The serum angiotensinogen concentration and variants of the angiotensinogen gene in white and black children.

The T235 allele of the angiotensinogen gene (AGT) has been associated with hypertension. Blood pressure increases faster over time in black children than in white children, and in adults hypertension is more prevalent in blacks. We sought evidence for a role for angiotensinogen to contribute to racial differences in blood pressure in a study of 148 white and 62 black normotensive children (mean age, 14.8 yr). The frequency of the T235 allele was 0.81 in blacks and 0.42 in whites (chi 2 = 77.3, P = 0.0001). The mean angiotensinogen level was 19% higher in blacks than in whites (P = 0.0001 for males, P = 0.004 for females). Genotype was positively related to serum angiotensinogen in white children (P = 0.0001 for males, P = 0.004 for females), but a similar relationship was absent in blacks where the frequency of M235 may have been too low to discern an association. Longitudinal blood pressure (measured twice yearly) adjusted for body mass index showed a marginally significant relationship to the angiotensinogen level (P = 0.07). An independent relationship of serum angiotensinogen with body mass index (P = 0.0001) and race (P = 0.0003) was also observed. In summary, T235 was more frequent, and the level of angiotensinogen was higher in blacks than in whites. Such a racial difference in the renin-angiotensin system may contribute to the disparity in blood pressure levels of white and black young people.

Doxorubicin clearance in the obese.

A study was carried out to examine the effect, if any, of obesity on doxorubicin pharmacokinetics. Body weight was found to be significantly related to doxorubicin clearance (r = -.75; P less than .001) and elimination half-life (r = .62; P = .003). Thus, the contribution of obesity on pharmacokinetics of antineoplastic agents should be taken into consideration in the analysis of clinical data with respect to toxicity and tumor response. Twenty-one patients were studied with their first course of doxorubicin (50 to 70 mg/m2) administered as a 60-minute intravenous (IV) infusion. Patients were divided into three groups on the basis of percentage of ideal body weight (IBW): normal (less than 115% IBW), mildly obese (115% to 130% IBW), and obese (greater than 130% IBW). Blood samples were collected up to 48 hours after the infusion and analyzed for doxorubicin and its metabolite, doxorubicinol, by high performance liquid chromatography. Doxorubicin area under the curve (AUC) was greater in obese than in normal patients (2,209 v 1,190 ng h/mL; P less than .05), yielding correspondingly reduced systemic clearance of the agent in obese patients (891 v 1,569 mL/min; P less than .001). The mean elimination half-life (T1/2) was 20.4 hours in the obese patients and 13.0 hours in the normal patients. The apparent volume of distribution (Vss) was not significantly different among the three groups of patients, indicating that the prolonged T1/2 in the obese patients is due to the reduction in clearance. The AUC and T1/2 of doxorubicinol were similar among all patient groups.

High molecular weight angiotensinogen levels in hypertensive pregnant women.

An apparent high molecular weight angiotensinogen (H-Aogen) can be separated from the usually predominant low molecular weight angiotensinogen (L-Aogen) by gel filtration of plasma. H-Aogen has been quantitated in plasma from normotensive menstruating women, estrogen treated women, normotensive pregnant women, women with pregnancy-induced hypertension (PIH), and women whose preexisting hypertension was exacerbated during pregnancy. When expressed as a percent of the total angiotensinogen, the H-Aogen levels were: menstruating women 4%, estrogen-treated women 10%, normotensive pregnant women 16%, women with PIH 25%, and pregnant women with exacerbated hypertension 28%. A significant difference (p less than 0.01) was found between H-Aogen concentration in normotensive pregnant women and women with PIH (1.10 +/- 0.12 and 1.73 +/- 0.16 micrograms angiotensin I/ml plasma respectively). In some hypertensive pregnant women, H-Aogen is the predominant form of angiotensinogen. Thus, H-Aogen should be recognized as a component of the renin-angiotensin system.

Association of the angiotensinogen gene to serum angiotensinogen in blacks and whites.

