Fig. 5. Transdifferentiation from mammary epithelial cells to keratinocytes due to the loss of Smad4. (A,B) BrdU labeling of normal (A) and hyperplasia (B) area of a L10 gland. We have counted BrdU⁺ cells in 12 equivalent normal (n=6) and hyperplasia (n=6) areas. Average percentages of BrdU⁺ cells are 0.85±0.63 and 8.5±1 in normal and hyperplasia, with the P<0.01 by Student's t-test. (C-E) Images of early stages of transdifferentiation revealed by H&E (C), BrdU (D) and K14 (E) immunohistochemical staining. Arrows indicate the center of the lesion. (F,G) Images of an abscess at later stages detected by H&E (F) and K14 (G) immunohistochemical staining. Arrow indicates to the wall of the abscess, which is K14 positive, and arrowhead indicates an alveoli, which is K14 negative. (H-L) Micro-dissection (H-J) and PCR genotype (K,L) of dissected abscesses. (H) Prior to dissection; (I) after dissection; and (J) the dissected sample. (K) Structure of floxed allele. (L) Two independent samples (1 and 2) collected by micro-dissection, were analyzed and both showed that the abscess is caused by the Cre-mediated deletion of Smad4. Primers a/c amplify about 500 bp from the recombined allele. Primers a/c amplify about 450 bp from conditional allele, which is absent in these samples. The sequences of the primers are: a, 5'-GACCCAAACGTCACCTTCAG-3'; b, 5'-GGGCAGCGTAGCATATAAGA-3'; and c, 5'-AAGAGCCACAGGTCAAGCAG-3'. All samples were from a 7-month-old Smad4^Co/CoWAP-Cre mouse. Scale bar: 180 µm in A,B; 35 µm in C-E; 60 µm in F,G.