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Relationship between functional Na+ pumps and mitogenesis in cultured coronary artery smooth muscle cells
T F Feltes1, C L Seidel, D K Dennison
1Department of Pediatrics, Baylor College of Medicine, Houston, Texas 77030.
Abstract:
An increase in functional sarcolemmal Na(+)-K(+)-ATPase (Na+ pump) precedes proliferation in vascular smooth muscle cells (VSMCs) seeded in 10% fetal bovine serum (FBS), but its role in mitogenesis is unresolved. Enzymatically dispersed canine coronary artery VSMCs were seeded in FBS and studied through confluence. Before a shift in cell cycle (G1-->S, G2 + M) and appearance of the nonmuscle isoform of myosin (MHCnm), intracellular Na+ content (Na+i) and cell volume (CV) increased (day 0 through day 3). Na+ pump number ([3H]-ouabain binding) increased at day 4 followed by a decrease in Na+i and CV. When Na+ pumps were inhibited by the addition of ouabain to FBS, VSMCs were arrested in G1, and MHCnm was not upregulated. Na+i increased similarly to that in FBS but failed to correct to day 0 levels. Withdrawal of ouabain at day 4 in culture led to an increase in Na+ pump number, a decrease in Na+i, entry of cells into S and G2 + M, and upregulation of MHCnm. These data suggest that Na+i, phenotypic modulation, and entry of cells into the cell cycle are temporally related, with Na+ pump-mediated correction of increased Na+i as a key event in the VSMC mitogenic process.
Insights
The Na(+)-K(+)-ATPase (Na+ pump) corrects increased intracellular sodium, a key event for vascular smooth muscle cell (VSMC) proliferation and cell cycle entry.
Area of Science:
- Cardiovascular Biology
- Cell Physiology
- Molecular Cardiology
Background:
- Vascular smooth muscle cell (VSMC) proliferation is crucial for cardiovascular health.
- The role of the sarcolemmal Na(+)-K(+)-ATPase (Na+ pump) in VSMC mitogenesis remains unclear.
- Understanding VSMC regulation is vital for treating cardiovascular diseases.
Purpose of the Study:
- To investigate the role of the Na+ pump in VSMC proliferation.
- To determine the relationship between intracellular sodium levels and cell cycle progression.
- To elucidate the signaling pathways involved in VSMC phenotypic modulation.
Main Methods:
- Enzymatic dispersion and culture of canine coronary artery VSMCs.
- Assessment of cell cycle progression using flow cytometry.
- Quantification of Na+ pump number via [3H]-ouabain binding.
- Measurement of intracellular sodium ([Na+]i) and cell volume (CV).
Main Results:
- Increased [Na+]i and CV preceded VSMC proliferation and cell cycle shift (G1 to S, G2+M).
- Na+ pump number increased, leading to decreased [Na+]i and CV.
- Inhibition of Na+ pumps with ouabain arrested VSMCs in G1 and prevented phenotypic modulation.
- Ouabain withdrawal restored Na+ pump activity, decreased [Na+]i, and promoted cell cycle entry and MHCnm upregulation.
Conclusions:
- Na+ pump-mediated correction of elevated intracellular sodium is a critical event in VSMC mitogenesis.
- Intracellular sodium levels, phenotypic modulation, and cell cycle entry are temporally linked in VSMCs.
- The Na+ pump plays a pivotal role in regulating VSMC proliferation and phenotypic switching.