Immunology Letters 168 (2015) 129–135

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Immunology Letters
journal homepage: www.elsevier.com/locate/immlet

BMSCs and hematopoiesis

Andrés García-García a,b , Carlos L.F. de Castillejo a,b , Simón Méndez-Ferrer a,b,∗
a
Stem Cell Niche Pathophysiology Group, Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), 28029 Madrid, Spain
b
Stem Cell Institute and Department of Haematology, University of Cambridge, and National Health Service Blood and Transplant, Cambridge Biomedical
Campus, CB20PT Cambridge, United Kingdom

a r t i c l e i n f o a b s t r a c t

Article history: Recent discoveries have significantly expanded our previous knowledge about the role of bone marrow
Received 23 June 2015 mesenchymal stem cells (BMSCs) in hematopoiesis. BMSCs and their derivatives modulate blood pro-
Accepted 23 June 2015 duction and immunity at different levels but a prominent role has emerged for BMSCs in the regulation
Available online 17 July 2015
of hematopoietic stem and progenitor cells (HSPCs). Additionally, BMSC-like cells regulate B and T cell
lymphopoiesis and also probably myelopoiesis. Furthermore, BMSCs might also exhibit key regulatory
properties in non-physiological conditions. BMSCs in extramedullary sites might provide a permissive
microenviroment to allow for transient hematopoiesis. BMSCs might be also involved in the manifesta-
tion and/or the development of hematopoietic diseases, as stemming from their emerging roles in the
progression of hematological malignancies. Here we review some key molecular pathways, adhesion
molecules and ligand/receptor interactions that mediate the crosstalk between BMSCs and hematopoi-
etic stem cells (HSCs) in health and disease. The development of novel markers to visualize and isolate
individual cells will help to dissect the stromal–hematopoietic interplay.
Crown Copyright © 2015 Published by Elsevier B.V. All rights reserved.

1. BMSCs & HSPCs
BMSCs and HSCs were described as the initiating cells responsi-
ble for bone marrow reconstitution after mechanical depletion of
marrow cavity (see Ref. [1] for a review). BMSCs lie at the top of the
hierarchy of bone marrow stromal elements giving rise to cartilage,
bone and adipose tissue. Since their discovery in the bone marrow
by Friedenstein et al. [2], it has been proposed that BMSCs support
hematopoiesis through direct and indirect mechanisms. Dexter
et al. devised murine long-term cultures in which bone marrow
mesenchymal stromal cells could maintain hematopoiesis in vitro
[3]. By preserving HSC properties in the bone marrow, BMSCs are
therefore key components of the so-called “stem cell niche” [4].
When the first evidences of the existence of a stem cell
niche in the bone marrow emerged, several studies suggested
that osteoblastic cells could have an active role in the regula-
tion of hematopoiesis given the proximity between HSPCs and
the inner bone layer (“endosteum”) of the bone marrow [5–7]. In
2003 a turning point took place as Calvi et al. provided for the
first time in vivo data suggesting that osteoblastic lineage cells
are regulatory components of the HSC niche via Notch pathway
∗ Corresponding author at: NHS Blood and Transplant. Cambridge Biomedical
Campus. Cambridge, CB2 0PT, UK.
E-mail address: sm2116@cam.ac.uk (S. Méndez-Ferrer).
signaling [8]. Simultaneously, Zhang et al. also identified bone-
lining osteoblasts as key regulators of bone marrow niche size.
In this case, the molecular pathway involved was BMP signaling
[9]. Angiopoietin-1 produced by endosteal cells was also found
to promote HSC quiescence by binding to Tie2 receptor in HSCs
[10]. Angiopoietin-1 has been recently found to be mainly pro-
duced by leptin receptor+ mesenchymal stromal cells and also
by HSPCs, which contribute in this way to regenerate their own
niche after irradiation [11]. Osteoblastic cells also regulate prim-
itive HSCs through thrombopoietin/MPL signaling [12], although
the megakaryocytes themselves have been recently proposed by
the same group as the main source of thrombopoietin the bone
marrow [13]. However, more primitive mesenchymal cells pro-
gressively acquired a more prominent role in the maintenance and
regulation of HSCs, compared with mature osteoblasts.
