Cytotherapy, 2014; 16: 1629e1642

Stem cells in animal asthma models: a systematic review

NADIM SROUR1,2,3,4,5 & BERNARD TH�EBAUD6,7,8

1Universit�e de Sherbrooke, Facult�e de M�edecine et des Sciences de la Sant�e, Department of Medicine, Division ofPulmonology, Sherbrooke, Canada, 2Hôpital Charles-LeMoyne, Department of Medicine, Division of Pulmonology,Montreal, Canada, 3McGill University, Department of Medicine, Montreal, Canada, 4Mount Sinai Hospital Centre,Montreal, Canada, 5The Ottawa Hospital Research Institute, Clinical Epidemiology Program, Ottawa, Canada, 6TheOttawa Hospital Research Institute, Regenerative Medicine Program, Ottawa, Canada, 7Children’s Hospital of EasternOntario, Ottawa, Canada, and 8The University of Ottawa, Faculty of Medicine, Ottawa, Canada

AbstractBackground aims. Asthma control frequently falls short of the goals set in international guidelines. Treatment options forpatients with poorly controlled asthma despite inhaled corticosteroids and long-acting b-agonists are limited, and newtherapeutic options are needed. Stem cell therapy is promising for a variety of disorders but there has been no human clinicaltrial of stem cell therapy for asthma. We aimed to systematically review the literature regarding the potential benefits of stemcell therapy in animal models of asthma to determine whether a human trial is warranted. Methods. The MEDLINE andEmbase databases were searched for original studies of stem cell therapy in animal asthma models. Results. Nineteen studieswere selected. They were found to be heterogeneous in their design. Mesenchymal stromal cells were used before sensiti-zation with an allergen, before challenge with the allergen and after challenge, most frequently with ovalbumin, and mainly inBALB/c mice. Stem cell therapy resulted in a reduction of bronchoalveolar lavage fluid inflammation and eosinophilia as wellas Th2 cytokines such as interleukin-4 and interleukin-5. Improvement in histopathology such as peribronchial and peri-vascular inflammation, epithelial thickness, goblet cell hyperplasia and smooth muscle layer thickening was universal. Severalstudies showed a reduction in airway hyper-responsiveness. Conclusions. Stem cell therapy decreases eosinophilic and Th2inflammation and is effective in several phases of the allergic response in animal asthma models. Further study is warranted,up to human clinical trials.

Key Words: animal studies, asthma, stem cells, systematic review, therapy

Introduction

Asthma is a worldwide problem [1]. In the developedworld, 20e30% of people are affected by allergicdisorders such as anaphylaxis, hay fever, eczema, andasthma [2]. Furthermore, asthma control frequentlyfalls short of the goals set in international guidelines[1]. Although definitions and estimates vary,approximately 15% of asthmatics may be classified ashaving severe asthma [3]. Unfortunately, there arefew treatment options for patients with poorlycontrolled asthma already receiving inhaled cortico-steroids and long-acting b-agonists. These optionsinclude leukotriene receptor antagonists, tiotropium,omalizumab or theophylline, which are eitherexpensive, cumbersome, of modest benefit or marredby potentially serious side effects. New therapeuticoptions are needed.

Correspondence: Nadim Srour, MD, Hôpital Charles-LeMoyne, 3120 boul. Tusherbrooke.ca

(Received 9 May 2014; accepted 12 August 2014)

http://dx.doi.org/10.1016/j.jcyt.2014.08.008ISSN 1465-3249 Copyright � 2014, International Society for Cellular Therapy. P

There has been much enthusiasm about thetherapeutic potential of mesenchymal stromal cells(MSCs) in several clinical disorders such as multiplesclerosis, stroke, myocardial infarction, diabetes,sepsis, hepatic and renal failure, as well as asthma[4,5]. However, to our knowledge, there has been nohuman clinical trial of MSC therapy for asthma. Wetherefore aimed to review the literature about thepotential benefits of MSC therapy in animal modelsof asthma.

Methods

Study selection

We sought to include studies of in vivo animal modelsof asthma, in which the effects of stem cell adminis-tration on clinical or biological outcomes relevant to

aschereau, Greenfield Park, QC J4V 2H1 Canada. E-mail: nadim.srour@

ublished by Elsevier Inc. All rights reserved.

1630 N. Srour & B. Th�ebaud

asthma were compared with the effects of controltherapy. We identified studies from two databases:Embase (1996 to 2014 week 24) and MEDLINE(Ovid MEDLINE In-Process & Other Non-IndexedCitations and Ovid MEDLINE, 1996 to June 13,2014). The search query “(exp Stem cells/ or expStromal Cells/ or exp Bone Marrow Cells/ or expStem Cell Transplantation/ or exp Bone MarrowTransplantation/) and (exp asthma/ or exp AirwayRemodeling/ or exp Bronchial Hyperreactivity/ or expBronchoconstriction/ or airway inflammation.ti,ab.)”was run on both databases. We then used Ovid’sdeduplication feature to identify unique studies, withhigher preference given to the Embase database.Limits were used to identify reviews, editorials andconference abstracts. The remaining abstracts and thefull text of selected abstracts were then reviewed forinclusion criteria: (i) an animal model of asthma wasused; (ii) there was administration of stem cells orprogenitor cells that were not used as a vector forother agents; (iii) the study reported on original data.

Query results: 1873Query results: 1873

Data extraction

Data were extracted from the selected studies. In a firststep, the following information was recorded: animalmodel; sensitizing agent, route, dose and time used tosensitize the animal and induce asthma; type of stemcells, dose, route and time of administration; outcomesreported; time of outcome measurement. Outcomeswere identified from the Methods section, the Resultssection, tables and figures and were classified intoquantitative, semi-quantitative and qualitative.

Abstracts reviewed: 874

DuplicatesReviewEditorialReview/EditorialConference

DuplicatesNo abstractChineseNon-originalNot of interestStem cells as vector

Non-original

Not stem cellsNot in vivo

19 articles selected

18940832

1369

4101

142686

1

Full text reviewed: 30

2531

Abstracts reviewed: 874

DuplicatesReviewEditorialReview/EditorialConference

DuplicatesNo abstractChineseNon-originalNot of interestStem cells as vector

Non-originalNot of interestNot stem cellsNot in vivo

19 articles selected

18940832

1369

4101

142686

1

Full text reviewed: 30

2531

Figure 1. Study selection.

