Bone Marrow Transplantation (2014), 1–7 & 2014 Macmillan Publishers Limited All rights reserved 0268-3369/14 www.nature.com/bmt

REVIEW

Stromal cells–are they really useful for GVHD? H Kaipe, T Erkers, B Sadeghi and O Ringde´n Mesenchymal stromal cells (MSCs) have immunomodulatory effects and are increasingly being used for the treatment of acute and chronic GVHD. Although they seem immuno-privileged, they induce alloresponses, but the risk of immunization is poorly characterized. After infusion, they first reach the lungs, liver and spleen, and are then difficult to trace. Several mechanisms are involved in stromal cells suppressing alloreactivity, such as induction of regulatory T cells, but whether or not this will also affect leukemic relapse or increase infections is not known. Although several encouraging pilot studies have been published, there have been few prospective randomized trials. There may be a bias in the literature, as negative results are seldom published, and there have been few comparative studies with other immunosuppressive regimens. Most animal models have failed to show any effect on GVHD. Several questions remain to be answered for optimization of stromal cell therapy. Which source is optimal–BM, fat, cord or decidua? Can stromal cells be replaced by exosomes, which culture conditions are most appropriate and at what passage and how frequently should cells be administered? More research is required to move stromal cell therapy forward to become an established treatment for acute and chronic GVHD. Bone Marrow Transplantation advance online publication, 27 January 2014; doi:10.1038/bmt.2013.237 Keywords: SCT; GVHD; mesenchymal stem cells; placenta; decidual stromal cells

INTRODUCTION Stromal cells support the integrity of tissues and provide growth factors for regeneration. Depending on their local distribution, stromal cells have different assignments. In the BM, mesenchymal stromal cells (BM-MSCs) support hematopoietic cell development,1 whereas decidual stromal cells (DSCs) in the placenta promote trophoblast invasion and therefore implantation of the developing fetus.2 DSCs are also important for the development of feto-maternal tolerance.3,4 Stromal cells have immunosuppressive capacity,5–7 which paved the way for using them in the treatment of inflammatory diseases. Owing to their low immunostimulatory effect, allogeneic third-party stromal cells are often used. After our first report on the treatment with BM-MSCs of a 9-year-old boy with refectory GVHD who miraculously improved,8 there were high expectations for these cells for the treatment of inflammatory disorders and in regenerative medicine. Unfortunately, this patient died of pneumonia 19 months after transplantation.9 Ten years later, we wonder whether stromal cells have solved anything. Have they really saved the lives of any GVHD patients? Several concerns have been raised regarding the use of allogeneic stromal cells (Table 1). It is often stated that MSCs are immunoprivileged, and are unable to induce alloresponses,10,11 but stromal cells constitutively express MHC class I, and do indeed induce proliferation of allogeneic lymphocytes.6 Beggs et al.12 showed that baboons produced alloantibodies after administration of allogeneic BM-MSCs, indicating that they can immunize the host. Furthermore, allogeneic BM-MSCs are rejected in MHC-mismatched murine recipients13 and they can induce memory T-cell responses, resulting in the rejection of allogeneic stem cell grafts.14 Thus, MSCs are not intrinsically immunoprivileged. Sundin et al.15 detected no anti-HLA Abs when

analyzing 12 GVHD patients after MSC infusion, but confirmatory studies are needed. Patients with acute GVHD are heavily immunosuppressed, but patients with functional immune systems may respond differently and develop multi-specific anti-HLA Abs. There are limited data on the homing capacity of MSCs in patients, and most studies on cell distribution have been confined to animal models. Intravenously infused cells first reach the lungs and then redistribute to the liver and spleen,16 whereas there is little proof of specific homing to inflamed tissue.9 Autopsy examinations have revealed that BM-MSCs can be detected in the lungs and lymph nodes of some patients, but at low levels.9,17 Thus, MSCs hardly seem to engraft, but rather may mediate their function through paracrine factors before they are rejected. As engraftment of BM-MSCs is rare, one might wonder if they even survive in the short term after infusion. Culture expanded fibroblast-like cells may not be adapted to survive in blood, which contains coagulation and complement factors. It has been shown that BM-MSCs are susceptible to complement activation after contact with human blood,18 which results in cell dysfunction or cell death.19 BM-MSCs also elicit the formation of clotting factors when in contact with blood.20 The immunosuppressive capacity of MSCs may not only be a positive feature when treating patients who are already highly susceptible to infections. Co-infusion of BM-MSCs with cord blood transplantation delays immune reconstitution, as shown by lower levels of T-cell receptor excision circles and serum IgG and IgM.21 It has further been found to be associated with the development of EBV-related post-transplant lymphoproliferative disorder,9,22 and invasive fungal infections.23 There are also worries that MSCs may promote cancer cell growth. Co-transplantation of BM-MSCs was found to be associated with a higher rate of occurrence of leukemic relapse.24 BM-MSCs have also been suggested to be

Division of Therapeutic Immunology and Centre for Allogeneic Stem Cell Transplantation, Karolinska Institutet, Karolinska University Hospital Huddinge, Stockholm, Sweden. Correspondence: Dr H Kaipe, Division of Therapeutic Immunology, F79 Department of Laboratory Medicine, Karolinska University Hospital Huddinge, Stockholm SE-141 86, Sweden. E-mail: [email protected] Received 29 October 2013; accepted 19 December 2013

Stromal cells for GVHD? H Kaipe et al

2 Table 1.

Putative problems with allogeneic stromal cell therapy

Problem

Published observations

References 12–14

Immunogenicity

B- and T-cell-mediated immunity to allogeneic MSCs in animal models

Immediate cell death

Complement activation Activation of coagulation factors and pulmonary embolism

18–20,104

Poor homing capacity

MSCs mainly target the lungs, liver, and spleen. Little or no confirmed homing to affected organs in patients

9,17,16,103

Infections

MSC infusion increases the risk of PTLD

9,22

Chromosomal aberrations

Sarcoma development in animal models. Chromosomal instability in mouse and human BM-MSCs

106–108

Promote malignant disease

Increased rate of leukemic relapse. Promote multiple myeloma cell growth in vitro

24–26,105

Long-term effects

Poor long-term survival of MSCtreated GVHD patients. Deprived thymic reconstitution

21,23,48

Lack of relevant animal models

Poor response in murine and canine models of GVHD. Different mechanisms of action in mice and humans