In the mouse, BMSCs identified based on the expression of the
intermediate filament protein nestin were found highly enriched
in fibroblastic colony-forming-unit activity and self-renewal capac-
ity in serial transplantations. Nestin+ BMSCs express high levels of
HSC maintenance genes, like Cxcl12 [14] and c-kit ligand, and their
depletion reduced HSC activity and bone marrow homing [15].
Mesenchymal progenitor activity was soon after found also
enriched in a bone marrow reticular stromal cell population charac-
terized by high expression of the key HSC maintenance chemokine
Cxcl12, therefore named Cxcl12-abundant reticular (CAR) cells

http://dx.doi.org/10.1016/j.imlet.2015.06.020
0165-2478/Crown Copyright © 2015 Published by Elsevier B.V. All rights reserved.
130 A. García-García et al. / Immunology Letters 168 (2015) 129–135
Fig. 1. Interactions between BMSCs identified using different markers and HSPCs. Diagram showing several pathways mediating the interaction of BMSCs identified using
markers like osterix (Osx), nestin, Leptin receptor, neural/glial antigen 2(NG2), Cxccl12::GFP knock-in (CAR), derivative of Paired related homeobox 1 (Prx1)+ lateral plate
mesoderm. CLP, common lymphoid progenitor; CMP, common myeloid progenitor; MPP, multipotent progenitor; BMPs, bone morphogenetic proteins; OPN, osteopontin;
HA, hyaluronic acid.
[16,17]. Later, several studies confirmed the importance of Cxcl12
expressing cells in HSPC regulation but suggested different roles
depending on their source and location. Cxcl12 expression by
osteoblastic cells would be important in B-cell lymphoiesis and in
HPC retention in the bone marrow; however, perivascular Cxcl12
expression would be key to directly support HSCs [18,19].
The precise identity of the mesenchymal stromal populations
that contain BMSC and HSC niche functions remains debated. In
2012 Ding et al. identified perivascular HSC-supporting stromal
cells using the expression of the HSC maintenance factor stem
cell factor (Scf; also known as kitl). This cell population over-
lapped only with Nestin–GFPlo cells, and not with Nestin–GFPhi
cells [20]. However, Kunisaki et al. proposed NG2+ Nestin–GFPhi
cells as components of an arteriolar HSC–BMSC niche. Depletion of
NG2+ cells promoted HSC proliferation and reduced its long-term
repopulating capacity in vivo [21]. Finally, Zhou et al. suggested that
leptin receptor positive cells contained all BMSC activity and rep-
resented the main source of adult bone formation [22], although
leptin receptor+/Lepr-cre-traced cells appeared more abundant
than previously described and could contain different cell types
with distinct functions.
In the human system, CD146+ perivascular BMSCs, reported
initially by Sacchetti et al. as the in vivo correlate of human CFU-
F-initiating cells [23], can support long-term persistence of human
myelolymphoid HSPCs through cell to cell contact and via Notch
pathway [24]. Other studies also used the CD105 antigen to identify
murine mesenchymal stem cells. In this sense, non-adherent mes-
enchymal spheres derived from human CD146+ CD105+ nestin+
cells could promote human umbilical blood HSC expansion [25].
Pinho et al. also showed HSC supporting activity using murine or
adult fetal PDGFR␣ + CD51+ nestin+ BMSCs [26]. However, adult
human BMSCs are highly enriched in the PDGFR␣-CD271+ cell frac-
tion [27].
There is now broad evidence suggesting that BMSCs have posi-
tive effects in HSC transplantation. Both human and murine BMSCs
seem to improve engraftment when co-transplanted with HSCs
[28,29]. Several works showed that ex vivo expanded BMSCs could
engraft in recipient marrow after intramedullary transplantation.
These BMSCs could give rise to most stromal component and
enhance engraftment and HSC function in the recipient marrow
[30]. Le Blanc et al. co-transplanted BMSC with HSC and achieved
faster engraftment of neutrophils and platelets with 100% donor
chimerism [31].
