Cytokine Production by Human Peripheral Blood Mononuclear Cells Stimulated by a Pseudomonas Aeruginosa Culture Filtrate: Role of Plasma and Polymyxin B

1996 ◽  
Vol 19 (5) ◽  
pp. 276-283 ◽  
Author(s):  
S. Sundaram ◽  
T.W. Barrett ◽  
N.K. Butt ◽  
R. Porat ◽  
A.J. King ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Peripheral Blood Mononuclear Cells ◽  
Culture Filtrate ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Cytokine Production ◽  
Polymyxin B ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Bacterial Products

The lack of consensus regarding the significance of transmembrane passage of bacterial products across hemodialysis membranes can be related to several methodological differences in the various studies, including the choice of circulating fluid in the blood compartment of the model, nature and concentration of the bacterial products employed to challenge the dialysate compartment and whether cytokine production by PMBC or the limulus amebocyte lysate (LAL) assay was used as the index of transfer and the cytokine used as the read-out. In this study, we examined the production of interleukin-1 alpha (IL-1α), interieukin-1 receptor antagonist (IL-1Ra) and interleukin-8 (IL-8) by peripheral blood mononuclear cells (PBMC) incubated with a Pseudomonas aeruginosa culture filtrate. Further, the effects of 10% autologous human plasma and Polymyxin B sulfate (PmB) on cytokine production by PBMC were also characterized. The results of our study indicate that the Ps. aeruginosa culture filtrate had both PmB suppressible and PmB non-suppressible components and that the addition of 10% human plasma significantly enhanced cytokine production by both PmB suppressible and PmB non-suppressible components. The enhancing effect of plasma was most evident at low concentrations of the filtrate. The inhibitory effect of PmB was most evident in samples cultured in the presence of 10% plasma. There was a direct correlation between the production of IL-1α and IL-1Ra suggesting that both pro-inflammatory cytokines and cytokine-specific inhibitory proteins are concurrently produced. There results have direct relevance to selection of study conditions for in vitro models used to study the transmembrane passage of bacterial products across hemodialysis membranes

Polymyxin-B Stimulates Tumor Necrosis Factor-α Production by Human Peripheral Blood Mononuclear Cells

1998 ◽  
Vol 21 (5) ◽  
pp. 269-273 ◽  
Author(s):  
B.L. Jaber ◽  
S. Sundaram ◽  
M. Cendoroglo Neto ◽  
A.J. King ◽  
B.J.G. Pereira
Keyword(s):  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Tumor Necrosis ◽  
Mononuclear Cells ◽  
Cytokine Production ◽  
Polymyxin B ◽  
Peripheral Blood Mononuclear ◽  
Gram Negative ◽  
Blood Mononuclear Cells

Gram-negative bacterial lipopolysaccharide (LPS) is a well known stimulus for cytokine production, particularly interleukin-1 (IL-1) and tumor necrosis tactor alpha (TNFα). Polymyxin B (PMX-B) is a cationic polypeptide that binds to LPS, neutralizing its biological effects. PMX-B also disrupts gram-negative bacterial cell membrane phospholipids but is highly toxic to mammalian cells, therefore is of limited use. PMX-B is used as additive to media, as a way to handle LPS contamination. To derive benefit from the ability of PMX-B to neutralize lipid A in vivo while avoiding its systemic toxicity, PMX-B was covalently bound to polystyrene-derivative fibers, creating a hemoperfusion column (PMX-F) for the selective removal of circulating ET In vitro PMX-F hemoperfusion studies have demonstrated effective ET removal, using either the Limulus amebocyte lysate assay or TNFα production by peripheral blood mononuclear cells (PBMC) as an index of ET removal. However, the question whether PMX-B itself could stimulate human PBMC to produce cytokines has not been adequately addressed. We examined the effect of increasing concentrations of PMX-B on cytokine production by PBMC in vitro. PBMC harvested from healthy volunteers were incubated for 24 hours at 37°C with control (tissue culture media RPMI), or 5 µg/ml, 10 µg/ml, 20 µg/ml or 100 µg/ml PMX-B. At the end of 24 hours, PBMC were subjected to three freeze-thaw cycles, and total TNFα production (pg/2.5x106 PBMC) was measured by radioimmunoassay. Total TNFα production by PBMC was 163 ± 3 pg, 171 ± 9 pg, 164 ± 4 pg, 323 ± 63 pg and 331 ± 58 pg, in the control, PMX-B 5 µg/ml, 10 µg/ml, 20 µg/ml and 100 µg/ml conditions, respectively. Compared to controls (RPMI), the percentage increase in TNFα production by PBMC was 5 ± 6% (P=0.23), 1 ± 3% (P=0.45), 99 ± 40% (P=0.03) and 103 ± 36% (P=0.02) in the presence of 5 µg/ml, 10 µg/ml, 20 µg/ml and 100 µg/ml of PMX-B, respectively. Furthermore, total TNFα production correlated significantly with increasing concentrations of PMX-B (R=0.53, P=0.007). We conclude that the use of PMX-B in in vitro studies as an LPS-neutralizing agent, or in the experimental treatment of endotoxic or septic shock can lead to erroneous interpretations of cytokine production by PBMC, and should be used cautiously in in vitro systems at high concentrations.

Probiotic supplement consumption alters cytokine production from peripheral blood mononuclear cells: a preliminary study using healthy individuals

2013 ◽  
Vol 4 (4) ◽  
pp. 313-317 ◽  
Author(s):  
N.J. Hepburn ◽  
I. Garaiova ◽  
E.A. Williams ◽  
D.R. Michael ◽  
S. Plummer
Keyword(s):  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Cytokine Production ◽  
Ex Vivo ◽  
Healthy Individuals ◽  
Probiotic Supplementation ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells ◽  
Preliminary Study

The objective of this study was to examine the effect of daily probiotic supplementation upon the immune profile of healthy participants by the assessment of ex vivo cytokine production. Twenty healthy adult volunteers received a multi-strain probiotic supplement consisting of two strains of Lactobacillus acidophilus (CUL60 and CUL21), Bifidobacterium lactis (CUL34) and Bifidobacterium bifidum (CUL20) and fructooligosaccharide for 12 weeks. Blood samples were collected at baseline, 6 and 12 weeks. Peripheral blood mononuclear cells (PBMCs) were isolated and cultured ex vivo in the presence or absence of lipopolysaccharide and cytokine production was assessed. Postintervention, a significant decrease in the production of interleukin-6 and interleukin-1β was apparent when PBMCs were incubated in the presence of lipopolysaccharide, whilst a significant increase in IL-10 and transforning growth factor-β production was seen when the cells were incubated without an additional stimulus. This preliminary study demonstrates the potential of a multi-strain probiotic supplement to alter the immune response as demonstrated by changes in ex vivo cytokine production. Such results demonstrate the potential benefit of probiotic supplementation for healthy individuals and warrants further investigation.

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