Fibulin-4 deficiency increases TGF-beta signalling in aortic smooth muscle cells due to elevated TGF-beta 2 levels

Fibulins are extracellular matrix proteins associated with elastic fibres. Homozygous Fibulin-4 mutations lead to life-threatening abnormalities such as aortic aneurysms. Aortic aneurysms in Fibulin-4 mutant mice were associated with upregulation of TGF-beta signalling. How Fibulin-4 deficiency leads to deregulation of the TGF-beta pathway is largely unknown. Isolated aortic smooth muscle cells (SMCs) from Fibulin-4 deficient mice showed reduced growth, which could be reversed by treatment with TGF-beta neutralizing antibodies. In Fibulin-4 deficient SMCs increased TGF-beta signalling was detected using a transcriptional reporter assay and by increased SMAD2 phosphorylation. Next, we investigated if the increased activity was due to increased levels of the three TGF-beta isoforms. These data revealed slightly increased TGF-beta 1 and markedly increased TGF-beta 2 levels. Significantly increased TGF-beta 2 levels were also detectable in plasma from homozygous Fibulin-4(R/R) mice, not in wild type mice. TGF-beta 2 levels were reduced after losartan treatment, an angiotensin-II type-1 receptor blocker, known to prevent aortic aneurysm formation. In conclusion, we have shown increased TGF-beta signalling in isolated SMCs from Fibulin-4 deficient mouse aortas, not only caused by increased levels of TGF-beta 1, but especially TGF-beta 2. These data provide new insights in the molecular interaction between Fibulin-4 and TGF-beta pathway regulation in the pathogenesis of aortic aneurysms.