Multiple studies have shown that BMSCs contribute to HSC
long-term maintenance, proliferation and differentiation through
different signalling pathways (Fig. 1). As indicated before,
Cxcl12–Cxcr4 axis is a key player in the interaction between BMSCs
and HSCs, but many other signaling pathways participate in this
crosstalk. For instance, Rac GTPases in nestin+ perivascular BMSCs
have been recently involved in HSC regulation [32,33]. Other stud-
ies suggest that long-term activation of Gs signaling in osteoblastic
cells negatively affect HSC maintenance [34,35]. Also, Wnt signal-
ing regulates the survival, self-renewal and differentiation of HSPCs
[36,37]. Agrin, a proteoglycan involved in neuromuscular junction,
also participate in the crosstalk between BMSCs and HSCs [38].
In addition to soluble secreted factors or extracellular proteins,
cummulative evidence suggests that direct cell-to-cell contact is
important in HSC regulation by BMSCs. As mentioned before, Notch
pathway is one example. Gottschling et al. reported that ␤1-
integrin expression in human BMSCs could promote self-renewing
divisions of HSCs [39]. There are discrepancies whether BMSCs
and/or osteobalstic cells can modulate HSPCs through the adhe-
sion molecule N-cadherin [40,41]. Other molecules that have been
involved in BMSC–HSC interactions will not be discussed here due
to space constrictions.
2. BMSCs, lymphopoiesis and myelopoiesis
As part of the study of the bone marrow stem cell niches, many
efforts have been directed towards understanding the interactions
 A. García-García et al. / Immunology Letters 168 (2015) 129–135
Fig. 2. BMSCs and/or their derivatives directly regulate HSPCs and mature blood
cells. Molecules involved in the crosstalk between mesenchymal stromal cells, lym-
phopoiesis and myelopoiesis. Mesenchymal stromal cells modulate T cell early
lymphopoiesis by cell-to-cell contact through CD49d and CD90. They also stimu-
late B cell lymphopiesis through VCAM-1, CXC12 and IL-7. However, Activin A and
G-CSF inhibit B cell formation, prompting myeloid differentiation. KITL, IL-6 and IL-3
directly contribute to myelopoiesis.
between BMSCs and HSCs. However, cummulative evidence sug-
gests that the BMSC derivatives could modulate hematopoiesis
by regulating committed hematopoietic progenitors. An in vitro
study suggested a role of mesenchymal stroma in early T-cell
lymphopoiesis through adhesion molecules [42]. Another study
proposed a negative effect of mesenchymal stroma in B-cell
lymphopoiesis mediated by activin A [43]. However, Ichii demon-
strated that human BMSCs in coculture with human CD34+ cord
blood cells can support B lymphopoiesis [44]. Zhu et al. showed
in vitro that osteoblasts are necessary and sufficient for B-cell com-
mitment and maturation [45]. Recently, it has been proposed that
granulocyte colony-stimulating factor (G-CSF) could modify BMSCs
properties resulting in suppression of B lymphopoiesis in mice [46].
As occurs with lymphopoiesis, some studies indicate that BMSCs
and/or their derivatives could affect myelopoiesis. Previous evi-
dence suggested the possibility that osteoblasts could stimulate
myelopoiesis by sustained G-CSF release [47]. Later, Angelopoulou
et al. showed that myelopoiesis and megakaryiopoiesis were
improved when CD34+ HSCs and human BMSCs were cotrans-
planted in immunodeficient mice [48]. However, it was described
later that the myelopoietic supporting capacity of BMSCs is inde-
pendent of their multipotency and it is more related to their
ostoegenic/adipogenic differentiation and their glycosylation sta-
tus [49].
Interestingly, mature hematopoietic cells can also modulate
hematopoietic progenitors through BMSCs. For instance, CD8+ T-
cells can activate early multipotent progenitors during acute viral
infection to promote temporarily myelopoiesis [50].
