Effects of hypoxia and ouabain on transcapillary exchange of [99mTc]hexakis (2-methoxyisobutylisonitrile) [SESTAMIBI, also known as MIBI or HEXAMIBI] and 201TI were investigated with indicator-dilution studies using isolated rabbit hearts. Peak myocardial extraction (Emax), permeability-surface area products (PScap), and net myocardial extraction (Enet) were compared among serial injections during constant coronary flows. Overall, measures of transcapillary transport (Emax and PScap) for SESTAMIBI were significantly lower (p less than 0.001) than those simultaneously determined for thallium, but estimates of tissue retention (Enet) for SESTAMIBI and thallium were not statistically distinguishable. Hypoxia had no significant effect on mean (+/- s.d.) Emax for SESTAMIBI (0.31 +/- 0.13) or thallium (0.59 +/- 0.11), nor on mean PScap or Enet values. Ouabain (1.5 X 10(-7) M and 1.5 X 10(-6) M) had no effect on SESTAMIBI or thallium Emax (respectively, 0.29 +/- 0.08 and 0.60 +/- 0.05) or on PScap for SESTAMIBI. Thallium PScap was depressed with higher ouabain dose (control, 1.22 +/- 0.40; high ouabain, 1.06 +/- 0.41 ml/min/g; p less than 0.01). Ouabain also caused a significant and progressive increase in average SESTAMIBI Enet (control, 0.23 +/- 0.10 to high ouabain, 0.33 +/- 0.12; p less than 0.05), but depressed thallium Enet (control, 0.38 +/- 0.14 to high ouabain, 0.32 +/- 0.18; p less than 0.01). These results suggest myocardial metabolic and/or functional status have minor influence on transcapillary transport of SESTAMIBI and thallium, but significantly affects cellular retention.

Adhesion is a crucial characteristic of epithelial cells to form barriers to pathogens and toxic substances from the environment. Epithelial cells attach to each other using intercellular junctions on the lateral membrane, including tight and adherent junctions, as well as the Na(+),K(+)-ATPase. Our group has shown that non-adherent chinese hamster ovary (CHO) cells transfected with the canine beta1 subunit become adhesive, and those homotypic interactions amongst beta1 subunits of the Na(+),K(+)-ATPase occur between neighboring epithelial cells. Ouabain, a cardiotonic steroid, binds to the alpha subunit of the Na(+),K(+)-ATPase, inhibits the pump activity and induces the detachment of epithelial cells when used at concentrations above 300 nM. At nanomolar non-inhibiting concentrations, ouabain affects the adhesive properties of epithelial cells by inducing the expression of cell adhesion molecules through the activation of signaling pathways associated with the alpha subunit. In this study, we investigated whether the adhesion between beta1 subunits was also affected by ouabain. We used CHO fibroblasts stably expressing the beta1 subunit of the Na(+),K(+)-ATPase (CHO beta1), and studied the effect of ouabain on cell adhesion. Aggregation assays showed that ouabain increased the adhesion between CHO beta1 cells. Immunofluorescence and biotinylation assays showed that ouabain (50 nM) increases the expression of the beta1 subunit of the Na(+),K(+)-ATPase at the cell membrane. We also examined the effect of ouabain on the activation of signaling pathways in CHO beta1 cells, and their subsequent effect on cell adhesion. We found that cSrc is activated by ouabain and, therefore, that it likely regulates the adhesive properties of CHO beta1 cells. Collectively, our findings suggest that the beta1 subunit adhesion is modulated by the expression levels of the Na(+),K(+)-ATPase at the plasma membrane, which is regulated by ouabain.