REVIEW ARTICLE
Structural and functional specificities of PDGF-C and
PDGF-D, the novel members of the platelet-derived growth
factors family
Laila J. Reigstad
1,2
, Jan E. Varhaug
2,3
and Johan R. Lillehaug
1
1 Department of Molecular Biology, University of Bergen, Norway
2 Department of Surgical Sciences, University of Bergen, Norway
3 Haukeland University Hospital, Bergen, Norway
Introduction
The platelet-derived growth factors PDGF-A and -B
have since the late 1970s been recognized as important
factors regulating embryonic development, differenti-
ation, cell growth and many diseases including malig-
nancies. The PDGFs have been classified as members
of the superfamily of growth factors characterized by
the strongly conserved pattern of six cysteine residues
making up intra- and intermonomer disulfide bridges,
the cystine knot family of proteins [1–3]. Examples of
cystine knot subfamilies are the glycoprotein hormone
family [4], the cyclotide family [5,6], and the TGFb
family and NGF family [2]. Extended information
about subfamilies can be obtained in the Cystine Knot
Database ( />This review focuses on the structure and function of
the two novel members, PDGF-C and -D, of the
PDGF subfamily of the cystine knot superfamily. The
PDGFs show high sequence identity with the vascular

PDGF-D also plays a role in the lung and in periodontal mineralization.
PDGF-C is expressed in Ewing family sarcoma and PDGF-D is linked to
lung, prostate and ovarian cancers. Both PDGF-C and -D play a role in
progressive renal disease, glioblastoma⁄ medulloblastoma and fibrosis in
several organs.
Abbreviations
CNS, central nervous system; CUB, Clr ⁄ Cls, urchin EGF-like protein and bone morphogenic protein 1; CVB3, coxsackievirus B3;
EGF, endothelial growth factor; PDGF, platelet-derived growth factor; VEGF, vascular endothelial growth factor.
FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS 5723
endothelial growth factors (VEGF) and the family is
therefore often referred to as the PDGF ⁄ VEGF
family.
The PDGF family of growth factors
The PDGF family consists of PDGF-A, -B, -C and -D
[7–12]. The cystine knot motif of the four PDGFs
contains two disulfide bridges linking the antiparallel
strands of the peptide chain forming a ring penetrated
by the third bridge [3]. This forces the protein to adapt
a three-dimensional arrangement that partly exposes
hydrophobic residues to the aqueous surroundings,
leading to the formation of either homo- or hetero-
dimers (PDGF-AA, -AB, -BB, -CC, -DD) [13,14]. In
addition to a conserved cystine knot motif, these four
growth factors show a high sequence identity. The four
PDGFs are inactive in their monomeric forms. They
share the same receptors; the PDGF receptor-a and -b.
These receptors dimerize when the dimeric PDGF
binds. The receptors may combine to generate homo-
or heterodimers, resulting in three possible combina-
tions, PDGFR-aa,-ab and -bb, having different

PDGF ⁄ VEGF family because they encode the highly
conserved cystine knot motif characteristic of the
growth factor family. While the classical PDGF-a and
PDGF-b mainly encode the growth factor domain,
PDGF-c and PDGF-d encode a unique two-domain
structure with an N-terminal ‘Clr ⁄ Cls, urchin endo-
thelial growth factor (EGF)-like protein and bone
morphogenic protein 1’ (CUB) domain [16] in addition
to the C-terminal growth factor domain (Fig. 1A).
The pdgf genes are located on four different chromo-
somes; PDGF-a and -b on chromosomes 7 and 22
[17,18], and PDGF-c and -d on chromosomes 4 and 11
[19], respectively. The genomic organization of the pdgf
genes is quite similar, although PDGF-c and -d genes
are significantly longer due to large intron sizes and
cover about 200 kb compared to approximately 20 kb
for PDGF-a and -b [19–21].
Each of the four pdgf genes contains a long 5¢
untranslated region and a verified (PDGF-a and -b)or
putative (PDGF-c and -d) signal peptide in exon 1
(Fig. 1B). In the PDGF-c and -d genes, exons 2 and 3
encode the CUB domains, while in PDGF-a and -b
these exons encode precursor sequences residing 5¢ to
the cystine knot encoding sequence. The hinge regions
of PDGF-c and PDGF-d connecting the CUB and the
cystine knot domains are encoded by exons 4 and 5,
respectively. These hinge region sequences encode con-
served basic motifs and similar motifs are found in
PDGF-A and -B. The motifs are identified as proteo-
lytic cleavage sites for proteases used in post-transla-

