Cell surface enzyme is responsible for the degradation
of substrates by cells. (A) Confluent HGF (donor, MH-12) were
stimulated with IL-1α (10 ng/ml) for 6 days in α-MEM supplemented
with 10% FCS in 96-well plates and then washed with PBS three times,
and confluent-monolayer cells were examined by DPPIV assay. Monolayer
cells were incubated with 2 mM substrate for DPPIV assay as described
in Materials and Methods. After a 60-min incubation, the supernatant
and the corresponding monolayer cells, which were resuspended in
peptidase medium, were incubated for an additional 90 min without
addition of the substrate. □, pNA formed by intact
fibroblasts; ◊, pNA formed by supernatant resulting from
60 min of preincubation of cells with the substrate; ○,
pNA released by cells preincubated for 60 min with the
substrate. (B) The percentage of lysed fibroblasts was determined by
trypan blue exclusion at each time point. The results are
representative of three different experiments with three different
donors (MH-12, KEK, and NIK).