Patients.
We investigated a total of 24 patients of either sex. Twelve patients
were newly diagnosed as having GH whereas the remaining 12 patients
were healthy controls. GH was diagnosed based on (1) increased
transferrin saturation (repeatedly higher than 50% at fast) and raised
serum ferritin level (standard assay); (2) hepatocellular hemosiderin
deposits of III-IV grade according to Scheuer et al.12;
(3) hepatic iron index (ratio of liver iron concentration (µmol/g)
and age (years) equal or greater than 2; and (4) no iron loading anemia
and no history of blood transfusions. All patients were homozygous for
cysteine to tyrosine mutation of the hemochromatosis gene (HFE) at
position 282 (C282Y), which is considered the causal mutation of
GH.13 All patients with GH (1) were normotensive (average
of 2 sphygmomanometric measurements, systolic and diastolic values
identified by the first and fifth Korotkoff sounds, respectively), (2)
had no history, physical, or laboratory evidence of cardiovascular
disease, including no visualization of atherosclerotic plaques in the
carotid and femoral arteries at echo color Doppler examination, and (3)
had no other clinically apparent disease and were under no type of
medical treatment. Liver histology was normal except for iron overload
and no other hemochromatosis-related complications were present
(i.e., diabetes, hypogonadism, arthrytes,
cardiomyopathy). All patients agreed to participate in the study after
explanation of its nature and purpose. The study protocol was approved
by the Ethics Committee of our Institution.
Evaluation of Radial Artery Wall Thickness.
Radial artery diameter and intima-media wall thickness were measured by
an A-mode ultrasonic echo-tracking device that recorded the
displacement of the radial artery over the cardiac cycle (NIUS 02;
Omega, Bienne Switzerland; and Capital Medical Services, Paris,
France).14-16 Briefly, the device made use of a highly
focalized transducer operating at a frequency of 10 MHz that was
stereotaxically positioned over the radial artery 2 to 4 cm above the
wrist, direct contact with the skin being prevented by use of a gel as
a medium. With the patient supine and the arm immobile at the heart
level, the transducer was oriented perpendicularly to the longitudinal
axis so that its focal zone was located in the center of the artery and
the backscattered echoes from both the anterior and the posterior walls
could be visualized. The arterial diameter and the posterior wall
thickness were measured when a double peak radiofrequency–ultrasound
signal from both the anterior and the posterior walls became visible,
that is when for both a first high-amplitude signal was followed up by
a relatively silent acoustic zone and then by a second high-amplitude
signal. The first electronic tracker was positioned on the inner
radiofrequency line of the second peak of the anterior wall whereas the
second electronic tracker that measured the intima-media thickness of
the posterior wall was positioned on the inner and outer radiofrequency
lines of the first and the second peaks of the posterior
wall.17,18
The internal diameter of the pulsating radial artery was measured at 50
Hz, the resolution allowing investigators to identify diameter
changes of 0.0025 mm during blood pressure changes from diastole to
systole. The wall thickness of the radial artery was measured by the
electronic tracker signals as seen on the screen. Measurements were
also made of the arterial wall cross sectional area, which was
calculated in mm2 by the formula cross sectional area
= 
(Re2 – Ri2), where Re is the mean
internal radius plus mean wall thickness (mm) and Ri is the mean
internal radius (mm).15,18 Because of the
incompressibility of the arterial mass this avoided the problem of the
influence of instantaneous blood pressure variations on wall thickness.
The technique has been shown to reflect the anatomic thickness of the
radial artery wall by in vitro comparisons.15
Evaluation of Radial Artery D.
