Human doppel protein and similar proteins PrP32-121SH-SY5Y cell toxicity.docVIP

 PAGE \* MERGEFORMAT 14 Human doppel protein and similar proteins PrP32-121SH-SY5Y cell toxicity Of: Xu Kun, Wang, Chan Tian, Shi Song, Gui-Rong Shiqi, Zhou Ruimin, Jiang Huiying, Chu Yong Lie, Xiao-Ping Dong [Abstract] Objective To observe the cells in vitro transient expression of human Dpl (doppel) protein in human neuroblastoma SH-SY5Y cells, and its structure is similar to proteins - truncated PrP (PrP32-121) for comparison. Methods PCR method to obtain human PRND PRNP gene and the truncated gene was cloned into the eukaryotic expression vector, transiently transfected SH-SY5Y cells, and localization of protein expression observed by MTT test to assay the growth of the state, with the flow cytometry, Western blot and other methods to assay apoptosis status. results of the two proteins can be expressed in transfected cells, and exist in the cell membrane, MTT results showed that 24h after transfection cells were apparent toxicity role in apoptosis was found in transfected cells Annexin V / PI double staining positive cells increased, pro-casepase-3 and reduce the level of Bcl-2 factor. Conclusion Transient expression of Dpl and truncated PrP proteins can have a similar toxic effects of nerve cells, and induced activation of apoptosis. [Keywords:] doppel protein, prion protein, cytotoxicity, apoptosis ChinaABSTRACT: Objective To observe the biological activities of human doppel (Dpl) protein transiently expressed and Dpl-like protein PrP32-121 on a human neuroblastoma cell line SH-SY5Y. Methods Recombinant mammalian expression plasmids containing human PRND gene and truncated PrP32-121 fragment were generated by PCR. The expression and location of Dpl and PrP32-121 post-transfection were observed by IFA. The cytotoxicity was measured by MTT analysis. Cellular apoptosis was investigated by flow cytometry and Western blot. Results Both Dpl and PrP32-121 protein were expressed and mainly located on the cell membrane. Remarkable cytotoxicity was dete