Constructing a recombinant adenovirus carrying human cytomegalovirus UL138 gene and evaluating the effects of UL138 gene on the functions of monocytes
Objective 
To construct a recombinant adenovirus carrying UL138 gene, which was related to the latent infection of human cytomegalovirus, and to investigate the effects of UL138 gene on the functions of THP-1 mononuclear cells. 
 
 
Methods 
The recombinant adenovirus expressing the UL138 gene was packaged. The titer of the recombinant adenovirus was determined by calculating 50% tissue culture infective dose (TCID50). THP-1 mononuclear cells were infected with the recombinant adenovirus at different multiplicity of infection (MOI) and the optimal MOI was determined (100 PFU/cell) by observing the expression of green fluorescent protein (GFP). Changes in the expression of proinflammatory cytokines by THP-1 mononuclear cells that was induced by overexpressed UL138 were analyzed by quantitative PCR. The expression of chemokines and their receptors were measured by quantitative PCR array. 
 
 
Results 
The recombinant adenovirus carrying the UL138 gene was successfully constructed with a titer of 1×1011 PFU/ml. The rate of THP-1 mononuclear cells that was infected with the recombinant adenovirus was 60% at the MOI of 1∶100. Results of the RT-PCR analysis and Western blot assay further confirmed that the recombinant adenovirus could infect THP-1 mononuclear cells successfully and the expression of UL138 protein increased gradually over time. The overexpressed UL138 in THP-1 mononuclear cells significantly inhibited the expression of IL-18, IL-1β, IL-6, IL-8 and TNF-α as indicated by the results of quantitative PCR. Results obtained from the quantitative PCR array analysis showed that most of the chemokines and their receptors were downregulated in the transfected THP-1 mononuclear cells except for the chemokines of CCL17, CCL21, CCL2 and XCL2 and the receptors of CCR2, CXCR1, CXCR2, CXCR4 and CX3CR1 which were upregulated. 
 
 
Conclusion 
We successfully constructed the recombinant adenovirus carrying UL138 gene which could be used to infect THP-1 mononuclear cells. Overexpressed UL138 in THP-1 mononuclear cells significantly affected the functions of THP-1 mononuclear cells. 
 
 
Key words: 
Human cytomegalovirus; Latent infection; UL138; Recombinant adenovirus; THP-1 mononuclear cell; Chemokine and receptor