Radial artery diameter was measured by the A-mode ultrasonic device
described in the previous section. Briefly, the highly focalized
transducer of 10 MHz was stereotaxically positioned over the radial
artery with gel as a medium, the transducer was oriented
perpendicularly to the longitudinal axis of the vessel based on the
acoustic Doppler signal, and the backscattered echoes from the inner
posterior and anterior walls of the artery were visualized on the
computer screen and electronically digitized (via an analogic/digital
fast transducer) to allow internal diameter variations to be derived at
50 Hz with a spatial resolution of 0.0025 mm.14-16
The device also made use of a photopletysmographic system (Finapres
2003; Ohmeda, Englewood, CO) that allowed blood pressure to be recorded
at 50 Hz from a finger ipsilateral to the radial artery examined, with
an accuracy similar to intra-arterial blood pressure
recording19 and a resolution of 2 mm Hg.19
The concomitant acquisition of continuous arterial diameter and blood
pressure signals allowed investigators to calculate diameter across the
diastosystolic pressure range. The diameter-pressure curve was analyzed
according to its fitting with the arctangent model of Langewouters et
al.,20 which is based on the formula
S = 
[
/2 + tan–1(P – 
/
)]
where S is the cross-sectional area, P is blood pressure
and , , and are 3 optimal parameters describing the spatial
position of the diameter/pressure curve.20 Cross-sectional
compliance (C = 
S/P) was calculated as:
C = / 1/1 + (P – /)2
The same formula was used to calculate cross-sectional D
(cross-sectional compliance divided by vessel area/pressure curve). D
was expressed both as the distensibility blood pressure curve, and as
the individual index, and obtained the integral of the D/blood pressure
curve. The technique and the formula used have been shown to provide
accurate distensibility values for the radial artery by mathematical
modeling approaches.14
Protocol and Data Analysis.
Each patient was asked to come to the outpatient clinic of the San
Gerardo University Hospital in the afternoon after a 24-hour abstinence
from alcohol and caffeine consumption. The protocol of the study was as
follows: (1) each patient was placed in the supine position and fitted
with the radial artery echo-tracking and blood pressure measuring
devices; (2) after a 20-minute interval, radial artery diameter, blood
pressure, and D were continuously measured for 15 minutes, and then
averaged for 5 periods of 30 seconds each taken at 3-minute intervals;
(3) radial artery wall thickness was measured in continuum over a
30-second period, the data collected being also averaged; (4) 9
patients with GH were reevaluated after iron depletion therapy. This
did not consist in chelation therapy but in phlebotomy, which was
performed at monthly intervals for 44 ± 26 months. Each
phlebotomy allowed removal of approximately 400 mL of blood in men and
350 mL in women, to achieve a serum ferritin lower than 30 mg/L, a
transferrin saturation below 30%, and a mild anemia that did not
promptly recover after phlebotomy cessation. The total iron removed was
calculated on the total amount of blood withdrawn during phlebotomies
(1 mL of blood removed corresponding to 0.5 mg of iron). The
revaluation of radial artery was performed after at least 2 months from
the last phlebotomy.
Data obtained in individual patients were averaged and shown as
mean ± SE. The statistical significance of the differences in
mean values was assessed by 2-way analysis of variance. The 2-tailed
t test for unpaired or paired observations was used to
locate differences, respectively, between patients with GH and control
patients, and between patients with GH before and after iron depletion.
A P value < .05 was taken as the level of statistical
significance.
Table 1
shows the demographic,
hemodynamic, and clinical data of the control healthy patients and of
the patients with GH. The 2 groups did not differ significantly for
age, sex representation, sphygmomanometric blood pressure measurements,
and heart rate values. Cardiovascular risk factors (body mass index,
percentage of smokers, lipid profile, diabetes, and family history of
cardiovascular disease) were also similar in the 2 groups. As expected,
serum ferritin concentration and transferrin saturation values were
strikingly greater in the GH group, which was also characterized by an
abnormal increase of liver iron concentration.
