New insights into the pathophysiology of diabetic nephropathy- from haemodynamics to molecular pathology

European Journal of Clinical Investigation (2004) 34, 785–796
Review
Blackwell Publishing, Ltd.
New insights into the pathophysiology of diabetic
nephropathy: from haemodynamics to molecular pathology
G. Wolf
University of Hamburg, Hamburg, Germany
Abstract
Although debated for many years whether haemodynamic or structural changes are more
important in the development of diabetic nephropathy, it is now clear that these processes
are interwoven and present two sides of one coin. On a molecular level, hyperglycaemia and
proteins altered by high blood glucose such as Amadori products and advanced glycation
end-products (AGEs) are key players in the development of diabetic nephropathy. Recent
evidence suggests that an increase in reactive oxygen species (ROS) formation induced by
high glucose-mediated activation of the mitochondrial electron-transport chain is an early
event in the development of diabetic complications. A variety of growth factors and cytokines
are then induced through complex signal transduction pathways involving protein kinase C,
mitogen-activated protein kinases, and the transcription factor NF-κB. High glucose, AGEs,
and ROS act in concert to induce growth factors and cytokines. Particularly, TGF-β is
important in the development of renal hypertrophy and accumulation of extracellular matrix
components. Activation of the renin-angiotensin system by high glucose, mechanical stress,
and proteinuria with an increase in local formation of angiotensin II (ANG II) causes many
of the pathophysiological changes associated with diabetic nephropathy. In fact, it has been
shown that angiotensin II is involved in almost every pathophysiological process implicated
in the development of diabetic nephropathy (haemodynamic changes, hypertrophy, extracellular matrix accumulation, growth factor/cytokine induction, ROS formation, podocyte
damage, proteinuria, interstitial inflammation). Consequently, blocking these deleterious
effects of ANG II is an essential part of every therapeutic regiment to prevent and treat
diabetic nephropathy. Recent evidence suggests that regression of diabetic nephropathy could
be achieved under certain circumstances.
Keywords Angiotensin II, diabetic nephropathy, growth factors, progression, reactive
oxygen species.
Eur J Clin Invest 2004; 34 (12): 785 –796
Introduction
Diabetic nephropathy is one of the major causes of endstage renal disease in the Western world. Although the incidence of nephropathy owing to type 1 diabetes is declining,
Department of Medicine, Division of Nephrology, Osteology,
and Rheumatology, University of Hamburg, Hamburg, Germany
(G. Wolf ).
Correspondence to: Gunter Wolf, MD, University of Hamburg,
University Hospital Eppendorf, Department of Medicine,
Division of Nephrology, Osteology, and Rheumatology,
Pavilion N26, Martinistraße 52, D-20246 Hamburg, Germany.
Tel.: +49 40/42803–5011; fax: +49 40/42803–5186;
e-mail: Wolf@uke.uni-hamburg.de
Received 11 October 2004; accepted 21 October 2004
© 2004 Blackwell Publishing Ltd
diabetes mellitus type 2, considered 20 years ago as a somewhat benign condition invariably associated with the ‘normal’
ageing process, is now the most common single cause of
renal insufficiency in the USA, Japan and Europe [1–3].
There are more than 10 million people with diabetes alone
in the USA and it has been estimated that this number will
double by 2030 [4]. Patients with type 2 diabetes undergoing
maintenance dialysis require significantly higher financial
resources than those suffering from nondiabetic end-stage
renal diseases. Furthermore, this group of patients has a very
poor prognosis on maintenance dialysis owing to extremely
high mortality caused by cardiovascular events [2,3]. To
develop innovative therapeutic concepts to prevent the
development and progression of diabetic nephropathy, a
comprehensive understanding of the pathophysiology of this
disease is mandatory [5]. Although initially thought that renal
786
G. Wolf
injury in diabetic nephropathy is mainly caused by haemodynamic alliterations such as hyperfiltration and hyperperfusion, there is now clear evidence that these changes are
only one aspect of a complex series of pathophysiological
alterations caused by disturbed glucose homeostasis. The
last few years have provided a better insight into the complex
pathophysiology of diabetic nephropathy on a molecular level.
Are there different mechanisms operative in
nephropathy caused by type 1 or 2 diabetes?
Although debated for some time that diabetic nephropathies
owing to either type 1 or 2 diabetes are specific entities,
there is now convincing evidence that basic pathophysiological mechanisms eventually leading to nephropathy are
similar in type 1 and 2 diabetes [2]. However, in type 2
diabetes other noxious factors, being related or not related
to diabetes, such as hypertension, obesity, dyslipidaemia
and ischaemic renal disease caused by arteriosclerosis, could
additionally injure the kidney resulting in complex patterns
of nephropathy. The pathophysiological changes before the
development of type 2 diabetes have been classified as the
metabolic syndrome [3]. This metabolic syndrome may
additionally harm the kidneys through hyperuricaemia and
obesity itself independently of hyperglycaemia [6,7].
The risk of nephropathy is strongly determined by genetic
factors and only approximately 40 –50% of patients with
either type 1 or type 2 diabetes will ultimately develop nephropathy [8]. Experimental data support this observation.
A recent study demonstrated that phenotypic changes in
bone marrow-derived mesangial cell progenitors transmit
diabetic nephropathy from donors with type 2 diabetes (db/
db mice) to naive, normoglycaemic recipients [9]. Genetic
factors may directly influence the development of diabetic
nephropathy and /or may be clustered with genes influencing
other cardiovascular diseases [8]. There is ongoing research
in identifying genetic loci for diabetic nephropathy susceptibility through genomic screening and candidate gene
approaches [10–12]. Although some potential genes have
been identified, linkage was only present in defined ethnic
subpopulations and not in the majority of patients. An
incomplete list of previously implicated genes is shown in
Table 1. The major problem with such studies is a nonsimplistic mendelian inheritance mode with several genes
likely involved [12]. A potential association between polymorphisms in candidate genes and the development and
progression of nephropathy has been widely studied. These
case–control retrospective studies are often problematic and
clear guidelines for such polymorphism studies have been
provided [13]. The complexity of this genetic linkage analysis
is exemplified by various studies with controversial results
investigating the insertion or deletion polymorphism of the
angiotensin-converting enzyme (ACE). The current opinion
is that the ACE polymorphism is, at best, in certain ethnic
population associated with progression of disease but not
as a predictor of the development of diabetic nephropathy
[8]. These discrepancies could be easily explained by genetic
Table 1 Some of the genes implicated in the susceptibility and /or
progression of diabetic nephropathy (modified after 12)
Gene
Gene variant
Promoter of RAGE
Histocompatibility antigen
Angiotensin-converting enzyme
Angiotensinogen
Aldose reductase
Transforming growth factor β1
Apolipoprotein E
Paraoxonase 1
Interleukin 1β
Atrial natriuretic peptide
Glucose transporter 1
Mannose-binding lectin
63-bp deletion (decreased risk)
DR3/ 4
D/ I
M235T
Z + 2 alleles
Leu10Pro, Arg25Pro
e2 allele
T107C, Leu54Met
T105C
C708T
Xba1/ HacIII
YA/ YA, XA/ YA
heterogeneity and by only small effects in limited cases in
most studies (often < 200 cases).
More recently, genome scan data become available for
diabetic nephropathy. This method allows for a comprehensive genetic survey of the entire genome for chromosomal
regions that are linked with a specific trait, in this case
diabetic nephropathy. A recent genome scan for diabetic
nephropathy in African Americans identified susceptibility
loci on chromosomes 3q, 7p and 18q [14]. Another scan in
Pima Indians also identified linkage to diabetic nephropathy
on chromosome 7 [15]. This powerful method may in the
future more clearly identify the genetic risk to develop diabetic
nephropathy.