28–32,66

involved in the progression of oncogenesis in multiple myeloma,25 which can be mediated by MSC-derived exosomes.26 The anticipation of using BM-MSCs as GVHD treatment dramatically declined in the scientific community when the negative results of the randomized clinical trial by Osiris Therapeutics were released.27 BM-MSC administration for treatment of GVHD did not improve long-term survival23 and have also failed to show effects on GVHD in animal models.28–32 Some studies from academic institutions have shown positive results, but there is most likely a selection bias in the literature as negative results are seldom published.33 With all these concerns in mind, should we really use stromal cells as treatment for GVHD? Before we condemn this kind of therapy, we will summarize what we know today. THE DEVELOPMENTAL ORIGIN OF MSCS The mesodermal precursors that give rise to the in vitro expanded multipotential MSC lines have not been identified, and where they reside in vivo is still not clear. The fact that MSCs and MSC-like cells can be isolated from many different tissues, including BM, fat and placenta, suggests that there is a common reservoir of these cells in the body (reviewed in Murray et al.34). However, whether all MSCs are phenotypically and functionally equivalent and how in vitro expansion affects these features is not known. Recent advances suggest that at least subsets of MSCs are anatomically and functionally associated with vascular niches.35,36 MSCs can also be isolated from avascular tissues, such as the amniotic membrane, but these still show an inherent capacity to promote angiogenesis.37 Pericytes surround endothelial cells in blood microvessels and contribute to vascular development, stabilization and remodeling.34 Pericytes have been shown to have MSC-like features, including multipotency38,39 and immunosuppressive capacities.40 A vascular origin of MSCs may be a positive trait in the context of blood compatibility after infusion, Bone Marrow Transplantation (2014) 1 – 7

Table 2.

Mechanisms for stromal cell-mediated suppression

Factors involved in suppression

Proposed mechanistic function

References

IDO

Depletion of tryptophan and production of suppressive metabolites, induction of M2 macrophages

56,61

Prostaglandin E2

Inhibition of NFAT and AP-1mediated IL-2 expression

50,109

HLA-G

Suppression of immune cells by ligation of immune inhibitory receptors ILT2 and ILT4

49

Exosomes and microvesicles

Carriers of suppressive factors, including miRNA

68

PD-L1

T-cell inhibitory molecule

62

ICAM-1 and VCAM-1

Promotes adhesion between stromal cells and T cells to potentiate suppression

64

Nitric oxide

Suppression of proliferation of T cells by inhibiting STAT5 activation

54,110

Galectins

Inhibition and apoptosis of T cells

51

CD39 and CD73

Adenosine-mediated suppression of T cells

65

but in vitro expansion of the cells can still affect the phenotype and function of MSCs. IMMUNE MODULATION BY MSCS The immunomodulatory properties of MSCs have been extensively studied over the past 10 years, and there are already several detailed reviews covering characterization, immune modulation and tissue regeneration by MSCs.41–45 In this section, we will briefly discuss some of these properties, which are summarized in Table 2. One prerequisite for the introduction of BM-MSCs as treatment for GVHD was their ability to suppress alloreactivity in mixed lymphocyte reactions.5,10,11,46,47 This anti-proliferative ability is shared by all stromal cells, including placental stromal cells6 and skin fibroblasts.7 Despite the fact that MSCs reduce alloreactivity in vitro, it has been difficult to correlate clinical outcome to successful inhibition of mixed lymphocyte reactions with BMMSCs and patient PBMCs as responder cells.48 MSCs use a variety of different factors in order to suppress immune responses, many of which are secreted. Some of these factors are constitutively produced by MSCs, such as HLA-G5,49 prostaglandin E2 (PGE2),50 and galectins.51 The immunomodulatory effects exerted by MSCs can be augmented if the cells are activated by cytokines such as IFN-g.52–54 It has also been shown that MSCs can be activated by stimulation of their Toll-like receptors.55 Another important mediator of suppression by stromal cells is the T-cell inhibitory enzyme indoleamine-2,3-dioxygenase (IDO).56 IDO expression can be induced by IFN-g57 and by activation of TLR-3.55,58 MSC-produced IDO is involved in the induction of regulatory T cells and inhibition of Th17 differentiation.59,60 MSCderived IDO can also promote differentiation of macrophages toward an M2 phenotype.61 Activated MSCs can modulate adaptive immune cells through contact-dependent mechanisms. These include activation of the PD-1 pathway,62 Fas-mediated T-cell apoptosis,63 engagement of & 2014 Macmillan Publishers Limited

Stromal cells for GVHD? H Kaipe et al

3 VCAM-1 and ICAM-1,64 and through upregulation of CD39 and increase in adenosine production.65 Although IDO is important in the T-cell suppression by MSCs in a human setting, synthesis of nitric oxide is also induced by IFN-g, which has been shown to be a main mediator of suppression in mice.54 This demonstrates the variation among different species regarding the mechanisms of immune suppression by MSCs and the translational difficulties that might occur when validating a new MSC therapy in animal models.66 MSCs may need to be licensed by IFN-g or nitric oxide, or transduced with IL-10 to ameliorate GVHD and improve survival in mouse models.31,54,67 STROMAL CELL-DERIVED EXOSOMES AND MICROVESICLES Exosomes and microvesicles are small but complex entities that contain both immunomodulatory proteins and microRNA, and have homing abilities.68 They are secreted by live cells and are involved in cell–cell communication and transfer of cellular material. Depending on the intracellular origin, secreted membrane vesicles can have distinct biochemical properties which probably also affect their function. True exosomes derive from multivesicular bodies and measure 50–100 nm in diameter.69 MSC-derived microvesicles have been shown to protect from acute kidney injury,70 myocardial ischemia71 and pulmonary hypertension72 in animal models (Table 3). The regenerative capacities of microvesicles appear to be at least partly dependent on the mRNA cargo, as treatment with RNAase to inactivate the mRNA content abrogated the protective effects.73 Some advantages of exosomes and microvesicles are that they may be more defined and with more controlled functions compared with viable cells, and that they can deliver target molecules intracellularly.74 Microvesicles derived from MSCs were also demonstrated to inhibit tumor growth.75 Exosomes may be ideal carriers of the intricate blend of paracrine factors that are needed for treatment of a complicated disease such as GVHD. One case where exosomes were used for treatment for severe acute GVHD has been reported in an abstract that was presented at the EBMT meeting in 2013.76 However, if therapeutic exosomes are to be translated into clinical use, there are still major concerns that need to be resolved. The isolation of

Table 3.