Table 1. Demographic, Hemodynamic, and Clinical Data
 | Hemochromatosis | Control |
| n | 12 | 12 |
| Age
(years) | 42 ± 2.3 | 40.6 ± 2.3 |
| Sex
(M/F) | 10/2 | 10/2 |
| BMI
(kg/m2) | 23.4 ± 0.86 | 24.2 ± 1 |
| Systolic blood pressure
(mm Hg)* | 136.8 ± 6.8 | 130.5 ± 4.3 |
| Diastolic blood
pressure (mm Hg)* | 82.2 ± 4 | 79.0 ± 2 |
| Heart rate
(beats/min)† | 64.6 ± 4.1 | 70.7 ± 3 |
| Smoking | 7 | 6 |
| Diabetes | 0 | 0 |
| Plasma cholesterol (mg/dL) | 181.2 ± 13.6 | 147.3 ± 9.3 |
| Plasma triglycerides (mg/dL) | 145.3 ± 19.1 | 107.4 ± 8.5 |
| Family history of cardiovascular disease | 1 | 3 |
| Serum ferritin (µg/L) | 1,390.8 ± 275.4 | 98.6 ± 22.1 |
| Transferrin saturation (%) | 84.9 ± 3.7 | 27.6 ± 3.2 |
| Liver iron concentration (µg/100
mg) | 2,042.4 ± 326.7 | |
NOTE. Data are shown as mean ± SE.
*Sphygmomanometry.
†Palpatory method over 30 seconds. |
Figure
1, left panels, shows that radial
artery wall thickness was significantly greater in patients with GH
than in control patients (+45%), this being the case also for radial
artery wall cross-sectional area (+56%). Radial artery diameter was
similar in the 2 groups and radial artery D was slightly less in
patients with GH than in control patients, the difference being,
however, not statistically significant (Figure
1, right panels), though
visible throughout systodiastolic pressure range
(Fig.
2, left panel).
Fig. 1. Wall thickness, cross-sectional area, diameter, and distensibility
index of the radial artery in patients with GH and in control subjects.
Data are shown as mean ± SE. *P < .05;
**P < .01.
|
|
|
Click on Image to view full size
|
Fig. 2. Distensibility/pressure curves of the radial artery in patients with GH
and in control subjects (left panel) and in patients with GH before and
after iron depletion (right panel). Data are shown as mean ± SE.
*P < .05; **P < .01.
|
|
|
Click on Image to view full size
|
In the 9 patients who reached iron depletion, serum ferritin
concentration decreased from 1,275 mg/L to 27 mg/L, and transferrin
saturation decreased from 86% to 23%, the total iron removed being
11.6 ± 1.7 g. Compared with the pretreatment condition,
blood pressure did not change significantly (systolic/diastolic values:
138.1 ± 9.1/83.7 ± 5.2 mm Hg vs. 137.5 ± 10.2
/80 ± 4.7 mm Hg), this being the case also for heart rate
(69.2 ± 3.2 vs. 68.6 ± 2.7 bpm). In contrast, whereas
radial artery wall thickness and radial artery cross-sectional area
decreased markedly (Fig.
3, left panels),
radial artery diameter did not change and radial artery distensibility
significantly increased (Fig.
3, right panels). The increase was
visible throughout the systodiastolic pressure range (Fig.
2, right
panel).
Fig. 3. Wall thickness, cross-sectional area, diameter, and distensibility
index of the radial artery in patients with GH before and after iron
depletion. Data are shown as mean ± SE. *P <
.05; **P < .01.
|
|
|
Click on Image to view full size
|
Our study shows that in patients with GH, no hypertension, and no
evidence of cardiovascular disease there is (1) a clear-cut increase in
radial artery wall thickness and cross-sectional area with no change in
vessel diameter and (2) this increase is accompanied by some reduction
of radial artery D. It also shows that after iron depletion, radial
artery wall thickness and cross-sectional area decrease whereas radial
artery D increases to values similar to those of normal controls. This
allows us to conclude that in patients with GH, midsize arteries are
characterized by an eccentric hypertrophy and that this structural
abnormality is accompanied by a functional abnormality as well. It also
allows us to conclude that this abnormality is iron dependent and that
it occurs at an early stage of this condition (i.e., before
its cardiovascular and noncardiovascular complications are clinically
visible). This study shows that iron overload causes an early
alteration of arterial wall structure and function in humans.
Our study was not aimed at investigating the precise mechanisms through
which iron causes arterial wall thickening in patients with GH.
