Induction of PAI-1 mRNA expression by overexpression of PKB.
Hepatocytes that were transfected either with PKB-K179A or myrPKB expression vectors or with the empty control vector were cultured for 24 hours under arterial pO2. At 24 hours the medium was changed, and cells were further cultured for the next 24 hours under normoxic (16% O2) and hypoxic (8% O2) conditions. (A) The PAI-1 mRNA levels were measured by Northern blotting. The mRNA level under normoxia (16% O2) was set equal to 100%. Values are expressed as means ± SEMs of 3 independent culture experiments. Student t test for paired values: *significant difference 16% O2 versus 8% O2, **significant difference 8% O2 + PKBK179A versus 8% O2 (control), ***significant difference 16% O2 + myrPKB versus 16% O2 (control),P ≤ .05. (B) Representative Northern blot. For Northern analysis 15 μg total RNA was hybridized to digoxigenin-labeled PAI-1 and β-actin antisense RNA probes (see “Materials and methods” for more information). Autoradiographic signals were obtained by chemiluminescence and scanned by videodensitometry.