Data analysis

We had planned for a meta-analysis of the 2 mostcommonly reported quantitative outcomes, whichwere bronchoalveolar lavage (BAL) total cell andeosinophil counts. This was not possible, mostlybecause of heterogeneity in study design. Further-more, the data were presented graphically, and only afew authors responded to our request for numericaldata. The remaining BAL total cell and eosinophilcount data were therefore extracted from the publishedvector or raster graphics. Although a meta-analysis wasnot performed, the data for these 2 outcomes arepresented by use of the ratio of means method [6]. Weassessed the risk of bias for these 2 outcomes throughthe use of funnel plots. For the latter purpose, only 1experimental group was included for each controlgroup; preference was given to groups that receivedbone marrowederived cells and syngeneic cells.Graphs were prepared with the use of ReviewManager(Version 5.2; Cochrane Collaboration, Oxford, UnitedKingdom). Publication bias refers to the tendency for

“negative” studies (in which the null hypothesis is notrefuted) to be less likely to be published than “posi-tive” studies (or to be published faster, in English, etc).For the current systematic review, this is relevantbecause it might be more interesting for a journal topublish a study in which MSC treatment improvedasthma outcomes than a study in which outcomeswere not improved. Thus published literature can thenoverestimate the effect of an intervention or show aneffect when in fact there is none.

A funnel plot is one technique than can behelpful to detect publication bias. It plots a measureof effect size on the x-axis with a measure of itsdispersion on the y-axis. In the absence of publi-cation bias, less precise studies should be scatteredsymmetrically around more precise studies. Thisclassically leads to a funnel appearance. An asym-metry can indicate publication bias, but there areother possible explanations.

Results

The search query returned 1873 entries (Figure 1),874 of which were not identified as duplicates, re-views, editorials or conference abstracts by use of theOvid system and were reviewed. Of these, 30 studieswere selected for full text review, 19 of which met theinclusion criteria. Two studies were excludedbecause bone marrowederived mononuclear cellswere used rather than stem cells [7,8] and another

Systematic review: stem cells in animal asthma models 1631

because soluble factors from bone marrow cells wereused [5].

Study design

All studies used mice asthma models (BALB/c mice:15 studies, C57BL/6 mice: 5 studies, Table 1). Themost common allergen used was ovalbumin (15studies), with other studies that used ragweed [9],toluene diisocyanate [10], house-dust mite [11,12],aspergillus fungal extract [13] and cockroach extract[11]. The studies varied in duration, up to 118 days[14]. Bone marrowederived mesenchymal stromalcells (BM-MSCs) were obtained from murine do-nors in 14 studies, rats in 1 study [10] and humans in4 studies [4,15e17]. One of the latter studies alsoused induced pluripotent stem cells (iPSCs) derivedfrom human fibroblasts [17]. One study used murineiPSCs without c-Myc [18]. One study comparedadipose tissue, umbilical cord and BM-MSCs [16].One other study used adipose tissue MSCs [19] andanother used compact bone MSCs [14]. MSCadministration occurred before allergen challengemost commonly but also occurred before sensitiza-tion [11,17,20,21] or after allergen challenge[12,14,15,18,22] in other studies. MSCs wereadministered intravenously in all but 3 studies[11,23,24] in which they were administeredintratracheally.

The two most commonly reported quantitativeoutcomes were BAL eosinophil count (16 studies,Table II) and BAL total cell count (13 studies, 1study reporting the differential count but not thetotal cell count). BAL cytokines were also commonlyreported. Histopathology was reported in all but 1study, sometimes semi-quantitatively (8 studies) orquantitatively (8 studies). Airway hyper-responsive-ness was reported in 11 studies that used varioustechniques.

The number of mice included in each experi-mental group was sometimes reported as a rangerather than an exact number.

BAL total cell count

All 13 studies reporting on this outcome found thatMSC treatment decreased BAL total cell count (P <0.0001 to P ¼ 0.05), except for 2 experiments. Oneexception was the group treated before sensitization inthe Sun et al. [17] study. However, treatment withMSCs before sensitization did decrease BAL total cellcount in the Kavanagh [25] and Goodwin [20]studies. The other exception was the recurrentinflammation group in the Lathrop [13] study [13].This study used Aspergillus fungal extract to induceTh17 airway inflammation rather than Th2. In one

experiment, MSCs were administered at the begin-ning of a second challenge at days 76e78, a firstchallenge having occurred at days 14e16. However, asignificant reduction in the BAL total cell count wasseen in other studies with repeated challenge such asthe Mariñas-Pardo [12] study, in which challengeoccurred 3 times weekly for 4 or 6 weeks, and theOgulur [14] study, in which challenge occurred twiceweekly for 12 weeks. The mean cell count ratio in theMSC group compared with the control group isillustrated in Figure 2. Studies with the most preciseestimates included some with a treatment effect closerto unity but also some with the largest effect sizes. Afunnel plot was used to help detect publication bias.In this case, an example of publication bias would bestudies in which MSC treatment led to a smaller or noeffect being less likely to be published, skewing theresults. The funnel plot for BAL total cell count,which only includes one of several experimentssharing a control group, is not a typical plot but doesnot suggest obvious publication bias.

BAL eosinophil count

BAL eosinophil count was reduced by MSC treat-ment in all but 2 of 16 studies reporting this outcome(P < 0.001 to P � 0.05). Although the 95% confi-dence interval of the ratio of means for the allogeneictreatment group in the Goodwin study includes unity(Figure 3), the treatment effect was reported as sig-nificant (P < 0.05). One exception was the Lathropet al.[13] Th17 inflammation study. The otherexception was the experiment in the Mariñas-Pardo[12] study, in which the outcome was assessed 72 hafter MSC administration, after 4 weeks of house-dust mite instillation. When BAL fluid was obtained2 weeks after MSC administration, a significantreduction in BAL eosinophils was seen [12]. Again,studies with the most precise estimates include somewith a treatment effect closer to unity but also somewith the largest effect sizes. The funnel plot appearsas expected and does not suggest publication bias.

Other biochemical outcomes

BAL interleukin (IL)-4 levels were assessed in 11studies [9,12,13,17e21,23,24,26] and were found tobe significantly decreased with MSC treatment in 8studies (P < 0.05), including with treatment beforesensitization [17,20,21], with treatment beforeallergen challenge [9,17,19,23,26] and with treatmentafter allergen challenge [18]. No significant effect wasseen in the Lathrop [13] study, the Mariñas-Pardo[12] study, one of the Ge studies [24] and in onegroup of the Sun study. BAL IL-5 levels were assessedin 9 studies [9,13,15e20,26] and were significantly

Table I. Characteristics of selected studies.

