Infusion of Mesenchymal Stromal Cells can Aid Hematopoietic Recovery Following Allogeneic Hematopoietic Stem Cell Myeloablative Transplant: A Pilot Study

Abstract

Mesenchymal stromal cells (MSCs) are important in the support of hematopoiesis. In this pilot study, we evaluated the safety and efficiency of donor-expanded MSC infusion after allogeneic hematopoietic stem cell transplantation (HSCT) in six patients with poor hematopoietic recovery. MSCs were infused without HSC and without conditioning at a dose of 1 × 10⁶/kg weight. Two patients displayed rapid hematopoietic recovery (days 12 and 21), and four patients showed no response. The two patients who showed hematopoietic recovery were in first complete remission (CR1) compared to the other heavily pretreated patients. There were no toxic side effects linked to MSC infusion. One patient developed cytomegalovirus (CMV) reactivation 12 days following the MSC infusion and died from CMV disease. We found that infusion of MSCs without HSC co-infusion can restore medullary function in some patients with poor hematopoietic recovery. Our data suggest that patients with a less damaged stroma could benefit from this approach.

Introduction

Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative approach for different hematological diseases [1]. Despite progress in the prevention and treatment of transplantation side effects, there is still a high rate of transplant mortality (TRM). The most life-threatening complications after allogeneic HSCT are graft versus host disease and infections. Poor hematological recovery or nonengraftment is also a serious complication that prolongs the duration of hospitalization and increases serious infectious complications, hemorrhagic risk, and TRM [2,3]. Expanded Mesenchymal stromal cells (MSCs) are characterized by their potential for differentiation into several tissues (eg, mesodermal, endodermal, or ectodermal lineages) [4,5], their phenotype (ie, CD34−, CD45−, HLADR−, CD29+, CD73+, CD90+, CD105+, and CD166+), their immunomodulatory capacities [6 –9], and their ability to support hematopoiesis [10]. In vivo phase I and II studies have stimulated interest in expanded MSCs for the treatment of acute graft versus host disease (GVHD) [11,12]. A recent report has shown that MSCs infusion could additionally repair therapy-induced tissue toxicity after HSCT such as hemorrhagic cystitis [13]. It has also been reported that MSC infusions can enhance hematopoietic recovery. Koç et al. described that co-infusion of HSCs and MSCs in patients undergoing autologous HSCT for breast cancer was safe and associated with a prompt hematopoietic recovery [14]. Le Blanc et al. reported that co-infusion of MSCs and HSCs in patients with graft failure or poor engraftment could enhance hematopoietic recovery [15].

We report herein the results of our pilot study that evaluated the benefit of infusion of donor MSCs expanded ex vivo without HSC co-infusion in patients with poor hematopoietic recovery after allogeneic HSC myeloablative transplant. The first objective of the study was to evaluate the safety of infusion of donor MSCs amplified ex vivo after allogeneic transplantation. The second goal was to evaluate the improvement of hematopoietic recovery in patients with poor engraftment after an allogeneic SCT.

Patients and Methods

Between January 2003 and July 2006, six patients were enrolled in the pilot study. The clinical protocol and consent form were both approved by the Institutional Review Board. Patients and donors signed written informed consent forms. The inclusion criteria were poor hematological recovery after allogeneic HSCT, defined as a persistent (>30 days post-transplantation) pancytopenia with a platelet level <50 × 10⁹/L and an absolute neutrophil count (ANC) <1.0 × 10⁹/L despite treatment of minimum 10 days with recombinant human-granulocyte colony-stimulating factor (G-CSF). Patients were required to have nonprogressive disease, no GVHD, no active infection or other causes of hypoplasia. Patients with graft failure or graft rejection—according to donor chimerism—were excluded since we infused MSCs without concomitant HSC infusion.

Patients

Six patients received MSCs according to the protocol. In three cases, the transplant was performed from an HLA-identical-related donor and, in three others, from a haploidentical-related donor. All patients had full donor chimerism before the MSC infusion and a marrow aspiration showing hypoplasia (<10% cellularity). All patients were in remission at the time of MSC infusion.