Haemodynamic changes
Glomerular haemodynamic changes including hyperfiltration and hyperperfusion have been viewed as pivotal in the
development of diabetic nephropathy and are found very
early in the disease process [16]. Principally, an elevation
in glomerular capillary pressure causing an enhanced transcapillary hydraulic pressure gradient as well as an increase
in glomerular plasma flow have been observed. These
changes result from a decrease in both afferent and efferent
arteriolar resistance, whereas the former is more dilated
than the latter leading to the increased glomerular capillary
pressure [16]. Formally, these changes could be described
as a defect in autoregulation because in the normal situation
an increase in perfusion pressure would result in preglomerular vasoconstriction in order to maintain glomerular
filtration rate at a constant rate [17]. Many diverse factors
including prostanoids, nitrogen oxide (NO), atrial natriuretic factor, growth hormone, glucagon, insulin, angiotensin
II (ANG II), and others have been implicated as agents
causing hyperperfusion and hyperfiltration [16]. Elevated
intraglomerular pressure has been linked to an increase
in mesangial cell matrix production and thickening of the
glomerular basement membrane, eventually leading to
glomerulosclerosis [18,19]. There has been a discussion
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
Diabetic nephropathy
going on for years about whether haemodynamic or structural changes are more important for the manifestation of
diabetic nephropathy [16]. However, it is now clear that
these processes are intimately interwoven. For example,
high glucose stimulates the synthesis of ANG II, which itself
exerts haemodynamic as well as trophic, inflammatory and
profibrogenic effects on renal cells [18,20]. Other factors
induced by the diabetic environment that could influence
glomerular haemodynamics are vascular endothelial growth
factors (VEGFs), and recent evidence indicates that even
cytokines, such as transforming growth factor beta (TGFβ), may mediate hyperfiltration by dilatation of the vas afferens by inhibiting calcium transients [20]. Furthermore,
TGF-β increases NO production in early diabetes, probably
by up-regulation of endothelial NO synthase (eNOS) mRNA
expression and by enhancing arginine resynthesis [21].Thus,
TGF-β could clearly play a role in diabetic vascular dysfunction and may mediate some of the haemodynamic
changes associated with early diabetic nephropathy [22].
On the other hand, shear stress and mechanical strain,
resulting from altered glomerular haemodynamics, induce
the autocrine and/or paracrine release of cytokines and
growth factors [18]. For example, exposing mesangial,
glomerular endothelial or podocytes to shear stress induces
specific cellular responses including activation of certain
signal transduction systems, growth responses, enhanced
synthesis of hormones and cytokines (e.g. ANG II, TGF-β),
and increased production of extracellular matrix proteins
[16]. These findings suggest that local haemodynamic stress
contributes to the structural changes of diabetic nephropathy by the local activation of cytokines and growth factors.
As an alternative explanation, a primary abnormality in
sodium reabsorption has been linked to glomerular hyperfiltration in diabetic nephropathy [23]. This explanation
suggests that an increase in reabsorption of sodium chloride
in proximal tubules or loops of Henle leads to an increase
in the glomerular filtration rate by an intact macula-densa
mechanism [23]. Diabetes-induced hypertrophy of tubules
that mediate stimulated sodium chloride reabsorption could
be pivotal in this process, linking again structural changes
with haemodynamic adaptation in diabetic nephropathy
[20].
Structural abnormalities
It is quite obvious that a systemic disease like diabetes
mellitus could result in injury of all renal compartments
[Table 2]. Consequently, glomerulosclerosis, vascular
diseases and changes of the tubulointerstitial architecture
with tubular atrophy and interstitial fibrosis have all been
described in diabetic nephropathy [24 –28]. Since the original
description by Kimmelstiel and Wilson, nodular glomerulosclerosis has been considered as a hallmark of diabetic
nephropathy [24]. However, neither is this lesion specific
for diabetic nephropathy because similar changes could be
also detected in light-chain nephropathy nor is it the most
frequent glomerular histological abnormality which is
787
Table 2 Renal structural abnormalities found in diabetic
nephropathy
Mesangial expansion
Glomerulosclerosis (diffuse, nodular)
Fibrin cap lesion
Capsular drop lesion
Basement membrane thickening (glomerular and tubular)
Endothelial foam cells
Podocyte abnormalities
Armanni-Ebstein cells (proximal tubules stuffed with glycogen)
Tubular atrophy
Interstitial inflammation
Interstitial fibrosis
Arteriosclerosis
diffuse intercapillary glomerulosclerosis. The earliest morphological change of diabetic nephropathy is expansion of
the mesangial area [26]. This is caused by an increase in
extracellular matrix deposition and mesangial cell hypertrophy. Structural–functional relationship investigations
indicate a highly significant inverse correlation between
glomerular filtration rate and mesangial expansion [26].
We have intensively studied the molecular mechanisms of
how hyperglycaemia induces mesangial hypertrophy [29 –
33]. After a short period of proliferation, mesangial cells
exposed to high glucose become arrested in the G1-phase
of the cell cycle [30]. This G1-phase arrest is mediated by
p27Kip1, an inhibitor of cyclin-dependent kinases [30,31].
High glucose via activation of mitogen-activated protein
kinases (MAPKs) lead to a post-transcriptional increase
in p27Kip1 expression [33]. Deletion of the p27Kip1 gene
attenuates high glucose-induced hypertrophy of mesangial
cells [32]. In addition, ANG II further enhances p27Kip1
induction and blockade of ANG II attenuates high glucosemediated mesangial cell hypertrophy [31].
Thickening of the glomerular basement membrane
(GBM) occurs early and is already found 1 year after onset
of type 1 diabetes [34]. As shown in Fig. 1, thickening of the
GBM is progressive over years; both increased extracellular
matrix synthesis and impaired removal contribute to GBM
thickening. Several biochemical alterations of the GBM
occur in diabetic nephropathy [34]. There is an increase in
collagen type IV deposition, whereas the expression of heparan
sulphate and the extent of sulphation decreases. In contrast
to the mesangial matrix in which the α1 and α2 of type IV
collagen are mainly expressed, the GBM contains α3, α4,
and α5 chains [35,36]. In diabetic nephropathy, there is an
up-regulation of α1(IV) and α2(IV) chains in mesangial
cells, whereas α3(IV) and α4 (IV) expression is increased
in the GBM [36]. Deposition of collagen type I and III in
the mesangial area occurs late in glomerulosclerosis and is
not an early event [35].
Glomerular epithelial cells (podocytes) directly cover the
GBM and there is recent evidence that alterations in structure and function of podocytes occur early in diabetic
nephropathy [37–39]. Podocytes adhere to the GBM by
α3β1 and α2β1 integrins. Hyperglycaemia induces a dysregulation in integrin expression, influencing the interaction
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
788
G. Wolf
Figure 1 Renal biopsies (electron-microscopy) from different patients suffering for various periods (3 month up to 12 years) from type
1 diabetes. There is a progressive increase in the thickness of the glomerular basement membrane from normal (left panel) until narrowing
of capillary loops and massive expansion of the mesangium by extracellular matrix (right panel). The process takes years until the right
stage is reached. Courtesy of Professor Helmchen, Institute of Pathology, University of Hamburg.
between podocytes and the GBM [35]. Longitudinal studies in humans with diabetic nephropathy demonstrated a
reduction in podocyte number during follow up which
closely correlated with proteinuria [37]. For example, Steffes
et al. determined the glomerular cell number with modern
morphometric techniques in patients with type 1 diabetes
and compared those numbers with age-matched normal
individuals [38]. They found a reduction in the podocyte
number in diabetic patients of all ages, with reduced podocytes per glomerulus even in diabetes of short duration [38].
In addition, renal biopsies from Pima Indians with type 2
diabetes showed a broadening in podocyte foot processes
and a concomitant reduction in the number of podocytes
per glomerulus [39]. In an European collective of White
type 2 diabetics, Vestra et al. found a significant reduction
in the numerical density of podocytes per glomerulus in
patients with type 2 diabetes that were normoalbuminuric
[40]. Abnormalities in podocyte structure are also observed
in animal models of diabetic nephropathy. Renal nephropathy of Zucker fa/fa rats, a model of type 2 diabetes with
ultimate development of segmental glomerulosclerosis, starts
with damage to podocytes, including foot process effacement and cytoplasmic accumulation of lipid droplets [41].
Early progressive podocyte damage antedates the development of glomerulosclerosis and tubulointerstitial damage in
this model [42].
