Abstract:      Background and purpose: Little about the function of p53 isoforms in gastric cancer was reported. This study was designed to explore the role of Δ133p53 in the effect of recombinant mutant human tumor necrosis factor (rmhTNF) on gastric cancer cells, and provide a new basis for the diagnostics and therapeutics of gastric carcinoma. Methods: MKN45 (with Δ133p53 expression) or SGC7901 (without Δ133p53 expression) cells were treated with rmhTNF of different concentrations only or combined with 5-FU (a traditional gastric cancer cellular killer), and the growth inhibition rate and apoptosis was detected by CCK-8 and flow cytometry. mRNA expressions of Δ133p53, Gadd45α and CyclinB1 were measured by nested reverse transcription-polymerase chain reaction (nRT-PCR) or real-time polymerase chain reaction(RT-PCR). Results: On MKN-45 cells with positive Δ133p53 expression, the inhibitory effect of rmhTNF was significant, the inhibition rates of 50 and 500 IU/mL rmhTNF were 24.82%, 72.33% after culturing for 24 h (t=-9.558, P<0.01); also, the inhibitory effect of 5-FU was improved by rmhTNF remarkably in time- and dose-dependence, the inhibition rates of 5-FU (25 μg/mL), rmhTNF (50 IU/mL) combined with 5-FU (25 μg/mL), rmhTNF (500 IU/mL) combined with 5-FU (25 μg/mL) were 18.20%, 48.66%, 59.83%, separately, after culturing for 24 h (F=82.742, P<0.01); the inhibition rates of rmhTNF (50 IU/mL) combined with 5-FU (25 μg/mL) were 48.66%, 68.20%, 85.23%, separately, after culturing for 24 h, 48 h and 72 h (F=128.583, P<0.01). However, on SGC-7901 cells with negative Δ133p53 expression, no growth inhibition was showed by rmhTNF only, the inhibition rates of 50 and 500 IU/mL were 2.74%, 3.25% after culturing for 24 h (t=-0.121, P>0.05). In apoptosis test, the apoptosis- enhancing effect of rmhTNF was significant on MKN45 cells, and the apoptosis-enhancing effect of 5-FU was further promoted significantly by rmhTNF, the apoptosis of rmhTNF (50 IU/mL), rmhTNF (50 IU/mL) combined with 5-FU (25 μg/mL), rmhTNF (500 IU/mL) combined with 5-FU (25 μg/mL) were 18.20%, 48.66%, 59.83%, separately, after culturing for 24 h (F=123.931, P<0.05). In mRNA measurement, down-regulation of Δ133p53 and CyclinB1, up-regulation of Gadd45α were significant in MKN45 cells treated by rmhTNF alone or combined with 5-FU. In nRT-PCR analysis, the mRNA levels of Δ133p53 were relatively 0.886, 0.499, 0.330, 0.161 (F=240.927, P<0.01); In real-time PCR analysis, the mRNA levels of Gadd45α were 1.227, 1.694, 3.394, and the mRNA levels of CyclinB1 were 1.221, 0.722, 0.316, relatively. The expression of Δ133p53 was positively related to CyclinB1 (r=0.977, P<0.01), but negatively related to Gadd45α (r=-0.950, P<0.01). Conclusion: In Δ133p53 positively expressed MKN45 cells, rmhTNF showed as an effective tumor inhibitor and an enhancer of 5-FU as well, and this effect might be helped by two p53 down-stream molecules CyclinB1 and Gadd45α. The results suggest that Δ133p53 might be a key target for the biological effect of rmhTNF against gastric cancer.

Key words: Recombinant mutant human tumor necrosis factor, 5-FU, Gastric cancer cell lines, Δ133p53