sequence of the PDGF-c promoter. FGF-2 stimulates
Egr1 expression through the Erk ⁄ MAPK pathway,
and Egr1 translocates to the nucleus where it binds to
the proximal PDGF-c promoter resulting in increased
PDGF-C expression.
Alternative splicing of the PDGFs
No alternative splicing of PDGF-c mRNA has been
demonstrated. However, alternative splicing is sugges-
ted because two shorter PDGF-C cDNAs have been
obtained [25]. Based on the variant PDGF-c sequences
isolated by PCR, the splice donor ⁄ acceptor sites are
located to nucleotides 719 ⁄ 720 and 988 ⁄ 989, resulting
in two alternative proteins; one short variant encom-
passing almost only the CUB domain, and the longer
variant containing the CUB domain and the final 30
residues in the C-terminal end of the growth factor
domain. These splice variants are also present in
human thyroid papillary carcinomas (L. J. Reigstad,
J. E. Varhaug and J. R. Lillehaug, unpublished results).
Based on mRNA analysis of PDGF-d, splice variants
have been reported to be present in mouse heart, liver
and kidney [26]. Interestingly, deletion of exon 6 cau-
ses a frame shift and an early stop codon in exon 7,
resulting in a protein lacking the growth factor domain
and without mitogenic activity (Fig. 1B). The PDGF-
D protein encoded by this splice variant could only be
detected in mouse tissues and not in human cell lines
or tissues. A second PDGF-d RNA splice variant lacks
A
B

18 bp within the CUB domain of both mouse and
human PDGF-D mRNA [26,27]. In the case of PDGF-a,
exon 6 may be present or not resulting in two splice
variants encoding a long and a short PDGF-A protein
(Fig. 1B) [28]. PDGF-B mRNA has not been reported
to be alternatively spliced.
The PDGF proteins
The PDGF-A and -B proteins contain only the growth
factor domains whereas PDGF-C and -D have a
unique two-domain structure containing the N-ter-
minal CUB domain separated from the C-terminal
growth factor domain by a hinge region (Fig. 1A).
PDGF-C and -D share an overall sequence identity of
42% with highest similarity in the CUB and cystine
knot-containing growth factor domain, whereas the
hinge region and the N-terminal region show less
identity [10,12].
While PDGF-A and -B can form both homo- and
heterodimers (PDGF-AA, -AB, -BB), PDGF-C and -D
exist only as homodimers (PDGF-CC and -DD). The
full-length PDGF-C and -D monomers are 54–55 kDa
and 49–56 kDa, respectively, differing from their theo-
retical sizes of 39 and 43 kDa based on their amino
acid sequences [8,12,29]. The divergence from the
theoretical values indicates that PDGF-C and -D may
be post-translationally modified. In addition to being
secreted to the extracellular space, the PDGF-C
protein is shown to be constitutively expressed in the
cytoplasm in rat smooth muscle cells residing in
arteries and arterioles [30]. Additionally, our data

