(A) β-Galactosidase activity was induced in Tet-off–inducible MEF/3T3 cells (control) transfected with TRE-PR39/LacZ vector in the absence of doxycycline (ON). In the presence of doxycycline (OFF), β-galactosidase expression was completely abolished. A similar extent of β-galactosidase activity was achieved with the non-inducible reporter vector pCMVβ. (B) RT-PCR analysis of PR39 transcript in mouse hearts. Note expression only in induced tTA⁺/PR39⁺ mice (ON) and resuppression of expression by doxycycline (ON/OFF). (C) Hearts of controls and tTA⁺/PR39⁺ mice after 3 weeks (top row) and 6 weeks of PR39 induction (middle and bottom rows). Note an increase in the heart size and LV chamber after 6 weeks ON and the extensive expression of the transgene as demonstrated by X-Gal staining (right column). Lower right panel: X-gal–stained LV cross-section. Scale bar: 2 mm; 1 mm (inset). (D and E) Total cardiac endothelial cell mass determined by in vivo anti-PECAM Ab labeling (D) or capillary counting (E). Note a significant increase in endothelial cell mass at week 3 that is maintained at week 6 (n = 5 mice/group). (F) Myocardial endothelial cell mass determined by in vivo anti-PECAM Ab labeling expressed per gram of heart tissue. Note normalization of the endothelial cell mass/g tissue at 6 weeks. (G) Heart weight measurements after 3 and 6 weeks of PR39 induction and 6 weeks ON followed by 3 weeks of suppression (6ON/3OFF), compared with controls. (H) Heart weight/body weight ratio determined after 3 weeks, 6 weeks ON, and 6ON/3OFF. *P < 0.05, **P < 0.01. 3 wk, n = 10 mice/group; 6 wk, n = 16 mice/group; 6ON/3OFF, n = 10 mice/group.