Oxidative stress as a common mediator
On a molecular level, at least five major pathways have been
implicated in glucose-mediated vascular and renal damage:
increased polyol pathway flux, increased hexosamine pathway
flux, activation of NF-κB, increased advanced glycation
end-product (AGE) formation, stimulation of ANG II
synthesis and activation of the protein kinase C (PKC) path-
Figure 2 Reactive oxygen species (ROS) as a common mediator
of pathophysiological effects of hyperglycaemia. Increased uptake
of glucose into cells leads to stimulated mitochondrial ROS
formation. This oxidative stress, in turn, activates different
processes involving protein kinase C (PKC), NF-κB, cytokines,
formation of advanced glycation end-products (AGEs), and others.
way [Fig. 2]. All of these pathways could reflect a single
hyperglycaemia-induced process of overproduction of reactive
oxygen species (ROS). Hyperglycaemia leads to an increase
in mitochondrial ROS formation [43]. The first step in this
process is the transport of glucose into the cells through
specific glucose transporters [Fig. 3]. Mesangial cells express
insulin-sensitive glucose transporters (GLUT-4) as well as
a brain type of glucose transporters (GLUT-1) through
which excessive extracellular glucose could easily enter the
cell in an insulin-independent manner [44,45]. The role of
GLUT-1 was clarified by overexpression of this transporter
in mesangial cells in which a stimulated production of
extracellular matrix proteins was then detected even under
normal external glucose concentrations [45]. An increase in
glucose uptake leads to overproduction of electron donors
(NADH and FADH2) from stimulated glycolysis and the
tricarboxylic acid cycle [43]. At the mitochondrial inner
membrane, where the electron-transport chain is localized,
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
Diabetic nephropathy
Figure 3 Overview of processes leading to the generation of
reactive oxygen species (ROS). Glucose is taken up into the cells
through insulin-sensitive glucose transporters (GLUT-4) that are
not regulated by insulin. Therefore, hyperglycaemia leads to an
increase in the intracellular glucose concentration, which is
subsequently metabolized to pyruvate through glycolysis. During
this process NADH is generated, which is transported into
mitochondria. In the tricarboxylic acid cycle (TCA) pyruvate is
further metabolized to CO2 and water with the generation of
additional electron donors (NADH and FADH2). This increase in
electron donors leads to a disturbance of the respiration chain with
the formation of ROS. Details compare with text.
the increase in electron donors (NADH, FADH2) generates
a high membrane potential by pumping protons across the
inner membrane [Fig. 3]. As a consequence, electron
transport is inhibited at complex III increasing the half-life
of free-radical intermediates of coenzyme Q, which finally
−
reduces O2 to superoxide ( O2 ). Experimental studies with
endothelial cells have demonstrated that overexpression of
manganese superoxide dismutase (MnSOD) abolished
the hyperglycaemia-induced generation of ROS, and overexpression of uncoupling protein-1 (UCP-1) collapsed the
proton electrochemical gradient and prevented generation
of ROS [43]. Furthermore, UCP-1 and MnSOD prevented, in this system, high glucose-activation of the polyol
pathway, stimulated AGE formation and increased PKC
activation, suggesting a pivotal role of mitochondrialgenerated ROS in these processes [43].
The formation of ROS does not only occur in mitochondria. Recent evidence suggests that the 12-lipoxygenase
pathway of arachidonic acid metabolism is activated by high
glucose in podocytes [46]. Stimulation of 12-lipoxygenase
−
generates O2 , making this an important additional pathway of ROS formation independent of mitochondria [47].
Finally, a reduction of antioxidants such as glutathione
contributes to oxidative stress in the diabetic state [48].
Glucose is converted to sorbitol by aldose reductase and
subsequently to fructose by sorbitol dehydrogenase in the
polyol pathway. An increase in cellular glucose uptake will
shift some of the glucose into this pathway. The rate of sorbitol production depends primarily on the intracellular
availability of glucose. Activation of the polyol pathway
could have several deleterious effects [44,48]. First, reduction
789
of glucose to sorbitol requires NADPH-depleting cells of an
important substrate for the regeneration of glutathione. As
a consequence, this exacerbates intracellular oxidative stress.
Second, the intermediate 3-deoxyglucone is a precursor of
AGEs. Thus, activation of the polyol pathway increases oxidative stress and also enhances AGE formation. Interestingly,
recent data suggest that the calcium antagonist amlodipine
inhibits sorbitol formation, indicating blockade of the polyol
pathway [49].
An increased flux of glucose through the hexosamine
pathway has also been linked to mechanisms of diabetic
nephropathy, particularly an increase in TGF-β [50].
Fructose-6-phosphate from glycolysis is converted to
glucosamine-6-phosphate in this pathway. Glycosylation of a
transcription factor such as Sp1 by N-acetylglucosamine
stimulates TGF-β transcription. In addition, an increase in flux
through the hexosamine pathway up-regulates the expression
of up-stream stimulatory factors (USFs) which transactivate
the TGF-β1 promoter [51].
Intracellular accumulation of glucose also increases de
novo formation of diacylglycerol (DAG) from glycolytic
intermediates such as dihydroxyacetone phosphate [44]. An
increase in DAG activates several isoforms of PKC. Inhibition of PKC-β, the major isoform induced in the kidney by
hyperglycaemia, ameliorates diabetic nephropathy. Moreover, activation of PKC could, in turn, further stimulate
MAPKs. Erk 1,2 as well as p38 MAPK have been implicated as signalling intermediates in diabetic nephropathy
[44]. MAPKs are additionally activated by ROS and there
is likely cross-talk between the various pathways [44]. The
importance of PKC in the development of some changes
of diabetic nephropathy is underscored by recent studies
demonstrating that albuminuria was absent in diabetic
PKC-α knockout mice [52]. However, glomerular hypertrophy or the up-regulation of TGF-β was not influenced
by the lack of PKC-α [52].
Advanced glycation end-products
Advanced glycation end-products (AGEs) are a complex
and heterogeneous group of compounds implicated in
diabetes-related complications including nephropathy [53].
Glucose reacts nonenzymatically with amino groups in proteins, nuclei acids and lipids through a series of steps to form
Schiff bases and Amadori products to finally produce AGEs
[54]. These processes occur over a period of weeks, thereby
affecting long-lived proteins.Therefore, structural components
of tissue matrix such basement membranes are important
targets [55]. For example, glycation inhibits the interactions
required for self-assembly of type IV collagen and laminin
[56]. Glycation of proteins can be accompanied by oxidation, a process called glycoxidation, which is obviously
enhanced by oxidative stress [57,58]. Some of the better
characterized AGEs such as pentosidine and N-carboxymethyllysine (CML) are examples of such glyoxidated products.
ANG II may further stimulate AGE formation. Interestingly, ACE inhibitors and AT1-receptor antagonist inhibit
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
790
G. Wolf
in vitro CML and pentosidine formation by interfering
with various oxidative steps, including carbon-centred and
hydroxyl radical [59]. Furthermore, products of the polyol
pathways forming intermediate products such as 3deoxyglucosone could further induce nonoxidative AGEs
(deoxyglucasone-lysine dimer). Although the majority of
AGEs is usually endogenously formed, it is apparent that
exogenous AGEs from food intake may contribute to the
overall burden, particularly in situations with reduced renal
function [55]. In early diabetic nephropathy, CML and
pentosidine accumulate in the expanded mesangial area and
the thickened glomerular capillary wall [57]. The receptor for
AGEs, called RAGE, is a multiligand member of the
immunoglobulin family of cell-surface receptors [60]. In
addition, other moieties such as galectin-3, CD36, macrophage scavenger receptor and others bind AGEs [58].
RAGE is the signal transduction receptor for AGEs and is
widely expressed on various renal cell types. Stimulation of
this receptor causes expression of proinflammatory cytokines
through NF-κB activation [58]. Besides induction of
cytokines, RAGE directly contributes to inflammation by
serving as an endothelial adhesion receptor promoting
leucocyte recruitment [61]. AGEs also induce TGF-β as a
profibrogenic cytokine [58]. AGEs are filtered by glomeruli
and are reabsorbed by proximal tubules [62]. Although not
directly binding to AGEs, megalin is involved in the tubular
uptake of filtered AGEs [62]. In tubular cells, RAGE activation leads to transition of these cells into myofibroblasts;
a response involved in the development of tubular atrophy
and interstitial fibrosis of ongoing diabetic nephropathy
[63]. RAGE expression itself is up-regulated in the diabetic
state, for example on podocytes. A novel therapeutic concept is the application of soluble RAGE (sRAGE), which
scavenges and neutralizes AGE [58]. In addition to being
a key player in the development of diabetic nephropathy,
AGEs also modify essential functions of leucocytes and contribute to the reduced immune function observed in diabetes
mellitus [64].
