Transcript Document 3053504
Asthmatic Phenotypes in the PhysGen Consomic Rats
UW-MCW Collaboration: Project Update L.A. Rosenthal, R.L. Sorkness, and R.F. Lemanske, Jr.
Morris Institute for Respiratory Research
Genetic Factors
Atopy (Cytokine Dysregulation) AGE
Environmental Factors
Respiratory Viruses Allergens
Asthma Phenotype
Post-bronchiolitis Asthma-like Phenotype aerosolized parainfluenza type 1 (Sendai) virus weanling rats (3-4 weeks old) Brown Norway (BN) high IgE producer “atopic” (Th2) Fischer 344 (F344) low IgE producer “nonatopic” (Th1) 8-10 weeks postinfection Abnormal Airways
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episodic, reversible airway obstruction
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chronic airway inflammation
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airway hyperresponsiveness to methacholine
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airway wall remodeling Normal Airways Model developed in collaboration with W.L. Castleman (U. of Florida)
Genetic Factor
Rat Strain AGE
Environmental Factor
Sendai Virus
Asthmatic Phenotype
Development of the Post-bronchiolitis Asthma-like Phenotype in Brown Norway Rats: Host Response
(Genetic Factor)
to Sendai Virus
(Environmental Factor)
During Early Life (
Developmental Factor
) Drives Process
Acute Phase
BN and F344: Bronchiolitis Dysregulation of innate immune response?
Chronic Phase
BN: YES F344: NO Sendai Virus Infection d2 Viral replication d6-8 Virus shedding Abnormal resolution of acute inflammatory changes?
d10-14 Resolution of acute inflammatory changes > Week 4 Development of asthma-like phenotype
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episodic, reversible airway obstruction
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chronic airway inflammation
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airway hyperresponsiveness
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airway wall remodeling
Development of Asthma-like Phenotype: Importance of IFN-
g
During Acute Phase
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IFN-
g
in bronchoalveolar lavage fluid during acute Sendai virus infection in weanling rats: F344 > BN
•
Aerosolized IFN-
g
prevents the development of post-bronchiolitis chronic airway dysfunction in BN rats.
Post-bronchiolitis Asthma-like Phenotype IFN-
g
Brown Norway (BN) high IgE producer “atopic” (Th2) aerosolized parainfluenza type 1 (Sendai) virus weanling rats (3-4 weeks old) Fischer 344 (F344) low IgE producer “nonatopic” (Th1) 8-10 weeks postinfection Normal Airways Sorkness et al. 1999. AJRCCM 160:705 Normal Airways
PhysGen Consomic Rats
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Parental strains for consomic rats:
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Dahl salt sensitive (SS; hypertension model)
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Fawn hooded hypertensive (FHH; hypertension and end-stage renal disease model)
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Brown Norway (BN; control strain: normotensive)
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Useful for studying the role of genetic factors in susceptibility/resistance to the development of the post-bronchiolitis asthma-like phenotype?
Balance Between Th1- and Th2-type Cytokines Th2 Th1 Th1 F344 SS?
FHH?
“non-atopic” BN “atopic” Th1-type: IFN-
g
, IL-12; Th2-type: IL-4, IL-5, IL-13 Th2
Are the Immune Responses of SS and FHH Rats Th1-biased Like Those of F344 Rats?
Screening Assay for Th1- and Th2-type Cytokine Production by Spleen Cells Con A 24 h
A B C D E F 10 9 8 7 6 5 4 3 2 1
Measure Cytokine Levels in Supernatant Fluids
IFN-
g
Production by Con A-stimulated Spleen Cells 8 7 6 5 4 3 2 1 0 BN SS FHH
IL-4 Production by Con A-stimulated Spleen Cells 16 14 12 10 8 6 4 2 0 BN SS FHH
IFN-
g
/IL-4 Ratio for Con A-stimulated Spleen Cells 1000 900 800 700 600 500 400 300 200 100 0 BN SS FHH
IL-13 Production by Con A-stimulated Spleen Cells 20 15 10 5 0 65 60 55 50 45 40 35 30 25 BN SS FHH
IFN-
g
/IL-13 Ratio for Con A-stimulated Spleen Cells 700 600 500 400 300 200 100 0 BN SS FHH
Balance Between Th1- and Th2-type Cytokines Th2 Th1 Th1 F344 SS FHH “non-atopic” BN “atopic” Th1-type: IFN-
g
, IL-12; Th2-type: IL-4, IL-5, IL-13 Th2
Are SS Rats, Like F344 Rats, Resistant to the Development of Post-bronchiolitis Airway Dysfunction?