In summary, it seems that BMSCs and/or their derivatives
(particularly osteoblasts) naturally support B-cell lymphopoiesis;
however, inhibiting this capacity could favor myelopoiesis (Fig. 2).
3. BMSCs in extramedullary hematopoiesis
Hematopoiesis outside the bone marrow is often associated
with pathological processes. Some studies indicate that BMSCs
could also favor hematopoiesis in these abnormal situations.
Khaldovanidi et al. reported that interleukin 5 induce spleen col-
onization with mesenchymal progenitor cells, which in turn favor
the establishment of hematopoiesis providing necessary signals to
131
originate a suitable microenvironment for HSPCs [51]. In addition,
mesenchymal stromal cells implanted subcutaneously together
with endothelial colony-forming cells can establish a functional
hematopoietic microenvironment. Human normal and leukemic
cells engrafted and remained functional in this extramedullary site
[52].
4. BMSCs in hematological malignancies
Research in the past ten years has definitely proved that
the microenvironment, and specially BMSCs, play more than a
bystander role in hematopoietic diseases. Alterations of stromal
cells have been implicated in the pathogenesis of myeloprolif-
erative neoplasms (MPNs), including chronic myeloid leukemias
(CMLs), and also in acute myeloid leukemias (AMLs). MPNs are
hematological malignancies arising from mutations in the HSCs,
and present with an overproduction and accumulation of myeloid
cells. Several mutations have been described in human patients,
including mutations in the Janus Kinase 2 gene (the most frequent
one being JAK2V617F) [53–56], the thrombopoietin receptor MPL
[57], and calreticulin [58,59]. Furthermore, the list of additional sec-
ondary mutations, mostly in epigenetic regulators, has increased as
of late. Mutations in ASXL1, CBL, IDH, IKZF1, and TET2 have been
linked to an increased frequency of leukemic transformation in
patients (see Ref. [60] for a review). In the case of leukemias, cytoge-
netic abnormalities and chromosomal translocation originate the
disease and promote the expansion of the leukemic stem cell (LSC)
clone (reviewed in Ref. [61]).
In 2007, Walkley et al. published two groundbreaking studies
providing (i) direct verification that niche alterations were required
for MPN-like disease inception and (ii) validation of the hypothesis
that cells from lineages other than the hematopoietic had leuke-
mogenic potential. Genetic deletion of the retinoblastoma gene
[62] or the retinoic acid receptor gamma (RAR␥) [63] provoked a
non-transplantable MPN-like phenotype. More strikingly, in both
models the disease was recapitulated only when cells from disease-
carrying mice were transplanted into recipients bearing the genetic
lesions, hence providing categorical confirmation of the necessity
of an altered niche for disease manifestation. Thereafter, several
reports have surfaced to substantiate the idea that modifications
in stromal cells are essential to obtain a transplantable pheno-
type. Specific deletion of Notch pathway component Mind bomb
1 in stromal cells [64], deletion of miRNA processing gene Dicer1
in BMSCs [65], and the presence of novel mutations in stromal
cells from human patients different from the HSC driver mutation
in patients [66] altogether suggest a prominent role of BMSCs in
myeloid neoplasias.
Some questions have arisen in the field: are the lesions of the
microenvironment secondary events? To what extent do stromal
cells switch from their normal physiology to a malignant one?
Could aberrant BMSCs actually drive and initiate disease? Many
advances have been made using human samples from patients
suffering from MPNs as well as an ample variety of murine mod-
els. Data from both of these sources portray consistent results:
mesenchymal stromal cells display aberrant secretory phenotypes
[67–69], disrupted signaling pathways [70–72] or atypical gene
expression patterns [73]. For instance, in myelodysplastic syn-
drome (MDS), BMSCs express anomalous levels of CXCL12 [74],
leukemic inhibitory factor (LIF), VEGF, and N-Cadherin or DICER
[75], among others. The altered expression profile of these BMSCs
has been proposed to be crucial for the remodeling of the neopla-
sic niche by creating a malignant microenvironment which favors
the development and maintenance of mutated MPN cells. Convinc-
ing data have been presented using xenograft models in which
CD34+ hematopoietic cells plus mesenchymal stromal cells from
132 A. García-García et al. / Immunology Letters 168 (2015) 129–135

Fig. 3. Some regulatory mechanisms in place within the malignant bone marrow niche. (A) Osteoblastic lineage bias. (B) Disruption of signal transduction pathways in BMSCs.
