Injury, Int. J.

Care Injured 45 (2014) 1816–1823

Contents lists available at ScienceDirect

Injury
journal homepage: www.elsevier.com/locate/injury

Review

The role of pleiotrophin in bone repair

Margarita Lamprou a,1, Angelos Kaspiris a,1, Elias Panagiotopoulos b, Peter V. Giannoudis c,d,
Evangelia Papadimitriou a,*
a
Laboratory of Molecular Pharmacology, Department of Pharmacy, School of Health Sciences, University of Patras, 26504 Patras, Greece
b
Academic Department of Trauma & Orthopaedics, School of Medicine, University of Patras, Greece
c
Academic Department of Trauma & Orthopaedic Surgery, University of Leeds, Clarendon Wing, Floor A, Great George Street, Leeds General Infirmary,
LS1 3EX Leeds, UK
d
NIHR Leeds Biomedical Research Unit, Chapel Allerton Hospital, LS7 4SA Leeds, West Yorkshire, UK

A R T I C L E I N F O A B S T R A C T

Article history: Bone has an enormous capacity for growth, regeneration, and remodelling, largely due to induction of
Accepted 7 October 2014 osteoblasts that are recruited to the site of bone formation. Although the pathways involved have not
been fully elucidated, it is well accepted that the immediate environment of the cells is likely to play a
Keywords: role via cell–matrix interactions, mediated by several growth factors. Formation of new blood vessels is
Angiogenesis also significant and interdependent to bone formation, suggesting that enhancement of angiogenesis
Bone could be beneficial during the process of bone repair. Pleiotrophin (PTN), also called osteoblast-specific
Bone repair
factor 1, is a heparin-binding angiogenic growth factor, with a well-defined and significant role in both
Osteoblasts
Osteoclasts
physiological and pathological angiogenesis. In this review we summarise the existing evidence on the
Pleiotrophin role of PTN in bone repair.
ß 2014 Elsevier Ltd. All rights reserved.

Contents

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1816
Materials and methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1817
Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1817
PTN receptors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1817
Regulation of PTN expression. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1817
PTN and angiogenesis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1818
PTN and bone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1819
PTN pathways in clinical skeletal diseases . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1819
Osteoporosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1819
Fracture healing. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1820
Osteoarthritis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1820
Discussion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1820
Conflict of interest statement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1821
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1821

Introduction

The formation of new blood vessels is vital to the functioning of all

tissues during development, as well as during regeneration process-
* Corresponding author at: Laboratory of Molecular Pharmacology, Department
es. In this context, at a bone fracture site, angiogenesis restores the
of Pharmacy, University of Patras, GR 26504 Patras, Greece. Tel.: +30 2610 962336.
E-mail address: epapad@upatras.gr (E. Papadimitriou). hypoxia and nutrient deprivation and facilitates osteogenesis. A
1
These authors have equally contributed to the work. number of therapeutic approaches that use angiogenic growth