A variant of the angiotensinogen gene (AGT) that encodes for threonine at codon 235 (T235) has been associated with a higher serum angiotensinogen concentration and with hypertension in white subjects. The frequency of T235 is about two times higher in blacks than whites, suggesting that AGT may contribute to the susceptibility to hypertension in blacks more than it does in whites. However, an association of T235 with angiotensinogen level or blood pressure has not been observed in blacks, possibly because the high prevalence of T235 makes it insufficiently informative as a marker. For this reason, we undertook to further differentiate the T235 carrier state by constructing haplotypes with alleles in the 5' upstream region of AGT. One such haplotype, -1074t;T235, showed a significant association with angiotensinogen level in a cohort of black and white children and adolescents (76 blacks, mean age = 12.3 +/- 2.0 [SD] years; 139 whites, mean age = 12.4 +/- 1.8 years). With a linear regression model, the level of serum angiotensinogen was significantly related to body mass index (P = .0017) and the haplotype (P = .0001). Within specific race groups, the haplotype was significantly related to serum angiotensinogen in both the blacks (P = .0277) and whites (P = .0001). The mean level of angiotensinogen was higher in the blacks carrying a single copy of the haplotype than in those without the haplotype (1472.2 +/- 68.4 versus 1274.9 +/- 46.7 nmol angiotensin I/L), a difference that was marginally significant (P = .0609). In the whites, the level of angiotensinogen was also higher in carriers of a single copy than in those with no copy (1527.9 +/- 71.2 versus 1099.2 +/- 20.1 nmol angiotensin I/L) (P = .0003). Serum angiotensinogen level did not increase with two copies of the haplotype, but in each racial group, there were only four individuals who were homozygous. The haplotype showed a marginally significant relation (P = .0757) to the mean of longitudinally determined diastolic pressures adjusted for body mass index, race, sex, and age. In summary, using a haplotype to differentiate further the T235 carrier state, we observed an association of genotype with serum angiotensinogen level and blood pressure in blacks and whites. The findings suggest that AGT may play an important role in blood pressure regulation in both racial groups.

Serum angiotensinogen concentration in relation to gonadal hormones, body size, and genotype in growing young people.

Multiple factors are thought to influence the level of circulating angiotensinogen (AGT). We showed previously that the serum AGT concentration was significantly related to body mass index (BMI) in a cohort of young people. In the present study, we studied whether levels of the gonadal hormones estradiol and testosterone might also predict the AGT level and might contribute to the BMI effect, since both the production of these hormones and BMI increase with age. In boys (n=127; mean+/-SD age, 14.7+/-1.9 years) and girls (n=104; age, 14.8+/-1.9 years) studied as a single group, we found a significant association of AGT level with level of estradiol (P=0.015) after adjustment for haplotype, age, race, testosterone concentration, and BMI. In girls studied alone, the level of AGT showed a significantly positive relation to level of testosterone (P=0.043), possibly a result of peripheral conversion of testosterone to estradiol, after adjustment for haplotype, age, race, estradiol concentration, and BMI. In boys, on the other hand, the level of testosterone was inversely related to AGT concentration (P=0.019), again after making adjustments for the other variables. Finally, in pairs of subjects matched for BMI, age, race, and gender where 1 member of each pair had either 1 or 2 copies of an AGT gene haplotype (T235 and -1074t) and the other member had no copy, the level of AGT was higher in the carrier of a haplotype in 24 of the 34 pairs (P<0.001). In conclusion, gonadal hormones are an additional influence on the circulating level of AGT in growing young people. In addition, with matching for BMI and other covariates, there is a strong association of AGT genotype with the serum level of AGT, emphasizing the importance of AGT gene expression as a determinant of the circulating level of AGT.

Plasma renin concentration and phenobarbital levels in patients with epilepsy.

Plasma renin activity, renin concentration, and renin substrate were measured in patients on long-term anticonvulsant medication. An inverse correlation was observed between phenobarbital levels and renin activity or concentration. There was a positive correlation between phenobarbital and renin substrate. Since elevation of renin substrate should have resulted in increased renin activity, it is possible that phenobarbital exerts 2 effects on the renin system: induction of renin substrate synthesis and suppression of renin release.

