[The effects of culture supernatant of human GRC-1 cells transfected with PF4 cDNA on the VEGF expression and the growth of ECV304 cells].

AIM

To construct the eukaryotic expression vector pcDNA3-PF4-SS, and to detect the effects of the culture supernatant of transfected GRC-1 cells on the VEGF expression in transfected GRC-1 cells and the growth of ECV304 cells.

METHODS

The eukaryotic expression vector pcDNA3-PF4-SS was constructed and identified with Bgl II/BamH I digestion. The pcDNA3-PF4-SS was transfected stably into GRC-1 cells with lipofectamine mediation. The VEGF expression in transfected GRC-1 cells was detected by immunohistochemical staining, and the effect of the culture supernatant of transfected GRC-1 cells on ECV304 cells was detected by MTT colorimetry.

RESULTS

Restrictive enzyme (Bgl II/BamH I)digestion analysis showed that the recombinant expression vector pcDNA3-PF4-SS had been constructed successfully. RT-PCR detection proved that hPF4 cDNA had been transfected into GRC-1 cells. The result of immunohistochemical staining showed that the VEGF expression could be seen in the cytoplasm and on cytomembrane of GRC-1 cells transfected with pcDNA3-PF4-SS, but the expression obviously weakened as comparison with that before transfection. Cell counting and MTT colorimetry manifested that the culture supernatant of transfected GRC-1 cells could inhibit markedly growth of ECV304 cells.

CONCLUSION

The eukaryotic expression vector pcDNA3-PF4-SS has been constructed successfully, and stably transfected into the GRC-1 cells. The culture supernatant of transfected GRC-1 cells has obviously inhibitory effect on the growth of ECV304 cells and the VEGF expression in the GRC-1 cells, which lays some foundation for exploring the mechanism for anti-tumor growth and developing tumor vaccine for kidney neoplasms.