Image analysis of p53 and cyclin D1 expression in premalignant lesions of the oral mucosa

OBJECTIVE: The expression of p53 and cyclin D1 proteins was analyzed by ima ge analysis in oral premalignant lesions and normal oral mucosa. STUDY DESIGN: Punch biopsies from the normal oral mucosa were obtained from 20 normal donors and 41 patients with oral dysplastic leukoplakias. After controlled formaldehyde fixation and paraffin embedding, immunohistochemist ry was used to detect cyclin D1 and p53. Image analysis was performed using stain intensity levels established by determining color thresholds (nuclea r score) in the basal and parabasal layers. RESULTS: Analysis of sections showed a similar pattern: only two normal don ors had a few intensely positive p53 cells in the basal layer of the floor of the mouth and the tongue epithelia. Similarly, only three donors had int ensely positive cyclin D1 cells in the normal epithelia of the same sites. Most cells fell in the range of negative or marginal stain (lower quartiles or terciles of nuclear score). These data on normal mucosa were compared w ith low grade oral leukoplakias (LGD) with mild to moderate dysplasia and w ith high grade leukoplakias (HGD) with severe dysplasia. Both markers were differentially expressed in precursor lesions versus normal epithelia. Stat istical analysis of our data shows that the intensity of the immunohistoche mical stain, as reflected in the nuclear scores of p53, is a reliable param eter that can differentiate between LGD and HGD of the oral mucosa. This wa s especially true when higher nuclear scores were compared. In contrast, lo w nuclear scores are more effective in differentiating normal epithelia fro m dysplastic epithelia. Although cyclin D1 immunohistochemistry does not st ain as intensely as p53 stain, similar conclusions can be derived from thos e data. CONCLUSION: Image analysis of these two markers proved useful in distinguis hing normal oral epithelia from low grade and high grade leukoplakias. With further developments in this field it is hoped that image analysis procedu res could be used in different types of studies in which variations of prot ein expression in tissue sections could have prognostic implications or cou ld be useful to determine subtle effects of curative or preventive treatmen t.