13 research outputs found
Human prorenin
Our interest in inactive renin originates from the observation that plasma renin
can be activated by several apparently unrelated modes of treatment. Dialysis
of plasma against a pH 3.3 buffer at 0 C for 24 hours followed by another
24 hours of dialysis at neutral pH causes a five-fold increase in renin activity. Renin activity of plasma is also increased, albeit to a lesser extent,
when plasma is stored at 0 C without prior acidification. Finally, the addition
of trypsin and some other proteases, such as plasma kallikrein and plasmin,
to plasma also increases renin activity.
Theoretically, the increase in the enzymatic activity of renin could occur by
several mechanisms:
1) changes of the fluid milieu of the enzyme, for instance changes in pH, certain
metal ions, chaotropic agents,
2) reversible binding of an activator to the active site or to an allosteric site
of the enzyme molecule,
3) destruction of an inhibitor of renin or dissociation of the inhibitor moiety
from a renin-inhibitor complex,
4) alteration of the primary structure of inactive renin by limited proteolysis.
The last possibility of activation by limited proteolysis is in agreement with
the concept of a proenzyme-enzyme conversion. The existence of inactive forms
of renin with a molecular weight greater than that of active renin has been
demonstrated in plasma but the Mr-figures that have been reported are widely
different. Such a concept is also in agreement with the presence of
an additional nucleotide sequence in mouse submandibulary renin and human kidney renin e-DNA, which may correspond toN-terminal amino acid sequences
that belong to the so-called prepropart of renin. Furthermore, there is recent
evidence that the propart amino acid sequence of human kidney prorenin is
also present in inactive renin of human plasma. Prorenin-renin conversion,
within or outside the juxtaglomerular cells where prorenin is synthesized, could
be an important regulatory step in the renin-angiotensin system. With this in
mind we have studied some biochemical aspects of the activation of inactive
renin and addressed the the possible physiological and clinical significance by
measurements of inactive renin in plasma and other body fluids. This thesis
describes our contributions to this subject in the past five year
Improved immunoradiometric assay for plasma renin
BACKGROUND: Our renin IRMA overestimated renin in plasmas with high
prorenin-to-renin ratios. We suspected that the overestimation of renin
was caused less by cross-reactivity of the renin-specific antibody with
prorenin than by a conformational change of prorenin into an enzymatically
active form during the assay. METHODS: Because the inactive form of
prorenin converts slowly into an active form at low temperature, we raised
the assay temperature from 22 degrees C to 37 degrees C, simultaneously
shortening the incubation time from 24 to 6 h. The former IRMA was
performed in <1 working day with these modifications. RESULTS: The
comeasurement of prorenin as renin was eliminated. Reagents were stable at
37 degrees C, and the new and old IRMAs were comparable in terms of
precision and accuracy. The functional lower limit of the assay (4 mU/L)
was below the lower reference limit (9 mU/L). The modified IRMA agreed
closely with the activities measured with an enzyme-kinetic assay. Results
were not influenced by the plasma concentration of angiotensinogen. At
normal angiotensinogen concentrations, the IRMA closely correlated with
the classical enzyme-kinetic assay of plasma renin activity. CONCLUSION:
The modified IRMA, performed at 37 degrees C, avoids interference by
prorenin while retaining the desirable analytical characteristics of the
older IRMA and requiring less time
Human renal and systemic hemodynamic, natriuretic, and neurohumoral responses to different doses of L-NAME
Experimental evidence indicates that the renal circulation is more
sensitive to the effects of nitric oxide (NO) synthesis inhibition than
other vascular beds. To explore whether in men the NO-mediated vasodilator
tone is greater in the renal than in the systemic circulation, the effects
of three different intravenous infusions of NG-nitro-L-arginine methyl
ester (L-NAME; 1, 5, and 25 microg. kg-1. min-1 for 30 min) or placebo on
mean arterial pressure (MAP), systemic vascular resistance (SVR), renal
blood flow (RBF), renal vascular resistance (RVR), glomerular filtration
rate (GFR), and fractional sodium and lithium excretion (FENa and FELi)
were studied in 12 healthy subjects, each receiving randomly two of the
four treatments on two different occasions. MAP was measured continuously
by means of the Finapres device, and stroke volume was calculated by a
model flow method. GFR and RBF were estimated from the clearances of
radiolabeled thalamate and hippuran. Systemic and renal hemodynamics were
followed for 2 h after start of infusions. During placebo, renal and
systemic hemodynamics and FENa and FELi remained stable. With the low and
intermediate L-NAME doses, maximal increments in SVR and RVR were similar:
20.4 +/- 19.6 and 23.5 +/- 16.0%, respectively, with the low dose and 31.4
+/- 26.7 and 31.2 +/- 14.4%, respectively, with the intermediate dose
