Table III. Effect of TPA pretreatment on IL-6 and IL-1 response of HepG2 cells.

HepG2 cells were transfected with pAGP(3×DRE)-140-CAT and pIE-MUP as in Table II. After a 24-h recovery period, the culture medium was replaced by serum-free MEM and to one set of cultures (24 h pretreatment) 0.15 μM TPA was added. All media were replaced again by fresh media after 8 h to collect secreted MUP. After an additional 4 h, TPA treatment of the second set of cultures (12 h pretreatment) began. Twelve h later, all media were collected and the cells were washed three times with MEM. Cells in each group of cultures were incubated for 8 h either with 1 ml MEM and 0.1 μM dexamethasone alone or with 0.5 μM TPA, 100 units/ml IL-6, and 250 units/ml IL-1α. Chloramphenicol acetyltransferase activity and MUP production were determined as indicated in Table II. The amounts of MUP secreted (nanograms per well during the final 24-h culture period) were 62 ± 16, 75 ± 14, and 92 ± 15 for the 0, 12, and 24 h pretreatment group, respectively. The specific chloramphenicol acetyltransferase activities represent mean values of duplicate wells.

Treatment	Specific chloramphenicol acetyltransferase activity
	0 ha	12 ha	24 ha
	% conversion/h × ng MUP
No addition	0.04	0.02	0.01
TPA	0.23	0.04	0.01
IL-6	1.13	0.76	0.32
IL-6 + TPA	3.33	0.82	0.34
IL-1	2.83	0.85	0.15
IL-1 + TPA	1.12	0.78	0.22

^aPretreatment with TPA.