tity among the sequences. The four cysteines conserved in the prototypical CUB domains are labelled in yellow. The two cysteines missing
in the PDGF-C and PDGF-D are marked by red circles while the two cysteines present are marked in blue circles. The two cysteines of
PDGF-C (accession no AAF80597) correspond to Cys104 and 124, whereas in PDGF-D (accession no AAK38840) these cysteines are
Cys109 and 131.
PDGF-C and -D, structure and function L. J. Reigstad et al.
5726 FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS
compact ellipsoidal b-sandwich, with a hydrophobic
core essential for the overall domain folding. The
b-sandwich is built up of two five-stranded b-sheets of
antiparallel b-strands [33–36]. Most CUB domains are
reported to contain four conserved cysteines that form
two disulfide bridges between nearest-neighbour cyste-
ines, resulting in disulfide bridges located on opposite
edges of the domain. As both the PDGF-C and -D
CUB domains contain only two cysteines [7] which,
compared to the classical CUB domains, are the two
most-C-terminally located cysteines, the CUB domains
of PDGF-C and -D may have only one disulfide
bridge. At present it is unclear how this may influence
their 3D structure. In the two crystallised CUB
domains of a serine protease associated with serum
mannose-binding proteins (MAPS), the N-terminally
located CUB domain contains only one disulfide
bridge, while the second CUB domain of MAPS has
two bridges. One disulfide bridge instead of two may
result in a slightly less tight b-sandwich in the N-ter-
minally located CUB domain, but the structural and
functional significance of only one bridge remains
unknown [35].
The CUB domain is found in several extracellular

Furthermore, a role for the PDGF CUB domains in
receptor binding is suggested based on studies of the
transmembrane receptor, neuropilin-1, which consists
of two CUB domains and a coagulation factor
domain, acting as coreceptors for VEGF-A and sem-
aphorins (reviewed in [40]). Crystallography studies of
the MAPS protein containing two CUB domains sug-
gests that CUB domains may also participate in pro-
tein heterodimer formations [35].
The hinge region and proteolytic cleavage for
growth factor activation
The hinge regions of PDGF-C and -D, separating the
CUB and the growth factor domains (Fig. 1), show no
homology to known sequences [41] but contain dibasic
cleavage sites for proteolytic removal of the CUB
domains and thereby activation of the growth factor
domains. PDGF-C and -D contain both the CUB and
growth factor domains when they are secreted and pro-
teolytic cleavage is therefore suggested to take place
extracellularly. Plasmin cleaves PDGF-C at RKSR234
[8,41], and PDGF-D at RKSK257 [12]. Tissue plasmi-
nogen activatior (tPA) cleaved PDGF-C at RKSR234
in vivo [42,43] and urokinase plasminogen activator
(uPA) was found to cleave PDGF-D at RGRS250,
thereby activating this growth factor [44]. PDGF-A is
cleaved by furin at RRKR86 [45], while PDGF-B is
cleaved at RGRR81 by a still unidentified protease [46].
The PDGF growth factor domain
The determination of the crystal structure of nerve
growth factor [47], transforming growth factor b2 [48],

(Fig. 3B). Six of the conserved cysteines are engaged in
three intrachain disulfide bonds (Cys I-VI, III-VII,
V-VIII) stabilizing the cystine knot structure, while
two cysteines (Cys II and IV) are involved in inter-
chain disulfide bonds (Fig. 3C,D) [55,56]. The three
intrachain disulfide bonds makes the cystine knot very
Fig. 3. The PDGF-C growth factor domain.
Ribbon presentations of the proposed
PDGF-C model [57] displaying the twisted
b-sheets and the N-terminal a-helix of the
PDGF-C monomer (A) and the PDGF-CC
dimer (B). The N-terminal (N) and C-terminal
(C) ends for the monomer are marked. The
three loops (loop 1-2-3) connecting the
b-strands are labelled. (C, D) Sequence
alignments of the growth factor domains of
PDGF-A (accession no P15692), PDGF-B
(accession no. 1109245 A), PDGF-C (acces-
sion no AAF80597), PDGF-D (accession
no AAK38840), VEGF-A (accession no
NP003367) and PIGF-1 (accession no.
1FZV). Red, squared areas show sequence
identity among the sequences. The eight
conserved cysteines are shown in yellow.
The extra cysteines of PDGF-C and -D are
labelled in blue. The green squares highlight
the area of disagreement in sequence align-
ment (see text). (C) Sequence alignment of
PDGF ⁄ VEGF family members where the
green square highlights the area containing

