NDT Advance Access published online on November 24, 2009
Nephrology Dialysis Transplantation, doi:10.1093/ndt/gfp643
© The Author 2009. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org
Endothelial progenitor cells in chronic kidney disease
Ferdinand H. Bahlmann,
Thimoteus Speer and
Danilo Fliser
Department of Internal Medicine IV, Saarland University Medical Centre, Homburg/Saar, Germany
Correspondence and offprint requests to:
Ferdinand H. Bahlmann; E-mail: ferdinand.bahlmann{at}uniklinikum-saarland.de
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Introduction
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The adequate supply of oxygen and nutrients is essential for
proper tissue function and survival. During evolution, blood
vessels arose to fulfil these fundamental tasks of life. Not
surprisingly, the integrity of the vascular system is crucial
also for repair of injured tissue, and the imbalance in growth
and/or preservation of blood vessels contribute to the pathogenesis
of numerous diseases.
In 1997, Asahara et al. described human peripheral blood cells that differentiate into mature endothelial cells and form new blood vessels in vivo [2]. Until then, it was thought that postnatal formation of blood vessels results exclusively from proliferation and remodelling of the pre-existing vascular network, a process referred to as angiogenesis. After this first report, many laboratories published their findings on identification and isolation of circulating primitive endothelial precursors or endothelial progenitor cells (EPCs) that participate in vasculogenesis, i.e. the de novo formation of blood vessels. Meanwhile, the role of EPCs in vascular and tissue regeneration and their therapeutic prospective in regenerative medicine has been intensively studied.
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Characterization and isolation of EPCs
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In their landmark paper, Asahara
et al. found that peripheral
blood mononuclear cells enriched for CD34+-expressing cells
could differentiate into endothelial-like cells [2]. After 7
days in culture on fibronectin, the majority of these cells
expressed the endothelial cell surface markers CD31 and/or Tie-2,
i.e. vascular endothelial growth factor (VEGF) receptor [2].
They also expressed endothelial nitric oxide synthase (eNOS)
mRNA and produced NO after stimulation with VEGF or acetylcholine
in a dose-dependent manner. In addition, they took up acetylated
low-density lipoprotein (acLDL), bound Ulex lectin and formed
tube-like structures
in vitro. Thus, these cultured CD34+-enriched
cells exhibited multiple antigenic and functional properties
that are typical for endothelial cells [2]. However, in many
subsequent studies, conflicting results on identification and
characterization of EPCs were reported due to the lack of a
unique marker to identify EPCs, their paucity in the circulation
and, most importantly, due to their phenotypic and functional
overlap with haematopoietic and mature endothelial cells. Several
studies revealed that many blood cells express the putative
endothelial cell antigens CD31 and Tie-2, and that there is
a considerable overlap in the endothelial cell/monocyte phenotype.
Moreover, monocytes also bind Ulex lectin and take up acLDL,
and inherently express CD31, CD105 or CD144, making them almost
phenotypically indistinguishable from endothelial cells [22].
So far, the only property identified to be specific for endothelial
cells is the expression of eNOS and release of NO in response
to physiologic stimuli [1,2,
19].
Several assays were developed to isolate EPCs from peripheral blood mononuclear cells. Two major cell types were harvested from these assays, the so-called early-outgrowth EPCs and the late-outgrowth EPCs, according to their time-dependent appearance in culture, and to other features such as cell surface markers, proliferation rate and function [25]. However, virtually all available experimental data is limited to early outgrowth EPCs, i.e. cultured spindle-shaped cells obtained from peripheral blood mononuclear cells as originally described by Asahara et al. [2], because the vast majority of laboratories used them in their experiments and because of absence of in vivo studies showing significant vasculogenesis and/or tissue regeneration by late-outgrowth EPCs. All later mentioned studies refer therefore on early-outgrowth EPCs.
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Mechanisms of neovascularization by EPCs
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The mechanism by which EPCs mediate neovascularization was initially
thought to be differentiation into endothelial cells and direct
incorporation into newly formed vessels. However, in studies
using different animal tissue injury models, only marginal EPC
incorporation into the vasculature was found in neo-vascularized
tissue that was analysed weeks (!) after the initial injury.
Based on these results, the idea that EPCs stimulate neovascularization
via paracrine effects gained increasing attention [27]. Although
the question how EPCs actually enhance neovascularization is
still not resolved, results from experimental studies indicate
that EPCs may be used as disease or prognostic marker or for
therapeutic implications, e.g. for blood vessel and renal tissue
regeneration.
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EPCs in chronic kidney disease
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Recent experimental and clinical data indicate that EPCs may
become incompetent in their ability to regenerate ischaemic
tissue thereby contributing to increased morbidity of patients
with cardiovascular risk factors such as hypertension or diabetes.
