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. 2021 Feb 17;11:3988. doi: 10.1038/s41598-021-82143-1

Figure 4.

Figure 4

HIV-induced BC expression of IL-8 mediates AM and neutrophil migration. BC were exposed to HIV or heat-inactivated HIV for 2 days. (A) Quantification of IL-8 in culture supernatants of untreated, heat-inactivated HIV and HIV- treated BC cultures by ELISA. (B) HIV-induced BC release of these mediators induce migration of human alveolar macrophages and neutrophils. Alveolar macrophages obtained from an HIV negative normal subject were plated in serum-free medium in the upper chamber and incubated with BC-conditioned media from untreated, NL4-3 treated and heat-inactivated NL4-3-treated BC culture in the lower chamber of transwell system. The number of migratory cells were quantified after 3 h. (C) Migration of human alveolar macrophages mediated by BC-release of IL-8. BC-conditioned media were pre-treated with mouse IgG1 control or anti-IL-8 neutralizing antibody (both at 2 µg/ml) for 30 min prior to addition to the lower chamber. Human alveolar macrophages were plated on the upper chamber and incubated with conditioned media from untreated BC, HIV-BC pretreated with mouse IgG1-treated or HIV-BC pretreated with anti-IL-8 neutralizing antibody. The number of migratory alveolar macrophages were counted after 3 h. Data shown are the mean ± SD of one representative of three independent experiments performed in triplicate. (D) Stimulation of migration of neutrophils. Human neutrophils in serum-free RPMI were plated in the upper chamber and incubated with BC-conditioned media from untreated, HIV-treated and heat-inactivated HIV-treated BC culture in the lower chamber of transwell system. The number of migratory cells were quantified after 1 h of incubation. (E) Migration of human neutrophils mediated by BC-released IL-8 in conditioned media. BC-conditioned media were pre-treated with mouse IgG1 control and anti-IL-8 neutralizing antibody (both at 2 µg/ml) for 30 min prior to addition to the lower chamber. Human neutrophils were plated on the upper chamber and incubated with conditioned media from untreated BC, HIV-BC pretreated with mouse IgG1-treated, HIV-BC pretreated with anti-IL-8 neutralizing antibody or HIV-treated BC. The number of migratory alveolar macrophages were counted after 1 h of incubation. Data are presented as mean ± SD of one representative of three independent experiments performed in triplicate.