http://dx.doi.org/10.1016/j.injury.2014.10.013
0020–1383/ß 2014 Elsevier Ltd. All rights reserved.
 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
factors and lead to enhanced angiogenesis seem to have a beneficial
effect on bone repair, as evidenced both in vitro and in vivo, including
the use of vascular endothelial growth factor (VEGF), fibroblast
growth factors (FGFs), transforming growth factor, platelet derived
growth factor (PDGF) and bone morphogenetic proteins (BMPs)
[1]. However, clinical data are limited and further research is required
in order to elucidate the interrelationships among the involved
molecules and pathways, and to apply such approaches to cases
where bone regeneration is desirable.
Pleiotrophin (PTN) is a secreted angiogenic growth factor with a
well defined regulatory role in physiological and pathological
angiogenesis [2,3] and a potential role in bone formation and
remodelling. The peptide sequence is highly conserved across
different species, with more than 90% identity observed among the
sequences of chicken, rat, bovine and human [4]. Pleiotrophin was
initially purified from bovine uterus and neonatal rat brain, while
homologues have been reported in different species such as
human, mouse, chicken, fish, frogs and insects [5]. The protein is
also known as heparin-binding growth-associated molecule,
heparin affin regulatory peptide, heparin binding growth factor-
8, protein 18 kDa, heparin binding neurotrophic factor and
osteoblast specific factor 1 (OSF-1) [2,3]. The term OSF-1 was
attributed to the factor in the early 90s, due to the fact that a cDNA
clone coding for PTN was isolated from the murine osteoblastic cell
line MC3T3-E1. The human counterpart was also found in cDNA
libraries from the human osteosarcoma cell line MG-63 [6].
Heteronuclear NMR studies have not concluded to PTN’s
conformation but have shown that PTN contains two b-sheet
domains connected by a flexible linker. Each of these domains
contains three antiparallel b-strands, which are homologous to the
thrombospondin type 1 repeat (TSR-1) [4], a motif that is found in a
wide variety of proteins that mediate interactions between cells
and/or between cells and the extracellular matrix. Thrombospon-
din and other TSR-containing proteins influence multiple aspects
of bone development and remodelling [7] supporting the
hypothesis that PTN may also be a significant regulator.
Materials and methods
The authors explored the MEDLINE/PubMed (1946 – present)
databases of the National Library of Medicine and EMBASE (1947 –
present) for appropriate articles addressing the significance of
pleiotrophin in bone repair. The key words, which were searched,
were the terms ‘‘angiogenesis’’, ‘‘bone’’, ‘‘bone repair’’, ‘‘osteo-
blasts’’, ‘‘osteoclasts’’ and ‘‘pleiotrophin’’, in different combina-
tions. Searching of the reference lists of potentially relevant origin
was also performed. Inclusion criteria were: papers written in
English, peer-reviewed journals, in vitro and in vivo studies that
evaluated the implication of PTN and/or its receptors in bone
regeneration process or its involvement in diseases of the skeletal
system which are associated with bone remodelling defects.
Exclusion criteria were: articles using language other than English,
letters and expert opinion publications. Articles that conformed to
the above mentioned criteria were retrieved and all of the studies
related to these were extensively searched.
Results
The search strategies yielded a total of over 200 potential
articles. During the selection process, the articles were excluded by
title or by abstract, because they were clearly irrelevant to the
study question. The papers, which fulfil the inclusion criteria, could
be distributed in four main categories. The first category comprised
the articles concerning the in vitro or in vivo expression of PTN
[6,8–15] or its receptors [8,9,16–20] in cells of the bone or the
cartilage tissue. The second category included the studies, which
1817
examined actions and possible mechanisms of PTN [8,21–27] or its
receptors [8,16,17,28–31] actions in bone cells. The third category
comprised of the studies, which investigate the macroscopic
effects of altered PTN expression on bone formation and/or
remodelling in animal models [8,32–40] or in clinical studies
[41]. Finally, the fourth category included articles on the regulation
of PTN expression in cells of the skeletal system and/or by factors
known to play a role in bone repair or remodelling [42–54], as well
as data on the PTN actions related to angiogenesis [55–78].
PTN receptors
The first identified PTN receptor was N-syndecan through its
heparan sulfate chains [79,80]. N-syndecan is expressed by
osteoblasts/osteoblast precursors and may participate in the
regulation of osteoblast recruitment [8]. Besides N-syndecan,
PTN also interacts with syndecans 1 and 4, which contain both
heparan and chondroitin sulfate chains. PTN binds to the highly
sulfated chondroitin sulfate (CS) chains of syndecan-4 with higher
affinity than to those of syndecan-1 and removal of the CS chains
decreased the association and dissociation rate constants of PTN
for both syndecans [81].
The best studied PTN receptor is receptor protein tyrosine
phosphatase beta/zeta (RPTPb/z), initially isolated from neural
tissue as a transmembrane protein-tyrosine-phosphatase that
consists of a large glycosylated extracellular domain, a transmem-
brane region and a cytoplasmic portion that contains two repeated
tyrosine phosphatase domains. Alternative splicing and proteolytic
processing give rise to secreted, transmembrane, or cytoplasmic
forms of not yet fully identified biological significance [82]. RPTPb/
z has a well described role in PTN-induced cell migration [83] and
may have a role in bone formation and remodelling based on the
data presented in Table 1.
An important PTN receptor that determines the stimulatory or
the inhibitory effect of PTN on cell migration is anb3 integrin,
which forms a functional complex with RPTPb/z on the cell surface
[75], and mediates PTN-induced cell surface nucleolin localization
[76]. PTN also interacts with several glycosaminoglycans (GAGs)
and such interactions contribute to PTN dimerization or storage
into the extracellular matrix [84,85], as well as protection of PTN
from proteolytic degradation [77]. PTN has a high affinity for both
low and over-sulfated CS chains, although the binding is more
intense with highly sulfated CS-D and CS-E [86]. An increase of
total sulfated GAGs was observed after supraspinatus tendon
overuse, which correlated with an increase in PTN protein levels,
suggesting that increased GAGs may sequester PTN, which may
thus initiate a shift towards the chondrocyte phenotype [20]. Data
that correlate anb3 or GAGs with bone remodelling are presented
in Table 1.
Nucleolin, a 100 kDa multifunctional protein, is considered to
be a low affinity receptor for PTN [87,88]. Cell surface nucleolin
interacts with both RPTPb/z and avb3 [76] and is required for PTN
[78]- and VEGF [89]-induced cell migration. Finally, anaplastic
lymphoma kinase (ALK), a 220 kDa receptor tyrosine kinase, has
been also mentioned as PTN receptor, although more recent
studies interrogate direct binding of PTN to ALK and favour the
notion that ALK is indirectly activated by PTN through PTN-
dependent inactivation of RPTPb/z [90]. There are no studies on
the role of nucleolin or ALK in bone pathophysiology.
Regulation of PTN expression
PTN expression has been described as regulated by several