identification of the cysteines that participate in the
conserved disulfide bridges difficult. Because of this,
two different sequence alignments of PDGF-C and -D
covering the area of conserved cysteines III to VI are
included here (Fig. 3C,D). Figure 3C shows the align-
ment of PDGF-C and -D to allow three residues
(NCA and NCG, respectively) between conserved
cysteines III and IV, an insert not present in the other
members of the PDGF ⁄ VEGF family. This alignment
also indicates that PDGF-D lacks the conserved cys-
teine V [7,8,10,12,15,25,57]. The alignment in Fig. 3D
has a different three-residue insert, which is located
between conserved cysteines V and VI. In this align-
ment, PDGF-D contains all eight conserved cysteines
of the cystine knot motif [20,21,41]. Crystallization or
NMR studies of PDGF-C and -D proteins will resolve
this debate, but our published 3D model of the
PDGF-C growth factor domain indicates the disulfide
bridges in PDGF-C to consist of Cys250 and 294,
Cys280 and 335, and Cys287 and 337, and the inter-
monomeric bonds to consist of Cys274 and 286 [57].
At present, analysis of PDGF ⁄ VEGF domains show
that PDGF-C is more similar to VEGFs than PDGFs
[8,15,57], all in all favouring the alignment in Fig. 3C.
The region C-terminal of the growth factor
domain
In PDGF-A and -B, the C-terminal regions contain a
basic sequence with a dual function. First, the
sequence mediates electrostatic interactions with com-
ponents of the extracellular matrix such as heparin [58]

SUMOylation or ubiquitinylation may be candidates.
Receptor binding of PDGFs
The PDGFs bind to the protein tyrosine kinase
receptors PDGF receptor-a and -b. These two recep-
tor isoforms dimerize upon binding the PDGF
dimer, leading to three possible receptor combina-
tions, namely -aa,-bb and -ab. The extracellular
region of the receptor consists of five immunoglo-
bulin-like domains while the intracellular part is a
tyrosine kinase domain. The ligand-binding sites of
the receptors are located to the three first immuno-
globulin-like domains (reviewed in [64]). The residues
in PDGF-A and -B responsible for receptor binding
reside in loop 2, in addition to RKK161 in PDGF-
AA and R27 and I30 in PDGF-BB. The residues
involved in PDGF-CC and -DD receptor binding
remain to be identified, but our published 3D model
of PDGF-C suggests, when compared to the crystal
structure of VEGF-AA complexed to domain 2 of
its receptor, that the region containing residues
W271 and LR312 might be involved [57].
L. J. Reigstad et al. PDGF-C and -D, structure and function
FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS 5729
PDGF-CC specifically interacts with PDGFR-aa and
-ab, but not with -bb, and thereby resembles PDGF-
AB [8,41]. PDGF-DD binds to PDGFR-bb with high
affinity, and to PDGFR-ab to a markedly lower extent
and is therefore regarded as PDGFR-bb specific [10,12].
PDGF-AA binds only to PDGFR-aa, while PDGF-BB
is the only PDGF that can bind all three receptor

PDGF-AA signalling [69]. Data on possible PDGF-CC
or -DD binding to SPARC have not been reported. The
major reversible PDGF-A and -B binding to extracellu-
lar protein is a
2
-macroglobulin [70]. The PDGF–a
2
-
macroglobulin complex serves multiple functions. It
makes PDGF-AA, -AB and -BB unable to bind their
receptors, it protects the PDGFs against proteolytic
degradation, and may remove the PDGFs from circula-
tion via a
2
-macroglobulin receptors. There are no data
currently available about interactions between the novel
PDGFs and a
2
-macroglobulin, but several other growth
factors, such as FGF-2, TGF-b and TNF-a, also bind
a
2
-macroglobulin. Fifth, the expression of highly speci-
fic proteases that proteolytically activate the PDGFs
will also influence the availability and activity of the dif-
ferent isoforms. This can be exemplified by the proteo-
lytic cleavage of PDGF-D. While the human prostate
carcinoma cell line LNCaP produces a specific protease
to process the full-length PDGF-D [66], there is no pro-
tease capable of cleaving the full-length PDGF-D secre-

