J ALLERGY CLIN IMMUNOL

VOLUME 133, NUMBER 2

Abstracts AB53

SATURDAY

188 TGF-beta1 Mobilizes Mesenchymal Stem Cells In Allergic

AsthmaTing Xu, MD1,2, Ling-Ling Xian, MD, PhD3, Yufeng Zhou, MD, PhD1,

Beverly Plunkett, MS1, Xu Cao, PhD3, Mei Wan, MD, PhD3, Peisong

Gao, MD, PhD1; 1Division of Allergy & Clinical Immunology, Johns

Hopkins University School of Medicine, Baltimore, MD, 2Department

of Respiratory Medicine, Southern Medical University, Guangzhou,

China, 3Department of Orthopedics Surgery, Johns Hopkins University

School of Medicine, Baltimore, MD.

RATIONALE: Mesenchymal stem cells (MSCs) have been suggested to

participate in airway repair/remodeling. Transforming growth factor b1

(TGFb1) is critical in the recruitment of stem/progenitor cells for tissue

repair, regeneration, and remodeling. We here sought to investigate

whether TGFb1 plays a role in MSC migration in allergic asthma.

METHODS: MSC migration was examined by using conditioned me-

dium (ECM) from cockroach extract (CRE)-challenged human bronchial

epithelial cells (HBECs) in air/liquid interface (ALI) culture. The

involvement of TGFb1 in ECM-induced MSC migration was investigated

by using ECM containing TGFb1 neutralizing antibody or control IgG

antibody. Furthermore, the migration of MSCs to CRE-challenged airway

and the effect of TGFb on MSC migration were examined in nestin-GFP

transgenic mice.

RESULTS: MSC migration was significantly enhanced by ECM from

cockroach extract-challenged HBECs when compared with those from un-

challenged cells. The increased migration was inhibited by antibodies

against TGFb1. Significant inhibition was also observed with TbR1

inhibitor use. MSCs from bone marrow of GFP-transgenic mice demon-

strated their ability to differentiate into fibro/myo-fibroblasts. Furthermore,

when these isolated GFP+-MSCs were injected systematically, the number

of migrated GFP+-MSCs into lung was significantly increased after CRE

challenge. Significantly greater numbers of endogenous GFP+ cells were

observed in allergen-challenged lung of nestin-GFP transgenic mice

when compared with those receiving saline alone. Interestingly, these

increased MSCs could be inhibited when TGFb1 neutralizing antibody

was utilized.

CONCLUSIONS: These results provide evidence supporting a role of

TGFb1 in MSCmigration, suggesting the importance of MSCs in allergen

induced airway repair/remodeling in asthma.

189 IL-10-Producing B Cells Are Increased After Grass PollenImmunotherapy Compared To Untreated Grass Pollen AllergicControls: A Blinded Cross-Sectional Study

Mr. James E. G. Charlesworth1,2, Dr. Guy W. Scadding, MD1,2,

Dr. Aarif Eifan, MD2, Ms. Rachel Yan, RN2, Ms. Andrea

Goldstone, RN2, Dr. Moises A. Calderon, MD, PhD1,2, Prof. Stephen R.

Durham, MA, MD, FRCP3,4, Dr. Mohamed H. Shamji, BSc, MSc,

PhD5; 1Medical Research Council and Asthma UK Centre for Allergic

Mechanisms of Asthma, United Kingdom, 2Imperial College London,

United Kingdom, 3Imperial College London, London, United Kingdom,4Medical Research Council and Asthma UK Centre for Allergic Mecha-

nisms of Asthma, London, United Kingdom, 5Imperial College London,

South Kensington, United Kingdom.

RATIONALE: Grass pollen immunotherapy (AIT) induces Foxp3 and/or

IL-10-expressing regulatory T cells and increases in allergen-specific

IgG4.We previously showed that in vitrostimulation with CpG induced IL-

10-competent B cells capable of suppressing Th2 responses. This study ex-

amines IL-10-producing B cells (Bregs) during AIT.

METHODS: 14 grass pollen allergics receiving AIT (8 subcutaneous

(SCIT) and 6 sublingual (SLIT) patients), 14 untreated allergics (SAR) and

14 non-allergic (NA) controls were studied. Blood was drawn for

quantification of IL-10-producing B cells following CpG stimulation by

FluoroSpot assay. IL-10 supernatant concentrations were measured by

ELISA and mRNA responses by qPCR. Blood was also obtained pre and 6

hours post nasal allergen challenge (NAC) for B cell phenotyping by flow

cytometry.

RESULTS: Allergic individuals trended towards decreased IL-10 mRNA

expression 5 hours post CpG stimulation (p50.06) compared to NA

controls, whereas no difference in IL-10+ B cell frequency (p50.5) nor IL-

10 concentration (p50.24) was observed. Overall, AIT patients showed a

trend towards increased proportions of IL-10+ B cells compared to SAR

(p50.06), whereas the SLIT group had significantly greater proportions

of IL-10+ B cells and IL-10 concentrations relative to SAR (p50.0009

and p50.046). Following NAC, Breg subsets (CD19+CD24hiCD38hi,

CD19+CD5hi, CD19+CD24hiCD27hi and CD19+CD25+) significantly

increased in NA and AIT groups (all p<0.05), but not in the SAR group.

CONCLUSIONS: Grass pollen immunotherapy is associated with an

increase in peripheral IL-10+ B cells and appears to restore a normal pe-

ripheral regulatory B cell response following local nasal allergen chal-

lenge, compatible to that observed in non-allergic controls.

190 Respiratory Syncytial Virus Induces IL-25 and IL-33Production In The Lungs

Matthew T. Stier1, Kasia Goleniewska2, R. Stokes Peebles1,2; 1Depart-

ment of Pathology, Microbiology, and Immunology, Vanderbilt University

School of Medicine, Nashville, TN, 2Allergy, Pulmonary, and Critical

Care Medicine, Department of Medicine, Vanderbilt University School

of Medicine, Nashville, TN.

RATIONALE: Respiratory syncytial virus (RSV) is the leading cause of

hospitalization in infants. Early, severe RSV infection is also a risk factor

for the subsequent development of asthma. RSV infects the airway

epithelium, causing mucus production, epithelial cell sloughing, peribron-

chiolar inflammation, and increased airway hyperresponsiveness.

Epithelial-derived cytokines IL-25 and IL-33 stimulate IL-13 and mucus

production in asthma models. RSV infection produces similar Th2-like

phenotypes, suggesting a role for epithelial-derived cytokines in this

system. We sought to analyze the role of IL-25 and IL-33 during RSV

infection.

METHODS: Six to eight week old BALB/c mice were infected with

1.5E6 PFU of RSV clinical isolate strain 01/2-20 or vehicle. Mice were

harvested at 3hr, 6hr, 12hr, 24hr, 48hr, 72hr, and 96hr with the collection of

left lung, right lung, and bronchoalveolar lavage fluid. Transcript levels

were measured by quantitative PCR and normalized to GAPDH. Protein

concentrations were evaluated by ELISA (R&D DuoSet).

RESULTS: IL-25 transcript and protein were most abundant at 3 and 6

hours post infection, respectively. Transcript and protein levels of IL-33

peaked at 12 hours post infection.

CONCLUSIONS: These data suggest a role for IL-25 and IL-33 in the

RSV-associated immune response, prompting the need for additional

studies on these epithelial-derived cytokines in regard to viral clearance

and immunopathology.