F. Merlin Bumpus

Cardiac Hypertrophy in Spontaneously Hypertensive Rats

Ventricular weight in spontaneously hypertensive rats (F26 generation, Okamoto-Aoki strain) was significantly higher (P < 0.001) than that in body weight-matched American Wistar and Kyoto-Wistar normotensive rats, not only among older groups of rats but also among younger groups that had not developed significant hypertension. Deoxyribonucleic acid (DNA) concentration in ventricular muscle was not different from normal in the youngest group (P < 0.4) but was significantly reduced in the older spontaneously hypertensive rats (P < 0.01). Plasma renin activity was significantly increased in younger spontaneously hypertensive rats before the development of established hypertension; moreover, ventricular weight and plasma renin activity were significantly correlated in younger rats (r = 0.788, P < 0.005 for all rats, r = 0.644, P < 0.01 for spontaneously hypertensive rats). Antihypertensive therapy with either α-methyldopa or hydralazine reduced blood pressure, especially in hypertensive rats; however, ventricular weight was reduced by methyldopa (P < 0.01) but not by hydralazine. Plasma renin activity was reduced by methyldopa but increased by hydralazine (P < 0.01). DNA concentration was reversed toward normal by methyldopa but not by hydralazine. Similar results were obtained when methyldopa and hydralazine were given to younger rats to prevent hypertension. The changes in ventricular weight with the onset of hypertension and with its reversal or its prevention suggest that blood pressure might not be the sole factor contributing to cardiac hypertrophy in the spontaneously hypertensive rat and that the renin-angiotensin system might play a permissive role enhancing myocardial hypertrophy.

Measurement of Renin Activity in Human Plasma

A method is described for estimating plasma renin activity by using renin substrate present in plasma. This method differs from other indirect renin assay methods by (1) incubation in the absence of ions thus establishing conditions for zero order kinetics for the reaction between endogeneous renin and substrate and (2) the use of angiotensinase inhibitors di-sodium ethylenediamine tetraacetic acid (EDTA) and d-isopropylfluorophosphate (DFP). Recoveries of renin added to plasma in levels similar to those occurring in plasma are 85% SD±7%. The incubation was done at pH 5.5 which was shown to be the optimum for human renin reacting with human substrate. By incubating human plasma samples with known quantities of human renin, evidence was obtained suggesting that factors other than enzyme or total substrate concentrations affect the velocity of angiotensin formation. This variability of reaction rate may be explained by the existence of an inhibitor or activator in this system or by a variation in the type of substrate.

Renin in Rats with Spontaneous Hypertension

An age-dependent study of plasma renin activity (PRA), kidney renin activity (KRA), and plasma renin substrate was carried out in rats with spontaneous hypertension during the prehypertensive, the early hypertensive, and the established hypertensive phases of their disease. PRA and KRA were both significantly elevated before and during the initial phase of hypertension and normal or subnormal during the established phase. In normal controls, neither KRA nor PRA was significantly different during the entire growth period. Plasma renin substrate was elevated throughout the growth period in rats with hypertension. This relationship between renin and blood pressure suggests that renin may play a primary role, possibly along with other factors, in the initiation of hypertension in rats with spontaneous hypertension.

Angiotensinase with a high degree of specificity in plasma and red cells.

A peptidase with a high degree of specificity for angiotensin II occurs in normal human plasma and red cells. Preparations from both sources have the same pH optimum, require calcium ions, and hydrolyze valyl5- or isoleucyl5-angiotensin II, but do not hydrolyze β-aspartyl1-angiotensin II, arginyl1-angiotensin II or deaminoangiotensin II. This enzyme, given the name angiotensinase A, requires α-L-aspartic acid or α-L-asparagine as the N-terminal amino acid in its angiotensin substrate, and thus differs from kidney leucine aminopeptidase. Other peptidases known to hydrolyze angiotensin also hydrolyze at least one of the other angiotensin analogs with substitution in the one position.