factors, including growth factors, cytokines, hormones, signalling
molecules, transcription factors and miscellaneous conditions. One
of the first studies showed up-regulation of PTN by PDGF-AB in
1818 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
Table 1
Existing evidence on a possible role of the known PTN receptors in bone
pathophysiology.
PTN Skeletal effect
receptor
Syndecans N-syndecan expression is increased parallel to PTN expression
in the periosteum accompanying the
regenerative bone response in an adjuvant-induced
injury model [8].
Syndecan 4 is functionally involved in endochondral
ossification, and fracture healing is markedly delayed in
syndecan 4 knockout mice and accompanied by
increased callus formation [16].
RPTPb/z RPTPb/z-deficient mice display a decreased trabecular bone
volume at the age of 50 weeks, caused by a
reduced bone formation rate [17].
RPTPb/z-deficient calvarial osteoblasts display decreased
expression of osteoblast markers and a
diminished potential to form osteocyte-like cellular
extensions on Matrigel-coated surfaces [17].
PTPRZ1 gene is over-expressed in samples of
osteosarcoma patients versus normal bone, but with
no correlation between PTPRZ1 gene expression profile,
clinicopathological parameters and survival [18].
It is expressed by chondrocytes and subchondral
osteocytes in osteoarthritis patients [9].
anb3 Mediates the anabolic effects of mechanical stimulation
in osteoblasts [28].
It is critical to osteoclast function and development
[19,29].
It is a central molecule for osteoclastic bone resorption
[30].
GAGs Artificial extracellular matrices composed of collagen I
and synthetically over-sulfated GAGs reveal significant pro-
osteogenic effect, as determined by tissue
nonspecific alkaline phosphatase activity and calcium
deposition, especially when tested in early osteoblasts
[31].
An increase of total sulfated GAGs was observed 4 weeks
after supraspinatus tendon overuse, which remained elevated
up to 16 weeks [20].
Nucleolin No studies available.
ALK No studies available.
NIH-3T3 cells [42]. PDGF-BB was also found to increase PTN mRNA
levels in hepatic stellate cells, one of the major effector cell types
during the repair process of liver tissue injury [43]. Basic FGF
(bFGF) also increases PTN mRNA levels in NIH3T3 cells [42], as well
as ptn gene transcription and PTN protein levels in human prostate
cancer LNCaP cells, through NAD(P)H oxidase-dependent hydro-
gen peroxide production, phosphorylation of ERK1/2 and p38, and
activation of AP-1. Interestingly, in these cells PTN seems to
mediate bFGF-induced cell proliferation and migration [44].
1a,25-Dihydroxyvitamin D3 decreases PTN mRNA levels in the
osteoblast-like cell lines MC3T3-E1 and ROS17/2.8, whereas in the
non-osteoblastic cell line, ROS 25/1, it increases PTN mRNA levels.
This effect was found specific to 1a,25-dihydroxyvitamin D3,
suggesting that PTN may play a role in vitamin D-dependent
regulation of calcium and phosphate homeostasis and for bone
remodelling [45]. Androgens and estrogens, which have direct
beneficial effects on bone, also increase PTN expression in prostate
cells [46–48]. In cultured mouse fibroblasts, PTN expression was
found to be up-regulated by tumour necrosis factor alpha (TNF-a)
and to a smaller extent by epidermal growth factor [49]. TNF-a
seems to accelerate bone fracture healing through the coordinate
regulation of the expression of specific matrix metalloproteinases
and angiogenic factors [50]. Hypoxia [43], signalling levels of
hydrogen peroxide [51] and endothelial nitric oxide synthase [52],
all important stimulators of angiogenesis, increase PTN expression
through activation of the transcription factor activator protein-1.
Treatment of odontoblast-like M06-G3 cells with recombinant BMPs
2 and 7 that are clinically available for improvement of fracture
healing, statistically increased PTN expression, while treatment with
BMP4 had the opposite effect [53]. Finally, mechanical loading
decreases PTN expression in primary osteoblasts and osteoblastic
SaOs-2 cells [54].
PTN and angiogenesis
Soluble PTN has been suggested to promote endothelial cells
proliferation [55,56], although this has not been observed in all
types of endothelial cells [57]. Besides soluble PTN, it has been also
shown that immobilised PTN stimulates proliferation of several
types of endothelial cells not responding to the soluble growth
factor [57]. On the other hand, PTN down-regulates the stimulatory
effect of VEGF on endothelial cell proliferation through direct
binding of the two growth factors [58].
PTN has been also shown to promote migration of endothelial
cells [56,59,60,75,76], and to mediate the stimulatory effect of
hydrogen peroxide [52], endothelial nitric oxide synthase/nitric
oxide [52] and aprotinin [70] on human endothelial cell
migration, through its receptor RPTPb/z. On the other hand,
PTN decreases VEGF-induced migration of human endothelial
cells to the levels of its own migratory effect [58,62,63,77],
suggesting that it may also play a regulatory role to limit
excessive angiogenic response.
Besides the effect of PTN on endothelial cells proliferation and
migration, PTN has been shown to induce embryoid body
angiogenesis [64] and promote in vitro formation of tube-like
structures in collagen gels [56,59], fibrin gels or matrigel [59] and
to stimulate in vivo angiogenesis of the chick embryo chorioallan-
toic membrane (CAM) [59,78] and in matrigel implants in mice
[65]. In the latter assay, PTN also limits VEGF-induced angiogenic
response to the levels of its own smaller stimulatory effect [58],
further supporting its regulatory effect.
It is also possible that besides a direct effect on endothelial cells,
PTN has an indirect effect, potentiating the angiogenic effect of other
molecules. For example, PTN up-regulates the mRNA levels of VEGF
isoforms 165 and 190, as well as activates metalloproteinase 2 in the
chick embryo CAM [78] and promotes VEGF expression and
cooperates with VEGF in promoting colorectal cancer angiogenesis
[66]. PTN induces proliferation of human peripheral blood
mononuclear cells [67] and increases the mRNA expression of the
VEGF receptor 1 in endothelial cell cultures [62]. Flt-1 is expressed
by monocytes and its expression is increased after monocyte
activation [68]. In the same line, PTN induces transformation of
monocytes into functional endothelial cells, thus supporting
angiogenesis [69,70].
The role of PTN in tumour angiogenesis has been initially
suggested by the observation that culture supernatants derived
from PTN transfected human adrenal carcinoma cells, lung cancer
cells or PTN transfected MCF-7 human breast cancer cells possess
mitogenic activities for endothelial cells. PTN also increases the
angiogenic potential of multiple myeloma. Ribozyme targeting of
PTN in a human melanoma cell line decreases vessel formation in
the primary tumour, as well as metastases [2]. Antisense PTN
expression in human prostate LNCaP cells decreases prostate
cancer cell-induced angiogenesis in vitro and in vivo [71,72]. On
the other hand, antisense PTN expression in rat glioma C6 cells
decreases glioma cell-induced angiogenesis in vitro and in vivo
[73] and PTN seems to act as an angiostatic factor in an in vivo
neuroblastoma model that is resistant to irinotecan [74]. In both
cases, its regulatory effect on VEGF actions has been discussed as
the most likely mechanism.
 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823 1819