Patient and transplantation characteristics are reported in Table 1.

Table 1.

Characteristics of Patients

Patients	Age/gender	Diseases/status	Donor	CD34 × 106/kg	Second infusion × 106/kg	Conditioning	MSC × 106/kg	Day infusion after transplantation
1	47/M	CML/h–CR1	HLA id sib	3	3.1	Bu–Cy	1	94
2	21/M	ALL/CR1	HLA id Sib	4.8	No	Cy–TBI	1	158
3	26/M	MDS	Haplo	3.4	No	Mel–Cy–Flu–TBI–ATG	1	50
4	48/M	CML/CR3	Haplo	3.4	3.4	Mel–Cy–Flu–TBI–ATG	1	161
5	24/M	AML/CR2	Haplo	2.1	No	Mel–Cy–Flu–TBI–ATG	1	184
6	27/M	AL L/CR5	HLA id sib	5	No	Cy–TBI	1	295

Abbreviations: ALL, acute lymphoblastic leukemia; AML, acute myeloid leukemia; ATG, anti-thymoglobulin; Bu, busulfan; CML, chronic myeloid leukemia; CR, complete remission; Cy, cyclophosphamide; Flu, fludarabine; haplo, haploidentical-related donor; HLA id sib, HLA-identical sibling; MDS, myelodysplastic syndrome; Mel, melphalan; MSCs, mesenchymal stromal cells; TBI, total body irradiation.

Conditioning regimen and allogeneic HSC infusion

The conditioning regimen for HLA-identical-related transplantation was cyclophosphamide (60 mg/kg/day for days 9 and 8) combined with total body irradiation (TBI) of 2 Gy (days 7 to 2) (n = 2) or busulfan (4 mg/kg/day) in four doses (days 8 to 5) and cyclophosphamide (60 mg/kg/day on days 3 and 2) for one patient. The conditioning regimen for the three haploidentical-related transplants consisted of melphalan (60 mg/m²/day on days 9 and 8), TBI (5 × 2 Gy on days 7 to 3), fludarabine (40 mg/m²/day for days 7 to 3), and antithymocyte globulin (ATG) fresenius (5 mg/kg/day on days 7 to 3) [16]. In all cases, the source of HSCs was peripheral blood stem cells (PBSCs) harvested by a standard apheresis procedure. Doses of PBSCs infused are reported in the Table 1. Prophylaxis for GVHD was cyclosporine combined with MTX injection for related transplantation. In haploidentical transplantation, PBSCs were T- and B-cells depleted, and patients received ATG in the conditioning regimen and cyclosporine in the pretransplantation period [16]. Five patients had grade II–IV GVHD, who required high doses of steroids before MSC infusion, and three of these patients were still on steroids at the time of MSC infusion.

Engraftment and chimerism at the time of MSC infusion

At the time of MSC infusion, two patients had a platelet level between 20 and 40 × 10⁹/L, requiring sporadic platelet transfusions and repeated red blood cell transfusions. Four patients were dependent on platelet (<20 × 10⁹/L) and red blood cell transfusions. All the patients had ANC below 1.0 × 10⁹/L with G-CSF stimulation. The median duration of G-CSF administration was 36 days (18–102).

Infectious or toxic causes of aplasia were excluded; patients had no active or treated GVHD; and all patients were in remission at the time of transplantation. Chimerism status in all patients showed 100% donor chimerism. The CD3 and CD33 chimerism was performed by real-time PCR.