Cytokines and growth factors
Various growth factors, cytokines, chemokines and vasoactive agents have been implicated in structural changes of
diabetic nephropathy [65 – 67]. A comprehensive review of
all growth factors potentially involved in diabetic nephropathy is beyond the scope of this article and excellent reviews
exist [65–69]. Insulin-like growth factors (IGFs) are among
the most widely and earliest studied growth factors in diabetic nephropathy but the exact role remains elusive [70].
An early and temporary increase in renal IGF-I protein after
the onset of diabetes is found in various animal models and
this increase is caused by hyperglycaemia [70]. Interference
with the IGF-I axis partly attenuates diabetic nephropathy
in some models [65].
A very important profibrogenic factor for the development of diabetic nephropathy is TGF-β [68,71]. High
glucose, Amadori products as well as AGEs increase TGF-β
mRNA and protein in renal tubular and mesangial cells in
vitro and in vivo [71–74]. Glomerular hypertension is an
additional potent stimulus for renal TGF-β expression linking metabolic and haemodynamic effects [68]. TGF-β and
its type II receptor expression are increased in models of
type 1 and 2 diabetes [75]. Finally, up-regulation of the
TGF-β system and production of TGF-β with a release in
the systemic circulation has been found in humans with
diabetes (for a review see [68]). A consensus nucleotide
sequence termed the ‘glucose response element’ has been
characterized in the TGF-β promoter [73]. Activation of
PKC and p38 MAP kinase plays an additional role through
AP-1 binding sites in transcriptional stimulation of the
TGF-β gene [73]. Additionally, a post-transcriptional activation of latent to active TGF-β is induced in mesangial cells
through thrombospondin-1 [74]. TGF-β stimulates the production of several extracellular matrix proteins including
fibronectin, and types I, III, and IV collagens [72]. TGF-β
also inhibits the degradation of extracellular matrix components through inhibition of matrix metalloproteinases.
Long-term treatment of mice with either type 1 or 2 diabetes
and nephropathy with neutralizing anti-TGF-β antibodies
prevented mesangial matrix expansion and attenuated the
decrease in renal function [75]. Bone morphogenic protein
(BMP)-7, a member of the TGF-β superfamily playing a
major role during embryonic development, can antagonize
TGF-β-dependent fibrogenesis in renal cells [76].
Connective tissue growth factor (CTGF) is one of the
more recently characterized growth factors and is a 36–38kDa cystein-rich peptide [77]. It may in fact be the direct
down-stream mediator of some effects hitherto thought to
be caused by TGF-β [66,77]. Further studies such as interference with CTGF expression and /or receptor blockade are
ultimately necessary to test its role in diabetic nephropathy.
Vascular endothelial growth factor is an approximately
34–46-kDa homodimeric glycoprotein which exists in at
least five different isoforms produced by differential exon
splicing [69]. Vascular endothelial growth factor itself is
expressed in podocytes, distal tubules and the collecting duct
[69,78]. It has been speculated that heavier VEGF isoforms
may be carried across the glomerular basement membrane
exerting than their effects on endothelial cells. Extracellular
matrix proteins regulate VEGF transcription in podocytes [79].
Circulating VEGF concentrations are significantly increased
in type 1 and 2 diabetes. Several studies indicate that VEGF
mRNA and protein expression are enhanced in kidneys with
early diabetic nephropathy (for a review see [69]). Furthermore,
AGEs contribute to expression of VEGF. A functional role
of VEGF in diabetic nephropathy was demonstrated by the
observation that monoclonal anti-VEGF antibodies administered to diabetic rats decreased hyperfiltration, albuminuria
and glomerular hypertrophy in these animals [80].
Hypoxia and diabetic nephropathy
Recent clinical studies have provided evidence that even
mild anaemia (Hb < 13·8 g dL−1) increases the risk for
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
Diabetic nephropathy
progression in patients with type 2 diabetes and nephropathy
[81]. Moreover, treating anaemia early in renal failure with
erythropoietin slows the decline of renal function [82,83].
The exact mechanisms by which anaemia increases the risk
for progression of diabetic nephropathy are incompletely
understood but a few suggestions could be made. Anaemia
likely causes renal hypoxia [84]. It has been described in
experimental models of chronic renal injury that hypoxia is
an important factor aggravating interstitial fibrosis, partly
by the induction of factors such as TGF-β and VEGF [84].
This induction of growth factors and cytokines is mediated
by hypoxia-inducing factor-1 (HIF-1), and ANG II can
further increase this important transcription factor [85]. On
the other hand, an erythropoietin application may have
additional effects besides correcting anaemia, and the mobilization of potential progenitor cells by this treatment has
become the focus of active research [86]. Certainly, more
experiments are necessary to decipher how anaemia may
exactly contribute to the development and progression of
diabetic nephropathy but this topic will be of great interest.
Inflammation and diabetic nephropathy
Biopsy studies from patients with diabetic nephropathy and
investigations in various animals models revealed the
presence of inflammatory cells in glomerular and tubulointerstitial compartments [87]. Mononuclear cell infiltrates
are often present [88]. Monocyte chemoattractant protein
1-(MCP-1) is an important chemokine for macrophages/
monocytes. High glucose stimulates MCP-1 expression in
mesangial cells. Increases in tubular expression of MCP-1
and RANTES, another chemokine, have been found in
renal biopsies from patients with diabetic nephropathy and
were adjacent to infiltrating immune cells [88 – 92]. The
proinflammatory transcription factor NF-κB was detected
mainly in tubular cells in biopsy specimens from 11 patients
with type 2 diabetes and overt nephropathy, indicating that
proteinuria may have contributed to this activation [90].
Fractalkine expression is induced in renal endothelial cells
from diabetic rats [91]. This induction promotes strong
adhesion of T cells and monocytes to the endothelium via
the receptor CX3CR1 [91]. It appears that circulating levels
of chemokines such as MCP-1 are already enhanced in the
metabolic syndrome and the adipocyte could be an important source [93]. The expression of proinflammatory molecules such as chemokines could be further stimulated in
podocytes and tubular cells by proteinuria acting in concert
with hyperglycaemia and AGEs. As infiltrating mononuclear
cells release proteases and profibrogenic cytokines, including TGF-β, such proinflammatory infiltrates contribute
to the destruction of nephrons in diabetic nephropathy.
Consequently, treatment of diabetic animals with antiinflammatory drugs such as mycophenolate mofetil prevents
the development of glomerular injury [92]. Diabetesassociated induction of proinflammatory cytokines is partly
dependent on ANG II [88]. Therefore, inhibition of ANG II
exerts anti-inflammatory effects also.
791
Pivotal role of the renin-angiotensin-aldosterone
system
Drugs interfering with the renin-angiotensin-aldosterone
system (RAAS) are a mainstay of therapy in preventing the
progression of diabetic nephropathy since the seminal
observation by Zatz and coworkers that an ACE-inhibitor
prevented the development of nephropathy in experimentally induced diabetes [94]. Although initially considered to
act solely through normalization of systemic and glomerular
hypertension, it is now clear that inhibition of the RAAS has
many effects, including antifibrotic and anti-inflammatory
mechanisms [95]. In fact, ANG II itself induces in renal
cells many proinflammatory and profibrogenic cytokines,
chemokines and growth factors [Fig. 4]. It has been demonstrated that many, if not all, RAAS components are
present in the kidney, for example in tubular cells [95]. High
glucose stimulates expression of renin and angiotensinogen
in mesangial and tubular cells [96–99]. This stimulation
results in an increase in local ANG II concentrations which
may, in turn, through autocrine and paracrine pathways
induce a whole battery of different cytokines and growth
factors [95]. Experimental studies indicate that a high
glucose-mediated generation of ROS is important in the upregulation of angiotensinogen in proximal tubular cells [98].
Inhibition of the RAAS reduces proteinuria in diabetic
nephropathy. Angiotensin-converting enzyme inhibitor or
AT1-receptor antagonist treatment attenuates podocyte
foot process broadening in rats with streptozotocin-induced
diabetes [100–103]. An AT1-receptor antagonist, but not
Figure 4 Summary of mediators involved in the pathophysiology
of diabetic nephropathy. High glucose leads to the generation of
reactive oxygen species (ROS) and increases advanced glycation
end-product (AGE) synthesis. Reactive oxygen species stimulate
local ANG II generation, which increases proteinuria, but
proteinuria also further enhances tubular angiotensin II (ANG II)
synthesis. Angiotensin II also increases AGE formation and is
pivotal in the induction of various cytokines and growth factors.
