Quantification of Residual Germinal Center Activity and HIV-1 DNA and RNA Levels Using Fine Needle Biopsies of Lymph Nodes During Antiretroviral Therapy

HIV-1 reservoirs are most often studied in peripheral blood (PB), but not all lymphocytes recirculate, particularly T follicular helper (Tfh) CD4(+) T cells, as well as germinal center (GC) B cells, in lymph nodes (LNs). Ultrasound-guided fine needle biopsies (FNBs) from inguinal LNs and PB samples were obtained from 10 healthy controls (HCs) and 21 HIV-1-infected subjects [11 antiretroviral therapy (ART) naive and 10 on ART]. Tfh cells and GC B cells were enumerated by flow cytometry. HIV-1 DNA and cell-associated (CA) RNA levels in LNs and PB were quantified by real-time polymerase chain reaction. FNBs were obtained without adverse events. Tfh cells and GC B cells were highly elevated in ART-naive subjects, with a median GC B cell count >300-fold higher than HCs, but also remained higher in 4 out of the 10 subjects on ART. GC B cell counts and Tfh cell counts were highly correlated with each other, and also with activated T cells in LNs but not in blood. Levels of HIV-1 DNA and CA RNA viral burden in highly purified CD4(+) T cells from FNBs were significantly elevated compared with those in CD4(+) T cells from PB in the ART-naive group, but only trended toward an increase in the ART patients. FNBs enabled minimally invasive access to, and parallel measurement of residual activated T and B cells and viral burden within LNs in HIV-1-infected patients. These FNBs revealed significant GC activity that was not apparent from corresponding PB samples.