Chemiluminescence of non-differentiated THP-1 promonocytes: developing an assay for screening anti-inflammatory milk proteins and peptides.
Human leukemic THP-1 promonocytes are widely used as a model for peripheral blood monocytes. However, superoxide production during respiratory burst (RB) of non-differentiated THP-1 (nd-THP-1) cells is very low. Here we present a rapid and low-cost method for measuring the chemiluminescence (CL) of opsonized zymosan (OZ) induced RB which allows detection of Escherichia coli lipopolysaccharide (LPS) induced priming of nd-THP-1 cells on the basis of CL reaction kinetics. Maximum CL intensity obtained was 2.20 ± 0.25 and 1.30 ± 0.11 relative light units, while CL peak time was achieved at 18.1 ± 2.6 and 28.7 ± 1.3 min in primed and non-primed cells, respectively. The priming of nd-THP-1 cells with LPS evoked typical TNF-α and IL-6 production. We tested the effects of bovine lactoferrin and protein fractions from Lactobacillus helveticus BGRA43 fermented milk for potential anti-inflammatory effects on LPS primed nd-THP-1 cells. Four fractions were found to inhibit the OZ-induced CL in a dose-dependent manner (IC(50) 3-30 µg/mL), while lactoferrin inhibited CL to a lesser extent (IC(50) 270 µg/mL). These results suggest that measuring CL response of nd-THP-1 cells can serve as a method for screening anti-inflammatory compounds which could be used in reducing the risk of phagocyte-mediated inflammatory diseases.
Tompa
Gorazd
G
University of Ljubljana, Biotechnical Faculty, Department of Animal Science, Domžale, Slovenia. gorazd.tompa@bf.uni-lj.si
Laine
Aire
A
Pihlanto
Anne
A
Korhonen
Hannu
H
Rogelj
Irena
I
Marnila
Pertti
P
D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002417:Cattle; D002460:Cell Line; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis; D007700:Kinetics; D007781:Lactoferrin / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000737:chemistry* / Q000473:pathology; D049449:Luminescence; D008894:Milk Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis; D015054:Zymosan / Q000032:analysis* / Q000037:antagonists & inhibitors
D015850:Interleukin-6; D008894:Milk Proteins; D010446:Peptide Fragments; D014409:Tumor Necrosis Factor-alpha; D015054:Zymosan; D007781:Lactoferrin