Some of these cytokines directly stimulate extracellular matrix
synthesis (e.g. TGF-β, CTGF), whereas other mediate
inflammation (MCP-1). As a consequence of these processes, tissue
destruction and fibrosis will result.
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
792
G. Wolf
the calcium-channel blocker amlodipine, normalized the
reduced nephrin expression in podocytes from spontaneously hypertensive rats with superimposed streptozotocininduced diabetes [103]. Thus, a local increase in ANG II
leads to suppression of nephrin expression in podocytes.
Podocytes express AT1- and probably AT2-receptors after
injury and could respond to a stimulation with ANG II
[104]. Transgenic rats with targeted overexpression of the
AT1-receptor to podocytes showed pseudocysts in podocytes, followed by foot process effacement and local detachments [105]. These changes subsequently progressed to
focal segmental glomerulosclerosis. It has been previously
shown that high ambient glucose concentrations induce
ANG II formation in podocytes through up-regulation of
angiotensinogen expression [106]. Furthermore, proteinuria and a likely transit of proteins through the ultrafiltration
barrier activates ANG II formation in podocytes [106;
Fig. 4]. Finally, mechanical stretch could increase ANG II
generation in podocytes [107]. Interestingly, ANG II
formation as a consequence of mechanical stress appears to
be independent of ACE [107]. In this regard it has been
demonstrated that chymase, an ANG II-forming enzyme
not inhibited by ACE inhibitors, is up-regulated in glomeruli of patients with nephropathy owing to type 2 diabetes
[108]. This observation suggests that local glomerular
formation of ANG II in the diabetic state may be partly
independent of ACE and could not be abolished by ACE
inhibition alone. As a consequence, an increase in local
ANG II concentrations in the podocyte microenvironment
suppresses nephrin expression, and thereby enhances ultrafiltration of proteins which further contributes to podocyte
damage.
Angiotensin II plays an important role in mediating proteinuria by various mechanisms including hyperfiltration,
opening of nonselective pores in the ultrafiltration barrier,
modifying the composition of the GBM and reducing nephrin
expression on podocytes (for a review see [18]). In addition,
it has been found that ANG II increases tubular reabsorption of ultrafiltered proteins, contributing thereby to tubular
inflammation and eventually fibrosis [18]. A recent study
has somewhat questioned this straightforward explanation
because targeted deletion of the AT1A receptor in mice does
not protect from progressive renal injury induced by overload proteinuria [109]. However, as it has been convincingly
demonstrated that the AT2-subtype of the ANG II-receptor
could induce proinflammatory effects through NF-κB activation [110,111], both subtypes of receptors are probably
involved in the deleterious effects of proteinuria on the
tubulointerstitium.
Recent experimental evidence also indicates a role of
aldosterone, working independently from ANG II, in the
development of diabetic nephropathy [99]. The aldosterone
antagonist spironolactone attenuates increased collagen
deposition in rats 3 weeks after streptozotocin administration [99]. Spironolactone also suppresses the enhanced
TGF-β1 expression in this model [99]. These findings have
been extended to patients. In a preliminary study, Schjoedt
and colleagues observed that an increase in the plasma
aldosterone level during long-term treatment with an AT1-
receptor blocker (aldosterone escape phenomenon) is associated with a decline in glomerular filtration rate in patients
with nephropathy owing to type 1 diabetes [113]. These data
demonstrate that aldosterone contributes to the progression
of diabetic nephropathy despite blockade of the AT1-receptor
[113]. On the other hand, spironolactone decreases proteinuria in patients with 2 diabetes and early nephropathy
[114]. As spironolactone is a nonspecific aldosterone receptor antagonist that may also bind to other steroid receptors
explaining adverse sexual effects, novel antagonists (e.g.
eplerenone) have been developed [115]. Preliminary studies
indicate that eplerenone decreases microalbuminuria in
patients with type 2 diabetes independent of its antihypertensive effects [115]. In addition to direct effects on renal
cells, aldosterone also potentates ANG II-mediated signal
processes such as MAPK activation, indicating that ANG
II and aldosterone act in concert [16].
Taking into account the many effects of the RAAS in the
pathophysiology of diabetic nephropathy, early treatment
with drugs interfering with the RAAS are a necessary
prerequisite to prevent development and progression of
nephropathy in diabetes mellitus. This treatment has entered
clinical practice and should be part of the optimal management of every diabetic patient.
Regression of diabetic nephropathy: a dream
comes true?
Common wisdom suggests that the morphological alterations of advanced diabetic nephropathy (e.g. glomerulosclerosis, interstitial fibrosis) are not reversible and represent a
one-way road to end-stage renal disease. Recent clinical
studies show, however, that this pessimistic scenario is not
necessarily true. Several studies indicate that aggressive
blood pressure control with agents that interfere with the
RAAS leads to remission of overt proteinuria (for a review
see [117]). High-dose ACE-inhibitor treatment induces
regression in experimental nondiabetic models of glomerulosclerosis [118]. Fioretto and colleagues performed a serial
renal biopsy in a collective of patients with type 1 diabetes
and nephropathy who received a pancreas transplant [119].
These investigators found that morphological alterations
of diabetic nephropathy including increased thickness of
basement membranes and mesangial expansion decreased
10 years after receiving the pancreas transplant. The data
clearly demonstrate that the control of blood glucose levels
can reverse the development of diabetic nephropathy, but
this process takes time. In addition, Perkins et al. showed
that the onset of microalbuminuria does not imply inexorably progressive nephropathy, at least not in patients with
type 1 diabetes [120]. This study provides indirect evidence
that aggressive management aiming at various risk factors
could lead to a remission of albuminuria. As no renal biopsies were performed in this study, it remains unproven
whether morphological changes of diabetic nephropathy are
also improved [120]. Nervertheless, there is light at the end
of the tunnel and a better understanding of the complex
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
Diabetic nephropathy
pathophysiology of diabetic nephropathy will likely improve
a multifactorial management to induce regression.
References
1 Ritz E, Rychlik I, Locatelli F, Halimi S. End-stage renal failure
in type 2 diabetes: a medical catastrophe of worldwide
dimensions. Am J Kidney Dis 1999;34:795 – 808.
2 Parving HH. Diabetic nephropathy: prevention and treatment.
Kidney Int 2001;60:2041–55.
3 Remuzzi G, Schieppati A, Ruggenenti P. Nephropathy in
patients with type 2 diabetes. N Engl J Med 2002;346:
1145 –51.
4 Harris MJ, Flegal KM, Cowie CC, Eberhardt MS,
Goldstein DE, Little R et al. Prevalence of diabetes, impaired
fasting glucose, and impaired glucose tolerance in U.S. adults.
Diabetes Care 1998;21:518 –24
5 Wolf G, Ritz E. Diabetic nephropathy in type 2 diabetes:
prevention and patient management. J Am Soc Nephrol
2003;14:1396 –405.
6 Wolf G. After all those fat years: renal consequences of obesity.
Nephrol Dial Transplant 2003;18:2471– 4.
7 Kanellis J, Watanabe S, Li JH, Kang DH, Li P, Nakagawa T
et al. Uric acid stimulates monocyte chemoattractant
protein-1 production in vascular smooth muscle cells via
mitogen-activated protein kinase and cyclooxygenase-2.
Hypertension 2003;41:1287– 93.
8 Berger M, Mönks D, Wanner C, Lindner TH. Diabetic
nephropathy: an inherited diseases or just a diabetic
complication? Kidney Blood Press Res 2003;26:143 –54.
9 Zheng F, Cornacchia F, Schulman I, Banerjee A, Cheng QI,
Potier M et al. Development of albuminuria and glomerular
lesions in normoglycemic B6 recipients of db/db mice bone
marrow. Diabetes 2004;53:2420 –7.
10 Rudofsky Jr G, Isermann B, Schilling T, Schiekofer S,
Andrassy M, Schneider JG et al. A 63 bp deletion in the
promoter of RAGE correlates with a decreased risk for
nephropathy in patients with type 2 diabetes. Exp Clin
Endocrinol Diabetes 2004;112:135 – 41.
11 Hansen TK, Tarnow L, Thiel S, Steffensen R, Stehouwer CD,
Schalkwijk CG et al. Association between mannose-binding
lectin and vascular complications in type 1 diabetes. Diabetes
2004;53:1570 – 6.
12 Susztak K, Sharma K, Schiffer M, McCue P, Ciccone E,
Böttinger EP. Genomic strategies for diabetic nephropathy.
