Tetraspanin blockage reduces exosome-mediated HIV-1 entry

Abstract

HIV-1 is one of the most studied retroviruses. The role of exosomes in HIV-1 entry and pathogenesis are beginning to be appreciated. Exosomes can incorporate host proteins that are also contained in viruses (e.g., tetraspanins).

Fig. 1

Western blot and nanoparticle tracking analysis (NTA) validation of exosomal samples. Western blots of plasma exosomes, breast milk exosomes, and 293 exosomes. 60 μg/lane, were probed with (A) anti-CD81 or anti-clathrin (B). Arrows indicate proteins of interest. NTA-generated size and concentration plots for (C) human plasma and (D) human breast milk-derived exosomes. The different colors lines represent different pools of exosomes

Fig. 2

Exosomes significantly enhance HIV-1 entry into a human T cell line. A3R5.7 cells were seeded at a density of 1x10⁵ cells/well with the addition of 5 µg/ml diethylaminoethyl-dextran [9, 24]. Virus was used at a multiplicity of infection (MOI) of 0.002. Virus entry into A3R5.7 cells was evaluated in the presence or absence of (A) plasma-derived exosomes (0.05 μg) (B) breast milk-derived exosomes (0.035 μg) or (C) 293-derived exosomes (0.1 μg). Viral entry was also evaluated in the presence of exosomes and anti-CD81 antibody (0.2 µg/well). All incubations were processed for one hour. Viral gene expression in all control and treatment groups was assessed by Renilla luciferase activity at 72 h post-infection. This time point was selected because we previously observed that it allows for optimal expression in a variety of cell types, including those in the present experiments. All conditions are identical through the following experiments unless otherwise noted

Fig. 3

Exosomes significantly enhance HIV-1 entry into a human macrophage cell line. THP2574 cells (1x10⁴ cells/well) were classically differentiated into macrophages with 1 ng/ml phorbol 12-myristate 13-acetate) for 96 h. Virus was used in THP2574 cells at an MOI of 0.18. Virus entry into THP2574 cells was evaluated in the presence or absence of (A) plasma-derived exosomes, (plasma exo) (B) breast milk-derived exosomes, (BM exo) or (C) 293-derived exosomes (293 exo). Viral entry was also evaluated in the presence of exosomes and anti-CD81 antibody

Fig. 4

Breast milk-derived exosomes significantly enhance HIV-1 entry into human immune cell lines. Virus entry into (A) A3R5.7 cells or (B) THP2574 cells was evaluated in the presence or absence of breast milk-derived exosomes (BM exo). Viral entry was also evaluated in the presence of exosomes and anti-CD9 antibody