However, an increased total collagen content has been found recently in
arteries of a man with -thalassemia major, suggesting a fibrotic
process that was ascribed to the presence of large amounts of tissue
iron and to iron-induced fibrogenesis.21 Furthermore, it
is well known that iron metabolism is causally involved in normal and
malignant cell proliferation and that active oxygen species, whose
production is strongly catalyzed by transition metals, may act as
vascular smooth muscle cell growth factor.4,5,7,22,23
Finally, studies on vascular smooth muscle proliferation in cultured
cells have shown that iron chelation may control vascular smooth muscle
cell proliferation.7 Thus, it is possible that the
arterial wall thickening that occurs in patients with GH originates
from iron-dependent growth of arterial wall tissue. It can be
speculated that this growth may be more evident for connective than for
smooth muscle tissue because connective tissue has a greater elastic
modulus and, thus, can account for the concomitant iron-dependent
reduction of arterial D.
Regardless of the mechanism involved, the arterial wall thickening and
stiffening we have observed in patients with GH has a pathophysiologic
significance. First, a thicker wall implies that the poorly
vascularized intimal layer is even less easily reached by oxygen and
other nutrients through passive diffusion, and, thus, even more easily
undergoes ischemic damage that may facilitate
atherosclerosis.24,25 Second, atherosclerosis is
facilitated also by a stiffer wall that enhances the traumatic effect
of intravascular pressure and makes development and progression of
atherosclerotic lesions more widespread than in vessels in which D is
greater.26 On a pathophysiologic ground this is in line
with the findings that in rabbits iron overload does accelerate the
development of atherosclerosis27,28 and that iron has an
adverse effect on endothelium.29 Epidemiologic data, on
the other hand, are somewhat more controversial because though recent
studies have reported iron to be a risk factor for patients with
coronary heart disease30,31 older pathology reports on
hereditary hemochromatosis do not provide descriptions of large and
midsize arteries either because they were not examined or because they
were normal.32,33 Furthermore, peripheral artery disease
has been found to be uncommon in patients with GH and insulin-dependent
diabetes.34,35 This could be explained by the fact that
iron is not invariably a significant factor in the genesis of
atherosclerosis and cardiovascular complications, or that in patients
with GH the adverse effects of iron are counteracted by some protective
factors. One of these factors could be the most common complication of
hemochromatosis (i.e., cirrhosis, which can reduce
cholesterol synthesis and increase nitric oxide
production).36,37 Indeed, in most older reports patients
had GH in an advanced stage and, thus, with a high prevalence of
cirrhosis.
In our study the only vessel examined was the radial artery. This
represents an advantage because muscle arteries of the size of the
radial muscle are devoid of atherosclerosis and, thus, their structural
and functional changes cannot just be a reflection of an initial
subclinical atherosclerotic process (i.e., a consequence and
not a causative factor in atherosclerosis). It also represents a
limitation, however, because data cannot be easily extrapolated to
larger elastic vessels where alterations in wall thickness and
distensibility sometimes correspond to the middle muscle
ones38 but sometimes do not.16,17 A more
generalized determination of arterial structure and function in GH is
therefore desirable.
Acknowledgments
We thank L'Associazione per lo studio dell'emocromatosi e delle
malattie da sovraccarico di ferro, Monza, for its kind collaboration
with our work.
1. Niederau C, Fisher R, Purschel A, Stremmel W, Haussinger D, Strohmeier G. Long term survival in patients with hereditary hemochromatosis. Gastroenterology 1996;110:1107-1119.
2. Cecchetti G, Binda A, Piperno A, Fargion S, Fiorelli G. Cardiac alterations in 36 consecutive patients with idiopathic haemochromatosis: polygraphic and echocardiographic evaluations. Eur Heart J 1991;12:224-230.
3. Wolfe L, Olivieri N, Sallan D, Colan S, Rose V, Propper R, Freedman MH, et al. Prevention of cardiac diseases by subcutaneous desferroxamine in patients with thalassemia major. N Engl J Med 1985;312:1600-1603.