(C) Defective sympathetic innervation on nestin+ cells. (D) Abnormal production or inhibition of healthy HSC maintenance factors. (E) Activation of niche remodeling led by
the bone-lining osteoblasts acting on the neoplasic clone. (F) Expansion of the mutated blood cell. (G) Reinforcement of the malignant microenvironment through paracrine
loops between the BMSCs and the mutated HSCs.

disease-carrying donors were transplanted into immunodeficient
NSG mice. Strikingly, mice transplanted with mesenchymal stromal
cells and CD34+ cells displayed a significantly higher reconstitu-
tion rate than mice transplanted only with CD34+ cells or together
with mesenchymal stromal cells from healthy donors [73]. These
cotransplation setups have aided to illustrate the remodeling prin-
ciple: mutant cells hijack the normal function of the stromal cells
and therefore a modified, disease-prone, and even drug-resistant
niche [76–80] is formed.
Drawing parallels to the homeostasis situation, mutant HSCs
reside within their particular microenvironment that works to
maintain the neoplasm and to suit the regulatory pathways
involved in the expansion and progression of disease. Multiple
reports have shed light on niche remodeling mechanisms. One is
based on the transcriptional reprogramming of BMSCs to (i) con-
fer an advantage to the mutant hematopoietic cell, or (ii) impair
healthy hematopoiesis; or possibly both concomitantly. Neopla-
sic cells rely most heavily on signaling cascades prompted by the
paracrine action of secreted factors or cytokines. In CML, mutant
cells have been reported to induce the upregulation and secretion
of placental growth factor (PIGF) in stromal cells, which in turn
stimulates the proliferation of CML cells in a positive loop fashion
[80]. In an additive manner, G-CSF secreted by CML cells leads to a
decrease in BMSC-produced CXCL12, leading to complications with
normal HSC maintenance and retention [81].
In an attempt to establish unified patterns for the whole spectra
of myeloid malignancies, one needs to underscore and highlight
the pivotal role of BMSCs and their progeny as major sources of
cytokine production. Paracrine loops are known to occur between
mutant HSCs and their healthy counterparts that are transformed
into “leukemic-like” pro-inflammatory cells [82,83]. Nevertheless,
several key studies have situated stromal elements in the midst of
fundamental niche transformations. One example of the stromal
alterations in myeloid diseases is the loss of bone mass occurring
in the blast phase of CML or in AML. Although partially explained
by an increase of osteaclastic bone resorption, the decrease in bony
matrix was the result of an increase in the levels of cytokine CCL3
produced by leukemic cells [84], which effectively decreased the
number of both osteoprogenitors and the more mature osteoblasts
[85]. Similar endocrine signals can regulate both osteoprogenitor
cells and HSCs. For instance, work from the Morrison lab and our
group has shown recently that the female sex hormones estro-
gens, known to be bone anabolic agents, also differentially regulate
HSC self-renewal, proliferation and survival [86,87]. Endogenous
estrogens might partially explain the higher incidence and sever-
ity of hematological malignancies in men, compared with women,
and selective estrogen receptor modulators might be useful in the
treatment of MPN [86]. Linked to the notion of a dysfunctional
mesenchyme compartment, alterations in gene expression in mes-
enchymal cells have been deemed to have oncogenic potential and
to increase the likelihood of leukemic transformation in human
patients. Specific deletion of the Schwachman–Bodian–Diamond
(sbds) gene in cells from mesenchymal origin prompted myelodys-
plastic symptoms in mice [65]. Furthermore, it has been proposed
that, in MPNs, BMSCs might develop a bias towards generation
of malfunctioning osteoblastic progenitors, ultimately leading to
faulty hematopoiesis and creating fertile grounds for marrow fibro-
sis and osteosclerosis [88]. Altogether, available data seem to agree
 A. García-García et al. / Immunology Letters 168 (2015) 129–135
with the hypothesis that mutated HSPCs cause damage to BMSCs
that is partially responsible for disease manifestation and progres-
sion (Fig. 3).
