300    

Background:  Our recent studies have shown that blood monocytes harbor heterogeneous HIV-1 variants that are genotypically distinguishable from those present in CD4⁺ T cells. However, the biological function of HIV-1 derived from monocytes, including association of genotype with phenotype, remains unclear. Here we analyzed the genotypes and co-receptor usage of HIV-1 derived from monocytes of 8 patients.

Methods:  CD14⁺ monocytes were purified from peripheral blood mononuclear cells by a two-step method of positive and negative selection. Nested polymerase chain reaction was used to amplify HIV-1 env from monocytes. Pseudoviruses were prepared in 293T cells by the cotransfection of pNL4-3 Luc-E^-R^- and patients’ env vectors. The U87 or ghost cells expressing different co-receptors were used to define the co-receptor usage.

Results:  Of  8 patients, 3 (P4, 5, and 6) were on highly active antiretroviral therapy (HAART) early at seroconversion; 2 patients (P2 and 8) began HAART 1520 and 3354 days post-seroconversion, and 3 patients (P1, 3, 7) had received no therapy. A total of 23 HIV-1 env clones isolated from monocytes at various time points spanning seroconversion to 9 years post-infection of 8 patients were tested. All pseudoviruses with monocyte-derived HIV-1 env utilized CCR5. However, 4 distinct phenotypes of co-receptor usage were observed: (1) All pseudoviruses with env from 3 patients (P5, 6, and 8) at different time points, including a time point of 9 years after infection, used CCR5 exclusively. (2) Env from 4 (50%) patients (P1, 2, 4, and 7) could use both CCR5 and CXCR4. Interestingly, the dual-R5X4 viruses appeared 7 days post-seroconversion (P1) and 25 days before seroconversion (P4) in the absence of HAART. (3) HIV-1 env from 4 patients, including all clones from P1 and P3, and 1 out of 3 clones from P2 and P7, utilized CCR3 in addition to CCR5 at acute infection. (4) Some HIV-1 env derived from monocytes showed a broader usage of co-receptors, including CCR1, CCR3, CCR5, and CXCR4 by monocyte viruses from P1 and P2 near seroconversion, P7 from year 3 of infection, and GPR15, CCR5, and CXCR4 by HIV-1 from P4.

Conclusions:  These findings indicate that blood monocytes could harbor viruses with multiple co-receptor usages at acute or late stages of HIV-1 infection, suggesting an important role that monocytes may play as a virus source of the transmission, establishment, and persistence of HIV-1 infection.

Keywords: Monocyte-derived HIV-1; Coreceptor; HIV-1 phenotype