(means +/- SD). With the high L-NAME dose, the increment in RVR was
greater than the increment in SVR. Despite a decrease in RBF, FENa and
FELi did not change with the low L-NAME dose, but they decreased by 31.2
+/- 11.0 and 20.2 +/- 6.3%, respectively, with the intermediate dose and
by 70.8 +/- 8.1 and 31.5 +/- 15.9% with the high L-NAME dose,
respectively. It is concluded that in men the renal circulation is not
more sensitive to the effects of NO synthesis inhibition than the systemic
circulation and that the threshold for NO synthesis inhibition to produce
antinatriuresis is higher than the threshold level to cause renal
vasoconstriction
High renin and prorenin in plasma and pleural exudate of a patient with the ovarian hyperstimulation syndrome
We present the case of a 35-year-old woman with a severe ovarian hyperstimulation syndrome (OHSS) as a complication of ovulation induction for primary infertility. The clinical picture showed massively enlarged ovaries, pleural effusion and haemoconcentration. She needed a thoracentesis for evacuation of the large pleural effusion. High levels of renin and prorenin were observed in plasma and pleural exudate
Free and total insulin-like growth factor I (IGF-I), IGF-binding protein-1 (IGFBP-1), and IGFBP-3 and their relationships to the presence of diabetic retinopathy and glomerular hyperfiltration in insulin-dependent diabetes mellitus
The existing literature on serum insulin-like growth factor I (IGF-I)
levels in insulin-dependent diabetes mellitus (IDDM) is conflicting. Free
IGF-I may have greater physiological and clinical relevance than total
IGF-I. Recently, a validated method has been developed to measure free
IGF-I levels in the circulation. Serum free and total IGF-I, IGF-binding
protein-1 (IGFBP-1), and IGFBP-3 levels were measured in 56
insulin-treated IDDM patients and 52 healthy sex- and age-matched
controls. Diabetic retinopathy was established by direct fundoscopy. In 54
IDDM patients, the glomerular filtration rate (GFR) and effective renal
plasma flow were calculated from the clearance rate of [125I]iothalamate
and [131I]iodohippurate sodium. Fasting free IGF-I, total IGF-I, and
IGFBP-3 levels were significantly lower in IDDM patients than in age- and
sex-matched healthy controls (free IGF-I, P < 0.005; total IGF-I, P <
0.001; IGFBP-3, P = 0.001), whereas IGFBP-1 levels were higher (P <
0.001). In IDDM subjects, decreases in free IGF-I, total IGF-I, and
IGFBP-3 levels with age were observed (free IGF-I, r = -0.27 and P = 0.05;
total IGF-I, r = -0.52 and P < 0.001; IGFBP-3, r = -0.37 and P = 0.005).
Free IGF-I was inversely related to fasting glucose in IDDM subjects (r =
-0.35; P = 0.01), whereas the relationship between total IGF-I and fasting
glucose did not reach significance (r = -0.27; P = 0.06). Age-adjusted
free IGF-I levels were significantly higher (P < 0.05) in IDDM subjects
with retinopathy than in subjects without retinopathy after adjustment for
age. Total IGF-I and IGFBP-3 levels were positively related to GFR (total
IGF-I, r = 0.35 and P < 0.05; IGFBP-3, r = 0.28 and P < 0.05). Both of
these differences lost significance after adjustment for age. Free IGF-I,
total IGF-I, and IGFBP-3 levels were lower and IGFBP-1 levels were higher
in insulin-treated IDDM subjects compared to those in age- and sex-matched
controls. Free IGF-I, total IGF-I, and IGFBP-3 levels decreased
significantly with age in IDDM subjects. Age-adjusted free IGF-I levels in
subjects with diabetic retinopathy were higher than those in subjects
without diabetic retinopathy. Total IGF-I and IGFBP-3 levels were
positively related to GFR in IDDM subjects, but these relations were lost
after adjustment for age. Measurement of serum free IGF-I levels in IDDM
subjects did not have clear advantages compared to that of total IGF-I,
IGFBP-1, and IGFBP-3 levels. Serum IGF-I and IGFBPs reflect their tissue
concentrations to a various degree. Consequently, extrapolations
concerning the pathogenetic role of the IGF/IGFBP system in the
development of diabetic complications at the tissue level remain
speculative
High-affinity prorenin binding to cardiac man-6-P/IGF-II receptors precedes proteolytic activation to renin
Mannose-6-phosphate (man-6-P)/insulin-like growth factor-II
(man-6-P/IgF-II) receptors are involved in the activation of recombinant
human prorenin by cardiomyocytes. To investigate the kinetics of this
process, the nature of activation, the existence of other prorenin
receptors, and binding of native prorenin, neonatal rat cardiomyocytes
were incubated with recombinant, renal, or amniotic fluid prorenin with or
without man-6-P. Intact and activated prorenin were measured in cell
lysates with prosegment- and renin-specific antibodies, respectively. The
dissociation constant (K(d)) and maximum number of binding sites (B(max))
for prorenin binding to man-6-P/IGF-II receptors were 0.6 +/- 0.1 nM and
3,840 +/- 510 receptors/myocyte, respectively. The capacity for prorenin
internalization was greater than 10 times B(max). Levels of internalized
intact prorenin decreased rapidly (half-life = 5 +/- 3 min) indicating
proteolytic prosegment removal. Prorenin subdivision into man-6-P-free and
man-6-P-containing fractions revealed that only the latter was bound.