Isolation and ex vivo expansion of MSCs

MSCs were derived from the bone marrow of HSC donors. A minimum of 40 mL of BM was collected with informed consent. Bone marrow mononuclear (BM-MNC) cells were isolated by layering on a density gradient (LinfoSep, Biomedics, Madrid, Spain) and washed in HBSS (Cambrex, Walkersville, MD, USA) medium. Cells were suspended in Ultra-culture medium (Cambrex) supplemented with 2% of serum substitute (Ultroser G, Pall Bio sepra, Cergy-saint-Christophe, France), 2 mM l-Glutamine (Cambrex), and 0.5% antibiotic–antimycotic solution (GibcoBRL, Grand Island, NY, USA) [17]. Cells were plated in a 75-cm² flask and incubated at 37° with 5% humidified CO₂. After 48 h, nonadherent cells were detached by treatment with TrypLE Select (Gibco) and adequate fresh medium was added. The medium was removed two times per week. When confluence was reached, cells were harvested and replated in the same medium at 1,000 cells/cm². Cells were harvested after two or three passages to obtain 1 × 10⁶ expanded cells/kg of recipient body weight. MSCs were characterized by flow cytometry analysis in order to establish their purity. The cells failed to express hematopoietic (CD14, CD34, and CD45) or endothelial markers (CD31) and expressed CD73, CD90, CD105, CD29, and CD166 antigens (>90%). To study their multilineage potential, MSCs were cultured in appropriate induction medium to assess adipogenic, osteogenic, and chondrogenic differentiation as previously described [18]. MSC suspensions were tested for viability and infection (eg, bacterial, mycotic, or mycoplasma contamination) before infusion.

MSC Infusion

MSCs were intravenously infused through a central venous catheter at the dose of 1 × 10⁶ cells/kg of recipient body weight over a period of 15 min. The patients were closely monitored during and after MSC administration, and we did not observed any immediate side effects or modifications in clinical parameters (eg, temperature, blood pressure, and heart rate).

Evaluation of engraftment and infection monitoring

Hematological parameters were evaluated a minimum of three times per week. Patients were monitored by polymerase chain reaction (PCR) for cytomegalovirus (CMV) reactivation before MSC infusion and twice a week until hematopoietic recovery or 3 months after MSC infusion, and then once a week for three consecutive months or during GVHD treatment. Presence of Aspergillus galactomannan antigen in the blood was evaluated twice a week during aplasia and once a week during treatment for GVHD.

Results

All patients received 1 × 10⁶ MSCs/kg. The median time between HSCT and the MSC infusion was 159.5 (50–295) days.

Immediate side effects

No immediate side effects were observed after MSC infusion, and there were no significant modifications in heart rate, blood pressure, or temperature during the first 12 h following the injection. We did not observe any skin reactions after MSC infusion.

Early infection (≤30 days after MSC infusion)

We observed one early CMV reactivation by PCR. In patient number 3, CMV PCR became positive 12 days after the MSC infusion and 62 days after the HSC transplantation (Table 2). This patient received a haploidentical-related HSC transplantation (CMV+/+). Before MSC infusion, he did not have GVHD, corticosteroid treatment, or previous CMV reactivation; CMV PCR was negative at the time of infusion. A preemptive treatment with ganciclovir was initiated. Fifteen days after the start of the treatment, however, and after an initial response in terms of PCR level, we observed an increase in the CMV PCR value. Thus, treatment was switched to foscavir for 2 weeks, and the PCR finally became negative. At days 77 and 102 after MSC infusion, the patient again required preemptive treatment with foscavir for CMV reactivation. Unfortunately, during this reactivation, the patient became resistant to foscavir treatment and developed a CMV disease (retinitis and encephalitis). In spite of bitherapy followed by cidofovir treatment, the patient died from CNS CMV disease.

Table 2.

Infectious Complications After MSC Infusion (Days 0–100)

Patients	CMVD/R	CMV PCR at MSC infusion	Viral PCR reactivation (day)	CMVD	Aspergillosis (day) RF
1	N/P	Neg	No	No	No
2	P/P	Neg	No	No	No
3	P/P	Neg	12	Encephalitis	No
4	P/P	Neg	No	No	Pulmonary (72) GVHD–HDS–Neutropenia
5	N/P	Neg	No	No	No
6	P/P	Neg	No	No	Pulmonary (84) GVHD–HDS– Neutropenia

Abbreviations: CMV, cytomegalovirus; CMVD, cytomegalovirus disease; D/R, donor/recipient; GVHD, graft versus host disease; HDS, high-dose steroid; MSC, mesenchymal stromal cell; RF, risk factor.

