1481 - Expression Profiling of Skin and Lung Tissue and Explanted Fibroblasts in a Transgenic Mouse Model of Scleroderma

Monday, November 7, 2011: 9:00 AM-6:00 PM
Hall F2 - Poster Hall (McCormick Place West)
Emma Derrett-Smith1, Rachel Hoyles1, Pia Moinzadeh1, Cecilia B. Chighizola1, Korsa Khan1, Voon Ong2, David J. Abraham1 and Christopher P. Denton1, 1UCL Medical School, London, United Kingdom, 2UCL Medical School, London, England
Presentation Number: 1481

Background/Purpose: Gene expression profiling of skin or lung tissue and fibroblasts in explant culture have been used to study intrinsic subsets and pathogenic mechanisms in systemic sclerosis (SSc) [1]. There are technical challenges integrating the results of whole tissue and fibroblasts, although the methods are often complementary.  We have applied a similar strategy to analysis of a transgenic mouse model that is a phenocopy of many of the histological and biochemical features of SSc.  This mouse strain has ligand-dependent upregulation of TGFb signalling due to altered TbRII receptor expression in fibroblasts.

Methods: In the present study we have analysed gene expression profiles of whole skin and lung from littermate TbRIIdk-fib mice and fibroblasts cultured from neonatal or adult skin and lung tissue (n≥3 in each group) using the illumina microarray platform. RNA was extracted, quantified and assessed for quality using standard methods. Technical validation of the data and additional quantitation of key gene expression was performed using quantitative RT-PCR assay using replicate samples.

Results: Cluster analysis identifies key gene profiles that are specific for skin or lung fibroblasts and also that are altered in whole tissues. In general the differential gene expression was much more marked in whole tissue and differences were more marked in neonatal compared with adult fibroblasts consistent with the higher levels of transgene expression previously described in younger mice. In particular, genes related to cytoskeletal and extracellular matrix structure and function (aSMA, troponin, tropomyosin 1, collagens type I, III, VI, VIII, XVII, matrix metalloproteinases 3, 9, 10, 13, 17, Timp3), endothelin (endothelin-1, Ednrb, Ednra), TGFb (Ltbp1, TGFb1, 2, 3, Ctgf), BMP (Bmp2, 4, Bmpr1) and VEGF (Vegfa, Vegfc) signalling axes and innate immunity (Il-6, Il-11, Il-13, Il-1r, Crp, Saa) were found to be differentially expressed both in transgenic whole skin, lung and explanted fibroblasts. In addition, genes coding for Pecam1 (p=0.03) and Elastin (p=0.003) were upregulated strongly in whole lung and skin. Some of these key genes that demonstrated significantly dysregulated expression in transgenic mouse skin and lung are summarised in more detail in Table 1.

Conclusions: These data are reminiscent of studies of human SSc tissue and illustrate another potential complementary strength for mouse models in better understanding the disease. 

1. Gardner H, Shearstone JR, Bandaru R, Crowell T, Lynes M, Trojanowska M, Pannu J, Smith E, Jablonska S, Blaszczyk M, Tan FK, Mayes MD. Gene profiling of scleroderma skin reveals robust signatures of disease that are imperfectly reflected in the transcript profiles of explanted fibroblasts. Arthritis Rheum. 2006 Jun;54(6):1961-73.

Table 1: Representative genes that demonstrate dysregulated expression in transgenic skin and lung

Gene

Target tissue

Relative transgenic expression

P value

Mus musculus matrix metalloproteinase 3 (Mmp3), mRNA

Lung fibroblast

1.94

0.02

Mus musculus transforming growth factor beta 1 (Tgfb1), mRNA

Lung fibroblast

1.14

0.01

Mus musculus  pleiotrophin (Ptn), mRNA

Skin fibroblast

-0.59

0.1

Mus musculus dual specificity phosphatase 1 (Dusp1), mRNA

Skin fibroblast

0.28

0.0004

Mus musculus homeo box B7 (Hoxb7), mRNA

Skin fibroblast

-0.49

0.01

Mus musculus annexin A1 (Anxa1), mRNA

Whole lung

-0.43

0.05

Mus musculus integrin alpha 6 (Itga6), mRNA

Whole lung

-1.89

0.02

Mus musculus collagen, type XII, alpha 1 (Col12a1), mRNA

Whole skin

0.82

0.01

Mus musculus similar to Fibrillarin, transcript variant 1 (LOC100044829)

Whole skin

0.40

0.02

Mus musculus vascular endothelial growth factor (Vegfa) transcript variant 2, mRNA

Whole skin

-1.86

0.05


Keywords: animal models, scleroderma and transforming growth factor

Disclosure: E. Derrett-Smith, None; R. Hoyles, None; P. Moinzadeh, None; C. B. Chighizola, None; K. Khan, None; V. Ong, None; D. J. Abraham, None; C. P. Denton, Actelion Pharmaceuticals US, 5 .