Cytokine profile of Plasmodium falciparum‐specific T cells in non‐immune malaria patients

Abstract

CD3+ T cells are important sources of both pro- and anti-inflammatory cytokines during Plasmodium falciparum malaria. We studied the frequency of interleukin-2 (IL-2), gamma interferon (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha) and IL-10 expressing CD3+ cells in 10 non-immune malaria patients with uncomplicated malaria and in one patient with cerebral malaria after P. falciparum-specific and non-specific mitogenic stimulation. Analysis by fluorescence-activated cell sorting was performed after drug-induced clearance of parasites to allow previously sequestered T cells to be detected in peripheral blood. CD3+ cells of patients responded to P. falciparum infected erythrocytes with significant increases in the percentage of IL-2, IFN-gamma, and TNF-alpha, but also IL-10, positive cells. CD3+ cells from malaria-naïve donors were also responsive to specific stimulation albeit to a much lesser extent. Mitogenic stimulation of PBMC revealed no significant differences between cells of patients and controls. CD3+ cells of the patient with cerebral malaria were hyporesponsive both to the infecting parasite isolate as well as to our laboratory-adapted P. falciparum isolate, whereas two patients with uncomplicated disease were more responsive to their infecting parasites than to the laboratory-adapted isolate. The results indicate that the increased responsiveness of in vivo primed compared to malaria-naïve CD3+ cells is Plasmodium-specific and biased towards production of IFN-gamma and TNF-alpha.

Increased specific T cell cytokine responses in patients with active pulmonary tuberculosis from Central Africa

“…It has been previously demonstrated that preferentially IFN-c producing, type 1 effector cells but not type 2 effectors undergo rapid apoptotic cell death when restimulated, which might explain to some extent the selective loss of PBMC derived IFN-c production in TB patients reported in earlier studies [53]. Second, we routinely use costimulation with MAb CD28 in addition to PPD to increase the yield of antigen-specific immune responses in T cells [49], which might overcome the reported downregulation of type 1 responses by MTB secretory antigen-differentiated and matured dendritic cells (DC), that display altered surface densities of co-stimulatory molecules [54].…”

“…Cells were cultured in 5 ml ultra culture medium (UCM) (Bio Whittaker, Walkersville, MD) supplemented with L-glutamine (2 mM/l; Sigma, St. Louis, MI), gentamicin (170 mg/l; Sigma) and 2-mercaptoethanol (3.5 µl/l; Merck, Darmstadt, Germany) for 18 h at 37°C in 5% CO 2 and stimulated with or without PPD (Statens Serum Institute, Copenhagen, Denmark) at a final concentration of 10 µg/ml. In order to amplify TCR signaling and to facilitate the initial phase of T cell activation the co-stimulatory MAb CD28 (Pharmingen, San Diego, CA) was added at a final concentration of 5 µg/ml to some wells [49]. Brefeldin A (10 µg/ml final concentration, Sigma) was added after 6 h to block protein secretion.…”

“…Brefeldin A (10 µg/ml final concentration, Sigma) was added after 6 h to block protein secretion. Cells were then harvested, fixed, and stained ( [49]. All MAb were purchased from Pharmingen.…”

“…Flow cytometric assessment of intracellular cytokine expression was performed as described previously [49,50]. Briefly, PBMC were isolated from heparinized blood and stimulated in UCM with phorbol 12-myristate 13-acetate (PMA, 10 ng/ml; Sigma) and ionomycin (1.25 µM; Sigma) in the presence of brefeldin A (1 µM; Sigma) for 4 h at 37°C in 5% CO 2 , Cells were harvested, fixed and stained with antihuman MAb.…”

Interferon-γ—central mediator of protective immune responses against the pre-erythrocytic and blood stage of malaria

“…[191]). In contrast, recall responses are markedly increased in malaria patients following a first clinical episode [192][193][194][195]. Indeed, even subclinical infections are sufficient to induce robust IFN-␥ responses to pRBC in previously naïve donors [19,20,42].…”