Late infection (1–3 months after MSC infusion)

One patient (number 6) had an EBV PCR reactivation on day 58. Preemptive treatment was started with two doses of rituximab (375 mg/m²), and the PCR remained negative for the rest of the follow-up. Two patients developed pulmonary aspergillosis (days 72 and 84) associated with prolonged neutropenia (>30 days) and with the development of GVHD requiring steroid treatment (at days 30 and 42 of MSCs infusion).

Late toxic side effects

We did not observe any late toxic effects.

Engraftment

Patient number 1 showed a normalization of platelet level and neutrophil count and an increase in reticulocytosis at 12 days after MSC infusion (Table 3). In brief, this patient was treated with an HLA-identical-related hematopoietic SCT for a chronic myeloid leukemia in partial cytogenetic remission after 7 months of treatment with imatinib. The patient had a poor hematological recovery 1 month after transplantation, with ANC varying between 0.5 and 0.1 × 10⁹/L without platelet recovery (<20 × 10⁹/L). On day 87, the patient received a second PBSC infusion and showed a slight increase in platelet level (20 to 40 × 10⁹/L). Fifty days after this second infusion, there was still no hematopoietic recovery, and the patient received expanded MSCs from his HSC donor.

Table 3.

Time to Hematopoietic Recovery for Patients 1 and 2

	Days to ANC >1,000 × 109/L	Days to platelets >50,000 × 109/L	Days to platelets >100,000 × 109/L	Transfusion needs after MSC infusion	Engraftment evolution
Patient 1	5	12	20	Plt: 1	Normal hematopoietic function at +4.8 years
Patient 1	5	12	20	RBC: 7U	Normal hematopoietic function at +4.8 years
Patient 2	15	21	30	Plt: 0	Normal hematopoietic function until relapse at +6 months
Patient 2	15	21	30	RBC: 2U	Normal hematopoietic function until relapse at +6 months

Abbreviations: ANC, absolute neutrophil counts; MSC, mesenchymal stromal cell; Plt, platelet; RBC, red blood cells.

Patient number 2 also showed normalization of hematopoietic function at 21 days after MSC infusion (Table 3). This patient was treated with an HLA-identical sibling-related PBSC transplantation for a Philadelphia-negative ALL in first complete remission (CR1). Five months after transplantation, the patient had no hematopoietic recovery and still required repeated red cell transfusions and had a maximum platelet level of 40 × 10⁹/L with occasional transfusion (maximum once weekly). At this time, the patient received an infusion of 1 × 10⁶/kg MSCs expanded from the bone marrow of his HSC donor. On day 21, we observed hematopoietic recovery. Six months after the MSC infusion, we observed a new degradation in hematopoietic function, indicating a relapse of the disease. The patient died 15 months after HSC transplantation due to ALL progression.

The other patients failed to show any significant hematologic improvement.

Discussion

Poor engraftment to various degrees is an infrequent but life-threatening side effect following HSC transplantation. A possible approach to this complication is the infusion of additional stem cells, but this procedure is associated with significant risk of GVHD [19]. In this study, we evaluated infusion of MSCs in six patients who had poor hematologic engraftment. The basis for this study was the known role of MSCs in supporting hematopoiesis. Previous studies on animal models have reported that human MSCs facilitate hematopoietic engraftment [20,21].

