Bcl-2 Expression and DNA Fragmentation in Breast Carcinoma, Pathologic and Steroid Hormone Receptors Correlates

Abstract

B-cell leukemia/lymphoma (bcl-2) expression can override the apoptosis development in lymphoid and hormonally regulated tissue-like breast. The presence of estrogen receptor (ER), progesterone receptor (PR), and androgen receptor (AR) have revealed in breast carcinomas, but they have not been correlated to the bcl-2 protein expression and DNA fragmentation markers. We evaluated the immunohistochemical expression of bcl-2 protein and hormonal receptors (ER, PR, AR) and differentiation grade in 37 infiltrating ductal carcinomas of the breast for which frozen tissues were available for DNA extraction. The immunohistochemical reaction for bcl-2 was considered positive if more than 50% of neoplastic cells had intense cytoplasmic staining, whereas for steroid receptor evaluation Battifora's criteria were used. The DNA was extracted according to the phenol-chloroform procedure and used for bcl-2 gene rearrangement study of the major breakpoint region (Southern blot) and for membrane-based end-labeling using digoxigenin-labeled nucleotides and E. coli DNA polymerase I (Klenow fragment). The results were quantified by three different observers. Low-grade carcinomas were positive for bcl-2 protein (27/28, 96.4%) and ER (15/28, 53.6%), whereas the remaining neoplasms were negative for bcl-2 (9/9, 100.0%) and ER (8/9, 53.6%) (p < 0.001). No statistically significant differences were revealed at the bcl-2, PR and AR comparisons. The Southern blot analysis for bcl-2 major breakpoint region showed neither rearrangement nor genetic amplification (densitometric study). Only the membrane-based end-labeling of DNA fragments showed correlation with bcl-2 protein and ER expressions: all except one bcl-2-negative tumor and two bcl-2-positive tumors had positive labeling using 7 pg of DNA at dot blot analysis (p < 0.002). The bcl-2 protein expression would allow both proliferation and cell progression by blocking apoptosis in well-differentiated, ER-positive breast carcinomas. In these neoplasms, DNA fragmentation as a molecular marker of apoptosis was prevented by bcl-2 expression.