Angiotensin Tachyphylaxis and its Reversal

Tachyphylaxis to angiotensin and some analogues has been demonstrated on spirally cut arterial strips from cat, dog, sheep and rat, and on venous strips from rabbit and cat. Rabbit and guinea pig arteries do not appear to become tachyphylactic. A free C-terminal carboxyl group of angiotensin is necessary for binding with receptor sites and development of tachyphylaxis. Tachyphylaxis seems to represent saturation of receptor sites. It can be reversed by plasma fractions rich in angiotensinase A, possibly by metabolizing the N-terminal part of angiotensin directly from the bound state. Dowex 50 can also reverse it, probably by physical adsorption and stronger binding of angiotensin. Angiotensinase A does not metabolize βaspartyl1-angiotensin and does not reverse tachyphylaxis to this peptide. A possible scheme of interaction between peptide and receptor site is presented.

Separation and characterization of the protein moiety of human α1-lipoprotein☆

Abstract A delipidation procedure was applied to human serum α 1 -lipoprotein samples prepared ultracentrifugally, to separate without denaturing its protein moiety. In the final protein residue, 0.6% lipides, predominantly as phospholipides, were present. Data on the physicochemical characterization of the delipidized protein residue are presented. No change in electrophoretic mobility was observed. Calculated values for the sedimentation constant and molecular weight were found to be slightly lower than those of albumin. The chromatogram was characterized by the absence of sulfur-containing amino acids. Aspartic acid was N-terminal.

The relationship of structure to pressor and oxytocic actions of isoleucine5 angiotensin octapeptide and various analogues

Abstract Isolated natural angiotensin I and synthetic angiotensin II have similar pressor activity but the latter has a much higher oxytocic activity. On a weight basis, the oxytocic activity of angiotensin II is about 1 8 that of oxytocin. Examination of analogues of angiotensin II demonstrate the following conditions must be met to have pressor and oxytocic activity: (a) the C-terminal amino acid must be l -phenylalanine, (b) the terminal carboxyl group must be free, (c) tyrosine must be present, and (d) the peptide must contain at least amino acids numbered 3 to 8. It is suggested that the spatial configuration of the molecule is an important aspect of the biological specificity of peptides in addition to the amino acid composition and sequence.

A proposed conformation of isoleucyl5-angiotensin II

Abstract Optical rotatory dispersion studies on isoleucyl 5 -angiotensin II show the peptide has a definite degree of order which is decreased by urea. Ultraviolet spectral studies on the phenolic hydroxyl group indicate it does not interact with any other groups in the molecule. A three-dimensional model of angiotensin II is suggested which is consistent with physical and biological data available.

Angiotensin II: An Intraovarian Regulatory Peptide

Mammalian ovarian follicles contain the enzymes and prohormones necessary to elaborate the active octapeptide hormone angiotensin II. In the rat ovary, angiotensin II receptors are located primarily in the theca interna and granulosa cell layers of a discrete subpopulation of follicles. Angiotensin II stimulates both androgen and estrogen secretion from rat ovarian slices. These findings suggest an autocrine/paracrine role for angiotensin II in ovarian follicular development.

Erythrocytosis in Spontaneously Hypertensive Rats

During the study of an inbred strain of Wistar rats which spontaneously develop hypertension when they reach a weight of approximately 150 g, it was found that these animals also develop an erythrocytosis. A significant increase in red cell count was observed in spontaneously hypertensive (SH) rats (8-11 x 10(6) RBC/mm(3)) when compared with normotensive rats (6-7 x 10(6) RBC/mm(3)) of the same strain. This increase in red cell count paralleled the increase in body weight and the rise in blood pressure. Since the plasma volume, as measured with labeled albumin was normal, there was an absolute increase in red cells. The hematocrit and hemoglobin content of the blood measured in SH rats were only slightly greater than those found in normotensive rats. However, the mean cell volume (MCV) of the red cells in the SH rats was 45-47 mu(3) as compared with 51-53 mu(3) in normotensive rats.A fourfold increase in 24 hr (59)Fe incorporation into the red cells was found in the SH rats when compared with normotensive controls. The bone marrow of the SH rats showed erythroid hyperplasia. When the SH rats were treated with alpha-methyldopa (Aldomet 200 mg/kg daily, i.p.) the red cell count fell in parallel with the drop in blood pressure. No change in red cell count or blood pressure was observed in normotensive rats treated in the same manner. The erythropoietin titer was high in SH rats, and was undetectable in normotensive rats. These observations suggest a direct relationship between the hypertension and the erythrocytosis mediated by erythropoietin; both are genetically controlled.