PTN and bone Osteoporosis

There are several studies that support a potential role of PTN in
the skeletal system, outlined in Table 2. Initially, PTN mRNA was
found during development in bone and cartilage progenitors and
in dental pulp [6,10]. Later, PTN was detected within the
epiphyseal plate of mouse suggesting a possible role in the
longitudinal bone growth and bone formation [11]. One biological
function that was early attributed to PTN is the promotion of
osteoblast attachment to the extracellular bone matrix through its
carboxy-terminal domain [21]. This was further verified in several
in vivo models, in which PTN was found to be prominently
expressed in the cell matrices that act as target substrates for bone
formation and to possibly play an important role in bone
formation, probably by mediating recruitment and attachment
of osteoblasts/osteoblast precursors to the appropriate substrates
for deposition of new bone [8]. Unpublished observations of
our research team in an animal model suggest that PTN is
expressed by chondrocytes in the hypertrophic and proliferating
zones of the growth plate and its expression depends on
mechanical loading.
PTN pathways in clinical skeletal diseases
The relevance of PTN implication in skeletal homeostasis and
diseases is currently under investigation. PTN is thought to affect
osteoblasts’ rather than osteoclasts’ functions; however, the
mechanisms involved are not yet fully elucidated.
The highest expression of PTN in PTN transgenic mice is
maintained in the periosteum, and the PTN transgenic mice
develop a phenotype characterised by an increased bone thickness
[8]. In transgenic mice over-expressing the human PTN gene driven
by the osteocalcin promoter, femoral bone mineral content was
increased compared with wild-type controls. In ovariectomised
mice, bone mass loss due to oestrogen deficiency was observed in
both transgenic and control mice but bone mass was higher in
transgenic mice [32]. In a later study using the same transgenic
line, the bone mass increase was found evident in female, but not
male mice, although no direct evidence supporting female-specific
mRNA synthesis of the transgene was obtained [34]. In both cases,
the mechanism seems to be through activation and recruitment of
osteoblasts. In a more detailed analysis, it was shown that PTN
over-expression enhanced intramembranous bone formation and
had multiple effects on long-term bone growth. The pubertal
growth spurt did not take place in transgenic mice, in which the
growth trajectory was steady and continuous until 25 weeks. By
30 weeks, transgenic and control mice were of the same size, but
the calcium content/mg bone was approximately 10% higher in the
transgenic group. PTN was also localised in growth plate and
articular chondrocytes, but only in transgenic mice. In the latter, an
encroachment of subchondral bone into the articular cartilage was
also observed [33].
PTN-deficient mice seem to have normal bone formation [37],
but they show growth retardation in the weight-bearing bones by
two months of age and low bone formation and osteopenia, as well

Table 2
Outline of evidence supporting a possible role for PTN in bone remodelling/repair.

Species Findings

PTN expression
Mouse PTN mRNA in calvarial osteoblast-enriched cells [6].
Human PTN mRNA in cDNA libraries from osteosarcoma cell line MG63 [6].
Rat PTN mRNA in bone and cartilage progenitors and in dental pulp during development [10].
Rat Abundant PTN expression in the secondary ossification centre, in the growth plate, and in the periosteum [8].
Rat Up-regulated PTN expression on the surface of adult damaged bone in a post arthritic ossification model [8].
Mouse Increased PTN expression in the periosteum of adult PTN transgenic mice and by the osteocytes dispersed in the
cortical bone [8].
Rat Immunolocalised PTN on both osteoblasts and endothelial cells in the well vascularised newly formed woven bone [11].
Mouse Increased PTN expression in the tibias of female animals after four days of mechanical stimulation [12].
Mouse Greater PTN expression in juvenile compared with the adult dura mater [13].
Human Increased PTN in serum of patients during fracture healing. Not observed in non union patients [14].
Human Over-expressed PTN in the synovial fluid of patients with osteoarthritis [15].
Human PTN in chondrocytes, osteocytes and bone lining cells in the joints of osteoarthritis patients [9].

In vitro cells/cell cultures

Human PTN binds on the surface of the osteoblast-like osteosarcoma cell lines HOS(TE85) and MG-63, and promotes osteoblast
attachment to the extracellular bone matrix [21].
Rat, Human PTN induces migration of osteoblast-type cells [8].
Human PTN promotes adhesion, migration, expansion and differentiation of osteoprogenitor cells [22].
Human PTN has chemotactic effects on both osteoblastic and endothelial cells [23].
Human PTN is an autocrine growth factor involved in the clustering and proliferation of chondrocytes [24].
Mouse PTN induces heparin-binding epidermal growth factor release to trans-activate epidermal growth factor receptor in primary
osteoblasts and osteoblast-like MC3T3-E1 cells, and through this pathway, it increases alkaline phosphatase activity and
inhibits dexamethazone-induced cell death in both cell lines [25].
Human PTN induces hypertrophy during chondrogenic differentiation of bone marrow stem cells [26].

