Late phase response of protein secretion; difference between the
low and high affinity activation
In order to assess mediator release during mast cell activation by
either low- or high affinity stimulation, we analyzed the expression of
91 soluble inflammation markers in the supernatant of mature mast cells
after 3, 6 and 24 hours of activation and detected 54 of these at at
least one of the four time points sampled. The early response after 3
hours was dominated by cytokines IL-8, IL-13, LIF, HGF and CCL4 produced
due to high affinity IgE crosslinking. PD-L1 was induced by both high
and low affinity and with no significant difference. A number of shedded
surface markers (TNFSF12, TNFSF14, CSF-1, TGF-a) and the cytosolic
protein STAMPB, were induced earlier and more persistently by high
affinity IgE than by low affinity IgE.
Most effects of signalling through low affinity IgE appeared after 6
hours, and many markers detected after low affinity activation were
shared with high affinity signalling. The cytosolic proteins CASP8 and
SIRT2 were specifically induced by low affinity IgE at 3 hours. The
markers that most notably appeared to be specific for low affinity
signalling were shedded CD40, SLAMF4 and CD5. This is consistent with
the observations that low affinity IgE signalling has slower onset than
high affinity signalling (2). CD40 is involved in the somewhat
controversial antigen presentation of mast cells (4). In mast cells, CD5
may not be expressed in the same way as it is on T cells (6), but
transcriptional activity at the promoter suggests it is expressed(7).
SLAMF4 is expressed as a regulatory molecule on mast cells (5) that may
have inhibitory as well as activating functions. Extensive shedding
after 6 hours of these molecules as well as IL10-RB and CD6 and
significantly more release of CCL3, TGF-b and CASP-8 differentiates the
low affinity response of human mast cells from a high affinity response.
One may speculate that the phenotype of the mast cell now adapts to the
low affinity stimulus. In addition, the membrane receptors released
become soluble signals that may modulate the physiologic response.
CSF-1, Fit3L, TGF-a, CD40 and TNFSF11 are known to be targets of ADAM17
that is expressed in mast cells, but there was no clear association of
ADAM17 and affinity of IgE or timing of release. CD6 and its ligand
CD318 (CDCP1) were shedded after low affinity IgE stimulation at 6 and
24 hours, respectively. While IL-18 and IL-10 were released at 3 and 6
hours under conditions of high affinity IgE, IL-18R1 and IL-10Rb were
shedded under low affinity IgE conditions at 6 and 24 hours, suggesting
the presence of a feedback loop. Shedding of a receptor may be a method
to quickly change the phenotype of the cell by releasing the surface
receptor, or to generate a signal molecule, or both (3).