Chemiluminescence of non‐differentiated THP‐1 promonocytes: developing an assay for screening anti‐inflammatory milk proteins and peptides

Abstract

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.

Meta‐Analysis for Correlating Structure of Bioactive Peptides in Foods of Animal Origin with Regard to Effect and Stability

“…Due to the complexity of mechanisms and to variability of in vivo responses (Leroux‐Roels et al., ; Table ), focusing on specific changes in either innate or adaptive immunity leads to incorrect conclusions about peptide activities, which can be corrected by measuring changes in both adaptive and innate immunities (Maestri et al., ). Anti‐inflammatory activity can be determined in vivo by the paw edema test, measuring swelling (as change in volume) after injection of inflammatory substance (Tavares et al., ), or in vitro by monitoring macrophage activity using chemiluminescence assays (changes in respiratory burst; Tompa et al., ), cytokine secretion (usually by ELISA; Tompa et al., ), or by monitoring monocyte adhesion to vascular endothelium (Aihara et al., ). Antiallergenic properties are measured in vitro determining IgE content by the reverse enzyme allergosorbent, ELISA, or enzyme allergosorbent tests (Falagiani et al., ; Wigotzki, Steinhart, & Paschke, ), or in vivo by measuring indexes of spleen and thymus status, such as changes in weight, hemolysin content, changes in optical density as a result of macrophage activity, and release of histamine from mast cells (Pan et al., ).…”

Technological and probiotic potential of BGRA43 a natural isolate of Lactobacillus helveticus

“…Interestingly, the obtained results show that one of the three major allergenic epitopes of BLG at the Val 41 , Tyr 42 , and Val 43 positions is hydrolyzed by the BGRA43 crude extract (Lozo et al, 2011; Strahinic et al, 2012). Moreover, four protein fractions released during BGRA43 milk fermentation exhibited an inhibitory effect on human monocyte RB, as well as anti-inflammatory effects on the Nd-THP-1 monocyte cell line due to the down-regulation of TNF-α or IL-6 production (Tompa et al, 2010). …”

Food‐derived immunomodulatory peptides

“…The corresponding humoral response resulted in an increased number of IgA + −secreting cells in bronchial tissue and the inhibition of fibrosarcoma growth . Additionally, peptide fractions from milk fermented by L. helveticus BGRA43 showed potentially anti‐inflammatory effects on lipopolysaccharide‐primed non‐differentiated THP‐1 cells . Further investigation could determine whether the effect of metabolities is due to exopolysaccharides or to the different components of bacteria present in fermented milk .…”