J Am Soc Nephrol 2003;14:S271– 8.
13 Couser WG. Revisions to instructions to JASN authors
regarding articles reporting studies using DNA arrays, DNA
polymorphisms, and randomized controlled clinical trials.
J Am Soc Nephrol 2003;14:2686 –7.
14 Bowden DW, Colicigno CJ, Langefeld CD, Sale MM,
Williams A, Anderson PJ et al. genome scan for diabetic
nephropathy in African Americans. Kidney Int
2004;66:1517–26.
15 Imperatore G, Hanson RL, Pettitt DJ. Sib-pair linkage analysis
for susceptibility genes for microvascular complications
among Pima Indians with type 2 diabetes. Diabetes
1998;47:821–30.
16 Hostetter TH. Hyperfiltration and glomerulosclerosis. Semin
Nephrol 2003;23:194 – 9.
17 Ruggenenti P, Schieppati A, Remuzzi G. Progression,
remission, regression of chronic renal diseases. Lancet
2001;357:1601– 8.
793
18 Wolf G, Butzmann U, Wenzel UO. The renin-angiotensin
system and progression of renal disease: from hemodynamics
to cell biology. Nephron Physiol 2003;93:P3–13.
19 Anderson S, Rennke HG, Brenner BW. Therapeutic
advantage of converting enzyme inhibitors in arresting
progressive renal disease associated with systemic
hypertension in the rat. J Clin Invest 1986;77:1993–2000.
20 Chen S, Wolf G, Ziyadeh FN. The renin-angiotensin system
in diabetic nephropathy. Contrib Nephrol 2001;135:212–21.
21 Sharma K, Deelman L, Madesh M, Kurz B, Ciccone E,
Siva S et al. Involvement of transforming growth factor-beta
in regulation of calcium transients in diabetic vascular smooth
muscle cells. Am J Physiol Renal Physiol 2003;285:F1258–70.
22 Tsuchida K, Cronin B, Sharma K. Novel aspects of
transforming growth factor-beta in diabetic kidney disease.
Nephron 2002;92:7–21.
23 Thomson SC, Vallon V, Blantz RC. Kidney function in early
diabetes: the tubular hypothesis of glomerular filtration. Am
J Physiol Renal Physiol 2004;286:F8–15.
24 Kimmelstiel P, Wilson C. Intercapillary lesions in the
glomeruli of the kidney. Am J Pathol 1936;12:82–97.
25 Adler S. Structure-function relationships associated with
extracellular matrix alterations in diabetic glomerulopathy. J
Am Soc Nephrol 1994;5:1165–72.
26 Mauer SM, Steffes MW, Ellis EN, Sutherland DER,
Brown DM, Goetz FC. Structural-functional relationships in
diabetic nephropathy. J Clin Invest 1984;74:1143–55.
27 DeFronzo RA. Diabetic nephropathy: etiologic and
therapeutic considerations. Diab Rev 1995;3:510–64.
28 Dalla Vestra M, Saller A, Bortolosco E, Mauer M, Fioretto
P. Structural involvement in type 1 and type 2 diabetic
nephropathy. Diab Metabol;200 (26):8–14.
29 Wolf G, Ziyadeh FN. Molecular mechanisms of diabetic
hypertrophy. Kidney Int 1999;56:393–405.
30 Wolf G, Schroeder R, Ziyadeh FN, Thaiss F, Zahner G,
Stahl RAK. High glucose stimulates expression of p27Kip1 in
cultured mouse mesangial cells: relationship to hypertrophy.
Am J Physiol 1997;273(Renal Physiol 42):348–56.
31 Wolf G, Wenzel U, Ziyadeh FN, Stahl RAK. Angiotensin
converting-enzyme inhibitor treatment reduces glomerular
p16INK4 and p27Kip1 expression in diabetic BBdp rats.
Diabetologia 1999;42:11425–32.
32 Wolf G, Schroeder R, Zahner G, Stahl RAK, Shankland SJ.
High glucose-induced hypertrophy of mesangial cells requires
p27Kip1, an inhibitor of cyclin-dependent kinases. Am J Pathol
2001;158:1091–100.
33 Wolf G, Reinking R, Zahner G, Stahl RAK, Shankland SJ. Erk
1,2 phosphorylates p27Kip1: functional evidence for a role in
high glucose-induced hypertrophy of mesangial cells.
Diabetologia 2003;46:1090–9.
34 Mason RM, Wahab NA. Extracellular matrix metabolism
in diabetic nephropathy. J Am Soc Nephrol
2003;14:1358–73.
35 Tsilibary EC. Microvascular basement membranes in diabetes
mellitus. J Pathol 2003;200:537–46.
36 Zeisberg M, Ericksen MB, Hamano Y, Neilson EG, Ziyadeh F,
Kalluri R. Differential expression of type IV collagen
isoforms in rat glomerular endothelial and mesangial cells.
Biochem Biophys Res Commun 2002;295:401–7.
37 White KE, Bilous RW, Marshall SM, El Nahas M, Remuzzi G,
Piras G et al. Podocyte number in normotensive type 1
diabetic patients with albuminuria. Diabetes 2002;51:3083– 9.
38 Steffes MW, Schmidt D, McCrery R, Basgen JM. Glomerular
cell number in normal subjects and in type 1 diabetic patients.
Kidney Int 2001;59:2104–13.
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
794
G. Wolf
39 Pagtalunan ME, Miller PL, Jumping-Eagle S, Nelson RG,
Myers BD, Rennke H et al. Podocyte loss and progressive
glomerular injury in type II diabetes. J Clin Invest
1997;99:342 –8.
40 Vestra MD, Masiero A, Roiter AM, Saller A, Crepaldi G,
Fioretto P. Is podocyte injury relevant in diabetic
nephropathy? Studies in patients with type 2 diabetes. Diabetes
2003;52:1031–5.
41 Gassler N, Elger M, Kränzlin B, Kriz W, Gretz N. Podocyte
injury underlies the progression of focal segmental
glomerulosclerosis in the fa/fa Zucker rat. Kidney Int
2001;60:106 –16.
42 Coimbra TM, Janssen U, Grone HJ, Ostendorf T, Kunter U,
Schmidt H et al. Early events leading to renal injury in obese
Zucker (fatty) rats with type II diabetes. Kidney Int
2000;57:167–82.
43 Brownlee M. Biochemistry and molecular cell biology of
diabetic complications. Nature 2001;414:813 –20.
44 Haneda M, Koya D, Isono M, Kikkawa R. Overview of
glucose signaling in mesangial cells in diabetic nephropathy.
J Am Soc Nephrol 2003;14:1374 – 82.
45 Heilig CW, Kreisberg JI, Freytag S, Murakami T, Ebina Y,
Guo L et al. Antisense GLUT-1 protects mesangial cells from
glucose induction of GLUT-1 and fibronectin expression. Am
J Physiol 2001;280:F657– 66.
46 Kang SW, Natarajan R, Shahed A, Nast CC, LaPage J,
Mundel P et al. Role of 12-lipoxygenase in the stimulation of
p38 mitogen-activated protein kinase and collagen α5(IV) in
experimental diabetic nephropathy and in glucose stimulated
podocytes. J Am Soc Nephrol 2003;14:3178 – 87.
47 Roy P, Roy SK, Mitra A, Kulkarni AP. Superoxide generation
by lipoxygenase in the presence of NADH and NADPH.
Biochim Biophys Acta 1994;1214:171– 9.
48 Morrison J, Knoll K, Hessner MJ, Liang M. Effect of high
glucose on gene expression in mesangial cells: upregulation of
the thiol pathway is an adaptational response. Physiol Genomics
2004;17:272 –82.
49 Bernobich E, Consenzi A, Campa C, Zennaro C,
Sasso F, Paoletti S et al. Antihypertensive treatment and
renal damage: amlodipine exerts protective effect through
the polyol pathway. J Cardiovasc Pharmacol
2004;4:401–6.
50 Kolm-Litty V, Sauer U, Nerlich A, Lehmann R,
Schleicher ED. High glucose-induced transforming growth
factor beta1 production is mediated by the hexosamine
pathway in porcine glomerular mesangial cells. J Clin Invest
1998;101:160 – 9.
51 Weigert C, Brodbeck K, Sawadogo M, Haring HU, Schleicher
ED. Upstream stimulatory factor (USF) proteins induce
human TGF-beta 1 gene activation via the glucose-response
element-1013/-1002 in mesangial cells: up-regulation of USF
activity by the hexosamine biosynthetic pathway. J Biol Chem
2004;279:15908 –15.
