Fig. 6. cAMP induces ß-casein expression in a GJIC-dependent and ß1-integrin-independent pathway. (A) Western blot analysis for ß-casein and Cx43, for CID-9 cells plated on (1) plastic, (2) plastic plus 50 µM 8-Br-cAMP, (3) plastic plus 50 µM 8-Br-cAMP and 10 µM 18 GA. ß-actin western blot demonstrates equal loading in (1), (2) and (3). Graph insets show that 8-Br-cAMP-treated cells on plastic had significantly higher ß-casein (P<0.05) and Cx43 (P<0.01) levels than untreated cells on plastic, whereas 18 GA significantly downregulated ß-casein (P<0.01) and Cx43 (P<0.01) levels in cells on plastic treated with 8-Br-cAMP. (B) Western blot analysis for ß-casein expression by CID-9 cells plated on (1) plastic plus 50 µM 8-Br-cAMP, (2) EHS-drip, (3) plastic plus 50 µM 8-Br-cAMP and 100 µg/ml anti-integrin, and (4) EHS-drip plus 100 µg/ml anti-integrin. ß-actin western blot was used to quantify ß-casein levels in (1), (2), (3) and (4) after normalization to equal ß-actin loading. A significant decrease in ß-casein levels (P<0.01) is noted when cells on EHS-drip are treated with anti-integrin antibody (4) compared with untreated cells (2). This is not the case when cells on plastic and supplemented with 8-Br-cAMP (1) are treated with the same antibody (3).