Identification of cell subsets expressing intracytoplasmic cytokines within HIV-1-infected lymph nodes

Objective: To describe the endogenous cytokine profile of HIV-l-infected lymph nodes (LN) and to identify the phenotype of individual cells expressing intracytoplasmic cytokines. Design and methods: Whole LN biopsies were collected from three HIV-seronegative controls and four HIV-I-positive individuals with persistent generalized lymphadenopathy. Three had established infection (Centers for Disease Control and Prevention 1993 criteria, stages A2, C1 and C3) and one was undergoing seroconversion illness. A combination of three methods was used to assess the impact of HIV-1 on LN architecture and endogenous cytokine expression. Immunocytochemistry was used to locate follicular dendritic cells (FDC), interdigitating cells and T and B cells. Reverse transcriptase-polymerase chain reaction was used to assess mRNA for interleukin (IL)-1 beta, IL-2, IL-4, IL-6, IL-10, tumour necrosis factor (TNF)-alpha and interferon (IFN)-gamma in collagenase-digested LN cells. Three-colour flow cytometry was used to identify intracytoplasmic cytokine expression within cell subsets. Results: Germinal center (GC) hyperplasia was observed in LN from two patients with established HIV-1 infection, and the third, coinfected with Mycobacterium tuberculosis, showed extensive necrosis. In the patient undergoing seroconversion, there was an extensive FDC network within the expanded and confluent GC which covered expansive areas of the LN. There was varied expression of IL-1 beta, IL-4, IL-6, IL-10 and TNF-a mRNA from the four HIV-l-infected LN and the patient undergoing serocon-version showed evidence for a mixed cytokine profile, which also included IL-2 and IFN-gamma. Flow cytometry revealed intracytoplasmic IL-IP protein restricted to cells expressing CD19, CD21 and CD38 antigens. Conclusions: Cytokines were detected in freshly isolated HIV-1-infected LN cells without requiring an exogenous stimulus. Seroconversion was associated with an expanded FDC network within enlarged GC, bounded by defined mantle zones containing B cells. There was diverse cytokine mRNA expression and IL-1 beta protein was restricted to cells expressing B-cell-associated antigens.