52 Menne J, Park JK, Boehne M, Elger M, Lindschau C,
Kirsch T et al. Diminished loss of proteoglycans and lack of
albuminuria in protein kinase C-α-deficient diabetic mice.
Diabetes 2004;53:2101– 9.
53 Singh R, Barden A, Mori T, Beilin L. Advanced glycation
end-products: a review. Diabetologia 2001;44:129 – 46.
54 Ziyadeh FN, Mogyorosi A, Kalluri R. Early and advanced
non-enzymatic glycation products in the pathogenesis of
diabetic kidney disease. Exp Nephrol 1997;5:2– 9.
55 Heidland A, Sebekova K, Schinzel R. Advanced glycation end
products and the progressive course of renal disease. Am J
Kidney Dis 2001;38 (Suppl. 1):S100 – 6.
56 Charonis AS, Tsilbary EC. Structural and functional changes
of laminin and type IV collagen after nonenzymatic glycation.
Diabetes 1992;41:49–51.
57 Horie K, Miyata T, Maeda K, Miyata S, Sugiyama S, Sakai H
et al. Immunohistochemical colocalization of glycoxidation
products and lipid peroxidation products in diabetic renal
lesions. J Clin Invest 1997;100:2995–3004.
58 Yang SF, Ramasamy R, Naka Y, Schmidt AM. Glycation,
inflammation, and RAGE. Circ Res 2003;93:1159–69.
59 Miyata T, van Yypersele de Strihou C, Ueda Y, Ichimori K,
Inagi R, Onogi H et al. Angiotensin II receptor antagonists
and angiotensin-converting enzyme inhibitors lower in vitro
the formation of advanced glycation end products:
biochemical mechanisms. J Am Soc Nephrol 2002;13:2478–
87.
60 Yamamoto Y, Kato I, Doi T, Yonekura H, Ohashi S, Takeuchi M
et al. Development and prevention of advanced diabetic
nephropathy in RAGE-overexpressing mice. J Clin Invest
2001;108:261–8.
61 Chavakis T, Bierhaus A, Al-Fakhri N, Schneider D, Witte S,
Linn T et al. The pattern recognition receptor (RAGE) is a
counterreceptor for leukocyte integrins: a novel pathway for
inflammatory cell recruitment. J Exp Med 2003;198:1507–15.
62 Saito A, Nagai R, Tanuma A, Hama H, Cho K, Takeda T
et al. Role of megalin in endocytosis of advanced glycation
end products: implications for a novel protein binding to both
megalin and advanced glycation end products. J Am Soc
Nephrol 2003;14:1123–31.
63 Li JH, Wang W, Huang XR, Oldfield M, Schmidt AM,
Cooper ME et al. Advanced glycation end products induce
tubular epithelial-myofibroblast transition through the RAGEERK1/2 MAP kinase signaling pathway. Am J Pathol
2004;164:1389–97.
64 Cohen G, Rudnicki M, Walter F, Niwa T, Hörl WH. Glucosemodified proteins modulate essential functions and apoptosis
of polymorphonuclear leukocytes. J Am Soc Nephrol
2001;12:1264–71.
65 Flyvbjerg A. Putative pathophysiological role of growth factors
and cytokines in experimental diabetic kidney disease.
Diabetologia 2000;43:1205–23.
66 Wolf G. Growth factors and the development of diabetic
nephropathy. Curr Diab Rep 2003;3:485–90.
67 Chiarelli F, Santilli F, Mohn A. Role of growth factors in the
development of diabetic complications. Horm Res
2000;53:53–67.
68 Ziyadeh FN. Mediators of diabetic renal disease: the case for
TGF-beta as the major mediator. J Am Soc Nephrol 2004;15
(Suppl. 1):S55–7.
69 Schrijvers BF, Flyvbjerg A, de Vriese AS. The role of vascular
endothelial growth factor (VEGF) in renal pathophysiology.
Kidney Int 2004;65:2003–17.
70 Sugimoto H, Shikata K, Makino H, Ota K, Ota Z. Increased
gene expression of insulin-like growth factor-I receptor in
experimental diabetic rat glomeruli. Nephron 1996;72:648–
53.
71 Wolf G, Sharma K, Chen Y, Ericksen M, Ziyadeh FN. High
glucose-induced proliferation in mesangial cells is reversed by
autocrine TGF-beta. Kidney Int 1992;42:647–56.
72 Ziyadeh FN, Sharma K, Ericksen M, Wolf G. Stimulation of
collagen gene expression and protein synthesis in murine
mesangial cells by high glucose is mediated by autocrine
activation of transforming growth factor-beta. J Clin Invest
1994;93:536–42.
73 Hoffmann BB, Sharma K, Zhu Y, Ziyadeh FN.
Transcriptional activation of transforming growth factor-β1 in
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
Diabetic nephropathy
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
mesangial cell culture by high glucose concentration. Kidney
Int 1998;58:1107–16.
Poczatek MH, Hugo C, Darley-Usmar V, Murphy-Ullrich JE.
Glucose stimulation of transforming growth factor-β
bioactivity in mesangial cells is mediated by thrombospondin1. Am J Pathol 2000;157:1353 – 63.
Ziyadeh FN, Hoffman BB, Han DC, Iglesias-de la Cruz M,
Hong SW, Isono M et al. Long-term prevention of renal
insufficiency, excess matrix gene expression, and glomerular
mesangial matrix expansion by treatment with monoclonal
antitransforming growth factor-β antibody in db/db diabetic
mice. Proc Natl Acad Sci USA 2000;97:8015 –20.
Wang S, Hirschberg R. BMP7 antagonizes TGF-β-dependent
fibrogenesis in mesangial cells. Am J Physiol 2003;284:1006–
13.
Weston BS, Wahab NA, Mason RM. CTGF mediates TGFβ-induced fibronectin matrix deposition by upregulating
active α5β1 integrin in human mesangial cells. J Am Soc
Nephrol 2003;14:601–10.
Onozaki A, Midorikawa S, Sanada H, Hayashi Y, Baba T,
Katoh T et al. Rapid change of glucose concentrations
promotes mesangial cell proliferation via VEGF. inhibitory
effects of thiazolidinedione. Biochem Biophys Res Commun
2004;317:23 –9.
Datta K, Li J, Karumanchi A, Wang E, Rondeau E,
Mukhopadhya D. Regulation of vascular permeability factor/
vascular endothelial growth factor (VPF/ VEGF-A) expression
in podocytes. Kidney Int 2004;66:1471– 8.
de Vriese AS, Tilton RG, Elger M, Stephan CC, Kriz W,
Lameire NH. Antibodies against vascular endothelial growth
factor improve early renal dysfunction in experimental
diabetes. J Am Soc Nephrol 2001;12:993 –1000.
Mohanram A, Zhang Z, Shahinfar S, Keane WF, Brenner
BM, Toto RD. Anemia and end-stage renal disease in patients
with type 2 diabetes and nephropathy. Kidney Int
2004;66:1131– 8.
Gouva C, Nikolopoulos P, Ioannidis JPA, Siampoulos KC.
Treating anemia early in renal failure patients slows the decline
of renal function: a randomized controlled trial. Kidney Int
2004;66:753 –60.
Bahlmann FH, Song R, Boehm SM, Mengel M, von
Wasielewski R, Lindschau C et al. Low-dose therapy with the
long-acting erythropoietin analogue darbepoetin alpha
persistently activates endothelial Akt and attenuates
progressive organ failure. Circulation 2004;110:1006–12.
Fine LG, Bandyopadhay D, Norman JT. Is there a common
mechanism for the progression of different types of renal
diseases other than proteinuria? Towards the unifying theme
of chronic hypoxia. Kidney Int 2000;75:S22– 6.
Wolf G, Schroeder R, Stahl RAK. Angiotensin II induces
hypoxia-inducible factor-1 alpha in PC 12 cells through a
posttranscriptional mechanism: role of AT2 receptors. Am J
Nephrol 2004;24:415 –21.
Bahlmann FH, de Groot K, Haller H, Fliser D.
Erythropoietin: is it more than correcting anaemia? Nephrol
Dial Transplant 2004;19:20 –2.
Noronha IL, Fujihara CK, Zatz R. The inflammatory
component in progressive renal disease-are interventions
possible? Nephrol Dial Transplant 2002;17:363 – 8.