Study identifierAnimals/group Animals Allergen Stem cell source

Sensitization(days) Challenge (days)

MSC injection(days) Outcome Timing

Abreu [26] 6 C57BL/6 mice Ovalbumin C57BL/6 BM-MSC 0, 2, 4, 6, 8,10, 12

40, 43, 46, 47 47 54 C

Ahmad [11] >5 BALB/c mice Ovalbumin Human/murine BM-MSC 0, 7, 14 21e27 27 32 A>5 BALB/c mice Cockroach extract Human/murine BM-MSC 0, 7, 14 27e30 30 31 A>5 BALB/c mice House-dust mite Human/murine BM-MSC 0, 7, 14 25e29 29 30 A

Bonfield [15] 16e24 BALB/c mice Ovalbumin Human BM-MSC 0 Every 2 days for 4 weeksstarting on day 14

During 6th week During 7thweek

Aa

Bonfield [4] 20e30 BALB/c mice Ovalbumin Human BM-MSC 0 14e18 14 or 16b 18 C/Ab

Firinci [22] 10 BALB/c mice Ovalbumin BALB/c BM-MSC 0, 14 �3x weekly for 8 weeksstarting on day 21

75 82 and 89 A

Ge [24] BALB/c mice Ovalbumin BALB/c BM-MSC 1, 14 24e26 23 27 (PFT), 28 CGe [23] 10 BALB/c mice Ovalbumin BALB/c BM-MSC 0, 7, 14 �3 weekly for up to 8 weeks

starting on day 2120 78 C

Goodwin [20] 4e5 C57BL/6 and BALB/cmice

Ovalbumin C57BL/6 BM-MSC 0, 7 14e16 0 and 7 18 S

Kavanagh [21] >15 BALB/c mice Ovalbumin FVB/NHanHsd miceBM-MSC

0, 7, 14 14, 25e27 7 and 14 28 S

Lathrop [13] 16e24 C57BL/6 mice Aspergillus hyphalextract

C57BL/6 BM-MSC 0, 7 14e16 � 76e78 (recurrent) 14 19, 81(recurrent)

C

Lee [10] 6 BALB/c mice TDI SD rat BM-MSC 1e5 9e11 5 13 CMariñas-Pardo

[12]5e8 BALB/c mice House-dust mite BALB/c ASC None �3 weekly for 4 or 6 weeks 28 31 or 42 A

Mathias [16] 9e15 BALB/c female mice Ovalbumin Human USC/ASC/BM-MSC

0 8e10 5e7 11,12 C

Nemeth [9] 4e9 C57BL/6 mice Ragweed C57BL/6J and BALB/cBM-MSC

0, 5 14 (IT), 15 (IN) 14 18 C

Ogulur [14] 4 BALB/c mice Ovalbumin BALB/C compact boneMSC

0, 14, 21 �2 weekly for 12 weeksstarting on day 26

104 118 A

Ou-Yang [25] 10 C57BL/6 mice Ovalbumin C57BL/6 BM-MSC 1, 8 15e17 14 18 CPark [19] 25 BALB/c female mice Ovalbumin BALB/c ASC 1, 14 21e23 18e20 24 (PFT), 25 CSun [17] 4e6 BALB/c mice Ovalbumin Human iPSC and

BM-MSC1, 3, 5, 7, 9,

11, 1321e27 20/0 29 S/C

Wang [18] 6 BALB/c mice Ovalbumin C57BL/6 iPSCc 0, 14 �3 during 4th week � day64

35 64 A

In the case of discrepancy, the number of animals/group refers to the experiment where BAL cell counts were measured. A, MSC administration after allergen challenge; C, before allergenchallenge; S, before allergen sensitization; ASC, adipose tissue MSC; IN, intranasal; IT, intratracheal; IMR90-IPSC and N1-IPSC, induced pluripotent stem cell lines; PFT, pulmonary functiontests; TDI, toluene diisocyanate; USC, umbilical cord MSC.aDuring last week of challenge.bseparate experiment, limited data.cWithout c-Myc (with Oct-3/4, Sox-2, Klf-4).

1632N.Srour

&B.Th �ebaud

Systematic review: stem cells in animal asthma models 1633

decreased by MSC treatment in 7 studies [15e20,26](P < 0.05), including with treatment before sensiti-zation [17,20], before allergen challenge [16,17,19,26]and after allergen challenge [15,18]. No significanteffect was seen in 2 studies, including the Lathropet al. study [9,13]. MSC treatment led to a significantdecrease in BAL IL-13 levels in 3 studies [9,15,17](P < 0.001 to P < 0.05), including with treatmentafter allergen challenge [15], whereas BAL IL-13levels were already low in controls in 1 study [25]and there was no significant change in 5 studies[12,13,20,23,24]. One short-term study thatreported on IL-9 levels in BAL found a reductionwith MSC treatment (P < 0.05) [26].

BAL IL-6 levels were significantly decreased in asyngeneic pre-sensitization model [20], whereaslevels were not detectable in the allogeneic model. Inthe Lathrop study [13], a significant decrease wasnoted in the recurrent challenge experiment and non-significant decrease in the unique challenge experi-ment. No significant effect was seen in 3 other studies[12,15,19]. MSC treatment before sensitization [21],before allergen challenge [24] and after allergenchallenge [18] led to a significant increase in BAL IL-10 levels (P < 0.05) in 3 studies, whereas no signifi-cant change was noted in 4 studies [9,12,13,23] thatused MSC treatment before [9,13,23] and after [12]allergen challenge.

The effect of MSC treatment on BAL interferon(IFN)-g was variable with a significant decreasenoted in 4 studies [4,15,18,26], including 2 studieswith treatment after allergen challenge [15,18] nosignificant change in 6 studies [9,12,13,17,23,24]and a significant increase in 3 studies [12,17,19]. Inthe Sun study [17], a significant increase was notedwith treatment with BM-MSCs but not withinduced progenitor cells. In the Mariñas-Pardostudy [12], a significant increased was noted 72 hafter MSC administration but not after 2 weeks.Levels were not detectable in the Goodwin study[20]. Serum IFN-g was increased in 2 studies withMSCs administered before or after allergen chal-lenge, but no effect was seen in 2 studies [23,24].The effect on BAL IL-12 levels was variable[12,13,23,24].

In one study that used ragweed as the allergen[9], MSC treatment before allergen challenge resul-ted in a significant increase in transforming growthfactor (TGF)-b levels (P < 0.05). Production ofTGF-b by MSCs in vitro was increased by exposureto BAL fluid from ragweed-sensitized and challengedmice [9]. However, BAL TGF-b was decreased byMSC administration before sensitization [20] andbefore allergen challenge [19] in 2 other studies. Noeffect was seen in 1 study with MSC administrationbefore allergen challenge [23].