Different pathways might explain the action of MSCs in hematopoietic function. MSCs are precursors of a majority of the constituents of the cellular microenvironment, which regulates hematopoiesis, and they could reconstitute the stroma altered by previous chemotherapy or radiotherapy. MSCs are also a source of several hematopoietic cytokines that promote self-renewal of HSCs and their differentiation [22]. In addition, the immunosuppressive properties of HSCs might promote engraftment. All of our patients had a partial but poor hematopoietic recovery with full donor chimerism; thus, we postulated that there was residual hematopoietic tissue and that the poor hematopoietic recovery could be explained by a stromal impairment. Many studies have indicated that the bone marrow stroma is damaged following transplantation, and its ability to support hematopoiesis might be impaired [23 –25]. In our study, MSCs were infused after HSC transplantation (50–294 days); thus, they were not given to enhance the engraftment of HSCs but to stimulate “residual” hematopoiesis and improve the function of the bone marrow microenvironment. Indeed, Muguruma et al. observed reconstitution of a functional human hematopoietic microenvironment derived from human MSCs in the murine bone marrow compartment [26]. Several studies have reported that human MSCs remain of host origin after HSCT, but whether culture-expanded MSCs result in detectable stromal chimerism in humans has not yet been fully established [27 –31]. In our study, only two patients recovered normal hematopoietic function following MSC infusion. One possible reason could be the number of MSCs infused. At present, there are still no data demonstrating the optimal dose of MSCs to obtain significant clinical responses. In the setting of allogeneic transplantation, different studies have reported on MSC infusion either concomitantly with HSC to improve hematopoietic recovery or alone to treat acute GVHD [11,12,15,32 –34]. In these reports, the dose of MSCs infused varied from 0.7 to 9 × 10⁶ cells/kg. In our study, we arbitrarily chose to administer 1 × 10⁶ cells/kg body weight to all of our patients. This level may be insufficient to restore a functional microenvironment. The two patients who recovered normal hematologic function were in their CR1, in contrast to the other patients who were heavily pretreated prior to HSC transplantation and likely had more severely impaired stromal function. The rapid hematopoietic response and the fact that the two patients with hematopoietic recovery were in CR1 could suggest the restoration of cytokine release by stromal cells required for HSC function rather than complete restoration of the hematopoietic niche. The absence of hematopoietic recovery in some patients could be due to the absence of sufficient residual hematopoietic tissue, suggesting that in those cases a co-infusion of HSC and MSCs might be the solution, as in the study of Le Blanc et al. [15]. This is supported by the fact that the patients with hematopoietic recovery had a lower transfusion need compared to patients who showed no hematological improvement following MSC infusion.

We also observed serious infectious complications. Some patients had late infectious complications, which were probably due to different parameters (eg, prolonged neutropenia, active GVHD, or prolonged corticosteroid treatment) and not primarily related to the MSC infusion. One of our patients, however, developed an early CMV disease and died from this complication. MSCs are well known for their immunosuppressive properties and are used to treat GVHD [11–12]. Their immunosuppressive activities may also inhibit responses to infectious complications, which are frequent in the setting of allogeneic transplantation. Sundin et al. reported that CMV could infect MSCs and could suppress lymphocyte responses to the infection [35]. In contrast to this study, Karlsson et al. suggested in their report that MSCs did not suppress virus-specific T-cell function in vitro [36]. Nevertheless, more data are needed to understand the immunosuppressive effects of MSCs on the response to active infections. Our patient was at day 62 following a haploidentical-related transplantation and was thus strongly immunosuppressed and it is impossible to assess the role of MSCs in this serious viral complication. It has also been reported that MSCs support tumor growth due to their immunosuppressive properties and by suppressing GVHD and the graft versus tumor effect in hematological malignancies [37 –39]. A recent study suggests that co-infusion of HSCs and MSCs after HLA-identical sibling-matched transplantation may increase the rate of relapse. The number of patients in the arm treated with MSCs (n = 10) was low, and these data still need to be confirmed in a larger study [39]. Our patient series was too small to draw any conclusions concerning the potential increase in relapse rate. Finally, we did not observe any toxic side effects that were potentially due to the MSC infusion.

Conclusions

Our study suggests that infusion of MSCs alone, without conditioning, is feasible in patients with poor medullary function and could help with hematologic recovery in some patients particularly in those with residual hematopoietic function. Patients with profound pancytopenia who are heavily pretreated and have a deeply damaged microenvironment may benefit from higher doses of MSCs or co-infusion of MSCs and HSC. Although it is still a subject of debate, MSCs may increase the risk of severe infections and patients should be closely monitored for infectious complications.

Footnotes

Author Disclosure Statement

The authors declare no competing financial interest.

