Nephrol Dial Transplant (2001) 16 wSuppl 5x:35±39
Erythropoietin and iron: the role of ascorbic acid
Der-Cherng Tarng1,2, Tung-Po Huang1,2 and Yau-Huei Wei3
1
Division of Nephrology, Department of Medicine, Taipei Veterans General Hospital, 2Faculty of Medicine and
Department of Biochemistry and Center for Cellular and Molecular Biology, National Yang-Ming University
School of Medicine, Taipei, Taiwan
3
Abstract. Provision of suf®cient available iron is a
prerequisite to ensure the optimal response to recombinant human erythropoietin (rHuEpo). Functional
iron de®ciency (a state when iron supply is reduced to
meet the demands for increased erythropoiesis) is the
common cause of rHuEpo hyporesponsiveness in
dialysis patients who have normal iron status, even
when they are iron-overloaded. Iron supplementation
is not justi®ed for this hyporesponsiveness in patients
with iron overload due to the potential hazards
of iron overload aggravated by intravenous iron
therapy. Furthermore, in vivo studies indicated that
the promising effect of intravenous iron medication to
overcome iron-de®cient erythropoiesis is not observed
in iron-overloaded haemodialysis (HD) patients.
Ascorbic acid, a water-soluble antioxidant as well as
a reducing agent, has a number of associations with
iron metabolism. Recent research highlights that
ascorbic acid can potentiate the mobilization of iron
from inert tissue stores and facilitates the incorporation of iron into protoporphyrin in iron-overloaded
HD patients being treated with rHuEpo. Interest has
turned towards the use of ascorbic acid as an adjuvant
therapy in this ®eld. This review focuses on the
improvement of rHuEpo response by administration
of ascorbic acid and discusses its clinical implications
and potential issues for nephrologists.
Keywords: ascorbic acid; functional iron de®ciency;
iron overload; recombinant human erythropoietin
Introduction
The potential role of adjuvant therapies in enhancing
the effectiveness of recombinant human erythropoietin
(rHuEpo) in patients with end-stage renal disease
Correspondence and offprint requests to: Der-Cherng Tarng, Division
of Nephrology, Department of Medicine, Taipei Veterans General
Hospital, No. 201, Section 2, Shih-Pai Road, Taipei 112, Taiwan.
#
(ESRD) has received increasing attention in recent
years. The important reason for adjuvant therapies
is that they may help to overcome rHuEpo hyporesponsiveness, allowing patients to achieve increased
haemoglobin concentration and derive more costeffectiveness and greater clinical bene®ts from rHuEpo
treatment. Recent research highlights the usefulness
of adjuvant therapies, including vitamins, hormones
and cytokines w1x. Some vitamins may generate an
ongoing supply of healthy red blood cells w2±5x. This
review emphasizes a role for vitamin C as an adjuvant
therapy in dialysis patients being treated with rHuEpo
and discusses its possible mechanisms and clinical
implications, and the potential issues of ascorbic acid
medication.
rHuEpo hyporesponsiveness in dialysis patients
with iron overload
Insuf®cient iron storageuavailability is the most
common cause of suboptimal response to rHuEpo.
Guidelines for the management of rHuEpo hyporesponsiveness suggest a serum ferritin level of 100 mgul
as the diagnostic threshold for iron de®ciency in haemodialysis (HD) patients w6,7x. Studies from Fishbane
et al. w8x and Tarng et al. w9x indicated that the quantity
of available iron for prompt erythropoiesis is inadequate in most HD patients when current guidelines
for serum ferritin and transferrin saturation (TS)
are used. Furthermore, cumulative data recommend
that it is crucial to maintain serum ferritin )300 mgul
for the optimal response since there is great need for
iron in Epo-stimulated erythroid progenitor cells
w10±13x. However, HD patients with increased iron
stores may exhibit rHuEpo hyporesponsiveness due
to impaired iron availability. The ferritin level to
indicate iron overload in dialysis patients is suggested
to be 500±1000 mgul w14±16x. Iron overload not only
increases cardiovascular and infectious morbidity
w17,18x, but may sometimes cause relative resistance
to rHuEpo in HD patients. The mechanism of this
hyporesponsiveness remains unclear, but inadequate
2001 European Renal Association±European Dialysis and Transplant Association
36
iron mobilization and defective iron utilization are
the possible reasons. El-Reshaid et al. found that the
rHuEpo dose required to maintain the target haemoglobin level is nearly twice as large in patients with
severe iron overload (ferritin )1100 mgul) as in controls
(ferritin 100±600 mgul) w19x. They further observed
that TS decreased to a level of -30% in six (55%) of
11 iron-overloaded patients. Ali et al. also showed that
marrow iron stores were depleted in 10 HD patients
with a mean ferritin of 1336 mgul w20x.
