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.
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.


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.


CONCLUSIONS
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.
Altfeld
M
M
Department of Internal Medicine I, University of Bonn, Germany. MALTFELD@partners.org
Addo
M M
MM
Kreuzer
K A
KA
Rockstroh
J K
JK
Dumoulin
F L
FL
Schliefer
K
K
Leifeld
L
L
Sauerbruch
T
T
Spengler
U
U
D018791:CD4 Lymphocyte Count; D016207:Cytokines / Q000097:blood* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D015658:HIV Infections / Q000276:immunology* / Q000821:virology; D015497:HIV-1 / Q000235:genetics / Q000502:physiology; D006801:Humans; D008213:Lymphocyte Activation / Q000276:immunology; D012333:RNA, Messenger / Q000097:blood / Q000235:genetics; D012367:RNA, Viral / Q000097:blood; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D018417:Th1 Cells / Q000378:metabolism*; D018418:Th2 Cells / Q000378:metabolism*
D016207:Cytokines; D012333:RNA, Messenger; D012367:RNA, Viral