Apoptosis or programmed cell death frequently parallels abnormalities in cell proliferation and differentiation. As hypertrophy/hyperplasia or remodeling occurs in organs affected by hypertension, we evaluated the degree of apoptosis in the heart, kidney, and brain in situ in genetically hypertensive mice and rats as well as in cultured vascular smooth muscle cells. Apoptosis was characterized by morphological features, DNA fragmentation, and laddering as well as by terminal deoxynucleotidyl transferase labeling of the 3' OH ends of both extracted DNA and tissue sections. The present report provides the first evidence of increased apoptosis in whole organs of genetically hypertensive rat and mouse strains: in the heart of spontaneously hypertensive rats (SHR) and in the heart (ventricular cardiomyocytes), kidney (inner cortex and medulla), and brain (cortex, striatum, hippocampus, and thalamus) of spontaneously hypertensive mice, with a higher effect of apoptotic inducers in cultured aortic smooth muscle cells derived from SHR. Both types of known apoptotic processes, oligonucleosomal cleavage and large DNA fragmentation, were observed in vascular smooth muscle cells, but only the former appeared to be increased in SHR. This study underlines the importance of cell death dysregulation in hypertension, reveals a new route for investigation of the pathogenesis of hypertension, and suggests novel targets of therapeutic intervention.

To accommodate expanding volume (V) during hyposmotic swelling, animal cells change their shape and increase surface area (SA) by drawing extra membrane from surface and intracellular reserves. The relative contributions of these processes, sources and extent of membrane reserves are not well defined. In this study, the SA and V of single substrate-attached A549, 16HBE14o(-), CHO and NIH 3T3 cells were evaluated by reconstructing cell three-dimensional topology based on conventional light microscopic images acquired simultaneously from two perpendicular directions. The size of SA reserves was determined by swelling cells in extreme 98% hypotonic (approximately 6 mOsm) solution until membrane rupture; all cell types examined demonstrated surprisingly large membrane reserves and could increase their SA 3.6 +/- 0.2-fold and V 10.7 +/- 1.5-fold. Blocking exocytosis (by N-ethylmaleimide or 10 degrees C) reduced SA and V increases of A549 cells to 1.7 +/- 0.3-fold and 4.4 +/- 0.9-fold, respectively. Interestingly, blocking exocytosis did not affect SA and V changes during moderate swelling in 50% hypotonicity. Thus, mammalian cells accommodate moderate (<2-fold) V increases mainly by shape changes and by drawing membrane from preexisting surface reserves, while significant endomembrane insertion is observed only during extreme swelling. Large membrane reserves may provide a simple mechanism to maintain membrane tension below the lytic level during various cellular processes or acute mechanical perturbations and may explain the difficulty in activating mechanogated channels in mammalian cells.

Long term elevation of the intracellular Na؉ /K ؉ ratio inhibits macromolecule synthesis and proliferation in the majority of cell types studied so far, including vascular smooth muscle cells (VSMC). We report here that inhibition of the Na ؉ ,K ؉ pump in VSMC by ouabain or a 1-h preincubation in K ؉ -depleted medium attenuated apoptosis triggered by serum withdrawal, staurosporine, or okadaic acid. In the absence of ouabain, both DNA degradation and Caspase-3 activation in VSMC undergoing apoptosis were insensitive to modification of the extracellular Na ؉ /K ؉ ratio as well as to hyperosmotic cell shrinkage. In contrast, protection of VSMC from apoptosis by ouabain was abolished under equimolar substitution of Na