In the ragweed study, treatment with MSCsresulted in decreased serum immunoglobulin (Ig)Elevels (P < 0.05) [9] with either syngeneic or allo-geneic MSCs. Treatment with MSCs also resulted indecreased serum IgE levels in another study withadministration after allergen challenge (P < 0.05)[15]. However, no significant changes were seen inanother study [12]. Five studies reported a decreasein serum allergen-specific IgE levels (P < 0.01 to P <0.05) [17,18,20,21,23] with MSC treatment, withone study reporting a significant decrease with allo-geneic but not syngeneic MSC treatment beforesensitization [20]. Serum allergen-specific IgG1 wasdecreased in some experiments [17,18,20,23] butnot others [17,20]. The effect on serum allergen-specific IgG2a was variable [18,20,23].

Treatment with MSCs before allergen challengewas also found to lead to a decrease in lung collagencontent in 3 studies in which MSC were administeredbefore allergen challenge (P < 0.05) [10,23,27]. LungPeriodic acideSchiff (PAS) cells were decreased in 6studies [10,12,14,17,22,23], including MSC admin-istration before sensitization [17] before [10,17,23]and after [12,14,22] allergen challenge, except in theexperiment in the Mariñas-Pardo [12] study, in whichBAL was sampled 2 weeks after MSC administration.

Regulatory T cells (Treg) were assessed in 7studies [9,13,14,16,21,23,24], in the lung tissue[9,13,14,16,21] pulmonary lymph nodes [23,24] andthe spleen [13,21]. They were increased by MSCtreatment in all but the Lathrop study [13].

Histopathology

Histopathology was not assessed in the Wang andAhmad studies. It was not described in the Sun study.All other studies reported improvement in lung histo-pathology in mice treated with MSCs, with theimprovement described as great, marked or dramatic.Visually, improvements in airway inflammation[4,9,10,15,25,26], peribronchial and perivascular infil-tration [9,12,13,16,19,26], epithelial lining thickening[4,15,22], subepithelial smoothmuscle layer thickening[22], basement membrane thickening [22] and gobletcell hyperplasia and mucus production [4,9,15,16,24]were seen. These improvements were seen in studies inwhich MSCs were administered before sensitization[20,21], before [4,9,10,13,16,19,23,24,26,27] or after[12,14,15,22] allergen challenge. In theMariñas-Pardo[12] study, the improvement in inflammation seen 72 hafter MSC administration rebounded 2 weeks afterMSC administration.

Ten of the studies [9e11,13,17,19,20,23,24,26]used a semi-quantitative index for lung histopathol-ogy. MSC treatment before challenge was found toresult in a significant reduction of an index of

Table II. Quantitative outcomes reported in at least 3 studies.

Study/experiment BAL Total BAL Eo BAL Lymph BAL Macro BAL Neutro BAL IFN-g BAL IL-4 BAL IL-5 BAL IL-6 BAL IL-10 BAL IL-12 BAL IL-13 BAL TGF-b

Abreu [26]Ahmad/Cockroach [11]Ahmad/house-dust mite [11]Ahmad/Ovalbumin [11]Bonfield [15] Y(0.05) Y(�0.05) e e [(0.05) Y(<0.05) Y(<0.05) e Y(<0.05)Bonfield [4] Y(0.03) Y(�0.05) e [(�0.05) Y(�0.05) Y(<0.05)Firinci [22]Ge [24] Y(<0.05) Y(<0.01) e e [(<0.05) [(<0.05) e e

Ge [23] Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) e Y(<0.05) e Y(<0.05) e

Goodwin/allogeneic [20] Y(<0.05) e [(<0.05) e ND Y(<0.05) Y(<0.05) ND e

Goodwin/syngeneic [20] Y(<0.05) e [(<0.05) e ND Y(<0.05) Y(<0.05) Y(<0.05) e Y(<0.05)Kavanagh [21] Y(<0.05) Y(<0.01) e Y(<0.05) e Y(<0.05) [(<0.05) ?Lathrop [13] Y(�0.01) e e e Y(�0.05) e e e eY e Y(�0.05) e

Lathrop/recurrent [13] e e e e [(�0.001) e e e Y(�0.05) e e e

Lee [10] Y(<0.05) Y(<0.05) e Y(<0.05) Y(<0.05)Mariñas-Pardo/2 weeks [12] Y(0.002) Y(0.003) Y(0.005) e e e e e [(0.025) e

Mariñas-Pardo/72 h [12] Y(0.037) e e e [(0.031) e e e [(0.047) e

Mathias/ASC [16] Y(�0.01) Y(�0.01)Mathias/BM-MSC [16] Y(�0.001)Mathias/USC [16] Y(�0.001)Nemeth/allogeneic [9] Y(<0.01) Y(<0.05) Y(<0.05) Y(<0.01)Nemeth/syngeneic [9] Y(<0.01) Y(<0.001) e Y(<0.01) e e Y(<0.001) [(<0.01)Ogulur [14]Ou-Yang [25] Y(<0.05) Y(<0.05) e e e Y(<0.05) Y(<0.05) Y(<0.05)Park [19] Y(<0.05) Y(<0.05) e Y(<0.05) e [(<0.05) Y(<0.05) Y(<0.05) e Y(<0.05)Sun/BM-MSC [17] Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) [(<0.05) Y(<0.05) Y(<0.05) Y(<0.05)Sun/iPSC-iMR90 [17] Y(<0.01) Y(<0.01) Y(<0.01) Y(<0.05) Y(<0.01) e Y(<0.05) Y(<0.05) Y(<0.05)Sun/iPSC-N1 [17] Y(<0.05) Y(<0.01) Y(<0.01) e Y(<0.05) e e Y(<0.05) Y(<0.05)Sun/pre [17] e Y(<0.01) e e e e Y(<0.05) Y(<0.05) Y(<0.05)Wang [18] Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) Y(<0.05) [(<0.05)

(continued)

1634N.Srour

&B.Th �ebaud

Study/experimentSerumIFN-g

Serum ovalbumin-specific IgG1

Serum ovalbumin-specificIgG2a

Serum ovalbumin-specific IgE

SerumIgE

Lung collagencontent

Lung PeriodicacideSchiff cells Treg AHR

Abreu [26] Y(0.05) Y(<0.05)a

Ahmad/cockroach [11] Y(<0.05)Ahmad/house-dust

mite [11]Y(<0.05)

Ahmad/ovalbumin[11]

Y(<0.05)

Bonfield [15] [(0.043) Y(<0.05)Bonfield [4] [(0.043)Firinci [22] Y(early: 0.010, late:

0.045)Ge [24] e [(LN: <0.01) Y(<0.05)Ge [23] e Y(<0.01) Y(<0.01) Y(<0.01) Y(<0.05) Y(<0.05)b [(LN: <0.05)Goodwin/allogeneic

[20]Y(<0.05) [(<0.05) Y(<0.05) Y(<0.05)

Goodwin/syngeneic[20]

e [(<0.05) e Y(<0.05)

Kavanagh [21] Y(<0.01) Y(<0.05)c [(LungþSpleen: <0.001) Y(<0.05)Lathrop [13] e(LungþSpleen) Y(�0.01)Lathrop/recurrent [13] e(LungþSpleen) Y(�0.05)d

Lee [10] Y(<0.05) Y(<0.05) Y(<0.05)Mariñas-Pardo/2 weeks

[12]e e Y(<0.05)

Mariñas-Pardo/72 h[12]

e Y(<0.05) e

Mathias/ASC [16] [(Lung: �0.001) Y(�0.01)Mathias/BM-MSC

[16]Y(�0.05)

Mathias/USC [16] Y(�0.05)Nemeth/allogeneic [9] Y(<0.05)Nemeth/syngeneic [9] Y(<0.05) [(Lung: <0.001)Ogulur [14] Y(<0.001) [(Lung: 0.037)Ou-Yang [25] Y(<0.01)Park [19] Y(<0.05)Sun/BM-MSC [17] Y(<0.05) Y(<0.05) Y(<0.01)Sun/iPSC-iMR90 [17] e Y(<0.01) Y(<0.01)Sun/iPSC-N1 [17] e Y(<0.05) Y(<0.01)Sun/pre [17] e Y(<0.05) Y(<0.05)Wang [18] Y(<0.005) Y(<0.05) Y(<0.05) Y(<0.005)

-, no significant change, Y, no significant change with a decreasing trend, AHR, airway hyper-responsiveness, ASC, amniotic fluid MSC, LN, lymph node, ND, not detectable, PAS, Periodic acid -Schiff, Treg, regulatory T cells, USC, umbilical cord MSC.aResistive pressure rather than airway hyper-responsiveness.bPeriodic acideSchiff positive goblet cell score.cSignificance not indicated.dSignificant for total tissue resistance, not airway resistance.

Table II. ContinuedSystem

aticreview

:stem

cellsin

animalasthm

amodels

1635

Study or SubgroupGe [23]Ge [24]Nemeth SyngeneicNemeth Allogeneic [9]Sun BM-MSCSun iMR90-iPSCSun N1-iPSCSun presensitization [17]Lee [10]Bonfield [15]Ou-Yang [25]Kavanagh [21]Park [19]Lathrop [13]Mariñas-Pardo 2w [12]Mariñas-Pardo 72h [12]Wang [18]Bonfield [4]

Ratio of Means, 95% CI0.19 [0.15, 0.24]0.20 [0.15, 0.27]0.28 [0.13, 0.60]0.30 [0.17, 0.54]0.32 [0.17, 0.61]0.32 [0.19, 0.54]0.46 [0.28, 0.75]0.63 [0.39, 1.02]0.37 [0.20, 0.68]0.50 [0.29, 0.87]0.51 [0.27, 0.95]0.55 [0.31, 0.97]0.54 [0.34, 0.87]0.58 [0.40, 0.85]0.61 [0.38, 0.99]0.61 [0.46, 0.82]0.64 [0.41, 1.02]0.77 [0.59, 0.99]

0.1 0.2 0.5 1 2 5 10Favors stem cell Favors control

0.1 0.2 0.5 1

0

0.1

0.2

0.3

0.4

0.5

Ratio of Means

SE

(log[

Rat

io o

f Mea

ns])

BA

Figure 2. Effect of stem cell treatment on BAL total cell count. (A) Forest plot. Brackets indicate study arms sharing a control group. (B)Funnel plot. IMR90-IPSC and N1-IPSC, induced pluripotent stem cell lines. Except for the pre-sensitization arm that used BM-MSCs,cells were administered before allergen challenge in the Sun et al. study. Periodic acideSchiff.

1636 N. Srour & B. Th�ebaud

peribronchial cuffing, perivascular cuffing, goblet cellhyperplasia and interstitial inflammation [P < 0.001[9]], an index of peribronchial and perivascularinflammation [P < 0.01 [26] and P < 0.05 [19,24]],an index of peribronchial inflammation [P <0.05[20] P � 0.001 [13] and P < 0.0001 [10]] and anindex of overall lung inflammation [P < 0.001 [17]and P < 0.05 [11]].

Study or SubgroupLee [10]Sun BM-MSCSun iMR90-iPSCSun N1-iPSCSun presensitization [17]Ge [24]Kavanagh [21]Bonfield [4]Nemeth AllogeneicNemeth Allogeneic [9]Mathias USCMathias BMMathias ASC [16]Bonfield [15]Wang [18]Ge [23]Park [19]Goodwin AllogeneicGoodwin Syngeneic [20]Ou-Yang [25]Mariñas-Pardo 2w [12]Lathrop [13]Mariñas-Pardo 72h [12]

Ratio of Means, 95% CI0.02 [0.01, 0.05]0.06 [0.03, 0.12]0.14 [0.06, 0.30]0.12 [0.05, 0.27]0.24 [0.14, 0.43]0.10 [0.06, 0.15]0.11 [0.08, 0.16]0.11 [0.04, 0.33]0.19 [0.04, 0.79]0.21 [0.06, 0.69]0.20 [0.12, 0.34]0.24 [0.14, 0.41]0.40 [0.28, 0.57]0.21 [0.08, 0.56]0.25 [0.02, 4.00]0.25 [0.20, 0.33]0.27 [0.13, 0.56]0.39 [0.10, 1.54]0.43 [0.26, 0.72]0.55 [0.34, 0.89]0.55 [0.36, 0.84]0.58 [0.33, 1.00]0.78 [0.37, 1.66]

0.01 0.1 1Favors stem cells Favors

A

Figure 3. Effect of stem cell treatment on BAL eosinophil count. (A) F(B) Funnel plot.

Six studies assessed histopathology quantita-tively. One study showed that treatment with MSCsafter challenge resulted in a reduction of basementmembrane thickness (P ¼ 0.00 and P ¼ 0.00), sub-epithelial smooth muscle thickness (P ¼ 0.047 andP ¼ 0.00) and number of mast cells (P ¼ 0.039 andP ¼ 0.002) after 1 and 2 weeks, respectively [22].Epithelial thickness was significantly reduced 2

10 100 control

0.01 0.1 1

0

0.5

1

1.5

2

Ratio of Means

SE

(log[

Rat

io o

f Mea

ns])

B

orest plot. Brackets indicate study arms sharing a control group.