exosomes must be improved to be cost effective, and the yield and purity of the product need to be increased.77,78 In addition, the exosome product obtained by the most common isolation protocol with differential centrifugation and a sucrose gradient yield a heterogeneous product. Therefore, an exosome or microvesicle product needs to be extensively characterized in order to assess its biological function before it can be used therapeutically. CLINICAL USE OF BM-DERIVED MSCS Needless to say, steroid-refractory severe acute or chronic GVHDs are terrible disorders where little therapeutic progress has been made over the last three decades.79–81 We were the first group to use MSCs for the treatment of life-threatening acute GVHD, and we reported dramatic responses in some patients, but unfortunately not all.8,9 These results encouraged a multi-centre phase-II study involving five centres and 55 patients.82 Complete response was seen in 68% of the children and in 53% of the adult patients. There was no difference whether the MSCs were from an HLA-identical sibling donor, a haploidentical donor or a third party. More than 200 patients have been reported to be treated for acute GVHD (Table 4). The MSC dose has ranged from 1 to 21 doses. Complete response has been seen in 52% of the patients, with partial response in 23% and no response in 25%.83 Most centres used MSCs expanded in FCS, but expansion of MSCs in platelet lysate medium and human AB serum was also reported.84,85 There have been few randomized studies using MSCs. Osiris Therapeutics, Inc. used expanded MSCs, Prochymal, for acute GVHD of grades II–IV.86 Initial response was 94%, with a complete response of 77%. Subsequently, a prospective double-blind placebo-controlled phase-III study was performed in patients with acute GVHD of grades II–IV using Prochymal in two patients for every one who received placebo.87 Durable complete response was 40% in the Prochymal group and 28% in the placebo group (P ¼ 0.08). Of 61 patients with acute GVHD of the liver, complete response was 76% in the Prochymal group and 47% in the placebo group (Po0.05). In patients with acute gastrointestinal GVHD, complete

Functions of stromal-cell-derived microvesicles and exosomes

Feature

Source

Isolation technique (size of product)

Biological effects

References

Promote angiogenesis Improve acute kidney injury Cardio-protective effects Promote oncogenic cell growth

Umbilical cord MSC

Ultracentrifugation (100 nm)

Increase endothelial cell growth and tube formation

Human BM-MSC ESC-derived MSC

Ultracentrifugation (80 nm–1 mm) Ultrafiltration (50–100 nm)

Human BM-MSC from MM patients

Ultracentrifugation and exosome precipitation (100 nm)

Anti-apoptotic, promote tubular epithelial cell proliferation in rat model Reperfusion of ischemic myocardium in mouse model Exosomes containing oncogenic proteins transferred to MM cells, supports tumor growth

111 73,112 71 26

Abbreviations: ESC ¼ embryonic stem cells; MM ¼ multiple myeloma; MSCs ¼ mesenchymal stromal cells.

Table 4.

A summary of published studies on MSCs for treatment of acute and chronic GVHD: 21 on acute GVHD and 10 on chronic GVHD83

Acute GVHD Chronic GVHD

Number of patients

Cell dose  106/kg

Range of doses

Complete response

Partial response

No response

190 61

0.4–9.0 0.2–20

1–21 1–11

98 (52%) 16 (26%)

44 (23%) 29 (48%)

48 (25%) 16 (26%)

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Stromal cells for GVHD? H Kaipe et al

4 response was also more common in the Prochymal group than in the placebo group (Po0.05). There have been few long-term reports using MSCs. At our centre, 31 patients treated with MSCs for acute GVHD or hemorrhagic cystitis were followed for 44 years.48 The patients who received MSCs from passage 1–2 had better survival (75% at 1 year) than those who received MSCs from passage 3–4 (21%) (Po0.01). Long-term survival was not better than in patients with severe acute GVHD who were not treated with MSCs.23

ALTERNATE SOURCES OF CELLS FOR ACUTE GVHD As an alternative to BM-derived MSCs, adipose tissue-, umbilical cord- or placental tissue-derived stromal cells have been used for the treatment of acute GVHD88,89 (Table 5). We recently introduced DSCs from term fetal membrane as a treatment for patients with GVHD.90 We first compared three different sources of cells from the placenta: DSCs, stromal cells from umbilical cord and stromal cells from placental villi in vitro.6 We found that DSCs exerted the most consistent inhibition of proliferation and inhibition of IFN-g and IL-17 production in mixed lymphocyte reactions compared with BM-MSCs and the other sources of stromal cells from placenta. Other major advantages with this cell population are the great expansion potential at low passage and high expression of integrins.6,52 We therefore used DSCs for treatment of severe steroid-refractory acute GVHD of grades III–IV in nine patients.90 Median age was 57 years and two patients had a complete response and four had a partial response. A number of other cell subsets have been suggested as treatment for GVHD, for example, CD4 þ regulatory T cells,91 myeloid-derived suppressor cells92 and CD8 þ regulatory T cells93 (Table 5). The target of these cell types is the suppression of alloreactivity, while MSCs and cells isolated from the placenta may also have regenerative abilities. The effect on tissue regeneration by MSCs in the GVHD setting has not been thoroughly investigated.

STROMAL CELLS FOR TREATMENT OF CHRONIC GVHD Stromal cells have also been used for treatment of chronic GVHD. In the first patient treated, we saw a partial response.9 The largest study was from Weng et al.94 who reported on 19 patients with

Table 5.

Cellular therapies for GVHD References

Stromal cells Sources BM Adipose tissue Umbilical cord Placenta-derived decidua Exosomes from stromal cells Expansion media FCS Platelet lysate medium AB serum Regulatory T cells CD4 þ CD8 þ Myeloid suppressor cells Suicide gene manipulated T cells in the graft

