T(H)1 to T(H)2 shift of cytokines in peripheral blood of HIV-infected patients is detectable by reverse transcriptase polymerase chain reaction but not by enzyme-linked immunosorbent assay under nonstimulated conditions

Abstract

Background: Dysregulation of cytokines has been implicated in the pathogenesis of HIV infection. Therefore, we determined tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-4, IL-10, and interferon-gamma (IFN-gamma) mRNA and serum levels in HIV-infected patients under nonstimulated conditions.

Material and methods: Blood samples of 32 HIV-infected patients and 10 healthy HIV-negative controls were analyzed. Cytokine serum levels were quantified by enzyme-linked immunosorbent assay (ELISA). Cytokine mRNA levels were determined semiquantitatively by competitive reverse transcriptase polymerase chain reaction (RT-PCR) and expressed as ratios relative to those of beta-actin.

Results: Competitive RT-PCR was shown to be more sensitive than protein ELISA in analyzing cytokine production. We found a significant correlation between steady-state mRNA ratios and serum protein levels for TNF-alpha. Significantly higher cytokine mRNA ratios were found in those patients with IL-10 and IFN-gamma levels detectable by ELISA. Steady-state mRNA ratios of TNF-alpha, IL-4, and IL-10 were significantly increased in patients with highly replicative HIV-infection. Furthermore, elevated IL-4:IFN-gamma ratios were related to both high viral load and loss of CD4 cells.

Discussion: Determination of steady-state mRNA ratios by semiquantitative RT-PCR represents a sensitive method to analyze cytokines in peripheral blood of HIV-infected patients under nonstimulated conditions. The data obtained with this technique provide further evidence for a T(H)1 to T(H)2 cytokine shift with progressive HIV disease.