References

Copelan

. (2006). Hematopoietic stem-cell transplantation. N Engl J Med, 354:1813–1826.

Davies

, Kollman

, Anasetti

, Antin

, Gajewski

, Casper

, Nademanee

, Noreen

, King

, Confer

and Kernan

. (2000). Engraftment and survival after unrelated-donor bone marrow transplantation: a report from the national marrow donor program. Blood, 96:4096–4102.

Dominietto

, Lamparelli

, Raiola

, Van Lint

, Gualandi

, Berisso

, Bregante

, Di Grazia

, Soracco

, Pitto

, Frassoni

and Bacigalupo

. (2002). Transplant-related mortality and long-term graft function are significantly influenced by cell dose in patients undergoing allogeneic marrow transplantation. Blood, 100:3930–3934.

Pittenger

, Mackay

, Beck

, Jaiswal

, Douglas

, Mosca

, Moorman

, Simonetti

, Craig

and Marshak

. (1999). Multilineage potential of adult human mesenchymal stem cells. Science, 284:143–147.

Jiang

, Jahagirdar

, Reinhardt

, Schwartz

, Keene

, Ortiz-Gonzalez

, Reyes

, Lenvik

, Lund

, Blackstad

, Du

, Aldrich

, Lisberg

, Low

, Largaespada

and Verfaillie

. (2002). Pluripotency of mesenchymal stem cells derived from adult marrow. Nature, 418:41–49.

Aggarwal

and Pittenger

. (2005). Human mesenchymal stem cells modulate allogeneic immune cell responses. Blood, 105:1815–1822.

Le Blanc

, Tammik

, Rosendahl

, Zetterberg

and Ringdén

. (2003). HLA expression and immunologic properties of differentiated and undifferentiated mesenchymal stem cells. Exp Hematol, 31:890–896.

Blanc K

, Tammik

, Sundberg

, Haynesworth

and Ringdén

. (2003). Mesenchymal stem cells inhibit and stimulate mixed lymphocyte cultures and mitogenic responses independently of the major histocompatibility complex. Scand J Immunol, 57:11–20.

Nauta

and Fibbe

. (2007). Immunomodulatory properties of mesenchymal stromal cells. Blood, 110:3499–3506.

10.

Maitra

, Szekely

, Gjini

, Laughlin

, Dennis

, Haynesworth

and Koç

. (2004). Human mesenchymal stem cells support unrelated donor hematopoietic stem cells and suppress T-cell activation. Bone Marrow Transplant, 33:597–604.

11.

Blanc K

, Frassoni

, Ball

, Locatelli

, Roelofs

, Lewis

, Lanino

, Sundberg

, Bernardo

, Remberger

, Dini

, Egeler

, Bacigalupo

, Fibbe

, Ringdén

; Developmental Committee of the European Group for Blood and Marrow Transplantation. (2008). Mesenchymal stem cells for treatment of steroid-resistant, severe, acute graft-versus-host disease: a phase II study. Lancet, 371:1579–1586.

12.

Ringdén

, Uzunel

, Rasmusson

, Remberger

, Sundberg

, Lönnies

, HU

Marschall

, Dlugosz

, Szakos

, Hassan

, Omazic

, Aschan

, Barkholt

and Le

Blanc K

. (2006). Mesenchymal stem cells for treatment of therapy-resistant graft-versus-host disease. Transplantation, 81:1390–1397.

13.

Ringdén

, Uzunel

, Sundberg

, Lönnies

, S

Nava

, Gustafsson

, Henningsohn

and Le Blanc

. (2007). Tissue repair using allogeneic mesenchymal stem cells for hemorrhagic cystitis, pneumomediastinum and perforated colon. Leukemia, 21:2271–2276.

14.

Koç

, Gerson

, Cooper

, Dyhouse

, Haynesworth

, Caplan

and Lazarus

. (2000). Rapid hematopoietic recovery after coinfusion of autologous-blood stem cells and culture-expanded marrow mesenchymal stem cells in advanced breast cancer patients receiving high-dose chemotherapy. J Clin Oncol, 18:307–316.