Bone Marrow Transplantation (2014) 1 – 7

8,9,23,48,82–87,96–100 83,88 83,89 83,90 76

8,9,82–87 83,85 84

91 93 92 113

refractory chronic GVHD. These patients were treated with 1–5 doses. A response was seen in 14 of the 19 patients (74%). Of the 61 patients reported to have been treated with MSCs for chronic GVHD, complete response was seen in 26% of the patients, partial response was seen in 48% of the patients and 26% of the patients did not respond at all (Table 4).83 STROMAL CELLS FOR PREVENTION OF GVHD Lazarus et al.95 were the first to show that it was safe to give HLA-identical MSCs to allogeneic HSCT recipients. No acute side effects were seen after infusion of MSC doses at a range of 1–5  106cells/kg in 46 HLA-identical sibling transplant recipients. Bernardo et al.96 used MSCs at the time of HSCT transplantation in children who were recipients of cord blood grafts. Ball et al.97 performed co-transplantation of MSCs in children undergoing haploidentical HSCT. Compared with retrospective controls, they saw less graft failure and faster engraftment of platelets using MSCs. In a prospective randomized study, patients undergoing HSCT from haploidentical donors were randomized to co-infusion with MSCs (3–5  105 cells/kg) or not. The MSC group reached platelets more than 50  109/L at a median of 22 days, as compared with 28 days in the control group (P ¼ 0.04).98 Acute GVHD and survival were not statistically significantly different between the groups.98 Kuzmina et al.99 performed a randomized prophylactic study with an MSC dose of 1  106/kg at the time of blood count recovery. Acute GVHD of grades II–IV developed in only 5% of the MSC patients as compared with 39% of the controls (P ¼ 0.002). DISCUSSION When a new experimental treatment is introduced, only severely ill patients–where there is little to lose–are included.8,9 A 50% response rate among end-stage acute GVHD patients may seem encouraging (Table 4). However, an improved survival was only noted in patients with complete response,82 for whom 2-year survival was 53% and long-term survival even worse.48 The results are more promising when used at an early stage of GVHD.86 Bernardo et al.96 treated grade-II acute GVHD with MSCs and reported no deaths that were attributable to acute GVHD as compared with 26% in retrospective control patients. According to animal data, not only early treatment, but also repeated doses may be needed. Although many patients respond well to one dose, some patients appear to benefit from several doses to maintain immunosuppression.100 Stromal cells may also be used for chronic GVHD, but they should probably be used before fibrosis occurs.101 Stromal cells have a profound effect on coagulation,18,20 which partly explains their effectiveness in stopping major hemorrhages.90,102,103 BM must be accessed by aspiration, and this source of MSCs has been challenged by MSCs from adipose tissue, umbilical-cordderived MSCs and stromal cells from the placenta–such as DSCs. Placental stromal cells are accessible without any invasive procedure and with little ethical consideration. To determine which source of stromal cells is superior would require large prospective randomized studies that may never be performed. Although no acute toxicity has been reported, the coagulation system is activated18,20 and there is a concern regarding pulmonary embolism.104 Patients treated with stromal cells need to be followed closely for the possible risk of tumor formation.25,26,105–108 When taking all clinical observations into consideration, we still believe that stromal cell therapy can be useful for GVHD, particularly in children. However, this treatment needs to be optimized, combining it with the right immunosuppressive therapies. We need to learn more about the therapeutic potential of stromal cells in order to understand their mechanistic functions & 2014 Macmillan Publishers Limited

Stromal cells for GVHD? H Kaipe et al

5 in the clinical setting. We have learnt that they are immunosuppressive and that they have regenerative properties, but so far these traits have only been confirmed in vitro. Whether or not the stromal cells need to localize to the affected organ or whether the systemic effect is sufficient is also unclear. Much more research– mechanistically in vitro, in vivo in experimental animals, and also in the clinic–is required to establish stromal cell therapy as an effective treatment for GVHD.

CONFLICT OF INTEREST The authors declare no conflict of interest.

ACKNOWLEDGEMENTS Helen Kaipe: The Swedish Research Council (K2012-99X-22013-01-3), the Cancer Society in Stockholm (121092), the Swedish Society of Medicine, the Childrens’ Cancer Foundation (PR2013-0020), Clas Groschinsky Foundation and Karolinska Institutet. Olle Ringde´n: The Swedish Cancer Society (CAN2011/419), the Swedish Research Council (K2011-64X-05971-31-6), the Childrens’ Cancer Foundation (PROJ12/021), the Cancer Society in Stockholm (111293) and Karolinska Institutet.

REFERENCES 1 Mendez-Ferrer S, Michurina TV, Ferraro F, Mazloom AR, Macarthur BD, Lira SA et al. Mesenchymal and haematopoietic stem cells form a unique bone marrow niche. Nature 2010; 466: 829–834. 2 Li MQ, Hou XF, Shao J, Tang CL, Li DJ. The DSCs-expressed CD82 controls the invasiveness of trophoblast cells via integrinbeta1/MAPK/MAPK3/1 signaling pathway in human first-trimester pregnancy. Biol Reprod 2010; 82: 968–979. 3 Nancy P, Tagliani E, Tay CS, Asp P, Levy DE, Erlebacher A. Chemokine gene silencing in decidual stromal cells limits T cell access to the maternal-fetal interface. Science 2012; 336: 1317–1321. 4 Oreshkova T, Dimitrov R, Mourdjeva M. A cross-talk of decidual stromal cells, trophoblast, and immune cells: a prerequisite for the success of pregnancy. Am J Reproduct Immunol 2012; 68: 366–373. 5 Le Blanc K, Tammik L, Sundberg B, Haynesworth SE, Ringden O. Mesenchymal stem cells inhibit and stimulate mixed lymphocyte cultures and mitogenic responses independently of the major histocompatibility complex. Scand J Immunol 2003; 57: 11–20. 6 Karlsson H, Erkers T, Nava S, Ruhm S, Westgren M, Ringden O. Stromal cells from term fetal membrane are highly suppressive in allogeneic settings in vitro. Clin Exp Immunol 2012; 167: 543–555. 7 Jones S, Horwood N, Cope A, Dazzi F. The antiproliferative effect of mesenchymal stem cells is a fundamental property shared by all stromal cells. J Immunol 2007; 179: 2824–2831. 8 Le Blanc K, Rasmusson I, Sundberg B, Gotherstrom C, Hassan M, Uzunel M et al. Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells. Lancet 2004; 363: 1439–1441. 9 Ringden O, Uzunel M, Rasmusson I, Remberger M, Sundberg B, Lonnies H et al. Mesenchymal stem cells for treatment of therapy-resistant graft-versus-host disease. Transplantation 2006; 81: 1390–1397. 10 Tse WT, Pendleton JD, Beyer WM, Egalka MC, Guinan EC. Suppression of allogeneic T-cell proliferation by human marrow stromal cells: implications in transplantation. Transplantation 2003; 75: 389–397. 11 Le Blanc K, Tammik C, Rosendahl K, Zetterberg E, Ringden O. HLA expression and immunologic properties of differentiated and undifferentiated mesenchymal stem cells. Exp Hematol 2003; 31: 890–896. 12 Beggs KJ, Lyubimov A, Borneman JN, Bartholomew A, Moseley A, Dodds R et al. Immunologic consequences of multiple, high-dose administration of allogeneic mesenchymal stem cells to baboons. Cell Transplant 2006; 15: 711–721. 13 Eliopoulos N, Stagg J, Lejeune L, Pommey S, Galipeau J. Allogeneic marrow stromal cells are immune rejected by MHC class I- and class II-mismatched recipient mice. Blood 2005; 106: 4057–4065. 14 Nauta AJ, Westerhuis G, Kruisselbrink AB, Lurvink EG, Willemze R, Fibbe WE. Donor-derived mesenchymal stem cells are immunogenic in an allogeneic host and stimulate donor graft rejection in a nonmyeloablative setting. Blood 2006; 108: 2114–2120. 15 Sundin M, Ringden O, Sundberg B, Nava S, Gotherstrom C, Le Blanc K. No alloantibodies against mesenchymal stromal cells, but presence of anti-fetal calf serum antibodies, after transplantation in allogeneic hematopoietic stem cell recipients. Haematologica 2007; 92: 1208–1215.