tractile phenotype, functioning as anchors for an
involuted epithelial sheet, the alveolar sac or the glom-
erulus. By losing this anchor in the knockout mice,
there is a failure of involution and the physiological
functions are impaired, as a result of decreased surface
area for gas exchange or glomerular filtration, in
PDGF-A and PDGF-B mutants, respectively.
Knockout studies on PDGF-c in mice clearly dem-
onstrate a role for PDGF-C in embryonal development
[73]. The knockout of PDGF-c results in mice dying
perinatally owing to difficulties in feeding and brea-
thing, as they have a complete cleft of the secondary
palate because the palate bones do not meet. Addition-
ally, the dorsal spinal cord was deformed in the
lower spine. The null mutant PDGF-C embryos had
PDGF-C and -D, structure and function L. J. Reigstad et al.
5730 FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS
subcutaneous oedema in the flank of the body between
the limbs lacking connective tissue, and showed several
blood-filled blisters in frontnasal and lateral forehead.
In the early embryo development, the features of the
knockout PDGF-c mice largely overlap with knockout
PDGF-a mice. PDGF-c ⁄ PDGF-a mice showed growth
retardation, pericardial effusion, a wavy neural tube
and subepidermal blisters, dying before E17. In total,
PDGF-C has specific roles in palatogenesis and in
morphogenesis of the skin tissue. PDGF-d knockouts
have not been reported.
Functions of the PDGF-C and PDGF-D proteins
The role of PDGF-A and -B proteins in normal pro-

increased mRNA and protein levels of metalloprotein-
ase-1 (MMP-1) and its inhibitor (TIMP-1), both being
important in the remodelling phase of tissues [78].
These results are further verified by in vivo experiments
showing that PDGF-C enhanced the repair of a full-
thickness skin excision in a delayed diabetic wound
healing mouse model by stimulation of fibroblast pro-
liferation, epithelial migration, extensive vasculariza-
tion and neutrophil infiltration [41].
PDGF-C in angiogenesis
The high PDGF-C expression in the angiogenic tissues
of placenta, ovary and embryo has led to several in vitro
and in vivo experiments defining PDGF-C as a potent
angiogenic factor, similar to VEGF and the classical
PDGFs. The underlying mechanisms are still to be
understood. In the aortic ring outgrowth assay, PDGF-
C mediated significant increased outgrowth of fibro-
blasts and smooth muscle cells, to a degree comparable
to that of VEGF, PDGF-AA and -BB [41]. PDGF-C
efficiently stimulated the formation of new blood vessels
with high vessel density growing towards the implanted
dish of the chorioallantoic membrane (CAM) assay
[79]. In addition, PDGF-C stimulated formation of new
branches and vessel sprouts from those initially formed.
Several reports show in vivo angiogenic PDGF-C
effects. When PDGF-C-coated micropellets were added
to mouse corneal micropockets, PDGF-C potently
induced neovascularization of the avascular corneal tis-
sue. In these experiments, PDGF-C was as potent as
PDGF-BB and more potent than PDGF-AA. PDGF-

mesenchyme and in the parietal epithelial cells [30,81].
During kidney development, PDGFR-a is expressed
in the glomerular epithelial mesenchyme, suggesting a
paracrine signalling pathway for both PDGF-A and
-C in kidney vascular and interstitial development [74].
In the embryonal rat CNS, PDGF-C mRNA was
expressed in the notochord (prestage of the spinal
cord) and subsequently in the maturing spinal cord,
while the adult spinal cord does not express PDGF-
C [27]. The presence of PDGF-C in the developing
spinal cord has also been shown in chicken [7].
PDGF-C mRNA is detected in the floor plate and
the ventricular zones of cortex and adjacent to the
floor plate of the embryonic brain, whereas in the
adult brain weak PDGF-C expression was observed
only in the olfactory nucleus and pontine nuclei [27].
Quantitative RT-PCR analysis did not detect PDGF-
C in human embryonic or adult brain tissues [82],
although this has been shown through northern blot
analyses [8,74].
PDGF-C mRNA has been detected in the develop-
ing ears of mouse and rat [9,74,83]. During rat
embryonic development, significant mRNA levels of
PDGF-C, PDGF-A and both PDGFRs are expressed
in cochlear progenitor hair cells of the inner ear [83].
PDGF-D in normal processes
Since its discovery four years ago, PDGF-D has been
linked to important functions both in embryogenesis
and in adult tissues (Fig. 4). In human adult tissue,
PDGF-D is highly expressed in heart, kidney, pan-