15.

Blanc K

, Samuelsson

, Gustafsson

, Remberger

, Sundberg

, Arvidson

, Ljungman

, Lönnies

, S

Nava

and Ringdén

. (2007). Transplantation of mesenchymal stem cells to enhance engraftment of hematopoietic stem cells. Leukemia, 21:1733–1738.

16.

Lewalle

, Delforge

, Aoun

, Crombez

, de Wilde

, Theunissen

, Lagneaux

, Nowak

, Misplon

, Bron

and Martiat

. (2004). Growth factors and DLI in adult haploidentical transplant: a three-step pilot study towards patient and disease status adjusted management. Blood Cells Mol Dis, 33:256–260.

17.

Meuleman

, Tondreau

, Delforge

, Dejeneffe

, Massy

, Libertalis

, Bron

and Lagneaux

. (2006). Human marrow mesenchymal stem cell culture: serum-free medium allows better expansion than classical alpha-MEM medium. Eur J Haematol, 76:309–316.

18.

Tondreau

, Lagneaux

, Dejeneffe

, Delforge

, Massy

, Mortier

and Bron

. (2004). Isolation of BM mesenchymal stems cells by plastic adhesion or negative selection: phenotype, proliferation kinetics and differentiation potential. Cytotherapy, 6:372–379.

19.

Remberger

, Ringdén

, Ljungman

, Hägglund

, Winiarski

, Lönnqvist

and Aschan

. (1998). Booster marrow or blood cells for graft failure after allogeneic bone marrow transplantation. Bone Marrow Transplant, 22:73–78.

20.

Almeida-Porada

, Porada

, Tran

and Zanjani

. (2000). Cotransplantation of human stromal cell progenitors into preimmune fetal sheep results in early appearance of human donor cells in circulation and boosts cell levels in bone marrow at later time points after transplantation. Blood, 95:3620–3627.

21.

Noort

, Kruisselbrink

, in’t Anker

, Kruger

, van Bezooijen

, de Paus

, Heemskerk

, CW

Löwik

, Falkenburg

, Willemze

and Fibbe

. (2002). Mesenchymal stem cells promote engraftment of human umbilical cord blood-derived CD34(+) cells in NOD/SCID mice. Exp Hematol, 30:870–878.

22.

Dazzi

, Ramasamy

, Glennie

, Jones

and Roberts

. (2006). The role of mesenchymal stem cells in haemopoiesis. Blood Rev, 20:161–171.

23.

Galotto

, Berisso

, Delfino

, Podesta

, Ottaggio

, Dallorso

, Dufour

, Ferrara

, Abbondandolo

, Dini

, Bacigalupo

, Cancedda

and Quarto

. (1999). Stromal damage as consequence of high-dose chemo/radiotherapy in bone marrow transplant recipients. Exp Hematol, 27:1460–1466.

24.

O’Flaherty

, Sparrow

and Szer

. (1995). Bone marrow stromal function from patients after bone marrow transplantation. Bone Marrow Transplant, 15:207–212.

25.

McMannus

and Weiss

. (1984). Busulfan-induced chronic bone marrow failure: changes in cortical bone, marrow stromal cells, and adherent cell colonies. Blood, 64:1036–1041.

26.

Muguruma

, Yahata

, Miyatake

, Sato

, Uno

, Itoh

, Kato

, Ito

, Hotta

and Ando

. (2006). Reconstitution of the functional human hematopoietic microenvironment derived from human mesenchymal stem cells in the murine bone marrow compartment. Blood, 107:1878–1887.

27.

Koç

, Peters

, Aubourg

, Raghavan

, Dyhouse

, DeGasperi

, Kolodny

, Yoseph

, Gerson

, Lazarus

, Caplan

, Watkins

and Krivit

. (1999). Bone marrow-derived mesenchymal stem cells remain host-derived despite successful hematopoietic engraftment after allogeneic transplantation in patients with lysosomal and peroxisomal storage diseases. Exp Hematol, 27:1675–1681.

28.