& 2014 Macmillan Publishers Limited

16 Gholamrezanezhad A, Mirpour S, Bagheri M, Mohamadnejad M, Alimoghaddam K, Abdolahzadeh L et al. In vivo tracking of 111In-oxine labeled mesenchymal stem cells following infusion in patients with advanced cirrhosis. Nuclear Med Biol 2011; 38: 961–967. 17 von Bahr L, Batsis I, Moll G, Hagg M, Szakos A, Sundberg B et al. Analysis of tissues following mesenchymal stromal cell therapy in humans indicates limited long-term engraftment and no ectopic tissue formation. Stem Cells 2012; 30: 1575–1578. 18 Moll G, Jitschin R, von Bahr L, Rasmusson-Duprez I, Sundberg B, Lonnies L et al. Mesenchymal stromal cells engage complement and complement receptor bearing innate effector cells to modulate immune responses. PLoS One 2011; 6: e21703. 19 Li Y, Lin F. Mesenchymal stem cells are injured by complement after their contact with serum. Blood 2012; 120: 3436–3443. 20 Moll G, Rasmusson-Duprez I, von Bahr L, Connolly-Andersen AM, Elgue G, Funke L et al. Are therapeutic human mesenchymal stromal cells compatible with human blood? Stem Cells 2012; 30: 1565–1574. 21 Uhlin M, Sairafi D, Berglund S, Thunberg S, Gertow J, Ringden O et al. Mesenchymal stem cells inhibit thymic reconstitution after allogeneic cord blood transplantation. Stem Cells Develop 2012; 21: 1409–1417. 22 Uhlin M, Wikell H, Sundin M, Blennow O, Maeurer M, Ringden O et al. Risk factors for Epstein Barr virus related post-transplant lymphoproliferative disease after allogeneic hematopoietic stem cell transplantation. Haematologica (e-pub ahead of print 20 September 2013; doi:10.3324/haematol.2013.087338). 23 Remberger M, Ringden O. Treatment of severe acute graft-versus-host disease with mesenchymal stromal cells: a comparison with non-MSC treated patients. Int J Hematol 2012; 96: 822–824. 24 Ning H, Yang F, Jiang M, Hu L, Feng K, Zhang J et al. The correlation between cotransplantation of mesenchymal stem cells and higher recurrence rate in hematologic malignancy patients: outcome of a pilot clinical study. Leukemia 2008; 22: 593–599. 25 Gupta D, Treon SP, Shima Y, Hideshima T, Podar K, Tai YT et al. Adherence of multiple myeloma cells to bone marrow stromal cells upregulates vascular endothelial growth factor secretion: therapeutic applications. Leukemia 2001; 15: 1950–1961. 26 Roccaro AM, Sacco A, Maiso P, Azab AK, Tai YT, Reagan M et al. BM mesenchymal stromal cell-derived exosomes facilitate multiple myeloma progression. J Clin Invest 2013; 123: 1542–1555. 27 Martin PJ, Uberti J, Soiffer R, Klingemann H, Waller EK, Daly AS et al. Prochymal improves response rates in patients with steroid-refractory acute graft versus host disease (SR-GVHD) involving the liver and gut: results of a randomized placebo-controlled multicenter phase III trial in GVHD. Biol Blood Marrow Transplant 2010; 16(suppl 2): S169–S170. 28 Badillo AT, Peranteau WH, Heaton TE, Quinn C, Flake AW. Murine bone marrow derived stromal progenitor cells fail to prevent or treat acute graft-versus-host disease. Br J Haematol 2008; 141: 224–234. 29 Jeon MS, Lim HJ, Yi TG, Im MW, Yoo HS, Choi JH et al. Xenoreactivity of human clonal mesenchymal stem cells in a major histocompatibility complex-matched allogeneic graft-versus-host disease mouse model. Cellular Immunol 2010; 261: 57–63. 30 Mielcarek M, Storb R, Georges GE, Golubev L, Nikitine A, Hwang B et al. Mesenchymal stromal cells fail to prevent acute graft-versus-host disease and graft rejection after dog leukocyte antigen-haploidentical bone marrow transplantation. Biol Blood Marrow Transplant 2011; 17: 214–225. 31 Sadeghi B, Ringden O. Mesenchymal stem cells for graft-versus-host disease in experimental animal models. In: Gross G, Ha¨upl T (eds). Stem Cell-Dependent Therapies: Mesenchymal stem cells in chronic inflammatory disorders. De Gruyter: Berlin, pp 125–141, 2013. 32 Sudres M, Norol F, Trenado A, Gregoire S, Charlotte F, Levacher B et al. Bone marrow mesenchymal stem cells suppress lymphocyte proliferation in vitro but fail to prevent graft-versus-host disease in mice. J Immunol 2006; 176: 7761–7767. 33 Paulson K, Saeed M, Mills J, Cuvelier GD, Kumar R, Raymond C et al. Publication bias is present in blood and marrow transplantation: an analysis of abstracts at an international meeting. Blood 2011; 118: 6698–6701. 34 Murray IR, West CC, Hardy WR, James AW, Park TS, Nguyen A et al. Natural history of mesenchymal stem cells, from vessel walls to culture vessels. Cell Mol Life Sci (e-pub ahead of print 25 October 2013). 35 Tang W, Zeve D, Suh JM, Bosnakovski D, Kyba M, Hammer RE et al. White fat progenitor cells reside in the adipose vasculature. Science 2008; 322: 583–586. 36 Crisan M, Yap S, Casteilla L, Chen CW, Corselli M, Park TS et al. A perivascular origin for mesenchymal stem cells in multiple human organs. Cell Stem Cell 2008; 3: 301–313. 37 Warrier S, Haridas N, Bhonde R. Inherent propensity of amnion-derived mesenchymal stem cells towards endothelial lineage: vascularization from an avascular tissue. Placenta 2012; 33: 850–858.