Systematic review: stem cells in animal asthma models 1637

weeks after treatment (P ¼ 0.002) but not after 1week (P ¼ 0.059) [22]. MSC administration afterallergen challenge decreased epithelial thickness,basement membrane thickness and smooth musclethickness (P < 0.05) in another study after 2 weeks[14]. The number of inflammatory nuclei wasdecreased by MSC administration after allergenchallenge in another study (P < 0.001) [15]. MSCadministration after allergen challenge resulted in areduction in airway contractile tissue after 2 weeks(P < 0.05) but not after 72 h in one study, with noeffect on extracellular matrix mass [12]. In anotherstudy, MSC treatment before allergen challengeresulted in reductions in airway inflammation, hy-perplasia of goblet cells and subepithelial fibrosis(P < 0.05 for all) [23]. In another study, MSCtreatment before challenge resulted in a decrease inalveolar collapse, neutrophilic infiltration and bron-choconstriction index (P < 0.05 for all) [26]. MSCadministration before sensitization resulted in areduction in airway mucin content (P < 0.05) andsubepithelial collagen (P < 0.05) [11].

Airway hyper-responsiveness and body weight

Twelve studies assessed airway hyper-responsive-ness to methacholine. All found a significantreduction with MSC treatment before sensitization[11,20,21] and before [10,13,16,19,24,25,26] andafter [12,18] allergen challenge, with the use of avariety of techniques such as invasive ventilation,the enhanced pause technique, forced oscillationand the airway pressure-time index. In the Mariñas-Pardo study [12], the significant improvement wasseen 2 weeks after MSC administration but not after72 h. In the Lathrop study [13], significantimprovement in airway resistance was seen in theexperiment in which MSCs were administeredbefore a first challenge on day 14, but, in theexperiment in which MSCs were administeredbefore a second challenge on day 76, the improve-ment was significant for total tissue resistance butnot for airway resistance.

In a study in which mice were challengedrepeatedly for 4 weeks, MSC administration at theend of the challenge period attenuated weight loss(P ¼ 0.05) [15].

Treatment group comparison

Although most studies used BM-MSCs, MSCs fromother sources such as compact bone [14], adiposetissue [12,16,19] and umbilical cord [16] and iPSCs[17,18] were effective as well. Allogeneic MSCs wereused in 2 studies. In the Nemeth [9]study, allogeneicMSCs had generally similar effects compared with

syngeneic MSCs, including BAL total cell andeosinophil count, BAL IL-4 and IL-13, as well asserum IgE levels [9]. In the Goodwin [20] study,allogeneic MSCs also had similar effects comparedwith syngeneic MSCs, including the effects on lunginflammation, airway hyper-responsiveness, BALeosinophils and BAL IL-4 and IL-5 [20].

Induced PSCs were used in 2 studies beforesensitization [17], before [17] and after allergenchallenge [18]. They were effective in reducing BALeosinophilia and generally in reducing Th2 cytokines.Airway hyper-responsiveness was only assessed in theWang [18] study, in which MSCs were administeredafter allergen challenge and were found to be signifi-cantly reduced. Adipose tissueederived stem MSCswere used before allergen challenge in the Park [19]study and after allergen challenge in the Mariñas-Pardo [12] study. They were compared with MSCsderived from bone marrow and umbilical cord in theMathias study [16], after allergen challenge. All thesetreatment groups were effective in reducing BALeosinophilia and airway hyper-responsiveness; how-ever, the effects were seen after 2 weeks in the Ma-riñas-Pardo study but not after 72 h.

Paraformaldehyde-fixed MSCs were used in theKavanagh [21] study, with a loss of effect on histology,BAL total cell and eosinophil count, ovalbumin-specific IgE levels and airway hyper-responsiveness. Inthe Goodwin study, the use of 1-ethyl-3-[3-dimethy-laminopropyl] carbodiimide hydrochloride (EDCI) toprevent the release of soluble mediators from MSCsresulted in loss of some effects such as the improve-ments in histopathology and BAL eosinophil counts,but other effects such as the reduction in BAL IL-4,IL-5 and IL-6 were preserved. Airway hyper-respon-sivenesswas only significantly different fromcontrols atthe lowest methacholine dose with EDCI-treatedMSCs. Similarly, in the Lathrop [13] study, sometreatment effects such as the reduction in IL3, IL12,keratinocyte chemoattractant, CXCL1 and RANTESBAL levels were preserved despite the use of EDCI toprevent the release of soluble mediators, whereas thereremained a nonsignificant trend toward improvementin airway hyper-responsiveness.

Murine dermal fibroblasts were used in 2 studies.In the Nemeth [9] study, there was still an effect onBAL total cell count and BAL IL-4 and IL-13, butthere was no effect on BAL eosinophils or serum IgEwith administration before allergen challenge. Therewas still a decrease in BAL-IL3. In the Goodwin [20]study, fibroblast administration before sensitizationdid improve airway hyper-responsiveness anddecrease the BAL total cell count but did not decreasethe BAL eosinophil count or histologic airwayinflammation. In one of the Bonfield studies [15],bone marrowederived macrophage administration

1638 N. Srour & B. Th�ebaud

after allergen challenge did not improve lung inflam-mation, BAL total cell count or differential, serumIgE or weight loss [15]. In the Abreu [26] study, bonemarrowederived mononuclear cell administrationbefore allergen challenge did improve several histo-logical parameters, lung IL-4 and IL-13. Lung TGF-band vascular endothelial growth factor as well as lungfunction were improved more than with MSCs.

Discussion

This review has identified several studies exploringthe effects of MSC administration in animal asthmamodels. Inflammation and particularly eosinophilicinflammation were reduced by MSC administrationin all studies. This included short-term and longerstudies, in which MSCs were administered beforesensitization with an allergen, before allergen chal-lenge and after allergen challenge. Th2 cytokines inBAL fluid such as IL-4 and IL-5 were decreased byMSC treatment, whereas the effects on IL-13 werenot as consistent. There was also evidence of adecrease on serum IgE or allergen-specific IgE. Allstudies reported improvement in histopathology suchas peribronchial and perivascular inflammation,epithelial thickness, goblet cell hyperplasia andsmooth-muscle layer thickening. A few studies re-ported on clinically relevant outcomes and showed areduction in airway hyper-responsiveness.