indicated [29]. In the human adult kidney, PDGF-D
protein expression was also detected in smooth mus-
cle cells of arteries, arterioles and vasa rectae. In
contrast to human and mouse adult kidney, the rat
adult kidney shows no PDGF-D protein in the
glomeruli [85]. As in the kidneys, lungs show spe-
cies-different PDGF-D expression. Cells in normal
human lungs do not express PDGF-D protein at
detectable levels [10], while in murine lungs PDGF-D
mRNA is constitutively expressed [84].
In the embryo, PDGF-D mRNA is hardly detect-
able in the spinal cord, but in the adult spinal cord
prominent expression is located to the motor neurons
[27]. In the brain, PDGF-D mRNA was registered
in the thalamus and in a ventricular zone of the
Kidney (9,30,74,81)
CNS (5,27,74)
Heart (74,100)
Ear (9,74,83)
Kidney (30,80,81)
CNS (27,74)
Embryonic development
Adult tissue
PDGF-C
PDGF-D
Kidney (27,29,80,85)
Eye (27,39)
CNS and brain (27)
Lung (84,101)
Peri. mineral. (86)

protein was found in its full-length form in all eye
tissues and cells investigated without sign of activation
by proteolytic removal of the CUB domain.
In periodontal mineralization, PDGF-D ⁄ PDGFR-b
appear to constitute an autocrine pathway when perio-
dontal ligament cells differentiate to cementoblasts
during in vitro formation of mineralized nodules [86].
During the mineralization process, the levels of both
PDGFR-b and PDGF-D in the ligament cells increase
significantly, whereas PDGF-B was not detected in
these cells.
PDGF-D in angiogenesis and wound healing
PDGF-D has been shown to stimulate angiogenesis
and deposition of the extracellular matrix, and thus
plays a part in the wound healing process [87–89].
Overexpression of PDGF-D in otherwise PDGF-
D-negative skin increased both the amount of macro-
phages in the dermis and the interstitial fluid pressure
[87]. The PDGF-D-mediated elevation of interstitial
fluid pressure is consistent with PDGFR-b being essen-
tial for the maintenance of steady state pressure level.
As the keratin 14 promoter is strongly upregulated
during wound healing, abundant PDGF-D was deliv-
ered when skin punch biopsy wounds were made in
mice. In this initial inflammation phase of wound heal-
ing, activated platelets secrete PDGF-A, -B and -C,
but not PDGF-D [76]. No endogen PDGF-D mRNA
could be detected [87], indicating that normally
PDGF-D does not play a role in the inflammatory
phase. A recent report supports the involvement of

made PDGF-B one of the earliest discovered onco-
genes, connecting PDGF ⁄ PDGFR to cellular transfor-
mation [96]. The finding that PDGF-C acts much like
PDGF-AB, the most mitogenic of all PDGFs [97], as
it activates both PDGFR-aa and -ab, indicates a role
for PDGF-C as an oncogene [41,82,98]. PDGF-C
induces tumours in nude mice, activates anchorage-
dependant growth, and is a potent transforming
growth factor of NIH ⁄ 3T3 cells [88]. The in vivo
tumourigenesis may partially be explained by PDGF-
C-mediated VEGF expression, promoting indirect sti-
mulation of tumour angiogenesis. The apparent roles
of PDGF-C in diseases and malignancies are discussed
below and are summarized in Fig. 5.
PDGF-C in Ewing family sarcomas
The first link between PDGF-C and malignancy was
provided when PDGF-C was identified as a secreted
transforming growth factor in Ewing family sarcomas
[99]. In these aggressive bone and soft tissue tumours,
L. J. Reigstad et al. PDGF-C and -D, structure and function
FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS 5733
arising due to aberrant EWS ⁄ FLI transcription factors,
PDGF-C was identified as an upregulated downstream
target. The expression of a dominant negative PDGF-
C in Ewing sarcoma cells caused a marked reduction
in anchorage-independent growth [99] but PDGF-C
was unable to fully reconstitute all features of the
EWS ⁄ FLI phenotype [100]. These findings, in combi-
nation with observations that selective inhibitors of
PDGFR inhibited anchorage-independent growth, sug-