Bone Marrow Transplantation (2014) 1 – 7

Stromal cells for GVHD? H Kaipe et al

6 38 Cossu G, Bianco P. Mesoangioblasts--vascular progenitors for extravascular mesodermal tissues. Curr Opin Genet Develop 2003; 13: 537–542. 39 Galli D, Innocenzi A, Staszewsky L, Zanetta L, Sampaolesi M, Bai A et al. Mesoangioblasts, vessel-associated multipotent stem cells, repair the infarcted heart by multiple cellular mechanisms: a comparison with bone marrow progenitors, fibroblasts, and endothelial cells. Arteriosclerosis, Thrombosis Vascular Biol 2005; 25: 692–697. 40 Ochs K, Sahm F, Opitz CA, Lanz TV, Oezen I, Couraud PO et al. Immature mesenchymal stem cell-like pericytes as mediators of immunosuppression in human malignant glioma. J Neuroimmunol 2013; 265: 106–116. 41 Keating A. Mesenchymal stromal cells: new directions. Cell Stem Cell 2012; 10: 709–716. 42 Le Blanc K, Mougiakakos D. Multipotent mesenchymal stromal cells and the innate immune system. Nat Rev Immunol 2012; 12: 383–396. 43 Nauta AJ, Fibbe WE. Immunomodulatory properties of mesenchymal stromal cells. Blood 2007; 110: 3499–3506. 44 Tolar J, Le Blanc K, Keating A, Blazar BR. Concise review: hitting the right spot with mesenchymal stromal cells. Stem Cells 2010; 28: 1446–1455. 45 Uccelli A, Moretta L, Pistoia V. Mesenchymal stem cells in health and disease. Nat Rev Immunol 2008; 8: 726–736. 46 Bartholomew A, Sturgeon C, Siatskas M, Ferrer K, McIntosh K, Patil S et al. Mesenchymal stem cells suppress lymphocyte proliferation in vitro and prolong skin graft survival in vivo. Exp Hematol 2002; 30: 42–48. 47 Di Nicola M, Carlo-Stella C, Magni M, Milanesi M, Longoni PD, Matteucci P et al. Human bone marrow stromal cells suppress T-lymphocyte proliferation induced by cellular or nonspecific mitogenic stimuli. Blood 2002; 99: 3838–3843. 48 von Bahr L, Sundberg B, Lonnies L, Sander B, Karbach H, Hagglund H et al. Long-term complications, immunologic effects, and role of passage for outcome in mesenchymal stromal cell therapy. Biol Blood Marrow Transplant 2012; 18: 557–564. 49 Selmani Z, Naji A, Gaiffe E, Obert L, Tiberghien P, Rouas-Freiss N et al. HLA-G is a crucial immunosuppressive molecule secreted by adult human mesenchymal stem cells. Transplantation 2009; 87(suppl 9): S62–S66. 50 Aggarwal S, Pittenger MF. Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood 2005; 105: 1815–1822. 51 Gieseke F, Bohringer J, Bussolari R, Dominici M, Handgretinger R, Muller I. Human multipotent mesenchymal stromal cells use galectin-1 to inhibit immune effector cells. Blood 2010; 116: 3770–3779. 52 Erkers T, Nava S, Yosef J, Ringden O, Kaipe H. Decidual stromal cells promote regulatory T cells and suppress alloreactivity in a cell contact-dependent manner. Stem Cells Dev 2013; 22: 2596–2605. 53 Krampera M, Cosmi L, Angeli R, Pasini A, Liotta F, Andreini A et al. Role for interferon-gamma in the immunomodulatory activity of human bone marrow mesenchymal stem cells. Stem Cells 2006; 24: 386–398. 54 Ren G, Zhang L, Zhao X, Xu G, Zhang Y, Roberts AI et al. Mesenchymal stem cellmediated immunosuppression occurs via concerted action of chemokines and nitric oxide. Cell Stem Cell 2008; 2: 141–150. 55 Opitz CA, Litzenburger UM, Lutz C, Lanz TV, Tritschler I, Koppel A et al. Toll-like receptor engagement enhances the immunosuppressive properties of human bone marrow-derived mesenchymal stem cells by inducing indoleamine-2,3-dioxygenase-1 via interferon-beta and protein kinase R. Stem Cells 2009; 27: 909–919. 56 Meisel R, Zibert A, Laryea M, Gobel U, Daubener W, Dilloo D. Human bone marrow stromal cells inhibit allogeneic T-cell responses by indoleamine 2,3dioxygenase-mediated tryptophan degradation. Blood 2004; 103: 4619–4621. 57 Ryan JM, Barry F, Murphy JM, Mahon BP. Interferon-gamma does not break, but promotes the immunosuppressive capacity of adult human mesenchymal stem cells. Clin Exp Immunol 2007; 149: 353–363. 58 Waterman RS, Tomchuck SL, Henkle SL, Betancourt AM. A new mesenchymal stem cell (MSC) paradigm: polarization into a pro-inflammatory MSC1 or an Immunosuppressive MSC2 phenotype. PLoS One 2010; 5: e10088. 59 Ge W, Jiang J, Arp J, Liu W, Garcia B, Wang H. Regulatory T-cell generation and kidney allograft tolerance induced by mesenchymal stem cells associated with indoleamine 2,3-dioxygenase expression. Transplantation 2010; 90: 1312–1320. 60 Tatara R, Ozaki K, Kikuchi Y, Hatanaka K, Oh I, Meguro A et al. Mesenchymal stromal cells inhibit Th17 but not regulatory T-cell differentiation. Cytotherapy 2011; 13: 686–694. 61 Francois M, Romieu-Mourez R, Li M, Galipeau J. Human MSC suppression correlates with cytokine induction of indoleamine 2,3-dioxygenase and bystander M2 macrophage differentiation. Mol Therapy 2012; 20: 187–195. 62 Augello A, Tasso R, Negrini SM, Amateis A, Indiveri F, Cancedda R et al. Bone marrow mesenchymal progenitor cells inhibit lymphocyte proliferation by activation of the programmed death 1 pathway. Eur J Immunol 2005; 35: 1482–1490.