Animal models
PTN transgenic mouse Increased thickness of the cortical bone, increased cortical and cancellous bone volume, and increased osteoblast surface.
No alteration in the bone marrow area [8].
Increased femoral bone mineral content compared with wild-type controls.
In ovariectomised mice, bone mass loss was lower in transgenic mice [32].
Enhanced intramembranous bone formation and multiple effects on long-term bone growth [33].
Bone mass increase in female but not male mice [34].
Bone healing process is impaired in the adult PTN mice [35].

PTN knockout mouse Growth retardation in the weight-bearing bones by two months of age and low bone formation and osteopenia, as well as
resistance to immobilization-dependent bone remodelling during adulthood [36].
The number, morphology, and function of osteoclasts, osteoblasts, and osteocytes are not altered in PTN-deficient mice
compared with wild-type littermates [37].
1820 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
as resistance to immobilization-dependent bone remodelling
during adulthood [36].
A microarray based identification of osteoporosis-related genes
in primary culture of human osteoblasts suggested that PTN is
among the candidate genes for further studies in the assessment of
the genetic susceptibility to osteoporosis [27]. In a recent clinical
study of 530 postmenopausal women, with and without hip
fractures, it was shown that the PTN gene promoter polymorphism
1227C > T and CT haplotype could contribute to the genetic
background of osteoporosis, affecting the bone mass density values
in the lumbar spine and in the femoral neck [41].
Interestingly, the microgravity-induced bone loss is found to be
due to an increased bone resorption and a decreased bone
deposition. PTN over-expressing mice exposed to normal gravity
present a poorer trabecular organization than wild type mice but the
expression of PTN during the flight offers some protection against
microgravity’s negative effects, as a result of a higher osteoblast
activity in the flight mice [38]. A subsequent study showed that the
reduction in the expression of collagen type I and osteocalcin in PTN
transgenic mice was less than in the samples from wild type mice,
possibly due to the higher level calcitonin expression that could
participate to the protective effect of PTN over-expression on the
bone damage [39].
Fracture healing
Fracture healing is a complex physiological process in which
PTN may play an important role. During fracture healing, PTN is
immunolocalised on both osteoblasts and endothelial cells in the
well vascularised, newly formed woven bone [40] and recombi-
nant human PTN has chemotactic effects on both human
osteoblastic and endothelial cells [23]. A role for PTN in bone
regeneration is also supported by data showing that PTN induces
hypertrophy during chondrogenic differentiation of human bone
marrow stem cells, as evidenced by increased expression of
collagen 2 protein and increased transcription of hypertrophic
chondrocyte markers, such as MMP13, collagen 10 and alkaline
phosphatase and enhanced calcification and content of collagen
10 protein [26]. Additionally, our unpublished data demonstrate
that PTN is expressed in chondrocytes during callus formation
(Fig. 1). PTN expression is greater in juvenile compared with the
adult dura mater and may be related with the reossification of
large calvarial defects that is possible in children but not adults
Fig. 1. Expression of PTN by chondrocytes during healing process, five weeks after a
rat’s vertebrae fracture. PTN shows both cytoplasmic (red arrows) and nuclear
(black arrows) localization (brown staining), with the latter being dominant.
Although there is no known physiological significance for the nuclear localization of
PTN, it has been also observed in endothelial and glioma cells [78], cardiomyocytes
[91] and osteoarthritic chondrocytes [9]. (For interpretation of the references to
color in this text, the reader is referred to the web version of the article.)
[13]. In the same line, PTN serum levels increased asymptotically in
timely fracture healing, while no such increase was detected
during delayed healing [14]. On the other hand, it has been also
shown that the bone healing process was impaired in adult mice
over-expressing PTN only in bone and this has been attributed to
inhibitory effects of PTN over-expression on BMP-2 mediated bone
induction [35].
Osteoarthritis
Studies on the possible involvement of PTN in the pathogenesis of
osteoarthritis are limited. PTN is over-expressed in the synovial fluid
[15], the serum, as well as in chondrocytes and subchondral bone
osteocytes of patients with osteoarthritis [9]. Specifically, PTN
expression was increased in OA patients graded Ahlback II and III,
localised in superficial chondrocyte clusters and chondrocytes in the
upper radial and tidemark zones. PTN was also immunolocalised in
the osteocytes, bone lining cells and the extracellular matrix of the
subchondral bone, as well as in the osteocytes of deeper trabeculae
[9]. These data make PTN a promising candidate for further studies
as far as developing therapeutic schemes for osteoarthritis.
Discussion
Bone is characterised by an increased potential for growth,
regeneration and remodelling throughout life. Bone repair is a
complex process of biological events regulated by specific cells, the
extracellular matrix, distinct growth factors and a variety of
hormones [92–100]. Despite the increased intrinsic capacity of
bone to restore fracture healing without scar formation, non-union
may still occur in 5–10% of the cases [101]. This impaired fracture
healing phenomenon has been attributed to a number of factors,
including poor mechanical stability, extensive soft tissue and
periosteal disruption, patient related co-morbidities, non-steroidal
anti-inflammatory drugs, vitamin D deficiency and malnutrition
amongst others [102,103]. Current research to understand better
and to treat successfully fracture non-union is focusing on
biological based approaches and involves the implantation of
growth factors, osteoprogenitor cells, biophysical stimuli, and the
promotion of local angiogenesis [104].
Angiogenesis is a critical step of bone healing. During the bone
repair process in humans, several angiogenic factors are expressed
and play key regulatory roles during the early phase of fracture
healing. PTN is a potent angiogenic factor that may also affect
several aspects of bone remodelling. Its importance in fracture
healing has been supported by the study of Weiss et al. (2009) in
patients with and without fracture union [14]. Compared to the
relatively low serum concentrations in non-injured adults,
systemic PTN values show a prolonged increase during physiolog-
ical fracture healing and remodelling, while no such increase could
be detected in non-union patients. Therefore, PTN merits further
investigation as a possible powerful bone-inducing component to
promote healing in orthopaedic and spinal surgery by affecting
bone cells themselves or indirectly, by regulating angiogenesis.
In all cases, since PTN may have multiple effects in other tissues,
as well, the perspective of exclusively targeting the bone tissue is
critical, so as to prevent potentially side effects at non skeletal sites.
PTN could be locally administered in implanted carriers such as
scaffolds, membranes for tissue-guided regeneration or hydrogels,
providing different schemes for release, in cases such as long bone
open fractures treated with intramedullary fixation, non-united
fractures, and in spine surgery for spinal fusion. Concerning the
required dose, clinical trials are desirable in order to monitor
carefully the appropriate local PTN concentration leading to a
sufficient bone formation. Alternatively, apart from direct admin-
istration of PTN, indirect ways of endogenous PTN regulation could
be another possible target.
 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
An important issue that should be taken into consideration is the