Indeed, these patients exhibit significantly lower numbers of
circulating EPCs than healthy controls [3,13,18]. Moreover,
impaired renal function and uraemia result in +AH4-30% decrease
of circulating EPCs (Table
1). In addition, these EPCs are also
malfunctioning [8,10]. In patients with chronic kidney disease
(CKD) stage V receiving haemodialysis therapy, a low level of
circulating CD34 cells, i.e. a population including also EPCs,
was shown to be an independent predictor for both prevalent
cardiovascular events and all-cause mortality [
20].
Several factors present in patients with CKD may have a negative
impact on EPC number and function: hypertension, dyslipidaemia,
glucose intolerance (even in the absence of frank diabetes),
micro-inflammation and increased C-reactive protein (which has
a direct effect on EPC number and activity), oxidative stress,
low levels of erythropoietin (EPO), known and yet unidentified
uraemic toxins etc. The latter is particularly intriguing, since
we and others could show that initiation of haemodialysis therapy
or successful kidney transplantation improves EPC number and
function. Interestingly, kidney graft function directly determines
EPC number [12,24]. In line with the hypothesis that attenuation
of the uraemic state restores EPC number and function, at least
in part, are results from a recent study by Chan
et al. [7]
showing that patients on prolonged and more frequent nocturnal
haemodialysis (i.e. 6–8 h a night, five to six nights
per week) circulating EPCs as significantly higher compared
to patients on conventional haemodialysis. Moreover, Choi
et al. [8] reported a positive association of dialysis dose (Kt/V)
and EPC function, suggesting that more intensive dialysis therapy
may improve EPC number and function in CKD patients on renal
replacement therapy. In this respect, findings from a recently
published study in 67 peritoneal dialysis and 142 haemodialysis
patients are of considerable interest [26]. Even after adjusting
for several potential confounders such as age, blood pressure,
history of cardiovascular disease, presence of diabetes, treatment
with drugs that stimulate EPCs etc., their number was significantly
higher in patients on peritoneal dialysis as compared to patients
on haemodialysis.
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Pharmacological modulation of EPCs
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Instead of cell therapy with EPCs (i.e. progenitor cell transplantation)
that is still hampered by many technical and regulatory obstacles,
pharmacological stimulation and/or modulation of EPCs might
be more promising in terms of clinical practicability. It is
known that physiological mobilization of EPCs from their niches
such as the bone marrow can be triggered by mechanical injury
and ischaemic stress via generation of hypoxia-inducible factor-1
regulated release of VEGF, EPO, SDF-1 and GM-CSF [4–6,11].
With respect to the possibility of their pharmacological mobilization,
various drugs like statins [17,28], glitazones [23] or insulin
[15] have been shown to stimulate EPCs. Studies on drugs that
are of special interest for nephrologists such as antihypertensive
agents or recombinant human EPO (rHuEPO) are summarized in Table
2.
View this table:
[in this window]
[in a new window]
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Table 2 Pharmacologic agents that are of particular interest for nephrologists and which stimulate/modify endothelial progenitor cell number and/or function
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The angiotensin-converting enzyme inhibitors ramipril [21] and
enalapril [29] were shown to increase EPC levels both in experimental
studies and in patients, probably by interfering with the CD26/dipeptidylpeptidase
IV system. Similar findings were shown for angiotensin receptor
blockers like olmesartan, irbesartan, losartan or telmisartan
[5,14,30]. The EPO receptor, which main function is to stimulate
the proliferation and the differentiation of erythroid precursor
cells, is also present on endothelial cells, suggesting a common
ontogenesis for the haematopoietic and endothelial lineage.
Both rHuEPO as well as its longer lasting derivate darbepoetin
alpha significantly enhance the number and functional properties
of EPC via the activation of the intracellular Akt protein kinase
pathway [4,5]. Interestingly, this effect was already observed
with a dose of 30 IU/kg per week and less, i.e. a dose which
does not induce erythropoiesis. Finally, some drugs such as
immunosuppressive agents may also negatively affect EPCs since
their number was reduced by >40% with addition of prednisolone
or cyclosporine A in concentrations corresponding to the usual
daily therapeutic doses into the cell culture media [9,16].
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Conclusion
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While stem cell therapy for tissue regeneration after kidney
injury is currently tested in numerous pre-clinical (animal)
studies, EPCs have attracted less attention so far. Most data
available support the notion that EPC numbers and function are
reduced in CKD patients, and this altered EPC biology may contribute
to the high cardiovascular burden of CKD patients due to compromised
reparative processes in the vascular system. EPC monitoring
might be useful for the clinician as a prognostic marker. Early
targeted pharmacological therapy in order to improve EPC number
and function in CKD patients might therefore be of interest
in order to improve cardiovascular outcome in this population.
Conflict of interest statement. None declared.
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References
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Received for publication: 3. 9.09
Accepted in revised form: 3.11.09
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Introduction
Characterization and isolation...