Bone Marrow Transplantation (2014) 1 – 7

63 Akiyama K, Chen C, Wang D, Xu X, Qu C, Yamaza T et al. Mesenchymal-stem-cellinduced immunoregulation involves FAS-ligand-/FAS-mediated T cell apoptosis. Cell Stem Cell 2012; 10: 544–555. 64 Ren G, Zhao X, Zhang L, Zhang J, L’Huillier A, Ling W et al. Inflammatory cytokine-induced intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 in mesenchymal stem cells are critical for immunosuppression. J Immunol 2010; 184: 2321–2328. 65 Saldanha-Araujo F, Ferreira FI, Palma PV, Araujo AG, Queiroz RH, Covas DT et al. Mesenchymal stromal cells up-regulate CD39 and increase adenosine production to suppress activated T-lymphocytes. Stem cell Res 2011; 7: 66–74. 66 Ren G, Su J, Zhang L, Zhao X, Ling W, L’Huillie A et al. Species variation in the mechanisms of mesenchymal stem cell-mediated immunosuppression. Stem Cells 2009; 27: 1954–1962. 67 Min CK, Kim BG, Park G, Cho B, Oh IH. IL-10-transduced bone marrow mesenchymal stem cells can attenuate the severity of acute graft-versus-host disease after experimental allogeneic stem cell transplantation. Bone Marrow Transplant 2007; 39: 637–645. 68 Thery C, Ostrowski M, Segura E. Membrane vesicles as conveyors of immune responses. Nat Rev Immunol 2009; 9: 581–593. 69 Thery C, Ostrowski M, Segura E. Membrane vesicles as conveyors of immune responses. Nat Rev Immunol 2009; 9: 581–593. 70 Bruno S, Grange C, Collino F, Deregibus MC, Cantaluppi V, Biancone L et al. Microvesicles derived from mesenchymal stem cells enhance survival in a lethal model of acute kidney injury. PLoS One 2012; 7: e33115. 71 Lai RC, Arslan F, Lee MM, Sze NS, Choo A, Chen TS et al. Exosome secreted by MSC reduces myocardial ischemia/reperfusion injury. Stem Cell Res 2010; 4: 214–222. 72 Lee C, Mitsialis SA, Aslam M, Vitali SH, Vergadi E, Konstantinou G et al. Exosomes mediate the cytoprotective action of mesenchymal stromal cells on hypoxiainduced pulmonary hypertension. Circulation 2012; 126: 2601–2611. 73 Gatti S, Bruno S, Deregibus MC, Sordi A, Cantaluppi V, Tetta C et al. Microvesicles derived from human adult mesenchymal stem cells protect against ischaemiareperfusion-induced acute and chronic kidney injury. Nephrol Dial Transplant 2011; 26: 1474–1483. 74 Andre F, Chaput N, Schartz NEC, Flament C, Aubert N, Bernard J et al. Exosomes as potent cell-free peptide-based vaccine. I. Dendritic cell-derived exosomes transfer functional MHC class I/peptide complexes to dendritic cells. J Immunol 2004; 172: 2126–2136. 75 Bruno S, Collino F, Deregibus MC, Grange C, Tetta C, Camussi G. Microvesicles derived from human bone marrow mesenchymal stem cells inhibit tumor growth. Stem Cells Develop 2013; 22: 758–771. 76 Kordelas L, Ludwig S, Radke P, Beelen D, Giebel B. Successful treatment of therapy-refractory acute graft-versus-host disease with mesenchymal stem cellderived exosomes. Bone Marrow Transplant 2013; 48: s176–s177. 77 EL Andaloussi S, Mager I, Breakefield XO, Wood MJ. Extracellular vesicles: biology and emerging therapeutic opportunities. Nat Rev Drug Discov 2013; 12: 347–357. 78 Lai RC, Yeo RW, Tan KH, Lim SK. Exosomes for drug delivery – a novel application for the mesenchymal stem cell. Biotechnol Adv 2013; 31: 543–551. 79 Deeg HJ. How I treat refractory acute GVHD. Blood 2007; 109: 4119–4126. 80 Hahn T, McCarthy Jr. PL, Zhang MJ, Wang D, Arora M, Frangoul H et al. Risk factors for acute graft-versus-host disease after human leukocyte antigenidentical sibling transplants for adults with leukemia. J Clin Oncol 2008; 26: 5728–5734. 81 Ringden O, Nilsson B. Death by graft-versus-host disease associated with HLA mismatch, high recipient age, low marrow cell dose, and splenectomy. Transplantation 1985; 40: 39–44. 82 Le Blanc K, Frassoni F, Ball L, Locatelli F, Roelofs H, Lewis I et al. Mesenchymal stem cells for treatment of steroid-resistant, severe, acute graft-versus-host disease: a phase II study. Lancet 2008; 371: 1579–1586. 83 Ringden O. Mesenchymal stem cells for treatment and prevention of graftversus-host disease and graft failure after hematopoietic stem cell transplantation and future challenges. In: Chase LG, Vemuri MC (eds). Mesenchymal Stem Cell Therapy. Humana Press: Springer Verlag, pp 173–206, 2013. 84 Perez-Simon JA, Lopez-Villar O, Andreu EJ, Rifon J, Muntion S, Campelo MD et al. Mesenchymal stem cells expanded in vitro with human serum for the treatment of acute and chronic graft-versus-host disease: results of a phase I/II clinical trial. Haematologica 2011; 96: 1072–1076. 85 von Bonin M, Stolzel F, Goedecke A, Richter K, Wuschek N, Holig K et al. Treatment of refractory acute GVHD with third-party MSC expanded in platelet lysatecontaining medium. Bone Marrow Transplant 2009; 43: 245–251. 86 Kebriaei P, Isola L, Bahceci E, Holland K, Rowley S, McGuirk J et al. Adult human mesenchymal stem cells added to corticosteroid therapy for the treatment of acute graft-versus-host disease. Biol Blood Marrow Transplant 2009; 15: 804–811.