Rieger

, Marinets

, Fietz

, Körper

, D

Sommer

, Mücke

, B

Reufi

, Blau

, Thiel

and Knauf

. (2005). Mesenchymal stem cells remain of host origin even a long time after allogeneic peripheral blood stem cell or bone marrow transplantation. Exp Hematol, 33:605–611.

29.

Stute

, Fehse

, Schröder

, S

Arps

, Adamietz

, Held

and Zander

. (2002). Human mesenchymal stem cells are not of donor origin in patients with severe aplastic anemia who underwent sex-mismatched allogeneic bone marrow transplant. J Hematother Stem Cell Res, 11:977–984.

30.

Wang

, Liu

and Lu

. (2005). Mesenchymal stem cells in stem cell transplant recipients are damaged and remain of host origin. Int J Hematol, 82:152–158.

31.

Villaron

, Almeida

, López-Holgado

, M

Alcoceba

, Sánchez-Abarca

, Sanchez-Guijo

, Alberca

, JA

Pérez-Simon

, JF

San Miguel

, MC

Del Cañizo

. (2004). Mesenchymal stem cells are present in peripheral blood and can engraft after allogeneic hematopoietic stem cell transplantation. Haematologica, 89:1421–1427.

32.

Lee

, Jang

, Cheong

, Kim

, Maemg

, Hahn

, Ko

and Min

. (2002). Treatment of high-risk acute myelogenous leukaemia by myeloablative chemoradiotherapy followed by co-infusion of T cell-depleted haematopoietic stem cells and culture-expanded marrow mesenchymal stem cells from a related donor with one fully mismatched human leucocyte antigen haplotype. Br J Haematol, 118:1128–1131.

33.

Lazarus

, Koç

, SM

Devine

, Curtin

, Maziarz

, Holland

, Shpall

, McCarthy

, Atkinson

, Cooper

, Gerson

, Laughlin

, FR

Loberiza

Jr , Moseley

and Bacigalupo

. (2005). Cotransplantation of HLA-identical sibling culture-expanded mesenchymal stem cells and hematopoietic stem cells in hematologic malignancy patients. Biol Blood Marrow Transplant, 11:389–398.

34.

Macmillan

, Blazar

, Defor

and Wagner

. (2008). Transplantation of ex-vivo culture-expanded parental haploidentical mesenchymal stem cells to promote engraftment in pediatric recipients of unrelated donor umbilical cord blood: results of a phase I-II clinical trial. Bone Marrow Transplant, 43:447–454.

35.

Sundin

, Orvell

, Rasmusson

, Sundberg

, Ringdén

and Le Blanc

. (2006). Mesenchymal stem cells are susceptible to human herpes viruses, but viral DNA cannot be detected in the healthy seropositive individual. Bone Marrow Transplant, 37:1051–1059.

36.

Karlsson

, Samarasinghe

, Ball

, Sundberg

, Lankester

, Dazzi

, Uzunel

, Rao

, Veys

, Le Blanc

, Ringdén

and Amrolia

. (2008). Mesenchymal stem cells exert differential effects on alloantigen and virus-specific T-cell responses. Blood, 112:532–541.

37.

Djouad

, Plence

, Bony

, Tropel

, Apparailly

, Sany

, Noël

and Jorgensen

. (2003). Immunosuppressive effect of mesenchymal stem cells favors tumor growth in allogeneic animals. Blood, 102:3837–3844.

38.

Zhu

, Xu

, Jiang

, Qian

, Chen

, Hu

, Cao

, Han

and Chen

. (2006). Mesenchymal stem cells derived from bone marrow favor tumor cell growth in vivo. Exp Mol Pathol, 80:267–274.

39.

Ning

, Yang

, Jiang

, Hu

, Feng

, Zhang

, Yu

, Li

, Xu

, Li

, Wang

, Hu

, Lou

and Chen

. (2008). The correlation between cotransplantation of mesenchymal stem cells and higher recurrence rate in hematologic malignancy patients: outcome of a pilot clinical study. Leukemia, 22:593–599.