Investigators reported that iron-de®cient erythropoiesis can be overcome by iv iron therapy in patients
with ferritin )500 mgul w15,21x. However, this bene®cial effect is not observed by Tarng et al. w22x,
RosenloÈf et al. w23x or Goch et al. w24x. Similar results
were also found in scorbutic animals w25x. Supplementation of iron did not improve the anaemia, but
increased deposition of haemosiderin in the spleen and
small intestine. After long-term iv administration of
iron dextran in 36 HD patients, Ali et al. found a
paradox of hepatosplenic iron overload and marrow
iron depletion w20x. They proposed that the bulk of iv
iron dextran is taken up by the liver and spleen, and
that the hepatosplenic iron stores fail to be mobilized
to the bone marrow. Iron supplementation is not
warranted in patients with iron overload since tissue
deposition of iron is suf®cient enough to cause organ
damage. In view of the risks related to iron overload
exaggerated by iv iron supplementation, an adjuvant
therapy is needed to improve the rHuEpo response
without hazardous side effects.
Intravenous ascorbic acid improves rHuEpo
responsiveness
Bothwell et al. reported that Bantu subjects with
scurvy had excessive iron deposits in tissues, which
modi®ed the metabolism of ascorbic acid w26x. Tissue
concentration of vitamin C is often decreased in
patients with iron overload, perhaps due to increased
vitamin oxidation catalysed by iron. Moreover, subclinical vitamin C de®ciency is frequently encountered
in HD patients owing to insuf®cient dietary intake,
loss through the dialyser and uraemia-associated
metabolic derangement w27x. Thus, systemic supplementation of ascorbic acid is essential. It has recently
been reported that resistant anaemia was corrected
by iv administration of ascorbic acid in HD patients
who had functional iron de®ciency, even when they
were overloaded with iron w2,3x. We treated 12 rHEporesistant, iron-overloaded patients with ascorbic acid
(300 mg iv post-dialysis, three times a week) w13x. After
8 weeks of treatment, the haematocrit had increased
signi®cantly, with a concomitant signi®cant rise in TS,
and a decrease in erythrocyte zinc protoporphyrin
(E-ZnPP). We also compared the effects of iv iron
administration with those of iv ascorbic acid in 27
rHEpo-resistant HD patients with serum ferritin
)500 mgul w22x. Patients were allocated to one of two
D-C. Tarng et al.
protocols: iv iron sucrose 100 mg 3 5 doses (ns15), or
iv ascorbic acid 300 mg three times weekly for 8 weeks
(ns12). Short-term iv iron supplementation in these
patients neither improved erythropoiesis nor reduced
rHuEpo dose. In contrast, iv ascorbic acid led to a
signi®cant increase in haematocrit and a reduction in
the requirement for rHuEpo.
Effect of iv ascorbic acid on iron metabolism
Ascorbic acid, a reducing agent, is able to release iron
from ferritin and mobilize iron from the reticuloendothelial system to transferrin w2,3,13x. This leads to
an increased iron availability. Protoporphyrin is an
intermediate in haem biosynthesis and the mechanism
of iron incorporation for haem synthesis is enzymatic.
Goldberg indicated that ascorbic acid plays a role
in maintaining haemoglobin iron in the reduced state
and potentiates the enzymatic iron incorporation into
protoporphyrin w28x. Possible explanations for the
effect of ascorbic acid on the improvement of rHuEpo
response include increased mobilization of iron from
inert tissue stores, and increased iron utilization in the
erythron (Figure 1).
To determine whether there is any index that can
predict which patients will require iv ascorbic acid
therapy, we further studied 65 iron-overloaded
patients w29x. The treatment group (ns46) comprised of patients hyporesponsive to rHuEpo who
received ascorbic acid 300 mg three times weekly for
8 weeks. Controls (ns19) had a haematocrit of )30%
and did not receive the adjuvant therapy. Thirteen
patients (four controls and nine ascorbic acid-treated
patients) had withdrawn by the end of the study.
The results showed that 18 of the remaining 37
ascorbic acid-treated patients had a good response,
with a dramatic rise in haematocrit (26.4"1.8% vs
31.6"2.8%; P-0.001) after 8 weeks and a concomitant 24% reduction in rHuEpo dose. The enhanced
erythropoiesis paralleled a rise in TS, from 27"10
to 48"19%, and a fall in E-ZnPP, from 123"44 to
70"13 mmolumol haem (P-0.05). In poor responders
and in controls, there were no signi®cant changes in
mean haematocrit, TS or E-ZnPP values. The receiver
operating curves used to analyse the 37 patients
(18 responders and 19 non-responders) showed that
two indices predicted the response to iv ascorbic
acid supplementation: E-ZnPP)105 mmolumol haem
and TS-25%. We concluded that when rHuEpo
hyporesponsiveness occurs in HD patients with iron
overload, the above values should be used to guide
treatment with iv ascorbic acid.