showed low levels of PDGFs and receptors. PDGFR-a
was endogenously activated in three out of five cell
lines, while PDGFR-b was not expressed, suggesting
an autocrine loop in medulloblastoma involving
PDGFR-a only. In cell lines having endogenously acti-
vated PDGFR-a, PDGF-C was highly expressed while
PDGF-A could not be detected or detected at very low
levels, indicating PDGF-C to play a significant role in
an autocrine loop of medulloblastoma tumours.
PDGF-C in progressive renal diseases
Characteristics of progressive renal diseases are a high
proliferation of interstitial cells (hypercellularity),
increased matrix accumulation and influx of macro-
phages to the glomeruli [105]. The autocrine loop of
PDGF-B ⁄ PDGFR-b has been shown in many progres-
sive renal diseases. From recent findings, PDGF-D
appears to act either in concert with, or compete with,
PDGF-B in autocrine signalling within the mesangium
[80]. Usually, PDGF-C mRNA is constitutively
expressed in parietal epithelial cells and smooth muscle
cells of human kidneys, but in injured renal states
Ewing family sarcoma
Progressive renal disease Glioblastoma/Medulloblastoma Fibrosis
Lung cancer
Prostate cancer
Ovarian cancer
PDGF-D
(98,99,100,101)
(82,94)
(30,80,81)

normal kidney and the expression is further increased in
the diseased kidney, suggesting that PDGF-C directly
interferes with the reported PDGF-A ⁄ PDGFR-a auto-
crine signalling present in the interstitium [80].
With respect to mesangioproliferative glomerulonep-
ritis in rats, PDGF-C is upregulated in the glomerul ar
mesangium but not in endothelial or epithelial glomer-
ular cells [30]. PDGF-C mRNA is markedly upregulated
in interstitial cells of rats suffering from severe tubulo-
interstitial fibrosis [85]. Although both PDGF-C
and PDGF-A are present in the interstitium, analyses
of rats with induced kidney disease show PDGF-A,
-B and -D mRNA levels to peak at days 7–9, while
PDGF-C mRNA has its maximum on day 28 after
disease induction. This suggests temporally different
levels of PDGF-C and PDGF-A possibly leading to a
dominating PDGF-C ⁄ PDGFR-a autocrine loop in later
stages of the disease.
PDGF-C in liver, lung, heart and pancreatic
fibrosis
A recent report shows that overexpression of PDGF-C
leads to liver fibrosis, steatosis (overproduction of mat-
rix) and hepatocellular carcinomas in transgene PDGF-
C mice [106]. PDGF-C is likely to be a hepatic fibrosis
inducer as the PDGF-C overexpression resulted in signi-
ficantly increased mRNA and protein levels of key pro-
fibrotic proteins like PDGFR-a and -b, TGFb1, and
TIMP-1 and -2 in the liver of transgenic mice. PDGF-C
treatment increased both mRNA and protein levels
of the metalloproteinase inhibitor, TIMP-1, in dermal

