Effects of maturation on cell water, protein, and DNA content in ovine cerebral arteries

Abstract

The present study examined the effects of maturation on base-soluble protein, DNA content, and intracellular water volume in ovine cerebral arteries to evaluate these variables as references for normalization of levels of cellular constituents in studies of vascular maturation. With maturation, base-soluble protein per unit wet weight (measured by using the Bradford method) increased by 27% to 46%, and cell volume (estimated as the ratio of cell water to DNA) increased by 53% to 97%. However, intracellular water per unit wet weight (calculated as the difference between total water measured by dehydration and extracellular water measured by using the extracellular marker [57Co]EDTA) increased by only 1% to 16%, because of maturation-related hypertrophy (quantitated histologically) combined with decreased cell number per unit wet weight (DNA content, quantitated by using Hoechst dye, decreased by 32% to 41%). In addition, the effects of maturation were artery specific: maturational increases in cell water and volume were more pronounced in common carotid than in cerebral arteries. These findings demonstrate that different methods of normalization can produce quantitatively opposite results in maturation-related studies. For cellular constituents, the most physiologically relevant normalization is relative to intracellular water volume, which yields units of apparent intracellular concentration. Because the relative content of intracellular water per unit wet weight changes little with maturation, normalization of levels of cellular constituents relative to wet weight provides a reasonable index of intracellular concentration. Clearly, no single approach is optimal for all studies, and the method of normalization should be carefully considered with regard to effects of maturation on vascular composition.