PROLIFERATING CELL NUCLEAR ANTIGEN DECREASES IN NORMAL HUMAN KERATINOCYTES WITH DIFFERENTIATION STIMULI BUT NOT IN AN HPV IMMORTALIZED CELL-LINE

Proliferating cell nuclear antigen (PCNA) is a co-factor for DNA polym erase delta, which replicates genomic DNA during cell growth and divis ion. Using a monoclonal antibody to PCNA (PC10) and conventional immun ofluorescent techniques, we have compared the effect of differentiatio n stimuli on PCNA expression in normal and HPV immortalised keratinocy tes. Two positive nuclear staining patterns were observed, a strong sp eckled form characteristic of proliferating cells and a weaker diffuse form. Strong nuclear staining was present in 44 +/- 4% (mean +/- SEM) of normal keratinocytes proliferating as a mono layer in 70 mu M calc ium serum-free medium but decreased to 13 +/- 3% after the differentia tion stimulus of 2 mM calcium medium for 2 days. An even greater reduc tion was observed following other differentiation agents, 1,25 dihydro xyvitamin D-3 the phorbol ester TPA and the non-specific protein kinas e C inhibitor staurosporine. Transforming growth factor-beta, which sl ows keratinocyte growth without inducing differentiation, reduced stro ng staining to 17 +/- 3% of cells, but with an increase in the diffuse pattern of staining from 39 +/- 4 to 57 +/-3%. HPV immortalised cells were resistant to the above agents except staurosporine, which inhibi ted growth and reduced the strong nuclear staining from 44 +/- 5% to 1 5 +/- 2%.