fact that it is becoming increasingly apparent that growth factors act
in a coordinated cascade of events to restore bone; therefore,
delivering combinations of growth factors may be of greater
significance in the clinic. Concerning PTN, its use in combination
with osteogenic growth factors, such as BMPs, may prove beneficial.
It has been shown that fracture healing was impaired in the adult
mouse with targeted PTN over-expression in bone, and this was
attributed to the inhibitory effects of PTN over-expression on BMP-2
mediated bone induction [35]. Taking into account that BMP-2 up-
regulates PTN expression [53], these data favour the notion of a
regulatory role of PTN in the pathways of primary signalling
molecules, such as BMPs, leading to the hypothesis that it may
regulate complications of BMPs, such as heterotopic ossifications or
hardware failure secondary to non-union. A similar regulatory role
of PTN has been shown for the key angiogenic growth factor VEGF
[2,3,58,63], further supporting a potential regulatory role of PTN for
both angiogenesis and bone formation and/or repair. A future
application of PTN in combination with other growth factors, e.g.
BMPs, may thus contribute to the clinically desired bone formation,
which has not been fully attained up to date with the application of a
single growth factor. Although the issues to be unravelled are
multiple in such a case and are related to the appropriate and safe
concentration of each growth factor, rate of release and timing of
application, this approach may better mimic the physiological
situation and deserves further consideration and appropriate
studies.
In conclusion, the heparin-binding angiogenic growth factor
PTN appears to play a well-defined and significant role in both
physiological and pathological angiogenesis. Emerging evidence
supports the view that it plays a crucial role in bone repair. Based
on its molecular properties, future studies must investigate the
potential of its application in clinical situations were bone
restoration is desirable.
Conflict of interest statement
The authors report no conflict of interest.
References
[1] Pountos I, Panteli M, Panagiotopoulos E, Jones E, Giannoudis PV. Can we enhance
fracture vascularity: what is the evidence? Injury 2014;45(Suppl. 2):S49–57.
[2] Papadimitriou E, Mikelis C, Lampropoulou E, Koutsioumpa M, Theochari K,
Tsirmoula S, et al. Roles of pleiotrophin in tumor growth and angiogenesis.
Eur Cytokine Netw 2009;20:180–90.
[3] Pantazaka E, Papadimitriou E. PTN (pleiotrophin). Atlas Genet Cytogenet
Oncol Haematol 2012;16:821–37.
[4] Kilpelainen I, Kaksonen M, Kinnunen T, Avikainen H, Fath M, Linhardt RJ, et al.
Heparin-binding growth-associated molecule contains two heparin-binding
beta-sheet domains that are homologous to the thrombospondin type I
repeat. J Biol Chem 2000;275:13564–70.
[5] Englund C, Birve A, Falileeva L, Grabbe C, Palmer RH. Miple1 and miple2
encode a family of MK/PTN homologues in Drosophila melanogaster. Dev
Genes Evol 2006;216:10–8.
[6] Tezuka K, Takeshita S, Hakeda Y, Kumegawa M, Kikuno R, Hashimoto-Gotoh
T. Isolation of mouse and human cDNA clones encoding a protein expressed
specifically in osteoblasts and brain tissues. Biochem Biophys Res Commun
1990;173:246–51.
[7] Hankenson KD, Sweetwyne MT, Shitaye H, Posey KL. Thrombospondins and
novel TSR-containing proteins, R-spondins, regulate bone formation and
remodeling. Curr Osteoporos Rep 2010;8:68–76.
[8] Imai S, Kaksonen M, Raulo E, Kinnunen T, Fages C, Meng X, et al. Osteoblast
recruitment and bone formation enhanced by cell matrix-associated heparin-
binding growth-associated molecule (HB-GAM). J Cell Biol 1998;143:1113–28.
[9] Kaspiris A, Mikelis C, Heroult M, Khaldi L, Grivas TB, Kouvaras I, et al.
Expression of the growth factor pleiotrophin and its receptor protein tyrosine
phosphatase beta/zeta in the serum, cartilage and subchondral bone of
patients with osteoarthritis. Joint Bone Spine 2013;80:407–13.
[10] Vanderwinden JM, Mailleux P, Schiffmann SN, Vanderhaeghen JJ. Cellular
distribution of the new growth factor pleiotrophin (HB-GAM) mRNA in
developing and adult rat tissues. Anat Embryol (Berl) 1992;186:387–406.
[11] Petersen W, Rafii M. Immunolocalization of the angiogenetic factor pleiotrophin
(PTN) in the growth plate of mice. Arch Orthop Trauma Surg 2001;121:414–6.
1821
[12] Xing W, Baylink D, Kesavan C, Hu Y, Kapoor S, Chadwick RB, et al. Global gene
expression analysis in the bones reveals involvement of several novel genes
and pathways in mediating an anabolic response of mechanical loading in
mice. J Cell Biochem 2005;96:1049–60.
[13] Wan DC, Aalami OO, Wang Z, Nacamuli RP, Lorget F, Derynck R, et al.
Differential gene expression between juvenile and adult dura mater: a
window into what genes play a role in the regeneration of membranous
bone. Plast Reconstr Surg 2006;118:851–61.
[14] Weiss S, Zimmermann G, Pufe T, Varoga D, Henle P. The systemic angiogenic
response during bone healing. Arch Orthop Trauma Surg 2009;129:989–97.
[15] Pufe T, Bartscher M, Petersen W, Tillmann B, Mentlein R. Pleiotrophin an
embryonic differentiation and growth factor, is expressed in osteoarthritis.
Osteoarthritis Cartilage 2003;11:260–4.
[16] Bertrand J, Stange R, Hidding H, Echtermeyer F, Nalesso G, Godmann L, et al.
Syndecan 4 supports bone fracture repair, but not fetal skeletal development,
in mice. Arthritis Rheum 2013;65:743–52.
[17] Schinke T, Gebauer M, Schilling AF, Lamprianou S, Priemel M, Mueldner C,
et al. The protein tyrosine phosphatase Rptpzeta is expressed in differen-
tiated osteoblasts and affects bone formation in mice. Bone 2008;42:
524–34.
[18] Toledo SR, Oliveira ID, Okamoto OK, Zago MA, de Seixas Alves MT, Filho RJ,
et al. Bone deposition, bone resorption, and osteosarcoma. J Orthop Res
2010;28:1142–8.
[19] Zou W, Teitelbaum SL. Integrins, growth factors, and the osteoclast cytoskel-
eton. Ann N Y Acad Sci 2010;1192:27–31.
[20] Attia M, Scott A, Duchesnay A, Carpentier G, Soslowsky LJ, Huynh MB, et al.
Alterations of overused supraspinatus tendon: a possible role of glycosami-
noglycans and HARP/pleiotrophin in early tendon pathology. J Orthop Res
2012;30:61–71.
[21] Gieffers C, Engelhardt W, Brenzel G, Matsuishi T, Frey J. Receptor binding of
osteoblast-specific factor 1 (OSF-1/HB-GAM) to human osteosarcoma cells
promotes cell attachment. Eur J Cell Biol 1993;62:352–61.
[22] Yang X, Tare RS, Partridge KA, Roach HI, Clarke NM, Howdle SM, et al.
Induction of human osteoprogenitor chemotaxis, proliferation, differentia-
tion, and bone formation by osteoblast stimulating factor-1/pleiotrophin:
osteoconductive biomimetic scaffolds for tissue engineering. J Bone Miner
Res 2003;18:47–57.
[23] Li G, Cui Y, McIlmurray L, Allen WE, Wang H. rhBMP-2, rhVEGF(165), rhPTN
and thrombin-related peptide, TP508 induce chemotaxis of human osteo-
blasts and microvascular endothelial cells. J Orthop Res 2005;23:680–5.
[24] Pufe T, Groth G, Goldring MB, Tillmann B, Mentlein R. Effects of pleiotrophin,
a heparin-binding growth factor, on human primary and immortalized
chondrocytes. Osteoarthritis Cartilage 2007;15:155–62.
[25] Fan JB, Liu W, Yuan K, Zhu XH, Xu DW, Chen JJ, et al. EGFR trans-activation
mediates pleiotrophin-induced activation of Akt and Erk in cultured osteo-
blasts. Biochem Biophys Res Commun 2014;447:425–30.
[26] Bouderlique T, Henault E, Lebouvier A, Frescaline G, Bierling P, Rouard H, et al.
Pleiotrophin commits human bone marrow mesenchymal stromal cells
towards hypertrophy during chondrogenesis. PLOS ONE 2014;9:e88287.
[27] Trost Z, Trebse R, Prezelj J, Komadina R, Logar DB, Marc J. A microarray based
identification of osteoporosis-related genes in primary culture of human
osteoblasts. Bone 2010;46:72–80.
[28] Rangaswami H, Schwappacher R, Marathe N, Zhuang S, Casteel DE, Haas B,
et al. Cyclic GMP and protein kinase G control a Src-containing mechanosome
in osteoblasts. Sci Signal 2010;3:ra91.
[29] Schneider JG, Amend SR, Weilbaecher KN. Integrins and bone metastasis:
integrating tumor cell and stromal cell interactions. Bone 2011;48:54–65.