Cells also bound and activated renal but not amniotic fluid prorenin. We
concluded that cardiomyocytes display high-affinity binding of renal but
not extrarenal prorenin exclusively via man-6-P/IGF-II receptors. Binding
precedes internalization and proteolytic activation to renin thereby
supporting the concept of cardiac angiotensin formation by renal prorenin
Prorenin accumulation and activation in human endothelial cells: importance of mannose 6-phosphate receptors
ACE inhibitors improve endothelial dysfunction, possibly by blocking
endothelial angiotensin production. Prorenin, through its binding and
activation by endothelial mannose 6-phosphate (M6P) receptors, may
contribute to this production. Here, we investigated this possibility as
well as prorenin activation kinetics, the nature of the
prorenin-activating enzyme, and M6P receptor-independent prorenin binding.
Human umbilical vein endothelial cells (HUVECs) were incubated with
wild-type prorenin, K/A-2 prorenin (in which Lys42 is mutated to Ala,
thereby preventing cleavage by known proteases), M6P-free prorenin, and
nonglycosylated prorenin, with or without M6P, protease inhibitors, or
angiotensinogen. HUVECs bound only M6P-containing prorenin (K(d) 0.9+/-0.1
nmol/L, maximum number of binding sites [B(max)] 1010+/-50
receptors/cell). At 37 degrees C, because of M6P receptor recycling, the
amount of prorenin internalized via M6P receptors was >25 times B(max).
Inside the cells, wild-type and K/A-2 prorenin were proteolytically
activated to renin. Renin was subsequently degraded. Protease inhibitors
interfered with the latter but not with prorenin activation, thereby
indicating that the activating enzyme is different from any of the known
prorenin-activating enzymes. Incubation with angiotensinogen did not lead
to endothelial angiotensin generation, inasmuch as HUVECs were unable to
internalize angiotensinogen. Most likely, therefore, in the absence of
angiotensinogen synthesis or endocytosis, M6P receptor-mediated prorenin
internalization by endothelial cells represents prorenin clearance
Do older hospital patients recognize adverse drug reactions?
OBJECTIVE: To establish the relationship between subjective complaints of
side effects of drugs and the objective presence of adverse drug reactions
in older patients. DESIGN: Observational cross-sectional study. SETTING:
Five medical wards at the University Hospital Rotterdam Dijkzigt.
SUBJECTS: Patients aged 70 and over admitted to the general medical wards
over a 3-month period. METHODS: Statistical comparison and Kramer's
algorithm. RESULTS: Of 106 patients, 102 used medication, and 93 of these
were able to report whether they believed they were experiencing drug side
effects. Thirty-six [39% (95% confidence interval 28.8-48.6)] believed
that they were experiencing side effects and the number of diagnoses per
patient and the proportion of patients with chronic obstructive pulmonary
disease was higher in these 36 'complainers' than in the group of the
'non-complainers'. We found a correct opinion (true positive and negative)
about the objective presence or absence of mild or severe adverse drug
reactions in 79% (95% confidence interval 70.2-86.8). Asking the patient
about side effects of drugs had a sensitivity of 0.70 and a specificity of
0.85 patients. The severe adverse drug reactions in 21 patients were not
recognized by 14 of them. CONCLUSION: At hospital admission, older
patients should be asked about drug side effects because they are often
correct in recognizing them. However, severe adverse drug reactions are
not easily recognized