Objective      To construct recombinant adenoviral vectors containing small interfering RNA (siRNA) targeting human SREBP cleavage activating protein (SCAP) gene and recombinant adenoviral vector containing SCAP gene

and investigate their effects on proliferation and apoptosis of ATDC5 stem cells.       Methods      By using pAdEasy

TM

adenovirus vector system

recombinant shuttle vector containing siRNA sequence targeting SCAP gene (pSES-HUS-SCAP siRNA) and recombinant shuttle vector containing SCAP gene (pAdTrack-SCAP) were constructed

and then transferred with pAdEasy-1 to generate recombinant adenovirus plasmids pAd-SCAPsiRNA and pAd-SCAP with an electroporation method. Subsequently

the plasmids were transferred into HEK-293 cells by lipofectamine-mediated transfection to package and amplify the recombinant adenoviruses Ad-siSCAP and Ad-SCAP. The ATDC5 cells were infected with the recombinant adenoviruses Ad-siSCAP and Ad-SCAP

respectively

and the expressions of SCAP at mRNA and protein levels were detected by RT-PCR and Western blotting. The effects of Ad-siSCAP and Ad-SCAP adenoviruses on the proliferation and apoptosis of ATDC5 cells were detected by flow cytometry. The expressions of Cleaved Caspase-3 and p-JNK were detected by Western blotting.       Results      Recombinant adenoviruses Ad-SCAPsiRNA with high titer of 2.5×10

10

PFU/mL and Ad-SCAP with 3.0×10

11

PFU/mL were successfully obtained. Both the SCAP mRNA and protein levels were significantly decreased in the ATDC5 cells after infection with Ad-siSCAP

and increased after infection with Ad-SCAP. Flow cytometry showed that the ATDC5 cells infected with Ad-siSCAP had more cells at S phase than the normal control and Ad-RFP (increased by 15.45% and 16.07%

respectively

P0.05)

and the apoptotic rate of the Ad-siSCAP group was decreased by 13.10% and 11.50% (P0.05) when compared with the 2 control groups. However

the ATDC5 cells infected with Ad-SCAP had less cells at S phase than the normal control and Ad-GFP (decreased by 14.27% and 13.45%

respectively

P0.05)

and the apoptotic rate of the Ad-SCAP group was increased by 10.51% and 10.17% (P0.05) when compared with the 2 control groups. Western blotting results of Cleaved Caspase-3 and p-JNK were in accordance with the results shown by flow cytometry.       Conclusion      Infection of ATDC5

cells with Ad-SCAPsiRNA promotes the proliferation and suppresses the apoptosis

while infection of ATDC5 cells with Ad-SCAP suppresses the proliferation and promotes the apoptosis.