Pharmacokinetics and pharmacodynamics of doxorubicin in patients with small cell lung cancer.

The pharmacokinetics and pharmacodynamics of doxorubicin and its metabolite, doxorubicinol, were studied in 35 adult (mean age, 66 1/2 years) patients with small lung cell cancer after a 1-hour intravenous infusion at a dose ranging from 45 to 72 mg/m2. All patients also received concomitant therapy with cyclophosphamide and vincristine. Serum concentrations were sampled to 48 hours after dosing. Wide interpatient variability was observed for all pharmacokinetic parameters with coefficients of variation for apparent volume of distribution, clearance, and area under the curve (AUC) of 62%, 65%, and 65%, respectively. Four patients with impaired liver function showed a significant (p < 0.05) decrease in clearance (239 versus 666 ml/min/m2) and increases in AUC (4610 versus 1834 ng.hr/ml) and elimination half-life (49.3 versus 25.6 hours) compared with patients with normal hepatic function. A significant relationship was found between systemic exposure of doxorubicin (defined by AUC) and surviving factor of white blood cells (r = 0.57, p = 0.0025). No relationships were noted between doxorubicinol exposure and surviving factor of white blood cells or platelets. These findings show the important relationship between systemic exposure of doxorubicin and the degree of myelosuppression in patients with small cell lung cancer.

An ocular renin-angiotensin system. Immunohistochemistry of angiotensinogen.

The circulating renin-angiotensin system (RAS) is an important determinant in maintaining adequate systemic blood pressure, and it also may modify organ-specific blood flow. All recognized RAS components have been identified in the eye. In this study, angiotensinogen (ANG) was localized using an affinity-purified antibody and paraffin sections of seven human eyes. An antibody for human serum albumin was used for comparison. The ANG was present selectively in the cytoplasm of the nonpigmented ciliary epithelium (NPCE), more prominently in the pars plana than in the pars plicata. Both ANG and albumin were present in the blood vessel lumina of the uvea and retina. Both antibodies also stained perivascular tissue in the uvea, but not in the retina, reflecting the relative tightness of blood-tissue barriers. The detection of ANG in the NPCE may be significant in view of previous descriptions localizing prorenin and angiotensin-converting enzyme in the same cell layer.

Regional distribution of the angiotensinogens in human placentae.

Angiotensinogen is the substrate for the enzyme renin. In addition to the well-characterized low molecular weight forms of angiotensinogen, a high molecular weight form (HMrA) comprises about 15 per cent of the total angiotensinogen in plasma of pregnant women. In the current study the different forms of angiotensinogen were quantitated in different regions of the placenta. The highest concentration of total angiotensinogen was found in the chorion laeve followed by the amnion. It was found that HMrA was the major form of angiotensinogen in all regions of the placenta, accounting for 72, 67, 63, and 59 per cent of the total angiotensinogen in the chorion laeve, chorion plate, amnion, and chorion frondosum, respectively. There was a significant correlation between the distribution of total renin and HMrA. This data in conjugation with previous data indicate that HMrA should be considered a significant component of the renin angiotensin system in the human pregnant state. It is suggested that HMrA may be a component of a tissue specific renin angiotensin system that occurs in the placenta.

Importance of preoperative myocardial glycogen levels in human cardiac preservation. Preliminary report.

