Acta Pharmacologica Sinica 2008 July; 29 (7): 847-852; doi: 10.1111/j.1745-7254.2008.00823.x

Acta Pharmacologica Sinica 2008 July; 29 (7): 847-852; doi: 10.1111/j.1745-7254.2008.00823.x

Original Article

Lentiviral vector-mediated siRNA knockdown of SR-PSOX inhibits foam cell formation in vitro¹

Li ZHANG², Hou-jia LIU³, Tie-jun LI⁴, Yang YANG², Xian-ling GUO², Meng-chao WU², Yao-cheng RUI^4,5, Li-xin WEI^2,5

²Tumor Immunology and Gene Therapy Center, Second Military Medical University, Shanghai 200438, China; ³Changhai Hospital and ⁴Department of Pharmacology, Second Military Medical University, Shanghai 200433, China

Aim: To investigate the expression of scavenger receptor that binds phosphatidylserine and oxidized lipoprotein (SR-PSOX)/CXC chemokine ligand 16 (CXCL16) in the human monocyte-derived cell line THP-1, and the effect of lentiviral vectors for the stable delivery of SR-PSOX/CXCL16 short hairpin RNA on foam cell formation.

Methods: A lentiviral expression vector containing enhanced green fluorescence protein (GFP) and SR-PSOX small interfering RNA (siRNA) (Lenti-SR-PSOXsi), or the control siRNA (Lenti-NC) gene was constructed. A human monocyte-derived cell line THP-1 was transfected with a different multiplicity of infection (MOI) of Lenti-SR-PSOXsi or Lenti-NC, and cultured to obtain stably-transfected THP-1KD and THP-1NC cells. After incubation with oxidatively-modified, low-density lipoprotein (Ox-LDL), the expression of SR-PSOX/CXCL16 mRNA was determined by real-time PCR. The expression of the SR-PSOX/CXCL16 protein was detected by flow cytometry analysis. The effect of Lenti-SR-PSOXsi on foam cell formation was assessed by Oil red O-stain analysis.

Results: Ox-LDL increased the expression of SR-PSOX/CXCL16 mRNA in a time- and dose-dependent manner in THP-1 cells. Four days after transfection with Lenti-SR-PSOXsi (MOI: 100), the percentage of GFP expression cells was over 89.3%. The expression of the SR-PSOX/CXCL16 mRNA and protein in THP-1KD cells significantly decreased compared with the parent cells, even the THP-1KD cells stimulated with 40 mg/L Ox-LDL. Ox-LDL uptake experiments in THP-1- and THP-1KD-derived macrophages indicated that SR-PSOX/CXCL16 deficiency decreased the development of macrophage-derived foam cell formation.

Conclusion: The above data showed that SR-PSOX siRNA delivered by using lentiviral vectors in THP-1 cells was a powerful tool for studying the effect of SR-PSOX, and decreased the expression of the SR-PSOX gene by inhibiting macrophage-derived foam cell formation.

Keywords: SR-PSOX; CXCL16; lentiviral vector; small interfering RNA; atherosclerosis; oxidatively-modified, low-density lipoprotein

¹This project was supported by the National Natural Science Foundation of China (No 30500615), the Young Scholars Foundation of Shanghai Public Health Bureau (No 2004-55-94), and the Young Scholars Foundation of SMMU (No 2005-SQ-11).

⁵Correspondence to Prof Yao-cheng RUI and Prof Li-xin WEI.
Phn/Fax 86-21-2507-4471.
E-mail ruiyc163@163.com (Yao-cheng RUI)
Phn 86-21-2500-0855.
Fax 86-21-6556-6349.
E-mail lixinwei@smmu.edu.cn (Li-xin WEI)
Received 2008-01-31 Accepted 2008-05-04

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