DCs may undergo a number of phenotypical changes, following maturation by the uptake and processing of an antigen. The process of DC maturation, in general, involves an increase in the surface expression of co-stimulatory molecules, like CD40, CD80, CD83, and CD86, which can be measured by flow cytometry. As a negative control unloaded DCs can be taken along, as well as a DC positively stimulated with a known maturation cocktail, like IL-1beta, IL-6, TNF and PGE-2 or TLR ligands.
Cytokine Production
Besides up-regulation of co-stimulatory molecules, DCs produce a large variety of cytokines that could be either pro/anti-inflammatory or skew the phenotype of T cells. Therefore, a quite simple procedure to test the activity of the DCs towards the food bioactives is the measurement of cytokine levels. A wide variety of cytokines may be expressed by mature DCs including IL-1 alpha, IL-1 beta, IL-4, IL-6, IL-8, IL-10, IL-12, IL-15, IL-16, IL-17, IL-18, IFN-alpha, IFN-beta, IFN-gamma, and TNF. These
cytokines can be measured by ELISA, or multiplex assays.
Other DC Readouts
Co-stimulation and cytokine production are effect parameters used in the vast majority of studies on effects of food bioactives on DCs (see Table 17.3 for an overview). Studying the induction of specific T cells is complicated due to the low
Table 17.3 In vitro studies using food-associated antigens in in vitro DC models
Peptic fragments of gliadin, pRQP and pDAC (decapeptides)
Yes: CD83, CD80, CD86, CCR7 and HLA-DR
Yes: IL-12,
TNF-a and IL-10
Yes
Giordani
et al. (2014)
MoDC
Proteins
Soluble peanut Ag (PNAg) and purified Ara h 1
LPS and CT (positive controls)
Yes: CD83, CD40,
CD86, and HLA-DR
No
Th-cytokine induction (IFN-g, IL4 and
IL-13)
Yes
Shreffler
et al. (2006)
MoDC
Proteins
Bet v 1, Mal d 1, Api g 1 and Dau c 1
Yes: CD40, CD83, CD80, CD86 and HLA-DR
No
Th-cytokine induction (IFN-g, IL4, IL-10
and IL-13)
No
Smole et al. (2010)
MoDC
Proteins
Peptic fragments of gliadin, soya protein, and OVA
Yes: CD83, CD80, CD86 and HLA-DR
Yes: various
Endocytosis and signal transduction
Yes
Palova-
Jelinkova
et al. (2005)
frequency of T cell precursors with the corresponding TCR and requires extensive culture conditions and broad read-out tools as the TCR specificity in these conditions is unknown. With regard to the T cell stimulatory potential of DCs the mixed lymphocyte reaction (MLR), in which DCs are used to stimulate alloreactive (naïve) T cells, can be considered. Proliferation in combination with cytokine production can be used as a read-out for T cell skewing.
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