apoptotic cells.

Figure 4

Effect of Nimesulide and 5-FU on PGE₂ content in mouse hepatoma. Each column is the mean ± SD of the sample. 1-4, control, nimesulide 20 mg/kg, 5-FU 20 mg/kg, and nimesulide 20 mg/kg + 5-FU 10 mg/kg. n = 3. ^-x ± s. ^aP < 0.01 vs control, ^cP < 0.01 vs nimesulide + 5-FU.

Figure 5

Effect of nimesulide and 5-FU on expression of c-fos, c-jun, and p53 in mouse hepatoma. 0-3, control, nimesulide 20 mg/kg, 5-FU 20 mg/kg, nimesulide 10 mg/kg + 5-Fu 10 mg/kg. The β-actin was used as an intrinsic reference molecule.