Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Aortic Smooth Muscle Cells Show a Pro-Fibrotic Phenotype in a TGF-Beta Dependent Mouse Model of Systemic Sclerosis
Derrett-Smith, Emma, Dooley, Audrey, Khan, Korsa, Shi-wen, Xu, Abraham, David, Denton, Christopher P.
In systemic sclerosis, the large elastic arteries have altered elasticity and compliance. We have examined the large systemic vessels in a mouse model of SSc characterised by ligand-dependent activation of TGF-b signalling in fibroblasts.
The transgenic mouse strain TbRIIDk-fib expresses a kinase-deficient type II TGF-b receptor linked to a fibroblast-specific promoter leading to balanced ligand-dependent upregulation of TGF-b signaling. Biological replicate samples from transgenic (n=6) or wildtype littermate control mice (n=6) were compared. Aortic and cardiac tissue were examined by histological, biochemical and isolated organ bath studies. Vascular and perivascular architecture was examined by H&E and special stains including immunostaining for TGF-b1 and pSmad 2/3. Confirmatory aortic smooth muscle cell proliferation, phenotype and functional assays, including signalling responses to exogenous TGF-b and endothelin-1 were performed. Aortic ring contractile responses to direct and receptor-mediated stimulation were assessed.
TGF-b1 and pSmad 2/3 staining were increased in transgenic aortic adventitia, which was increased in diameter with an associated reduction in smooth muscle layer. Non cross-linked collagen content of transgenic thoracic aortic tissue measured by Sircol assay was increased compared to wildtype (mean transgenic collagen content 22.5±1.87 mg/ml, mean wt 12.4±0.45 mg/ml, P<0.05). Transgenic aortic smooth muscle cells showed upregulation of TGF-b responsive genes important for cytoskeletal function, such as transgelin (mean transgenic copy number 1.06×106±6.6 x104, mean wildtype copy number 7.6×105±5.5×104, P<0.05) and smoothelin (mean transgenic copy number 11406±1306, mean wildtype copy number 5627±758, P<0.05), which were then resistant to further stimulation with exogenous TGF-b1. Consistent with an activated phenotype, transgenic SMC promoted significantly more contraction of free floating type I collagen lattices when compared with wildtype, but there was no further contraction after TGF-b1 stimulation (see Figure 1).
Aortic ring responses to receptor-mediated contraction with endothelin were reduced in the transgenic animals: for instance, mean change in tension from baseline with 1×10-9endothelin-1 treatment in wildtype animals (n=3) was 21.4±9.0 mg, compared with -5.7±2.9 in transgenic animals (n=3), P<0.05. Bosentan reduced endothelin-mediated contraction in wild-type animals, but had no effect in transgenic animals. Endothelin receptor A gene expression was reduced in transgenic animals (mean wildtype copy number 2517±1261, mean transgenic copy number 315±73, p<0.05).
The histological, biochemical and functional phenotype of this transgenic mouse model of scleroderma offers insight into the altered biomechanical properties previously reported for large elastic arteries in human SSc, and supports a potential role for perturbed TGF-b and endothelin activity in this process.
To cite this abstract, please use the following information:
Derrett-Smith, Emma, Dooley, Audrey, Khan, Korsa, Shi-wen, Xu, Abraham, David, Denton, Christopher P.; Aortic Smooth Muscle Cells Show a Pro-Fibrotic Phenotype in a TGF-Beta Dependent Mouse Model of Systemic Sclerosis [abstract]. Arthritis Rheum 2009;60 Suppl 10 :1061