During chronic HIV/SIV infection, the B-cell follicle is a major site of virus replication, and large numbers of infectious viral particles accumulate on follicular dendritic cells (FDCs) in the form of immune complexes (ICs). Despite the importance of the B-cell follicle in HIV/SIV persistence, the establishment and composition of FDC-bound HIV/SIV ICs remains poorly characterized. In this study we analyzed FDC-bound viral products during HIV/SIV infection.
Formalin fixed paraffin embedded (FFPE) lymph nodes (LNs) from acutely infected (7 dpi) and chronically infected (Non SAIDS, 69-118dpi) rhesus macaques or chronically HIV-infected humans were evaluated. Rhesus macaque tissue sections were stained with antibodies to CD20 (B-cells) and smooth muscle myosin heavy chain (FDC networks) and combined with in situ hybridization for SIV RNA (RNAScope ACD). For Human/HIV studies, 1-2 sections of FFPE or snap frozen LNs and tonsils from chronically infected untreated and healthy participants were evaluated. FFPE sections were analyzed for HIV RNA (RNAscope) and snap frozen sections were stained with antibodies to IgD (rings FDCs) and Nef. Imagescope (Leica) was used for quantification. FDC-bound SIV RNA was quantified by positive pixel counts on FDC networks with RNA+ cells excluded. Data were analyzed using Mann-Whitney tests and Spearmen coefficients (r).
The percent LN comprised of FDC networks tended to be larger in chronic (n=8) compared to acute (n=6) SIV infection (p=0.08; medians, acute= 5.24% chronic=18.73%). FDC-bound SIV RNA was not observed during acute infection, but was detected in all chronic infections. The size of the FDC-bound RNA reservoir strongly correlated with plasma SIV load in chronic infection (r=0.857, p=0.01). In chronic HIV infection, FDC-bound HIV RNA was detected in all LNs tested. Additionally, HIV-Nef+ staining was evident on FDC networks in most LNs (6/8) and tonsils (1/1), but was absent in healthy controls.
No FDC-bound SIV RNA is established during the first 7 days of acute SIV infection despite high viral loads, consistent with the notion that antibodies are necessary for the establishment of this reservoir. The strong relationship between FDC-bound RNA and plasma viral load suggests that they are in equilibrium with each other. Because Nef is a known immune-modulator, the accumulation of Nef on FDC networks may contribute to dysfunction of Tfh and other follicular cell subsets.