Objective To observe heat shock protein 22 (HSP22) gene eukaryotic expression vector pEGFP-N1 transfected HSP22 expression levels in human umbilical vein endothelial cells in hyperlipidemia environment.
Methods HSP22 cDNA was obtained from MCF-7 by reverse transcription-polymerase chain reaction (RT-PCR) and linked to pEGFP-N1, and then recombinant plasmid pEGFP-N1/HSP22 was transformed into DH5α. Positive clones were identified by double enzyme digestion and sequencing. Recombinant plasmid pEGFP-N1/ HSP22 was transfected into HUVECs with liposome. The cell model of endothelial cells (EC) hyperlipemia was established to simulate environment of hyperlipemia injury. To observe heat shock protein 22 (HSP22) gene eukaryotic expression vector pEGFP-N1 transfected HSP22 expression levels in human umbilical vein endothelial cells by hyperlipidemia environmental.
Results (1) Agarosegel electrophoresis detection showed that HSP22 DNA segment was about 211 bp, which accorded with the expectation. Positive clones were identified by enzyme digestion after HSP22 gene was cloned into pEGFP-N1, and the sequencing results indicated it was correct. (2) RT-PCR methods showed HSP22 mRNA was expressed in transfected HUVECs. Western blot detected a specific protein band of HSP22. (3) HSP22 expression was significantly increased in the high-fat environment.
Conclusions The recombinant plasmid pEGFP-N1/HSP22 is successfully constructed and expressed positively with hyperlipemia in HUVECs.