& 2014 Macmillan Publishers Limited

Stromal cells for GVHD? H Kaipe et al

7 87 Lucchini G, Introna M, Dander E, Rovelli A, Balduzzi A, Bonanomi S et al. Plateletlysate-expanded mesenchymal stromal cells as a salvage therapy for severe resistant graft-versus-host disease in a pediatric population. Biol Blood Marrow Transplant 2010; 16: 1293–1301. 88 Fang B, Song Y, Liao L, Zhang Y, Zhao RC. Favorable response to human adipose tissue-derived mesenchymal stem cells in steroid-refractory acute graft-versushost disease. Transplant Proc 2007; 39: 3358–3362. 89 Wu KH, Chan CK, Tsai C, Chang YH, Sieber M, Chiu TH et al. Effective treatment of severe steroid-resistant acute graft-versus-host disease with umbilical cordderived mesenchymal stem cells. Transplantation 2011; 91: 1412–1416. 90 Ringden O, Erkers T, Nava S, Uzunel M, Iwarsson E, Conrad R et al. Fetal membrane cells for treatment of steroid-refractory acute graft-versus-host disease. Stem Cells 2013; 31: 592–601. 91 Di Ianni M, Falzetti F, Carotti A, Terenzi A, Castellino F, Bonifacio E et al. Tregs prevent GVHD and promote immune reconstitution in HLA-haploidentical transplantation. Blood 2011; 117: 3921–3928. 92 Highfill SL, Rodriguez PC, Zhou Q, Goetz CA, Koehn BH, Veenstra R et al. Bone marrow myeloid-derived suppressor cells (MDSCs) inhibit graft-versus-host disease (GVHD) via an arginase-1-dependent mechanism that is up-regulated by interleukin-13. Blood 2010; 116: 5738–5747. 93 Zheng J, Liu Y, Liu M, Xiang Z, Lam KT, Lewis DB et al. Human CD8 þ regulatory T cells inhibit GVHD and preserve general immunity in humanized mice. Sci Transl Med 2013; 5: 168ra9. 94 Weng JY, Du X, Geng SX, Peng YW, Wang Z, Lu ZS et al. Mesenchymal stem cell as salvage treatment for refractory chronic GVHD. Bone Marrow Transplant 2010; 45: 1732–1740. 95 Lazarus HM, Koc ON, Devine SM, Curtin P, Maziarz RT, Holland HK et al. Cotransplantation of HLA-identical sibling culture-expanded mesenchymal stem cells and hematopoietic stem cells in hematologic malignancy patients. Biol Blood Marrow Transplant 2005; 11: 389–398. 96 Bernardo ME, Ball LM, Cometa AM, Roelofs H, Zecca M, Avanzini MA et al. Co-infusion of ex vivo-expanded, parental MSCs prevents life-threatening acute GVHD, but does not reduce the risk of graft failure in pediatric patients undergoing allogeneic umbilical cord blood transplantation. Bone Marrow Transplant 2011; 46: 200–207. 97 Ball LM, Bernardo ME, Roelofs H, Lankester A, Cometa A, Egeler RM et al. Cotransplantation of ex vivo expanded mesenchymal stem cells accelerates lymphocyte recovery and may reduce the risk of graft failure in haploidentical hematopoietic stem-cell transplantation. Blood 2007; 110: 2764–2767. 98 Liu K, Chen Y, Zeng Y, Xu L, Liu D, Chen H et al. Coinfusion of mesenchymal stromal cells facilitates platelet recovery without increasing leukemia recurrence in haploidentical hematopoietic stem cell transplantation: a randomized, controlled clinical study. Stem Cells Dev 2011; 20: 1679–1685. 99 Kuzmina LA, Petinati NA, Parovichnikova EN, Lubimova LS, Gribanova EO, Gaponova TV et al. Multipotent mesenchymal stromal cells for the prophylaxis of

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105

106

107

108

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acute graft-versus-host disease-a phase II study. Stem Cells Int 2012; 2012: 968213. Ball LM, Bernardo ME, Roelofs H, van Tol MJ, Contoli B, Zwaginga JJ et al. Multiple infusions of mesenchymal stromal cells induce sustained remission in children with steroid-refractory, grade III-IV acute graft-versus-host disease. Br J Haematol 2013; 163: 501–509. Ringden O, Keating A. Mesenchymal stromal cells as treatment for chronic GVHD. Bone Marrow Transplant 2011; 46: 163–164. Ringden O, Leblanc K. Pooled MSCs for treatment of severe hemorrhage. Bone Marrow Transplant 2011; 46: 1158–1160. Ringden O, Uzunel M, Sundberg B, Lonnies L, Nava S, Gustafsson J et al. Tissue repair using allogeneic mesenchymal stem cells for hemorrhagic cystitis, pneumomediastinum and perforated colon. Leukemia 2007; 21: 2271–2276. Jung JW, Kwon M, Choi JC, Shin JW, Park IW, Choi BW et al. Familial occurrence of pulmonary embolism after intravenous, adipose tissue-derived stem cell therapy. Yonsei Med J 2013; 54: 1293–1296. Chauhan D, Uchiyama H, Akbarali Y, Urashima M, Yamamoto K, Libermann TA et al. Multiple myeloma cell adhesion-induced interleukin-6 expression in bone marrow stromal cells involves activation of NF-kappa B. Blood 1996; 87: 1104–1112. Miura M, Miura Y, Padilla-Nash HM, Molinolo AA, Fu B, Patel V et al. Accumulated chromosomal instability in murine bone marrow mesenchymal stem cells leads to malignant transformation. Stem Cells 2006; 24: 1095–1103. Tarte K, Gaillard J, Lataillade JJ, Fouillard L, Becker M, Mossafa H et al. Clinicalgrade production of human mesenchymal stromal cells: occurrence of aneuploidy without transformation. Blood 2010; 115: 1549–1553. Tolar J, Nauta AJ, Osborn MJ, Panoskaltsis Mortari A, McElmurry RT, Bell S et al. Sarcoma derived from cultured mesenchymal stem cells. Stem Cells 2007; 25: 371 9. Paliogianni F, Boumpas DT. Prostaglandin E2 inhibits the nuclear transcription of the human interleukin 2, but not the Il-4, gene in human T cells by targeting transcription factors AP-1 and NF-AT. Cell Immunol 1996; 171: 95–101. Sato K, Ozaki K, Oh I, Meguro A, Hatanaka K, Nagai T et al. Nitric oxide plays a critical role in suppression of T-cell proliferation by mesenchymal stem cells. Blood 2007; 109: 228–234. Zhang HC, Liu XB, Huang S, Bi XY, Wang HX, Xie LX et al. Microvesicles derived from human umbilical cord mesenchymal stem cells stimulated by hypoxia promote angiogenesis both in vitro and in vivo. Stem Cells Dev 2012; 21: 3289–3297. Bruno S, Grange C, Collino F, Deregibus MC, Cantaluppi V, Biancone L et al. Microvesicles derived from mesenchymal stem cells enhance survival in a lethal model of acute kidney injury. PLoS One 2012; 7: e33115. Ciceri F, Bonini C, Stanghellini MT, Bondanza A, Traversari C, Salomoni M et al. Infusion of suicide-gene-engineered donor lymphocytes after family haploidentical haemopoietic stem-cell transplantation for leukaemia (the TK007 trial): a non-randomised phase I-II study. Lancet Oncol 2009; 10: 489–500.

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