One hundred seventeen patients undergoing elective coronary bypass were divided into four groups according to prebypass myocardial glycogen levels and the use of potassium chloride cardioplegia. Myocardial glycogen levels were enhanced with a preoperative fat loading diet and overnight glucose loading. The control group (n = 27) which had mean cardiac glycogen levels of 750 mg/100 gm heart weight and no cardioplegia, had a transmural myocardial infarct rate of 14.4%; 35% had severe atrial arrhythmias 65% had severe ventricular arrhythmias, and 31% had severe vasopressor dependence. The group (n = 30) with low cardiac glycogen (736 mg/100 gm) and with potassium chloride cardioplegia had an infarct rate of 6.4%; 6.7% had severe atrial arrhythmias, 18% had severe ventricular arrhythmias, and 16.7% had severe vasopressor dependence. However, the group (n = 26) which had high cardiac glycogen levels (1,208 mg/100 gm) and no cardioplegia had no myocardial infarctions; 3.8% had severe atrial arrhythmias, 27% had severe ventricular arrhythmias, and only 7.8% had severe vasopressor need. The group (n = 34) which had high glycogen levels (1,516 mg/100 gm) and potassium chloride cardioplegia did best of all with no myocardial infarctions or no severe atrial arrhythmias; 14% had severe ventricular arrhythmias and 2.81% severe vasopressor need. The lessening of vasopressor dependence and severe atrial and ventricular arrhythmias were significant by chi square contingency tables at p less than 0.05 and p less than 0.001, respectively. One cardiac-related death each occurred in the two groups with low glycogen and none in those with high glycogen levels. This suggests that better preoperative cardiac nutrition as represented by enhanced cardiac glycogen helps that heart tolerate anoxic stress whether cardioplegia is utilized or not and is additive to potassium chloride cardioplegia.

Angiotensin-converting enzyme in farmer's lung.

Serum levels of angiotensin-converting enzyme were measured in a group of patients with farmer's lung and a group of precipitin-positive subjects with no history of farmer's lung. The levels did not differ significantly from control subjects matched for age and smoking history. The mean serum level of angiotensin-converting enzyme in a group of acutely ill patients with farmer's lung was significantly reduced. An acute challenge of three patients with Micropolyspora faeni did not produce an increase in serum levels of angiotensin-converting enzyme. These studies suggest that an increased serum level of angiotensin-converting enzyme can be a diagnostic aid in making a differential diagnosis between sarcoidosis and farmer's lung.

Quantitation of five forms of high molecular weight angiotensinogen from human placenta.

In addition to the well characterized predominant form of plasma angiotensinogen, which will be termed low molecular weight angiotensinogen (LMrA), significant quantities of a high molecular weight angiotensinogen (HMrA) occur in the human pregnant state. HMrA is the predominant form of angiotensinogen in the placenta and amniotic fluid. The ratio of HMrA/LMrA is elevated in approximately half of the women who develop pregnancy induced hypertension (PIH). This report documents a simple two step methodology for the separation and quantitation of different forms of HMrA. The two steps are gel filtration and ion exchange chromatography. The application of this methodology to amniotic fluid and plasma from normotensive pregnant women and extracts of amnion, chorion, and placental tissue has shown five distinct forms in the tissue extracts and amniotic fluid, but only three significant forms in plasma. The elution position of all forms of HMrA are highly reproducible and are the same for each tissue or fluid studied, thus lending support to the concept that the three forms seen in plasma of normotensive pregnant women are the same as three of the forms of HMrA seen in extracts of placental tissue.

Immunochemical comparison of high molecular weight angiotensinogen from amniotic fluid, plasma of men, and plasma of pregnant women.

A high molecular weight form of angiotensinogen (HMrA) is present in low quantities in plasma from men, in moderate quantities in plasma from pregnant women, and in larger quantities in amniotic fluid. It was shown that immobilized IgG from anti-low molecular weight angiotensinogen (LMrA) sera effectively removed HMrA from all three sources. Immunoblots of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) using anti-LMrA sera showed that HMrA from each of the three sources contained a subunit that was identical to LMrA with respect to molecular weight and heterogeneity. Thus, no difference in the HMrA from these three sources could be demonstrated. It is concluded that HMrA contains at least one subunit that is very similar if not identical to LMrA.

A study of the temporal relationship between plasma high molecular weight angiotensinogen and the development of pregnancy-induced hypertension.