Recent reports have added the peripheral nervous system as
yet another layer of regulation in normal hematopoiesis [89–91]
but also in the development of MPNs, as neuropathy-triggered
inflammatory damage to BMSCs seems to be required for MPN
manifestation [92]. In a murine model of JAK2V617F polycythemia
vera, we have shown that the overproduction of interleukin-1␤
by mutant HSPCs triggers the death of Schwann cells and causes
neuropathy in sympathetic nerve terminals in the bone marrow,
sensitizing nestin+ BMSCs to undergo apoptosis. Acceleration of
the polycythemic phenotype can be directly attributed to the loss
of BMSCs in the marrow, since in vivo depletion of nestin+ BMSCs
or their Cxcl12 production fueled disease progression, whereas the
pharmacological rescue of the sympathetic innervation of BMSCs
through administration of sympathomimetic agents was able to
prevent nestin+ BMSC loss and thus block mutant HSC expan-
sion [92]. Similar findings were soon after reported in a MLL-AF9
acute myeloid leukemia (AML) murine model [93], although the
relevance of bone marrow neuropathy in this setting is unclear
since, unlike in MPN [92], adrenergic signals did not seem to affect
significantly AML progression [93]. Due to its clinical relevance
and impact on public health, AML has been extensively studied.
Many relevant insights have been made on the crosstalk between
leukemic stem cells (LSCs) and their niche, both in xenograft and
murine models. Whereas the niche in acute leukemias can assist in
LSC engraftment and homing [94,95] and promote chemotherapy-
resistance [96], it remains to be determined whether it can also
regulate other aspects of LSC biology in AML. Dissecting the involve-
ment of stromal cells in leukemic maintenance and development
will be vital to design meaningful and powerful therapeutic strate-
gies.
In summary, multiple studies using genetically-modified mouse
strain converge in the notion that perivascular BMSCs cooper-
ate with endothelial cells in regulating HSCs. Similar conclusions
have also been obtained in phylogenetically distant species, such
as zebrafish [97]. Parallels are emerging in the skeletal and the
hematopoietic systems, since the physiological turnover of both
bone [98,99] and blood [100] seem to rely mostly in committed
progenitors or short-term stem cells, but not in the most primitive
stem cells. However, during stress situations MSCs and HSCs can be
activated and contribute to organ turnover, such as bone fracture
or myeloablation, respectively.
Also, BMSCs have emerged as key contributors and partici-
pants in hematopoietic malignancies. Despite the ever-increasing
amount of sound reports, we are in need of a thorough characteri-
zation of the chronological events that end in niche remodeling in
order to obtain a clear picture of the disease-initiating or disease-
propagating mechanisms. With the advent of improved models and
technological advances to study individual cells, the field of BMSCs
and hematopoiesis will surely witness its landscape modified in
coming years.
Acknowledgements
A.G.-G. is the recipient of La Caixa-Severo Ochoa PhD fellow-
ship. C.L.F. de C. has been awarded a PhD fellowship by Boehringer
Ingelheim Fonds. This work was supported by the Fundación Cen-
tro Nacional de Investigaciones Cardiovasculares Carlos III, the
Spanish Ministry of Economy and Competitiveness (TerCel, Span-
ish Cell Therapy Network; Plan Nacional grant SAF-2011-30308;
Ramón y Cajal Program grant RYC-2009-04703), Marie Curie Career
Integration Programme grant (FP7-PEOPLE-2011-RG-294096) and
ConSEPOC-Comunidad de Madrid S2010/BMD-2542 grant to S.M.-
133
F., who is also supported in part by an International Early Career
Scientist grant of the Howard Hughes Medical Institute. We apolo-
gize for omission of relevant literature due to space contraints.
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