surgically resected pancreatic cancerous lesions showed
PDGF-C mRNA to be upregulated compared to non-
neoplastic pancreatic tissue. The upregulation correla-
ted with collagen type I and III expression, indicating
that PDGF-C is a potent inducer of matrix overpro-
duction and thereby fibrosis in the pancreas.
Two reports show the effect of in vivo PDGF-C
overexpression using transgenic techniques in which
the full-length PDGF-c gene was targeted to the mouse
heart where PDGF-C was overexpressed [8,38]. The
overexpression induced hyperproliferation of myocar-
dial interstitial cells, giving expanded heart interstitium
and resulting in animals with progressive fibrosis and
cardiac hypertrophy [8]. The expanded heart intersti-
tium gave drastic disorganized myocardial fibres, pro-
gressive collagen accumulation and increased thickness
of the ventricular walls.
Additionally, recent reports on major histocompati-
bility complex class II knockout mice shows elevated
expression of PDGF-C in the hearts after infection
with coxsackievirus B3 (CVB3), but not in hearts of
immunocompetent mice infected with CVB3 [67,109].
In the immunodeficient mice, the CVB3 infection led
to chronic myocarditis characterized by marked cell
infiltration to the myocardium, and necrosis of cardio-
myocytes, resulting in heart fibrosis. In these hearts,
upregulation of both PDGF-C and PDGFR-a was
located to the areas of inflammatory cell invasion.
Taken together, in myocardial, liver, lung and pancre-
atic fibrosis, PDGF-C plays a role in fibrosis develop-

The glomerular and systemic in vivo overexpression
of PDGF-D in the pathogenesis of mesangio-prolifera-
tive changes is responsible for a large glomerular macro-
phage influx, for significant increase in glomerular
size, and an increased proliferation of the mesangial
cells that have gained a myofibroblast phenotype
(expression of a-smooth muscle actin) which is respon-
sible for the increased accumulation of collagen type I
and IV matrix [80,85,110]. Furthermore, PDGF-D is
a likely candidate for controlling the progression of
metastatic renal cell carcinoma since its overexpression
enhanced tumour progression and metastasis in an
orthotropic tumour model in SCID mice [111].
PDGF-D in fibrosis
Progressive renal diseases often end as interstitial fibro-
sis, and data on human and murine PDGF-D protein
expression at this endpoint have been reported [80].
After induction of interstitial fibrosis in mice, expres-
sion of PDGF-D, -B and the PDGFRs was detected in
the interstitial fibroblasts, in contrast to the interstitial
cells of normal kidney. The PDGFR-b protein was dis-
tributed in the tubointerstitial fibrosis areas with an
immunostaining pattern corresponding to the sites of
PDGF-D but less so to the PDGF-B positive sites.
Similar results were obtained in humans suffering from
chronic renal nephropathy, suggesting that PDGF-D
exerts its fibrogenic effect by activating PDGFR-b,
and furthermore to be as significant as PDGF-B in
progressive renal diseases. PDGF-D may act in concert
with, or compete with, PDGF-B in autocrine signalling

PDGF-D levels were also detected in sera of lung and
ovarian cancer patients [89]. This high level of circula-
ting PDGF-D is also detected in mice infected with
adenovirus-containing PDGF-D, resulting in peri-
vascular lymphoid cell infiltrates of the lung and fibro-
sis in the liver [110].
Conclusions
The novel members, PDGF-C and -D, of the PDGF
subfamily of the cystine knot family of growth factors
are potent cytokines important for normal embryogen-
esis and maintenance of adult tissues in several species.
Both PDGF-C and -D are involved in various malfunc-
tions such as progressive renal diseases, fibrosis in
many organs, with specific functions (PDGF-C) in
Ewing family sarcomas, and (PDGF-D) in lung,
PDGF-C and -D, structure and function L. J. Reigstad et al.
5736 FEBS Journal 272 (2005) 5723–5741 ª 2005 The Authors Journal compilation ª 2005 FEBS
prostate, and ovarian cancer. Structure models of
PDGF-C suggest it to resemble VEGF-A but conclu-
sive determination of the 3D structures of both PDGF-
C and -D is lacking. The current information on their
activities points to a wide scope of biological effects;
however further understanding of how these factors
interplay with other members of the cystine knot family
and in particular the PDGF-A, -B, and VEGF growth
factors, must be the focus of future investigations.
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