The study design, that is, the point of MSCadministration within the experiments, is a crucialissue. One pitfall of many studies is that MSCs wereadministered before allergen challenge. We wouldcontend that MSC administration before sensitiza-tion, before allergen challenge and after allergenchallenge are all relevant to human disease.Regarding MSC administration before allergensensitization, patients will be sensitized to newagents in their daily life from time to time. Perhapsmore importantly, the prevention of a deleteriousreaction to future allergen challenges is important inasthma control because most asthmatics arerepeatedly exposed to allergens. Given the parox-ysmal natural history of asthma, it is just as impor-tant to prevent further clinical events (such asasthma exacerbations) and further remodeling(MSC administration before allergen challenge)than to try to reverse established damage (MSCadministration after allergen challenge). A hypo-thetical advantage of MSC therapy is the potentialefficacy in all phases of the allergic response,possibly by different mechanisms in each case.

In the Mariñas-Pardo study, the improvement inhistopathology seen 72 h after MSC administration(which occurred after allergen challenge) bouncedback after 2 weeks. However, the decrease in BAL

total cell count was still present, whereas a decreasein BAL eosinophil count, an improvement in airwayhyper-responsiveness and a decrease in airway con-tractile mass was only seen after 2 weeks [12].Furthermore, in the Ogulur study [14], in whichchallenge occurred repeatedly until day 104, whenMSC were administered, and when outcomes wereassessed 2 weeks later, an improvement in histopa-thology was still seen.

The Lathrop study [13] used Aspergillus fungalextract to induce Th17 inflammation. In this study,there was no decrease in BAL eosinophils and Th1cytokines in BAL such as IL-4, IL-5 or IL-13 butdecreased BAL IL-17 levels with decrease in lunginflammation and airway hyper-responsiveness.

Although most studies used BM-MSCs, it mayfeasible to use adipose tissue [12,16,19,27], umbilicalcord [16] and compact bone [14] as sources, or eveniPSCs [17,18], but corroboration is needed. Somestudies have used nonestem cell types [9,15,20,26] oreven soluble factors [5], but the evidence at this timeis not sufficient to support efficacy of these treatmentsin all phases of the allergic response.

Proposed pathways

Multiple pathways have been proposed for the effectsof MSCs in asthma. Nemeth et al. [9] have shownthat TGF-b production in vitro increased whenMSCs were co-cultured with serum from ragweed-challenged mice. This phenomenon was not presentwhen IL-4eneutralizing antibodies were added tothe medium or when IL-4R knock-out MSCs wereused [9]. Furthermore, the phenomenon was notpresent when STAT6-deficient MSCs were used,illustrating the dependence of this phenomenon onthe IL4-Ra/STAT6 pathway [9]. In vivo, thedecrease in BAL total cell counts and eosinophilcounts seen with MSC treatment before allergenchallenge was eliminated when treating the animalswith TGF-beneutralizing antibodies or when TGF-b1 knock-out MSC or STAT6-deficient MSCs [9]were used. However, TGF-b levels were decreasedby MSC administration before sensitization; thisdiscrepancy may result from the timing of MSCadministration and suggests that the response ofMSC depend on their micro-environment [20].TGF-b is thought to be involved in Treg differenti-ation, and an increase in Treg in MSC-treated micewas observed by Nemeth et al. [9], Kavanagh et al.[21] and Ge et al. [23]. Furthermore, Treg depletionthrough the use of cyclophosphamide nullified thebeneficial effects of MSCs on peribronchial inflam-mation, mucus production, IgE levels or the shiftaway from a Th2 profile [21]. However, the effect onBAL eosinophilia persisted, which suggests the

Systematic review: stem cells in animal asthma models 1639

existence of a Treg-independent pathway. One studyco-cultured iPSCs with peripheral blood mono-nuclear cells of human subjects with allergic rhinitis[28]. Decreased lymphocyte proliferation and a shiftaway from Th2 and toward Th1 was observed with adecrease in supernatant IL-4, IL-5 and IL-13 and anincrease in IFN-g. An increase in supernatant IL-10with increased Treg expansion with allergen stimu-lation was also seen. Prostaglandin E2 might bemediating these effects, because inhibition of pros-taglandin E2 production with a Cox-2 inhibitorsignificantly reversed the immunomodulatory effecton lymphocyte proliferation. The effects of MSCs,in a study in which they were injected beforesensitization, might also be partly dependant onIFN-g because the effects on BAL eosinophilia andIL-4, IL-5 and IL-13 were not seen in IFN-gknock-out mice [20]. In that study, MSCs did notinhibit in vivo, antigen-specific T-cell proliferationbut rather altered antigen-specific T-cell differenti-ation toward a Th1 response. In an ovalbumin-induced Th1-mediated lung inflammation, MSCshad no effect on the Th1-mediated increase in BALneutrophils and lymphocytes [20]. Similarly, adecrease in BAL and spleen IL-4 with an increase inIFN-g in the Park study suggests a shift away from aTh2 response. Alveolar macrophages appear to beimportant in mediating the effects of MSCs. In theMathias study [16], although there was no increasein macrophage TGF-b that would increase the Tregpopulation and no shift in the M1/M2 macrophageprofile, depletion of alveolar macrophages led to aloss of effects on airway hyper-responsiveness andhistopathology.

MSCs exert their effects through a paracrine effectrather than by lung engraftment [12,23,29]. In fact,Abreu et al. [7,8] have shown that bone marrowmononuclear cell administration before allergenchallenge also reduced eosinophilic infiltration, Th2cytokines, airway remodeling and airway hyper-responsiveness whether injected intravenously orintratracheally. With the use of MSCs treated with across-linker to prevent release of soluble factors,Goodwin et al. [20] demonstrated that the effects ofMSCs were not due to surface antigens either, but tosoluble factors. In fact, Ionescu et al. [5] used mediumconditioned by the use of plastic-adherent bonemarrow cells differentiated along mesenchymal line-ages rather than actual cells in murine ovalbumin-induced acute and chronic asthma models. This wasadministered intranasally after each challenge [5]. Inboth acute and chronic models, conditioned mediumprevented airway hyper-responsiveness and inflam-mation [5]. In the chronic model, conditioned me-dium prevented airway smooth-muscle thickening andperibronchial inflammation [5]. Significant reductions

in the Th2 cytokines IL-4 and IL-13 and a significantincrease in IL-10 were found along with an increasein IL-10esecreting Treg lymphocytes [5]. Adipo-nectin was found in greater concentrations in bonemarrow celleconditioned medium than in fibroblast-conditioned medium, and administration of adipo-nectin rather than conditioned medium did preventairway hyper-responsiveness, airway smooth musclethickening and peribronchial inflammation. Further-more, the effects of conditioned medium from adi-ponectin knock-out mice or in which adiponectin wasneutralized were blunted [5]. However, treatment witha cross-linker did not entirely eradicate the effects ofMSC treatment in the Goodwin [20] and Lathropstudies [13]. Thus, it appears that the observed MSCeffects are mostly (but not entirely) mediated throughsoluble factors rather than by MSCs acting as pre-cursors to mesenchymal structural cells.