Other potential bene®ts of iv ascorbic acid
Nowadays, hyperferritinaemia in dialysis patients is
always attributed to previous multiple transfusions
Ascorbic acid, iron and erythropoietin
37
Fig. 1. Associations between ascorbic acid and iron metabolism: ascorbic acid mobilizes iron from the crystal core of ferritin, promotes iron
release to transferrin, and facilitates the enzymatic iron-incorporation into protoporphyrin. (a) Iron-de®cient erythropoiesis in state of iron
overload. (b) Possible mechanisms of ascorbic acid administration to combat the iron-de®cient erythropoiesis.
and over-treatment with iv iron if a state of in¯ammation, liver disease or malignancy is excluded. Cellular
iron overload plays an important role in a number
of pathological states. It produces a variety of deleterious effects, including haemosiderosis, hepatic and
cardiac dysfunction, and cardiovascular and infectious
complication w17,18,30x. Anaemia improved and serum
ferritin levels declined simultaneously after 8 weeks
of iv ascorbic acid medication w29x. This implies that
iv ascorbic acid may also reduce the magnitude of iron
overload in uraemic patients. Confronted with the
potential hazards of iron overload, iv ascorbic acid,
mimicking an iron-chelating agent, provides an additional bene®t for HD patients. Ascorbic acid combined
with desferrioxamine (DFO) results in very ef®cient
removal of iron in non-uraemic patients with iron
overload w31,32x. Studies indicated that DFO can
enhance burst-forming unit-erythroid proliferation w33x
and improve anaemia in iron-overloaded HD patients
w34x. Therefore, long-term iv ascorbic acid alone or
combined with DFO appears to be promising in the
treatment of HD patients with iron overload.
It has been shown that endothelial vasomotor
function is abnormal in the setting of atherosclerosis.
Increased oxidative stress is implicated as one potential
mechanism for this observation. Vitamin C, the main
water-soluble anti-oxidant in human plasma, effectively scavenges reactive oxygen species and plays a
pivotal role in the regulation of intracellular redox
state. Cumulative data showed that vitamin C can
improve endothelial vasomotor dysfunction in patients
with risk factors for coronary heart disease and in
those with coronary artery disease w35,36x. Accelerated
atherosclerosis is a well-known complication in HD
patients. Also, there is evidence of increased oxidative
stress and enhanced susceptibility of low-density
38
lipoprotein to oxidation in these patients. These factors
have been associated with endothelial dysfunction,
which is regarded as an important early event in atherogenesis w35±37x. Accordingly, improved vascular endothelial function with supplementation of ascorbic acid
could potentially reduce the risk of atherosclerotic
disease and coronary events in ESRD patients.
Potential problems of ascorbic acid treatment
in ESRD patients
Certain laboratory and clinical observations proposed
concern about the safety of using ascorbic acid in
patients with severe iron overload. Cardiac death w38x
and deterioration of cardiac function w39x have been
reported in thalassaemic children receiving DFO and
ascorbic acid. However, similar ®ndings were not
observed in non-thalassaemic adults with transfusional
iron overload during chelation therapy w40x. Despite
the fact that untoward cardiac events associated
with ascorbic acid may not be a potential problem in
uraemic patients, we recommend the evaluation and
follow-up of cardiac function, particularly when ascorbic acid is being administered intravenously on a
weekly basis.
There are concerns that ascorbic acid promotes
electron exchange and it can be shown in vitro to
enhance the iron toxicity to cellular constituents.
However, Proteggente et al. showed no compelling
evidence for a pro-oxidant effect of ascorbic acid
supplementation, in either the presence or absence of
iron, on DNA base damage in healthy subjects w41x.
Berger et al. further indicated that ascorbic acid exhibits antioxidant activity in iron-overloaded human
plasma w42x. In our preliminary study (unpublished
data), we also observed that leukocyte DNA damage
was not exaggerated by iv ascorbic acid medication
in HD patients with serum ferritin of )500 mgul.
Vitamin C overdose may cause secondary oxalosis
w43x, leading to plasma levels of oxalate at which
deposition of calcium oxalate in tissue can occur. The
use of excessive amounts of vitamin C should be
avoided in ESRD patients. The available data suggest
that a daily dose of ascorbic acid not exceeding 150 mg
is considered safe in uraemics w44x. The dose of ascorbic acid in our studies was 300 mg three times weekly,
which is less than the recommended dosage. Furthermore, plasma oxalate concentration did not signi®cantly change, although a trend towards a difference
between baseline and 8 weeks was noted w29x.
Conclusions
Haemodialysis may be associated with subclinical
vitamin C de®ciency, and some investigators recommend routine vitamin C supplementation for HD
patients, whether or not they are receiving rHuEpo.
A daily dose of 100±200 mg is recommended in
D-C. Tarng et al.
some reports w44x, but prescription of ascorbic acid in
different dialysis facilities varies between 55 and
1000 mg daily w45± 47x. Based on the consensus of
the workshop chaired by Professor HoÈrl in the Second
European Epoetin Symposium w1x, the recommendations are summarized as follows:
. When subclinical vitamin C de®ciency is suspected,
ESRD patients not on dialysis should receive oral
ascorbic acid 1±1.5 guweek, and HD patients should
receive the same regimen, or iv ascorbic acid 300 mg
three times weekly at the end of dialysis.
. In iron-overloaded HD patients with rHuEpo
hyporesponsiveness, E-ZnPP of )105 mmolumol
haem and TS of -25% should be used to guide
the iv ascorbic acid treatment.
. In the long term, high doses of ascorbic acid could
lead to oxalate accumulation. However, preliminary
data suggest that this potential hazard can be
prevented by avoiding vitamin B6 de®ciency w48x.
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