In addition to the normally prevalent low molecular weight angiotensinogen (LMrA), significant quantities of a high molecular weight angiotensinogen (HMrA) are present in the human pregnant state. Previous studies have documented that 47% of women who develop pregnancy-induced hypertension (PIH) have a significantly elevated plasma HMrA/LMrA ratio. The purpose of this study is to establish whether or not the increase in the HMrA/LMrA ratio precedes the development of hypertension. Serial plasma samples were collected from a group of women throughout their pregnancy. High molecular weight angiotensinogen and LMrA levels in the samples from these women were determined. Fifteen of these women developed PIH. Seven women in the PIH group had a significantly elevated plasma HMrA/LMrA ratio. There was no consistent relationship between the elevation of the HMrA/LMrA ratio and the onset of hypertension. Three women had an elevated HMrA/LMrA ratio before the development of hypertension. In one woman the two events occurred simultaneously, and in three women the HMrA/LMrA ratio was elevated only after the development of hypertension. The current study shows that the development of hypertension during pregnancy is not the primary biologic signal for elevation of the plasma HMrA/LMrA ratio. Other parameters associated with fetal distress or abnormal development of placental circulatory systems must be involved in increasing the HMrA/LMrA ratio. It is proposed that the elevation of the HMrA/LMrA ratio is a mechanism by which the placental tissue specific renin-angiotensin system is attenuated.

Quantitation of the five forms of plasma high molecular weight angiotensinogen in women with pregnancy-induced hypertension.

In the human pregnant state a high molecular weight form of angiotensinogen (HMrA) is present in significant quantities in addition to the usual low molecular weight angiotensinogen (LMrA). In a previous study involving a small number of white women, it was found that women who had developed pregnancy-induced hypertension (PIH) had significantly higher levels of plasma HMrA. It has been determined that there are five isoforms of HMrA. The objectives of this study were to expand the previous study with the inclusion of black women and to determine which isoform(s) of plasma HMrA are elevated in PIH. Plasma LMrA and HMrA were quantitated in 24 normotensive pregnant women and 65 women with PIH. The PIH group had higher levels of HMrA and somewhat lower levels of LMrA than the normotensive group. The HMrA/LMrA ratio was elevated in 47% of the PIH group. The five isoforms of HMrA were quantitated in plasma from 10 white women with PIH, 10 black women with PIH, and 6 normotensive pregnant white women. Half of both the white and black women with PIH had an elevated HMrA/LMrA ratio. The relative proportion of the HMrA isomers was similar in all groups. These studies show that half the women with PIH have a distinct abnormality in their renin angiotensin system. Both white and black women show this abnormality. In those women who have an elevated total HMrA, all five isoforms of HMrA are equally elevated.

The disposition of doxorubicin on repeated dosing.

Twelve cancer patients (aged 49-74 years) receiving doxorubicin (66 +/- 8 mg/m2) as a 1-hour intravenous infusion had serial serum samples (0-48 hours) obtained after the first and second courses of therapy. Mean number of days between courses was 24.3, and all patients had normal liver function. Patients received the same concomitant antineoplastic agents and doses in both courses. Doxorubicin and doxorubicinol concentrations were assayed by high-performance liquid chromatography and fitted to a two- or three-compartment infusion model. White blood cell and platelet toxicity were evaluated as (initial--nadir/initial)* 100. Differences in pharmacokinetic parameters were determined by a paired t test. Wide intrapatient and interpatient variability was seen between therapeutic courses. A significant decrease in the apparent volume of distribution of the central compartment (Vc = 16.6 versus 10.4 L/m2; P < .05), and a nonsignificant decrease in clearance (CL = 748 versus 658 mL/min/m2) was observed on the second course of therapy. Doxorubicinol area under the curve and elimination half-life were similar between courses. Extensive chemotherapy-induced changes in white blood cell and platelet counts were observed but were similar in degree for courses 1 and 2. These data suggest that higher initial doxorubicin concentrations on the second course of therapy are secondary to an alteration in distribution volume (Vc). In this subset of patients, however, these changes were not associated with an increase in hematologic toxicity.

Can the frequency of myocardial infarction be reduced during coronary artery operations.