A possible explanation for the partial effects withcross-linkers is effect mediation through cell contact,as proposed by several authors [13,20,30]. Mostcompelling are data from the study co-culturingiPSCs with peripheral blood mononuclear cells fromhuman subjects with allergic rhinitis [28]. Withtranswell separation of iPSCs from the mononuclearcells, the effects on lymphocyte proliferation andTreg expansion were lost [28]. Some data suggestthat microsomes are involved either in signaling bylung epithelial cells injured by radiation towardmarrow-derived cells [31] or as mediators of MSCeffects in a pulmonary hypertension model [32].Ahmad et al. [11] have used rotenone to inducemitochondrial stress in epithelial cells. Theydemonstrated and produced a movie of mitochon-drial transfer from MSCs to epithelial cells throughtunneling nanotubes. They produced MSC under-or over-expressing Miro1, a mitochondrial transportprotein that was decreased by rotenone in this study.In an ovalbumin mouse model, MSC over-express-ing Miro1 were more effective in reducing airwayhyper-responsiveness, decreasing lung IL-5 and IL-13 and decreasing inflammatory cell infiltration,collagen deposition and mucus hypersecretion. Inhouse-dust mite and cockroach extract models,MSCs overexpressing Miro1 were more effective inreducing airway hyper-responsiveness and airwayremodeling, whereas MSCs under-expressing Miro1were less effective than control MSC.

Study limitations

It was unfortunate that data were often presented ingraphs but not in the numerical form that wouldfacilitate a systematic review or a meta-analysis.Furthermore, the sample size was often unclear andreported as a range rather than an exact number. We

1640 N. Srour & B. Th�ebaud

attempted to obtain the required data from the au-thors, but few replied. The significance of this isunclear. Sample size and patient selection and groupassignment is of utmost importance in clinicalstudies. Issues such as these in a clinical study wouldraise concern about selection bias (for instance, wasone subject excluded from an experimental group fora reason that could affect the outcome?), but theseanimal models involve genetically identical in-dividuals with a fairly unchanging environment.Regardless, inadequate reporting has been describedas wasteful [33], and more rigorous reporting andcompliance with the ARRIVE guidelines [34] shouldcertainly be encouraged.

The studies included in this review were het-erogeneous in their design, including the type ofMSC used (murine, rat, human, induced pluripo-tent cells, syngeneic or allogeneic cells), the timingof MSC administration (before sensitization with anallergen, before allergen challenge and after allergenchallenge), the duration of the study and the sensi-tizing agent. For all these reasons, a meta-analysiswould not have been valid. However, given thateosinophilic inflammation was clearly decreased bymeans of MSC administration in almost all studies,the heterogeneity in study design, rather than beingdetrimental, only serves to strengthen this conclu-sion. The funnel plot for BAL total cell count isatypical and does not suggest obvious publicationbias. More importantly, the funnel plot for BALeosinophil count is as expected and does not suggestpublication bias. In any case, the validity of funnelplot analysis is questionable, given the heterogeneityof the study design. For the same reason, the use ofthe I2 statistic to assess heterogeneity in effect size isirrelevant.

Another limitation is that many studies did notuse nonestem cell types as controls. There arestudies showing partial responses to fibroblasts[9,20], and one study shows a better response tomononuclear cells than to MSCs [26]. However, theonly study that used nonestem cells (macrophages)as controls administered after allergen challengefound that they had no effect [15]. Further data aretherefore needed to determine whether other celltypes could be used instead of MSCs, during allphases of the allergic response.

We note that histology was assessed visually,qualitatively or semi-quantitatively inmost studies butwere only assessed quantitatively in 6 studies. Blindingof the histology evaluator, as would be expected inclinical studies, would have been ideal. Nevertheless,the conclusion that MSC treatment decreases eosin-ophilic inflammation is supported by the demonstra-tion in several studies of downregulation of Th2

inflammation and by the plausible mechanisms thathave been uncovered.

In contrast to embryonic stem cells and iPSCs,MSC treatment does not tend to result in teratomaformation [35]. The Wang [18] study used iPSCswithout c-myc; no tumor formation was found 6months after transplantation. Other concerns mayinclude clumping of the stem cells in the pulmonaryvasculature when administered intravenously and theintroduction of foreign antigens present in culturemedia. We note that intratracheal administration isfeasible because it was used successfully in a fewstudies [8,11,23,24]. MSC treatment has beenevaluated in human clinical trials for several otherconditions including acute myocardial infarction[36], stroke, diabetes and multiple sclerosis [37] andfor cartilage repair [38], with no particular safetyconcerns emerging, including tumors. This includes85 patients with a follow-up of 5 years [39] and 41patients with follow-up of 5e137 months (mean, 75months) [38].

Conclusions

In summary, MSCs decrease eosinophilic and Th2inflammation and are effective in several phases ofthe allergic response. MSCs derived from sourcesother than the bone marrow may also be effective.The mechanism may be different, depending on thephase. There is some evidence that soluble factorsmay be used to reproduce the effects of MSCs duringthe sensitization and challenge phases, but there areno data on their effectiveness during the post-allergen challenge phase. Some data suggest thatnonestem cell types may also be useful, but furtherdata are needed. We note that a randomized,controlled trial of MSCs for patients with chronicobstructive pulmonary disease has been completed[40]. There was no evidence of clinical efficacy,although the study was not powered for that, butMSC treatment did result in a reduction of C-reac-tive protein levels and, most importantly, appeared tobe safe. This was the first human trial of stem cellsfor any lung disease. The observed reduction ininflammation in this trial is relevant to asthma andpaves the way for future stem cell trials in lung dis-ease, including asthma.

Acknowledgments

The authors wish to thank Dr Qing-Ling Fu, Dr EvaMezey, Dr Daniel Weiss, Dr Meagan Goodwin, DrMelissa Lathrop and D. Anurag Agrawal forproviding the numerical data for their respectivestudies. This study was not funded.

Systematic review: stem cells in animal asthma models 1641

Disclosure of interests: The authors have nocommercial, proprietary, or financial interest in theproducts or companies described in this article.

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