[30] Nakamura I, Duong le T, Rodan SB, Rodan GA. Involvement of alpha(v)beta3
integrins in osteoclast function. J Bone Miner Metab 2007;25:337–44.
[31] Hempel U, Preissler C, Vogel S, Möller S, Hintze V, Becher J, et al. Artificial
extracellular matrices with oversulfated glycosaminoglycan derivatives pro-
mote the differentiation of osteoblast-precursor cells and premature osteo-
blasts. Biomed Res Int 2014;2014:938368.
[32] Masuda H, Tsujimura A, Yoshioka M, Arai Y, Kuboki Y, Mukai T, et al. Bone
mass loss due to estrogen deficiency is compensated in transgenic mice
overexpressing human osteoblast stimulating factor-1. Biochem Biophys Res
Commun 1997;238:528–33.
[33] Tare RS, Oreffo RO, Sato K, Rauvala H, Clarke NM, Roach HI. Effects of targeted
overexpression of pleiotrophin on postnatal bone development. Biochem
Biophys Res Commun 2002;298:324–32.
[34] Hashimoto-Gotoh T, Ohnishi H, Tsujimura A, Tsunezuka H, Imai K, Masuda H,
et al. Bone mass increase specific to the female in a line of transgenic mice
overexpressing human osteoblast stimulating factor-1. J Bone Miner Metab
2004;22:278–82.
[35] Li G, Bunn JR, Mushipe MT, He Q, Chen X. Effects of pleiotrophin (PTN) over-
expression on mouse long bone development, fracture healing and bone
repair. Calcif Tissue Int 2005;76:299–306.
[36] Imai S, Heino TJ, Hienola A, Kurata K, Buki K, Matsusue Y, et al. Osteocyte-
derived HB-GAM (pleiotrophin) is associated with bone formation and
mechanical loading. Bone 2009;44:785–94.
[37] Lehmann W, Schinke T, Schilling AF, Catala-Lehnen P, Gebauer M, Pogoda P,
et al. Absence of mouse pleiotrophin does not affect bone formation in vivo.
Bone 2004;35:1247–55.
[38] Tavella S, Ruggiu A, Giuliani A, Brun F, Canciani B, Manescu A, et al. Bone
turnover in wild type and pleiotrophin-transgenic mice housed for three
months in the International Space Station (ISS). PLoS ONE 2012;7:e33179.
1822 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
[39] Albi E, Curcio F, Spelat R, Lazzarini A, Lazzarini R, Cataldi S, et al. Loss of
parafollicular cells during gravitational changes (microgravity, hypergravity)
and the secret effect of pleiotrophin. PLoS ONE 2012;7:e48518.
[40] Petersen W, Wildemann B, Pufe T, Raschke M, Schmidmaier G. The angiogenic
peptide pleiotrophin (PTN/HB-GAM) is expressed in fracture healing: an
immunohistochemical study in rats. Arch Orthop Trauma Surg 2004;124:
603–7.
[41] Mencej-Bedrac S, Prezelj J, Komadina R, Vindisar F, Marc J. 1227C > T
polymorphism in the pleiotrophin gene promoter influences bone mineral
density in postmenopausal women. Mol Genet Metab 2011;103:76–80.
[42] Li YS, Gurrieri M, Deuel TF. Pleiotrophin gene expression is highly restricted
and is regulated by platelet-derived growth factor. Biochem Biophys Res
Commun 1992;184:427–33.
[43] Antoine M, Tag CG, Wirz W, Borkham-Kamphorst E, Sawitza I, Gressner AM,
et al. Upregulation of pleiotrophin expression in rat hepatic stellate cells by
PDGF and hypoxia: implications for its role in experimental biliary liver
fibrogenesis. Biochem Biophys Res Commun 2005;337:1153–64.
[44] Hatziapostolou M, Polytarchou C, Katsoris P, Courty J, Papadimitriou E.
Heparin affin regulatory peptide/pleiotrophin mediates fibroblast growth
factor 2 stimulatory effects on human prostate cancer cells. J Biol Chem
2006;281:32217–26.
[45] Tamura M, Ichikawa F, Guillerman RP, Deuel TF. Nodal M.1a,25-dihydrox-
yvitamin D(3) down-regulates pleiotrophin messenger RNA expression in
osteoblast-like cells. Endocrine 1995;3:21–4.
[46] Vacherot F, Laaroubi K, Caruelle D, Delbe J, Barritault D, Caruelle JP, et al.
Upregulation of heparin-affin regulatory peptide by androgen. In Vitro Cell
Dev Biol Anim 1995;31:647–8.
[47] Orr B, Vanpoucke G, Grace OC, Smith L, Anderson RA, Riddick AC, et al.
Expression of pleiotrophin in the prostate is androgen regulated and it
functions as an autocrine regulator of mesenchyme and cancer associated
fibroblasts and as a paracrine regulator of epithelia. Prostate 2011;71:
305–317.
[48] Lee JY, Jeong W, Lim W, Kim J, Bazer FW, Han JY, et al. Chicken pleiotrophin:
regulation of tissue specific expression by estrogen in the oviduct and distinct
expression pattern in the ovarian carcinomas. PLoS ONE 2012;7:e34215.
[49] Pufe T, Bartscher M, Petersen W, Tillmann B, Mentlein R. Expression of
pleiotrophin, an embryonic growth and differentiation factor, in rheumatoid
arthritis. Arthritis Rheum 2003;48:660–7.
[50] Lehmann W, Edgar CM, Wang K, Cho TJ, Barnes GL, Kakar S, et al. Tumor
necrosis factor alpha (TNF-alpha) coordinately regulates the expression of
specific matrix metalloproteinases (MMPS) and angiogenic factors during
fracture healing. Bone 2005;36:300–10.
[51] Polytarchou C, Hatziapostolou M, Papadimitriou E. Hydrogen peroxide sti-
mulates proliferation and migration of human prostate cancer cells through
activation of activator protein-1 and up-regulation of the heparin affin
regulatory peptide gene. J Biol Chem 2005;280:40428–35.
[52] Polytarchou C, Hatziapostolou M, Poimenidi E, Mikelis C, Papadopoulou A,
Parthymou A, et al. Nitric oxide stimulates migration of human endothelial
and prostate cancer cells through up-regulation of pleiotrophin expression
and its receptor protein tyrosine phosphatase b/z. Int J Cancer
2009;124:1785–93.
[53] Erlandsen H, Ames JE, Tamkenath A, Mamaeva O, Stidham K, Wilson ME, et al.
Pleiotrophin expression during odontogenesis. J Histochem Cytochem
2012;60:366–75.
[54] Liedert A, Augat P, Ignatius A, Hausser HJ, Claes L. Mechanical regulation
of HB-GAM expression in bone cells. Biochem Biophys Res Commun
2004;319:951–8.
[55] Courty J, Dauchel MC, Caruelle D, Perderiset M, Barritault D. Mitogenic
properties of a new endothelial cell growth factor related to pleiotrophin.
Biochem Biophys Res Commun 1991;180:145–51.
[56] Souttou B, Raulais D, Vigny M. Pleiotrophin induces angiogenesis: involve-
ment of the phosphoinositide-3 kinase but not the nitric oxide synthase
pathways. J Cell Physiol 2001;187:59–64.
[57] Papadimitriou E, Heroult M, Courty J, Polykratis A, Stergiou C, Katsoris P.
Endothelial cell proliferation induced by HARP: implication of N or C terminal
peptides. Biochem Biophys Res Commun 2000;274:242–8.
[58] Héroult M, Bernard-Pierrot I, Delbé J, Hamma-Kourbali Y, Katsoris P, Barri-
tault D, et al. Heparin affin regulatory peptide binds to vascular endothelial
growth factor (VEGF) and inhibits VEGF-induced angiogenesis. Oncogene
2004;23:1745–53.
[59] Papadimitriou E, Polykratis A, Courty J, Koolwijk P, Heroult M, Katsoris P.
HARP induces angiogenesis in vivo and in vitro: implication of N or C terminal
peptides. Biochem Biophys Res Commun 2001;282:306–13.
[60] Polykratis A, Katsoris P, Courty J, Papadimitriou E. Characterization of heparin
affin regulatory peptide signaling in human endothelial cells. J Biol Chem
2005;280:22454–61.
[61] Koutsioumpa M, Hatziapostolou M, Mikelis C, Koolwijk P, Papadimitriou E.
Aprotinin stimulates angiogenesis and human endothelial cell migration
through the growth factor pleiotrophin and its receptor protein tyrosine
phosphatase beta/zeta. Eur J Pharmacol 2009;602:245–9.
[62] Kokolakis G, Mikelis C, Papadimitriou E, Courty J, Karetsou E, Katsoris P.
Effect of heparin affin regulatory peptide on the expression of vascular
endothelial growth factor receptors in endothelial cells. In Vivo
2006;20:629–35.
[63] Koutsioumpa M, Poimenidi E, Theodoropoulou C, Pantazaka E, Skoura A,
Megalooikonomou V, et al. A role of receptor protein tyrosine phosphatase
beta/zeta in vascular endothelial growth factor-induced endothelial cell
migration (Abstract). Angiogenesis 2014;17:754.
[64] Magnusson PU, Dimberg A, Mellberg S, Lukinius A, Claesson-Welsh L. FGFR-1
regulates angiogenesis through cytokines interleukin-4 and pleiotrophin.
Blood 2007;110:4214–22.
[65] Bernard-Pierrot I, Delbe J, Caruelle D, Barritault D, Courty J, Milhiet PE. The
lysine-rich C-terminal tail of heparin affin regulatory peptide is required for
mitogenic and tumor formation activities. J Biol Chem 2001;276:12228–34.
