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Na(+)-H+ exchange inhibitors decrease neointimal formation after rat carotid injury. Effects on smooth muscle cell
1Department of Medicine, Emory University School of Medicine, Atlanta, Ga 30322.
Abstract:
The presence of multiple growth stimuli at the sites of vascular injury following angioplasty suggests that therapies targeted toward common growth pathways will be more effective than therapies that inhibit only a single growth factor. We tested this hypothesis using amiloride and ethyl isopropyl amiloride (EIPA), which are inhibitors of the Na(+)-H+ exchanger, whose activity is required in many cells for proliferation and migration. In the rat carotid injury model, EIPA (100 micrograms/h for 15 days) significantly decreased intimal area and the ratio of intimal to medial area, whereas amiloride (25 micrograms/h) showed an inhibitory trend that was similar to that observed for captopril (80 mg/kg per day) and heparin (25 U/h). EIPA and amiloride inhibited rat vascular smooth muscle cell DNA synthesis, with IC50 values of 8.8 and 82.2 microM, respectively. Using platelet-derived growth factor as a chemoattractant, EIPA caused a concentration-dependent inhibition of migration (IC50, approximately 60 microM). Because amiloride and EIPA have nonspecific effects on cellular function (especially inhibition of tyrosine kinases), we sought to characterize the specific role of the Na(+)-H+ exchanger in vascular smooth muscle cell proliferation and migration. We generated a Na(+)-H+ exchanger-deficient mutant cell line [RNHE(-)]. Studies with these cells suggested that the inhibitory effects of EIPA and amiloride were mediated only in part via Na(+)-H+ exchange because (1) RNHE(-) cells grew well at pH 6.8 to 7.5 in bicarbonate-containing medium, and (2) there was no difference in migration in response to platelet-derived growth factor in the RHNE(-) cells. In summary, these data indicate that amiloride and EIPA inhibit neointimal formation in the rat carotid after injury. However, the mechanism of inhibition is likely to involve cellular events other than Na(+)-H+ exchange, such as an effect on tyrosine kinases.
Insights
Amiloride and EIPA inhibit neointimal formation after vascular injury. However, their mechanism may involve more than just Na(+)-H+ exchange, potentially affecting tyrosine kinases.
Area of Science:
- Cardiovascular Research
- Cell Biology
- Pharmacology
Background:
- Vascular injury following angioplasty involves multiple growth stimuli.
- Targeting common growth pathways may be more effective than single growth factor inhibition.
- The Na(+)-H+ exchanger is crucial for cell proliferation and migration.
Purpose of the Study:
- To test the hypothesis that therapies targeting common growth pathways are effective.
- To investigate the role of Na(+)-H+ exchanger inhibitors (amiloride, EIPA) in neointimal formation.
- To elucidate the specific mechanism of amiloride and EIPA in vascular smooth muscle cell proliferation and migration.
Main Methods:
- In vivo rat carotid injury model treated with EIPA, amiloride, captopril, or heparin.
- In vitro studies assessing rat vascular smooth muscle cell DNA synthesis and migration.
- Generation and use of a Na(+)-H+ exchanger-deficient mutant cell line (RNHE(-)) to study inhibition mechanisms.
Main Results:
- EIPA significantly reduced intimal area in the rat carotid injury model.
- Amiloride showed an inhibitory trend, similar to captopril and heparin.
- EIPA and amiloride inhibited vascular smooth muscle cell DNA synthesis and EIPA inhibited migration; however, these effects were only partially mediated by Na(+)-H+ exchange, as shown by studies with RNHE(-) cells.
Conclusions:
- Amiloride and EIPA effectively inhibit neointimal formation post-vascular injury in rats.
- The inhibitory mechanism of these drugs likely involves cellular processes beyond Na(+)-H+ exchange, including potential effects on tyrosine kinases.
- These findings suggest complex therapeutic targets for post-angioplasty vascular remodeling.