Susceptibility to human immunodeficiency virus-1 infection of human foreskin and cervical tissue grown in explant culture

Abstract

Numerous studies have indicated a protective effect of male circumcision against acquisition of human immunodeficiency virus (HIV)-1. We investigated mechanisms responsible for the possible increased HIV-1 susceptibility of human foreskin. Foreskins from eight pediatric and six adult patients with (n = 3) and without (n = 11) histories of sexually transmitted disease were evaluated. Six cervical biopsies from HIV-1-seronegative women were included as controls. CD4(+) T cells, macrophages, and Langerhans' cells (LCs) were quantified using image analysis. Cells expressing HIV-1 co-receptors CCR5 and CXCR4 were quantified using immunofluorescence and image analysis. Foreskin biopsies were infected ex vivo in organotypic culture with HIV-1. HIV-1 DNA copies in foreskin and cervical mucosal tissue were compared and the infected cell phenotype was determined. Foreskin mucosa contained higher mean proportions of CD4(+) T cells (22.4%), macrophages (2.4%), and LCs (11.5%) in adults than in children (4.9%, 0.3%, and 6.2%, respectively) or in cervical mucosa (6.2%, 1.4%, and 1.5%, respectively). The highest proportions of CD4(+) T cells and LCs occurred in patients with a history of infection. Foreskin immune cells expressed predominantly the CCR5 HIV-1 co-receptor. Adult foreskin mucosa had greater susceptibility to infection with HIV(bal) than cervical mucosa or the external surface of foreskin tissue. Circumcision likely reduces risk of HIV-1 acquisition in men by decreasing HIV-1 target cells.

Figure 1.

Immunohistochemical staining of LCs (CD1a), T cells (CD4), and macrophages (CD68) in foreskin tissue from an adult (Table 1 ▶ , no. 12) and pediatric (Table 1 ▶ , no. 8) patient. Cells expressing the appropriate antigen appear brown. Tissues were counterstained with hematoxylin. Images were stored and scanned for quantitative image analysis. Scale bar, 50 μm.

Figure 2.

Representative laser confocal image of two-color immunofluorescence staining for the HIV-1 co-receptors CCR5 and CXCR4 in adult male foreskin. Red, CCR5⁺ cells (arrowheads); green, CXCR4⁺ cells (arrows). The dotted line represents the epithelial (Epi)/submucosa (Sm) junction. Scale bar, 20 μm.

Figure 3.

A: Representative laser confocal image from the inner (mucosal) surface of foreskin (Table 1 ▶ , no. 10) infected by HIV-1_Bal in explant culture demonstrating infection of T cells (small, green, round cells; large arrowheads) and LCs (large, wavy, yellow cells; arrows). Uninfected CD4 T cells are red (small arrowheads). The majority of LCs in this field are infected. B: Representative laser confocal image from the outer surface of foreskin (Table 1 ▶ , no. 10) infected by HIV-1_Bal in explant culture showing uninfected T cells (small, red, round cells; small arrowheads) and uninfected LCs (large, wavy, orange cells; arrows). T cells and LCs expressing HIV-1 gag-pol mRNA were identified using ultrasensitive fluorescence in situ hybridization with simultaneous CD4-APC and CD1a-PE immunofluorescence in a three-color laser confocal microscope. The green color in A represents red CD4 plus blue-green HIV-1. The yellow color in A represents orange CD1a plus blue-green HIV-1. The red color in A and B represents CD4 alone and the orange color in B represents CD1a alone. Dotted lines represent the epithelial (Epi)/submucosa (Sm) junction. Scale bar, 20 μm.

Figure 4.

Representative photomicrograph of H&E-stained foreskin tissue from the outer surface (left) and the inner surface (right) of the foreskin. The extent of the keratin layer is identified by the black and white arrows.