[66] Kong Y, Bai PS, Nan KJ, Sun H, Chen NZ, Qi XG. Pleiotrophin is a potential
colorectal cancer prognostic factor that promotes VEGF expression and
induces angiogenesis in colorectal cancer. Int J Colorectal Dis 2012;27:
287–98.
[67] Achour A, Laaroubi D, Caruelle D, Barritault D, Courty J. The angiogenic factor
heparin affin regulatory peptide (HARP) induces proliferation of human
peripheral blood mononuclear cells. Cell Mol Biol (Noisy-le-grand)
2001;47 Online Pub:OL73–7.
[68] Shibuya M. Structure and dual function of vascular endothelial growth factor
receptor-1 (Flt-1). Int J Biochem Cell Biol 2001;33:409–20.
[69] Sharifi BG, Zeng Z, Wang L, Song L, Chen H, Qin M, et al. Pleiotrophin induces
transdifferentiation of monocytes into functional endothelial cells. Arterios-
cler Thromb Vasc Biol 2006;26:1273–80.
[70] Chen H, Campbell RA, Chang Y, Li M, Wang CS, Li J, et al. Pleiotrophin
produced by multiple myeloma induces transdifferentiation of monocytes
into vascular endothelial cells: a novel mechanism of tumor-induced vascu-
logenesis. Blood 2009;113:1992–2002.
[71] Hatziapostolou M, Delbe J, Katsoris P, Polytarchou C, Courty J, Papadimitriou
E. Heparin affin regulatory peptide is a key player in prostate cancer cell
growth and angiogenicity. Prostate 2005;65:151–8.
[72] Tsirmoula S, Dimas K, Hatziapostolou M, Lamprou M, Ravazoula P, Papadi-
mitriou E. Implications of pleiotrophin in human PC3 prostate cancer cell
growth in vivo. Cancer Sci 2012;103:1826–32.
[73] Parthymou A, Lampropoulou E, Mikelis C, Drosou G, Papadimitriou E. Heparin
affin regulatory peptide/pleiotrophin negatively affects diverse biological
activities in C6 glioma cells. Eur J Cell Biol 2008;87:17–29.
[74] Calvet L, Geoerger B, Regairaz M, Opolon P, Machet L, Morizet J, et al.
Pleiotrophin a candidate gene for poor tumor vasculature and in vivo
neuroblastoma sensitivity to irinotecan. Oncogene 2006;25:3150–9.
[75] Mikelis C, Sfaelou E, Koutsioumpa M, Kieffer N, Papadimitriou E. Integrin
alpha(v)beta(3) is a pleiotrophin receptor required for pleiotrophin-induced
endothelial cell migration through receptor protein tyrosine phosphatase
beta/zeta. FASEB J 2009;23:1459–69.
[76] Koutsioumpa M, Polytarchou C, Courty J, Zhang Y, Kieffer N, Mikelis C, et al.
Interplay between alphavbeta3 integrin and nucleolin regulates human
endothelial and glioma cell migration. J Biol Chem 2013;288:343–54.
[77] Polykratis A, Delbé J, Courty J, Papadimitriou E, Katsoris P. Identification of
heparin affin regulatory peptide domains with potential role on angiogenesis.
Int J Biochem Cell Biol 2004;36:1954–66.
[78] Koutsioumpa M, Drosou G, Mikelis C, Theochari K, Vourtsis D, Katsoris P, et al.
Pleiotrophin expression and role in physiological angiogenesis in vivo:
potential involvement of nucleolin. Vasc Cell 2012;4:4.
[79] Kinnunen T, Raulo E, Nolo R, Maccarana M, Lindahl U, Rauvala H. Neurite
outgrowth in brain neurons induced by heparin-binding growth-associated
molecule (HB-GAM) depends on the specific interaction of HB-GAM with
heparan sulfate at the cell surface. J Biol Chem 1996;271:2243–8.
[80] Raulo E, Chernousov MA, Carey DJ, Nolo R, Rauvala H. Isolation of a neuronal
cell surface receptor of heparin binding growth-associated molecule (HB-
GAM), identification as N-syndecan (syndecan-3). J Biol Chem 1994;269:
12999–3004.
[81] Deepa SS, Yamada S, Zako M, Goldberger O, Sugahara K. Chondroitin sulfate
chains on syndecan-1 and syndecan-4 from normal murine mammary gland
epithelial cells are structurally and functionally distinct and cooperate with
heparin sulfate chains to bind growth factors. A novel function to control
binding of midkine, pleiotrophin, and basic fibroblast growth factor. J Biol
Chem 2004;279:37368–76.
[82] Pantazaka E, Papadimitriou E. Chondroitin sulfate-cell membrane effectors as
regulators of growth factor-mediated vascular and cancer cell migration.
Biochim Biophys Acta 2014;1840:2643–50.
[83] Koutsioumpa M, Papadimitriou E. PG receptors with phosphatase action in
cancer and angiogenesis. In: Karamanos N, editor. Extracellular matrix:
pathobiology and signalling. Berlin/Boston: Walter de Gruyter GmbH &
Co. KG; 2012. p. 813–23.
[84] Bernard-Pierrot I, Héroult M, Lemaı̂tre G, Barritault D, Courty J, Milhiet PE.
Glycosaminoglycans promote HARP/PTN dimerization. Biochem Biophys Res
Commun 1999;266:437–42.
[85] Vacherot F, Delbé J, Heroult M, Barritault D, Fernig DG, Courty J. Glycosami-
noglycans differentially bind HARP and modulate its biological activity. J Biol
Chem 1999;274:7741–7.
[86] Mizumoto S, Fongmoon D, Sugahara K. Interaction of chondroitin sulfate and
dermatan sulfate from various biological sources with heparin-binding
growth factors and cytokines. Glycoconj J 2013;30:619–32.
[87] Take M, Tsutsui J, Obama M, Ozawa M, Nakayama T, Maruyama I, et al. Identifi-
cation of nucleolin as a binding protein for midkine (MK) and heparin-binding
growth associated molecule (HB-GAM). J Biochem 1994;116:1063–8.
[88] Said EA, Courty J, Svab J, Delbe J, Krust B, Hovanessian AG. Pleiotrophin
inhibits HIV infection by binding the cell surface-expressed nucleolin. FEBS J
2005;272:4646–59.
 M. Lamprou et al. / Injury, Int. J. Care Injured 45 (2014) 1816–1823
[89] Destouches D, El Khoury D, Hamma-Kourbali Y, Krust B, Albanese P,
Katsoris P, et al. Suppression of tumor growth and angiogenesis by a
specific antagonist of the cell-surface expressed nucleolin. PLoS ONE
2008;3:e2518.
[90] Deuel TF. Anaplastic lymphoma kinase: ligand independent activation me-
diated by the PTN/RPTPb/z signaling pathway. Biochim Biophys Acta
2013;1834:2219–23.
[91] Li J, Wei H, Chesley A, Moon C, Krawczyk M, Volkova M, et al. The pro-
angiogenic cytokine pleiotrophin potentiates cardiomyocyte apoptosis
through inhibition of endogenous AKT/PKB activity. J Biol Chem
2007;282:34984–93.
[92] Buschmann J, Härter L, Gao S, Hemmi S, Welti M, Hild N, et al. Tissue
engineered bone grafts based on biomimetic nanocomposite PLGA/amor-
phous calcium phosphate scaffold and human adipose-derived stem cells.
Injury 2012;43:1689–97.
[93] Calori GM, Colombo M, Mazza E, Ripamonti C, Mazzola S, Marelli N, et al.
Monotherapy vs. polytherapy in the treatment of forearm non-unions and
bone defects. Injury 2013;44(Suppl. 1):S63–9.
[94] Blokhuis TJ, Calori GM, Schmidmaier G. Autograft versus BMPs for the
treatment of non-unions: what is the evidence? Injury 2013;44(Suppl.
1):S40–2.
[95] Giannoudis PV, Calori GM, Begue T, Schmidmaier G. Bone regeneration strate-
gies: current trends but what the future holds? Injury 2013;44(Suppl. 1):
S1–2.
1823
[96] Papanna MC, Al-Hadithy N, Somanchi BV, Sewell MD, Robinson PM, Khan SA,
et al. The use of bone morphogenic protein-7 (OP-1) in the management of
resistant non-unions in the upper and lower limb. Injury 2012;43:1135–40.
[97] Reyes R, De la Riva B, Delgado A, Hernández A, Sánchez E, Évora C. Effect of
triple growth factor controlled delivery by a brushite-PLGA system on a bone
defect. Injury 2012;43:334–42.
[98] Jungbluth P, Hakimi AR, Grassmann JP, Schneppendahl J, Betsch M, Kröpil P,
et al. The early phase influence of bone marrow concentrate on metaphyseal
bone healing. Injury 2013;44:1285–94.
[99] Neunaber C, Yesilkaya P, Pütz C, Krettek C, Hildebrand F. Differentiation
of osteoprogenitor cells is affected by trauma-haemorrhage. Injury
2013;44:1279–84.
[100] Giannoudis PV, Ahmad MA, Mineo GV, Tosounidis TI, Calori GM, Kanakaris
NK. Subtrochanteric fracture non-unions with implant failure managed with
the diamond concept. Injury 2013;44(Suppl. 1):S76–81.
[101] Tzioupis C, Giannoudis PV. Prevalence of long-bone non-unions. Injury
2007;38(Suppl. 2):S3–9.
[102] Copuroglu C, Calori GM, Giannoudis PV. Fracture non-union: who is at risk?
Injury 2013;44:1379–82.
[103] Pountos I, Georgouli T, Blokhuis TJ, Pape HC, Giannoudis PV. Pharmacological
agents and impairment of fracture healing: what is the evidence? Injury
2008;39:384–94.
[104] Pountos I, Georgouli T, Pneumaticos S, Giannoudis PV. Fracture non-union:
can biomarkers predict outcome? Injury 2013;44:1725–32.