A prospective experiment was carried out in 56 patients undergoing coronary artery revascularization to determine whether those having a glucose-insulin-potassium (GIK) perfusion during the procedure would have fewer myocardial infarctions (MI) compared with patients given a control perfusion of Normosol-R. Six patients (11%) developed an MI, defined as a 24-hour creatine phosphokinase MB isoenzyme value of 100 IU per liter or greater. Four (13%) had control perfusions and 2(8%) had GIK perfusion. One MI in a double-graft recipient who had GIK perfusion occurred because of a technical surgical error; therefore, the corrected MI rate was 13% in control patients compared with 4% for the GIK group. These data, as well as the more frequent spontaneous defibrillation in patients who had GIK perfusion, suggest that GIK was of benefit.

Multi-drug resistance in chronic lymphocytic leukemia.

We evaluated 45 chronic lymphocyte leukemia (CLL) patients for the presence of multi-drug resistance (MDR) by the ex vivo techniques: 1) a functional assay utilizing doxorubicin (dox) retention with modulation; 2) a cytotoxicity assay (MTT) with modulation; 3) and four monoclonal antibodies. Ex vivo tests were correlated with disease stage and prior treatment, and were repeated as patients became resistant to alkylating agents, fludarabine and VAD chemotherapy (infusion of vincristine, dox, and oral dexamethasone). The majority of patients (64.4%) were in early stage and were untreated (62.2%). P-glycoprotein (p-gp 170) was detected most frequently by the monoclonal antibody MRK-16 (48%) and by functional modulation of dox retention by PSC-833 (40.6%) and by functional modulation of the MTT assay with vincristine (0.29) and dox (0.39) with PSC-833 at 1.0 microg/mL. Functional modulation of dox retention with PSC-833 was significantly associated with stage, but not with either the MTT assay or any of the monoclonal antibodies. None of the tests correlated with prior chlorambucil treatment. Correlation of dox retention with the monoclonal antibodies was mild to moderate and became stronger following chlorambucil treatment. Three patients who became resistant to VAD were found to express p-gp 170. We conclude that MDR can frequently be detected in patients with CLL. Furthermore, the expression of p-gp 170 increases with advancing stage, but not prior alkylating agent therapy. The functional expression of p-gp 170 increases with advancing stage and prior treatment and correlates well with monoclonal antibody detection (especially MRK-16). Patients who become resistant to VAD more frequently express p-gp 170 by a variety of techniques. PSC-833 is a more potent modulator of MDR than cyclosporin-A (CsA) ex vivo, and correlates better with stage of disease.

The effect of proteolytic activity on plasma renin activity assay.

Carrier proteins are used in the radioimmunoassay of angiotensin I. It has been shown that the proteolytic activity found in commercial lysozyme will falsely elevate the observed plasma renin activity. This effect is probably due to the fact that human plasma contains proteinase inhibitors which inhibit this activity during radioimmunoassay of angiotensin I. This proteolytic activity can be inhibited by diisopropyl fluorophosphate or by heating lysozyme.

High molecular weight angiotensinogen: a pregnancy associated protein.

Although it was previously recognized that human amniotic fluid contained appreciable quantities of angiotensinogen, it remained unknown what fraction of the total is high molecular weight angiotensinogen. Fractionation of human amniotic fluid showed that high molecular weight angiotensinogen represents the major component of total angiotensinogen; 58% for 11 normotensive pregnant women and 67% for 12 hypertensive pregnant women. In contrast to plasma where high molecular weight angiotensinogen is elevated in hypertensive pregnant women as compared to normotensive pregnant women, no such difference exists for amniotic high molecular weight angiotensinogen. Serum and amniotic fluid high molecular weight angiotensinogen were compared in six subjects, and no significant correlation was found. In fetal cord blood, high molecular weight angiotensinogen represented only 5.8% of the total angiotensinogen. The site of synthesis of high molecular weight angiotensinogen remains unknown but these data suggest that it is produced in the placenta or in the maternal uterine tissue. Therefore, we propose that human high molecular weight angiotensinogen should be classified as a pregnancy-associated protein.