P53 EXPRESSION IN ODONTOGENIC KERATOCYST EPITHELIUM

The expression of p53 protein was studied in odontogenic keratocysts ( OKC, 11 solitary, 5 recurrent and 6 NBCCS cysts), radicular (RC, n=5) and dentigerous (DC, n=5) cysts, using a panel of antibodies to p53 (c lone BP53-12, clone 1801 and polyclonal CM1) and a sensitive biotin-st reptavidin method on paraffin embedded sections. Of the three antibodi es tested, clone BP53-12 gave the most intense and consistent nuclear staining pattern. Clone 1801 and polyclonal CM1 stained only 38% and 7 1% OKC linings, respectively, but not RC and DC linings. However, BP53 -12(+) cells were detected in the epithelial linings of all cyst types . Quantification of BP53-12(+) cells was performed by manual counting and by relating cell number to unit length of basement membrane as det ermined by TV image analysis. BP53-12(+) cell counts in solitary OKC l inings (25.5 +/- 11.0 cells/mmBM) were significantly greater than thos e in DC (9.3 +/- 4.9 cells/mmBM, P<0.01) and RC (6.7 +/- 2.6 cells/mmB M, P<0.01) linings. The epithelial distribution of positive cells in O KC was predominantly suprabasal, which also varied from that of DC and RC linings (P<0.005). There were no detectable differences in BP53-12 reactivity between the different subtypes of OKC (i.e., solitary, rec urrent and NBCCS-associated OKC; P>0.1). When data for the NBCCS-relat ed OKC group were excluded, there was a significant correlation (r=0.5 5, P<0.01) between p53 and Ki67 labelling. To detect the presence of p 53 gene mutations, genomic DNA, extracted from paraffin sections of OK C (4 solitary, 2 recurrent and 4 NBCCS cysts), RC (n=3) and normal ora l mucosa (n=1), was subjected to a combination of polymerase chain rea ction and single-stranded conformation polymorphism (PCR-SSCP) analysi s for exons 5-10 of the p53 gene. Exon 4 was not analysed because of c ompromised DNA quality. No abnormality in banding patterns was found a nd all samples gave results similar to DNA from known, sequenced, norm al p53 gene controls. Absence of p53 mutations within exons 5-9 was co nfirmed by the direct sequencing of 2 fresh frozen OKC samples (1 soli tary and 1 NBCCS cyst). These results suggest that overexpression of p 53 protein in OKC epithelium, detected by immunocytochemistry, is not reflected by alteration of the p53 gene and presumably reflects overpr oduction and/or stabilisation of normal p53 protein.