4. Halliwell B. Current status review: free radicals, reactive oxygen species and human disease: a critical evaluation with special reference to atherosclerosis. Br J Exper Pathol 1989;70:737-757.
5. McCord JM. Oxygen derived free radicals in post ischemic injury. N Engl J Med 1985;312:159-163.
6. Williams RE, Zwieier JL, Flaherty JT. Treatment with desferroxamine ischemia improved functional and metabolic recovery and reduces reperfusion-induced oxygen radical generation in rabbit hearts. Circulation 1991;83:1006-1014.
7. Porreca E, Ucchino S, Di Febbo C, Di Bartolomeo N, Angelucci D, Napolitano AM, Mezzetti A, et al. Antiproliferative effect of desferroxiamine on vascular smooth muscle cells in vitro and in vivo. Arterioscler Thromb Vasc Biol 1994;14:299-304.
8. Abdalla DSP, Campa A, Monteiro MP. Low density lipoprotein oxidation by stimulated neutrophils and ferritin. Atherosclerosis 1992;97:149-159.
9. Pang J, Jiang MJ, Chen YL, Wang FW, Wang DL, Chu SH. Increased ferritin gene expression in atherosclerotic lesions. J Clin Invest 1996;97:2204-2212.
10. Kiechl S, Aichner F, Gerstenbrand F, Egger G, Mair A, Rungger G, Spogler F, et al. Body iron stores and presence of carotid atherosclerosis. Results from the Bruneck Study. Arterioscler Thromb Vasc Biol 1994;14:1625-1630.
11. Salonen JT, Nyyssonen K, Korpela H, Tuomilehto J, Seppanen R, Salonen R. High stored iron levels are associated with excess risk of myocardial infarction in Eastern Finnish men. Circulation 1992;86:803-811.
12. Scheuer PJ, Williams R, Muir AR. Hepatic pathology in relatives of patients with hemochromatosis. J Pathol Bacteriol 1962;84:53-64.
13. Piperno A. Classification and diagnosis of iron overload. Haematologica 1998;83:447-455.
14. Tardy Y, Meister JJ, Perret F, Brunner HR, Arditi M. Non-invasive estimate of the mechanical properties of peripheral arteries from ultrasonic and photoplethysmographic measurements. Clin Phys Physiol Meas 1991;12:39-54.
15. Girerd X, Mourad JJ, Acar C, Heudes D, Chiché S, Bruneval P, Mignot JP, et al. Non-invasive measurement of medium-sized artery wall thickness in humans: in vitro validation. J Vasc Res 1994;31:114-120.
16. Giannattasio C, Rivolta MR, Failla M, Mangoni AA, Stella ML, Mancia G. Large and medium sized abnormalities in untreated and treated hypothyroidism. Eur Heart J 1997;18:1492-1498.
17. Laurent S. Arterial wall hypertrophy and stiffness in essential hypertensive patients. Hypertension 1995;26:355-362.
18. Girerd X, Mourad JJ, Copie X, Moulin C, Acar C, Safar M, Laurent S. Non invasive detection of vascular hypertrophy in untreated hypertensive patients. Am J Hypertens 1994;7:1076-1084.
19. Parati G, Casadei A, Groppelli A, Di Rienzo M, Mancia G. Comparison of finger and intra-arterial blood pressure monitoring at rest and during laboratory testing. Hypertension 1989;13:645-647.
20. Langewouters GJ, Zwart A, Busse R, Wesseling KH. Pressure diameter relationship of segments of human finger arteries. Clin Phys Physiol Meas 1986;7:43-45.
21. Cardoso LEM, Mourao PAS. Compositional and structural alterations of arterial glycosaminoglycans associated with the complications brought about by thalassemia major. A case report. Angiology 1996;47:175-183.
22. Halliwell B, Gutteridge JMC. Oxygen toxicity, oxygen radicals, transition metals and disease. Biochem J 1984;219:1-14.
