Characterization of peripheral blood human immunodeficiency virus isolates from Hispanic women with cognitive impairment

Abstract

Human immunodeficiency virus type 1 (HIV-1) tropism plays an important role in HIV-associated dementia. In this study, aimed at determining if the tropism and coreceptor usage of circulating viruses correlates with cognitive function, the authors isolated and characterized HIV from the peripheral blood of 21 Hispanic women using antiretroviral therapy. Macrophage tropism was determined by inoculation of HIV isolates onto monocyte-derived macrophages and lymphocyte cultures. To define coreceptor usage, the HIV isolates were inoculated onto the U87.CD4 glioma cell lines with specific CCR5 and CXCR4 coreceptors. HIV isolates from cognitively impaired patients showed higher levels of replication in mitogen-stimulated peripheral blood mononuclear cells than did isolates from patients with normal cognition (P < .05). The viral growth of HIV primary isolates in macrophages and lymphocytes did not differ between patients with and those without cognitive impairment. However, isolates from the cognitively impaired women preferentially used the X4 coreceptor (P < .05). These phenotypic studies suggest that cognitively impaired HIV-infected women receiving treatment may have a more highly replicating and more pathogenic X4 virus in the circulation that could contribute to their neuropathogenesis.

Figure 1

Isolation of HIV-1 variants from peripheral blood of HIV-positive women characterized for cognitive function. Peripheral blood mononuclear cells (PBMCs) were isolated from women with normal cognition and women with cognitive impairment and cocultured with HPV-1-negative PBMCs. Culture supernatants were tested for HIV p24 antigen concentration at day 14 post infection to detect virus replication. Isolates from HIV patients with cognitive impairment showed significantly higher HIV p24 titers (* P = .049) than those from HIV patients with normal cognition.

Figure 2

Coreceptor usage of peripheral blood primary isolates. Coreceptor specificity was defined through the transfected glioma cell lines U87.CD4.GCR5 and U87.CD4.CXCR4, which express CCR5 and CXCR4 coreceptors, respectively. Cells were inoculated with 25 ng of HIV p24 antigen of primary HIV isolates from women with normal cognition (isolates 1 to 9) and with cognitive impairment (isolates 10 to 21). Infection was monitored by HIV p24 antigen ELISA at 8 and 13 days post infection. The laboratory-adapted isolates, HIV_BAL, and HIV_LAI, were used as the R5 and X4 positive controls, respectively. CXCR4 and CCR5 coreceptor utilization by HIV-1 primary isolates from patients with normal cognition is shown in A and B, respectively. CXCR4 and CCR5 coreceptor usage of HIV-1 primary isolates from patients with cognitive impairment is shown in C and D, respectively. Results are representative of one experiment in duplicate. There was no significant difference between HIV p24 titers in the R5 line between isolates from patients with (D) and those without (B) cognitive impairment at 8 (P = .538) or 13 (P = .537) days post infection. Isolates from patients with cognitive impairment (C) showed higher HIV p24 levels in the X4 line than the isolates from women with normal cognition (A) at 8 (P = .048) and 13 (P = .047) days post infection.

Figure 3

Cytopathic effects of HIV-1 primary isolates in MT-2 and U87.CD4 cell lines. Cytopathic effects for lymphocyte tropism was determined in the MT-2 cell line with the uninfected negative control (A) and the T-tropic isolate as positive control, HIV-1_LAI (B). Isolates were characterized as not syncytia-inducing (NSI) (isolate 9) (C) or syncytia-inducing (SI) (isolate 12) (D) variants. Syncytia formation was also used as a parameter to determine coreceptor usage in the U87.CD4 glioma cells with laboratory-adapted isolates HIV-1_LAI (H) and HIV-1_BAL (I) used as positive controls for the U87.CD4.CXCR4 and U87.CD4.CCR5 lines, respectively. The negative control of the assay is shown in (E). Cells were screened every 3 days for cytopathic effects on U87.CD4.CCR5 (isolate 11) (F) and U87.CD4.CXCR4 (isolate 3) (G) lines. Data are representative from three different experiments performed in duplicate.