Mezzano S, Droguett A, Burgos ME, Ardiles LG, Flores CA,
Aros CA et al. Renin-angiotensin system activation and
interstitial inflammation in human diabetic nephropathy.
Kidney Int 2003;86 (suppl.):S64 –70.
Lee FTH, Cao Z, Long DM, Panagiotopoulos S, Jerums G,
Cooper ME et al. Interactions between angiotensin II and
90
91
92
93
94
95
96
97
98
99
100
101
102
103
104
795
NF-κB-dependent pathways in modulating macrophage
infiltration in experimental diabetic nephropathy. J Am Soc
Nephrol 2004;15:2139–51.
Mezzano S, Aros C, Droguett A, Burgos ME, Ardiles L,
Flores C et al. activation and overexpression of regulated genes
in human diabetic nephropathy. Nephrol Dial Transplant
2004;19:2505–12.
Kikuchi Y, Ikee R, Hemmi N, Hyodo N, Saigusa T,
Namikoshi T et al. Fractalkine and its receptor, CX3CR1,
upregulation in streptozotocin-induced diabetic kidneys.
Nephron Exp Nephrol 2004;97:e17–25.
Utimura R, Fujihara CK, Mattar AL, Malheiros DM,
Noronha IL, Zatz R et al. Mycophenolate mofetil prevents the
development of glomerular injury in experimental diabetes.
Kidney Int 2003;63:209–16.
Takahashi K, Mizuarai S, Araki H, Mashiko S, Ishihara A,
Kanatani A et al. Adiposity elevates plasma MCP-1 levels
leading to the increased CD11b-positive monocytes in mice.
J Biol Chem 2003;278:46654–60.
Zatz R, Dunn BR, Meyer TW, Anderson S, Rennke HG,
Brenner BM. Prevention of diabetic glomerulopathy by
pharmacological amelioration of glomerular capillary
hypertension. J Clin Invest 1986;77:1925–30.
Wolf G, Ziyadeh FN. The role of angiotensin II in diabetic
nephropathy: emphasis on nonhemodynamic mechanisms.
Am J Kidney Dis 1997;29:153–63.
Vidotti DB, Casarini DE, Cristovam PC, Leite CA,
Schor N, Boim MA. High glucose concentration stimulates
intracellular renin activity and angiotensin II generation
in rat mesangial cells. Am J Physiol Renal Physiol
2004;286:F1039–45.
Singh R, Sing AK, Alavi N, Leehey DJ. Mechanism of
increased angiotensin II levels in glomerular mesangial
cells cultured in high glucose. J Am Soc Nephrol
2003;14:873–80.
Hsieh TJ, Zhang SL, Filep JG, Tang SS, Ingelfinger JR,
Chan JS. High glucose stimulates angiotensinogen gene
expression via reactive oxygen species generation in rat kidney
proximal tubular cells. Endocrinology 2002;143:2975–86.
Zhang SI, To C, Chen X, Filep JG, Tang SS, Ingelfinger JR
et al. Essential roles (s) of the intrarenal renin-angiotensin
system in transforming growth factor-β1 gene expression and
induction of hypertrophy of rat kidney proximal tubular cells
in high glucose. J Am Soc Nephrol 2002;13:3002–12.
Mifsud SA, Allen TJ, Bertram JF, Hulthen UL, Kelly DJ,
Cooper ME et al. Podocyte foot process broadening in
experimental diabetic nephropathy: amelioration with
renin-angiotensin blockade. Diabetologia 2001;44:878–82.
Gross ML, El-Shakmak A, Szabo A, Koch A, Kuhlmann A,
Munter K et al. ACE-inhibitors but not endothelin receptor
blockers prevent podocyte loss in early diabetic nephropathy.
Diabetologia 2003;46:856–68.
Kelly DJ, Aaltonen P, Cox AJ, Rumble JR, Langham R,
Panagiotopoulos S et al. Expression of the slit-diaphragm
protein, nephrin, in experimental diabetic nephropathy:
differing effects of anti-proteinuric therapies. Nephrol Dial
Transplant 2002;17:1327–32.
Bonnet F, Cooper ME, Kawachi H, Allen TJ, Boner G,
Cao Z. Irbesartan normalizes the defiency in glomerular
nephrin expression in a model of diabetes and hypertension.
Diabetologia 2001;44:874–7.
Gloy J, Henger A, Fischer KG, Nitschke R, Mundel P,
Bleich M et al. Angiotensin II depolarizes podocytes in the
intact glomerulus of the rat. J Clin Invest
1997;99:2722–81.
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796
796
G. Wolf
105 Hoffmann S, Podlich D, Hahnel B, Kriz W, Gretz N.
Angiotensin II type 1 receptor overexpression in podocytes
induces glomerulosclerosis in transgenic rats. J Am Soc Nephrol
2004;15:1475 – 87.
106 Abbate M, Zoja C, Morigi M, Rottoli D, Angioletti S,
Tomasoni S et al. Transforming growth factor-beta1 is upregulated by podocytes in response to excess intraglomerular
passage of proteins: a central pathway in progressive
glomerulosclerosis. Am J Pathol 2002;161:2179 – 93.
107 Durvasula RV, Petermann AT, Hiromura K, Blonski M,
Pippin J, Mundel P et al. Activation of a local tissue
angiotensin system in podocytes by mechanical strain.
Kidney Int 2004;65:30 – 9.
108 Huang XR, Chen WY, Truong LD, lan HY. Chymase is
upregulated in diabetic nephropathy: implications for an
alternative pathway of angiotensin II-mediated diabetic renal
and vascular disease. J Am Soc Nephrol 2003;14:1738–47.
109 Benigni A, Corna D, Zoja C, Longaretti L, Gagliardini E,
Perico N et al. targeted deletion of angiotensin II type 1A
receptor does not protect mice from progressive nephropathy
of overload proteinuria. J Am Soc Nephrol 2004;15:2666–74.
110 Esteban V, Lorenzo O, Ruperez M, Suzuki Y, Mezzano S,
Blanco J et al. pathway, regulates the inflammatory response
in unilateral ureteral obstruction. J Am Soc Nephrol
2004;15:1514 –29.
111 Wolf G, Wenzel U, Burns KD, Harric RC, Stahl RAK,
Thaiss F. Angiotensin II activates nuclear transcription
factor-κB through AT1 and AT2 receptors. Kidney Int
2002;61:1986 – 95.
112 Fujisawa G, Okada K, Muto S, Fujita N, Itabashi N,
Kusano E et al. Spironolactone prevents early renal injury in
streptozotocin-induced diabetic rats. Kidney Int
2004;66:1493–502.
113 Schjoedt KJ, Andersen S, Rossing P, Tranow L, Parving HH.
Aldosterone escape during angiotensin II receptor blockade
in diabetic nephropathy is associated with enhanced decline
in GFR (Abstract). J Am Soc Nephrol 2003;14:7A.
114 Sato A, Hayashi K, Naruse M, Saruta T. Effectiveness of
aldosterone blockade in patients with diabetic nephropathy.
Hypertension 2003;41:64 – 8.
115 Hollenberg NK. Aldosterone in the development and
progression of renal injury. Kidney Int 2004;66:1–9.
116 Mazak I, Fiebeler A, Muller DN, Park JK, Shagdarsuren E,
Lindschau C et al. Aldosterone potentiates angiotensin
II-induced signaling in vascular smooth muscle cells.
Circulation 2004;109:2792–800.
117 Makimo H, Nakamura Y, Wada J. Remission and regression
of diabetic nephropathy. Hypertens Res 2003;26:515–9.
118 Adamczak M, Gross ML, Krtil J, Koch A, Tyralla K, Amann
K et al. Reversal of glomerulosclerosis after high-dose enalapril
treatment in subtotally nephrectomized rats. J Am Soc Nephrol
2003;14:2833–42.
119 Fioretto P, Steffes MW, Sutherland DE, Goetz FC,
Mauer M. Reversal of lesions of diabetic nephropathy after
pancreas transplantation. N Engl J Med 1998;339:69–75.
120 Perkins BA, Ficociello LH, Silva KH, Finkelstein DM,
Warram JH, Krolewski AS. Regression of microalbuminuria
in type 1 diabetes. N Engl J Med 2003;348:2285–93.
© 2004 Blackwell Publishing Ltd, European Journal of Clinical Investigation, 34, 785–796