Post-bronchiolitis Airway Dysfunction in BN, but not SS, Rats
Therefore, in vitro cytokine data and in vivo physiology data suggest that the PhysGen consomic rat strains will be useful tools for studying the role of genetic factors in susceptibility/resistance to the development of the post-bronchiolitis asthma-like phenotype.
Screening of Consomic Rat Strains
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SS.BN13
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SS.BN16
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SS.BN18
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SS.BN9
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SS.BN20
Screening Assay for Th1- and Th2-type Cytokine Production by Spleen Cells Con A 24 h
A B C D E F 10 9 8 7 6 5 4 3 2 1
Measure Cytokine Levels in Supernatant Fluids
IFN-
g
Production by Con A-stimulated Spleen Cells 10 1 BN SS 13 16 18 SS.BN
9 20
Th1 and Th2 Cytokine Production
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No strain differences were observed among SS and these five SS.BN strains with regard to Con A-stimulated IFN-
g
production [or IL-4 or IL-13 production (not shown)] .
Chr. 20: Major Histocompatibility Complex
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SS.BN20
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Histocompatible with BN?
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Histoincompatible with SS?
Strain A Strain B Two-way Mixed Lymphocyte Cultures 72 h
A B C D E F 10 9 8 7 6 5 4 3 2 1
Measure IL-2 and IFN-
g
Levels in Supernatant Fluids
IL-2 Production in 2-way Mixed Lymphocyte Cultures: BN and SS.BN20 Splenocytes are Histocompatible BN + BN F344 + F344 SS.BN20
+ SS.BN20
BN + F344 F344 + SS.BN20
BN + SS.BN20
15 20 25 30 35 IL-2 (pg/ml) 40 45
IL-2 Production in 2-way Mixed Lymphocyte Cultures: SS and SS.BN20 Splenocytes are Histoincompatible BN + BN SS + SS SS.BN20
+ SS.BN20
BN + SS SS + SS.BN20
BN + SS.BN20
16 18 20 22 24 26 IL-2 (pg/ml) 28 30 32 34
IFN-
g
Production in 2-way Mixed Lymphocyte Cultures: SS and SS.BN20 Splenocytes are Histoincompatible BN + BN SS + SS SS.BN20
+ SS.BN20
BN + SS SS + SS.BN20
BN + SS.BN20
0 100 200 300 IFN-
g
(pg/ml) 400 500
Potential Uses of the Histocompatible SS.BN20 Strain for Studies in BN Rats
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in vitro
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cell mixing studies
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in vivo
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adoptive cell transfer studies
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organ transplant studies
PhysGen Consomic Rat Development Timeline
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SS.BN12
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FHH.BN1 (IL-4R
a
)
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SS.BN6 (Immunoglobulin)
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SS.BN7 (IFN-
g
)
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SS.BN8
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Fischer CDF(F-344)CrlBR
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BN/SsNHsd/Mcw http://pga.mcw.edu
Future Directions
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Continue screening spleens from adult consomic rats.
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PhysGen consomic rat strains to be distributed through Charles River Laboratories.
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Availability of weanling consomic rats will significantly increase options with regard to screening.
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Level of characterization of weanlings from each consomic strain? Resources that will inform these decisions include:
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Rat Genome Database; Rat Genome Sequencing Project
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Virtual Comparative Maps
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MCW-TIGR microarray expression profiling of rat consomic panels (lung)
Funded by: NIH/NIAID Asthma and Allergic Disease Research Center Grant AI050500