[Research of cloning, expression of human endostatin gene and its inhibition effects to the glioma in vivo].

OBJECTIVE

To clone human endostatin gene, detect its biological activities of endostatin protein and use it to cure human SHG44 gliomas of nude mice in vivo.

METHODS

The mRNA from the human liver tissue was extracted. And the functional fragment of endostatin gene was amplified by RT-PCR. It was cloned into pUC19 and sequenced according to Dye primer sequencing kit. The non-fusion expression vector pDH-endo was constructed. The recombinant human endostatin gene was expressed in DH5alphaat the condition of temperature induction. The protein activities were tested by Chicken Chorio-Allantoic Membrane (CAM) assay and endothelial cell proliferation inhibitory assay. Endostatin protein was applied to the nude mice through hypodermic injection in order to cure the SHG44 glioma.

RESULTS

The acquired endostatin gene is 551 bp, its sequence is correct and the expressed protein is 20 kDa. The protein possesses the anti-angiogenesis activity. Hypodermic injection of endostatin at the dose of 5 mg/kg/d, 10 mg/kg/d or 20 mg/kg/d can inhibit the human glioma angiogenesis and tumor growth(the inhibition rate of the tumor are seperately 34.5%, 76.1% and 80.2%).

CONCLUSION

The cloning, expression and preliminary application of human endostatin protein lay the foundation for the antiangiogenesis therapy of glioma and the other solid tumors.