23. Rao GN, Berk BC. Active oxygen species stimulate vascular smooth muscle cell growth and proto-oncogene expression. Circ Res 1992;70:593-599.
24. Ross R. The pathogenesis of atherosclerosis: an update. N Engl J Med 1986;314:488-500.
25. Ip JH, Fuster V, Badimon L, Badimon J, Taubman MB, Chesebro JH. Syndromes of accelerated atherosclerosis: role of vascular injury and smooth muscle cell proliferation. J Am Coll Cardiol 1990;15:1667-1687.
26. Booth RFG, Martin JF, Honey AC, Hassal DG, Beesley JE, Moncada S. Rapid development of atherosclerotic lesion in the rabbit carotid artery induced by perivascular manipulation. Atherosclerosis 1989;76:257-268.
27. Arraujo JA, Romano EL, Brito BE, Parthe V, Romano M, Bracio M, Montano RF, et al. Iron overload augments the development of atherosclerotic lesions in rabbits. Arterioscler Thromb Vasc Biol 1995;15:1172-1180.
28. Jacob HS. Newly recognized causes of atherosclerosis: the role of microorganisms and of vascular iron overload. J Lab Clin Med 1994;123:808-816.
29. Lekakis J, Papamicheal C, Stamatelopulos K, Cimiponeriu A, Voutsas A, Vemmos K, Mavrikakkis M, et al. Haemocromotasosis associated with endothelial dysfunction: evidence for the role of iron stores in early atherogenesis. Vasc Med 1999;4:147-148.
30. de Valk B, Marx JJ. Iron, atherosclerosis, and ischemic heart disease. Arch Intern Med 1999;159:1542-1548.
31. Horwitz LV, Rosenthal EA. Iron-mediated cardiovascular injury. Vasc Med 1999;4:93-99.
32. Powell L, Jazwinska E, Hallidau J. Primary iron overload In: J Brock, J Halliday, M Pippard, L Powell eds. Iron Metabolism in Health and Disease. London: Saunders. 1994;227-270.
33. MacDonald RA, Mallory K. Haemochromatosis and hemosiderosis: study of 211 autopsied cases. Arch Intern Med 1960;105:686-700.
34. Miller M, Hutchins GM. Haemochromatosis, multiorgan hemosiderosis and coronary artery disease. JAMA 1994;272:231-233.
35. Passa P, Rousselie F, Gauville C, Canivet J. Retinopathy and plasma growth hormone levels in idiopathic haemochromatosis with diabetes. Diabetes 1977;26:113-120.
36. Cicoganani C, Malavolti M, Morselli-Labate AM, Zamboni L, Sama C, Barbara L. Serum lipid and lipoprotein patterns in patients with liver cirrhosis and chronic active hepatitis. Arch Intern Med 1997;157:792-796.
37. Galley HF, Coomansingh D, Webster NR, Brunt PW. Nitric oxide synthase activity is increased in relation to the severity of liver dysfunction. Clin Sci 1998;95:355-359.
38. Giannattasio C, Failla M, Piperno A, Grappiolo A, Gamba P, Paleari F, Mancia G. Early impairment of large artery structure and function in Type 1 diabetes mellitus. Diabetologia 1999;42:987-994.
- From the 1Clinica Medica,
Università di Milano Bicocca and Ospedale San Gerardo,
Monza (MI), Italy; and 2IRCCS
Ospedale San Luca, Milan, Italy.
- Received February 22, 2000
- Accepted June 15, 2000
- Address Reprint request to: Giuseppe Mancia, M.D.,
Clinica Medica, Ospedale S. Gerardo, Via Donizetti 106, 20052 Monza
(MI), Italy. Fax: (39) 39 32 22 74.
Copyright © 2000 American Association for the Study of Liver Diseases
- 0270-9139/00/3203-0019$3.00/0
- doi:10.1053/jhep.2000.16265
| Articles with References to this Article |
TOP |
This article is referenced by these articles:
Iron Overload and Atherosclerosis
Hepatology
September 2000 • Volume 32 • Number 3
Claus Niederau, M.D.
)
)
)
