The findings we have presented here reveal a novel mechanism by which the multifunctional receptor LRP1 functions as an integrator of two distinct cellular signaling pathways in the vascular wall. They also suggest a functional relationship between the molecular events that underlie MFS, the formation of aneurysms and atherosclerosis. Our results show that LRP1 controls Smad2/3-as well as PDGF-dependent signaling in VSMC in a coordinated fashion and by distinct mechanisms.
LRP1 is a target of PDGFRβ-dependent Src kinase activation 
and we showed earlier that loss of LRP1 expression in VSMCs results in increased PDGFRβ expression and activation 
. In the present study we have demonstrated, that loss of LRP1 expression also results in increased Smad2/3-dependent signaling in VSMCs. A potent inducer of Smad2 phosphorylation and nuclear translocation is TGFβ 
. Moreover, induction of PDGF as well as PDGFRβ expression in response to TGFβ stimulation has been recognized as an important component of epithelial-mesenchymal transition, a process that underlies metastasis of some tumors 
and increases the aggressiveness of others 
. Our results show that increased pSmad2/3 signaling in the LRP1-deficient vascular wall results in increased PDGFRβ expression and activation, making it likely that TGFβ is directly involved.
Furthermore, LRP1/TGFβR(V), can directly bind TGFβ1 
, as well as enter into a complex with TGFβR(I) 
. By binding and endocytosis of the active growth factor and also by sequestering TGFβR(I) away from TGFβR(II), LRP1 is in a central position where it can suppress the normal mode of TGFβ signaling and Smad2/3 activation through the TGFβR(I)/R(II) complex. In addition, LRP1 mediates the endocytosis of TSP1 as well as several matrix metalloproteinases (MMP) from the extracellular space 
, and TSP1 as well as active MMP2 and MMP9 levels are increased in smLRP deficient aortas (, and 
). TSP1 and MMPs potently and independently activate TGFβ1 
. Loss of smLRP would thus be predicted to activate TGFβ/Smad signaling in VSCM by multiple independent mechanisms, and this is consistent with the dramatically increased nuclear pSmad2/3 levels in smLRP−
aortas (). The resulting disruption of elastic layers, vascular fibrosis, elongation and distension of the aorta, and high incidence of aortic aneurysms directly support the pivotal role of TGFβ signaling in the pathogenesis of MFS and reveal LRP1 as a critical regulator of this pathway in vivo. As such, LRP1 could conceivably contribute as a genetic modifier to the variable severity of expression of MFS in humans.
We have further demonstrated the importance of TGFβ signaling through pharmacological intervention and reversal of the smLRP−
phenotype with the PPARγ agonist rosiglitazone. Activation of PPARγ with thiazolidinediones potently inhibits TGFβ signaling in vitro and in vivo 
. The striking effect of rosiglitazone on the nuclear accumulation of pSmad2/3 and the near complete reversal of the aortic wall thickening and fibrosis, and prevention of atherosclerotic lesion progression confirm the central importance of TGFβ pathway activation for this phenotype. PPARγ agonists might thus complement and enhance the potential therapeutic use of the angiotensin II type 1 receptor blocker Losartan in the treatment of MFS 
Increased susceptibility to atherosclerosis is not a part of the MFS complex, but a striking feature of smLRP−
. Likewise, PPARγ agonists are only partially effective against diet-induced atherosclerosis in LDLR deficient mice when LRP1 is functional 
. By contrast, we have shown that rosiglitazone (this study) as well as the PDGFR tyrosine kinase inhibitor Gleevec 
are highly potent in protecting smLRP−
mice from atherosclerosis. Our findings thus suggest that abnormal activation of PDGFRβ signaling, which has so far not been reported in MFS, is the major mediator of the increased atherosclerosis in the smLRP−
mice, and that increased PDGFRβ activity is brought about by two independent and synergistic mechanisms. First, activation of Smad/TGFβ signals in the absence of LRP1 results in the disruption of elastic layers and a tortuous aorta 
, as well as increased expression of PDGF receptors ( and ) 
. However, since LRP1 also controls PDGFRβ activity and recycling at the level of the plasma membrane 
, loss of LRP1 in smooth muscle cells also results in a greatly increased mitogenic response, VSMC proliferation and the promotion of atherosclerotic lesion formation. This second event is prevented in MFS by the presence of LRP1, which suppresses the abnormal PDGFRβ activation through a posttranslational mechanism 
. Thus, rosiglitazone (as a Smad/TGFβ antagonist) and Gleevec (as a PDGFRβ inhibitor) act at different steps in a linear sequence of events () that result in a Marfan-like syndrome and culminate in extensive atherosclerosis in smLRP−
Control of TGFβ and PDGF signaling and protection of vascular wall integrity by LRP1.
Nevertheless, blockade of PDGFRβ signaling alone by administration of Gleevec was sufficient to reduce atherosclerotic lesion progression, as well as vascular wall thickness, in smLRP−
, suggesting that signaling through this pathway is necessary for elastic layer disruption and wall thickening. This raises the possibility that PDGFRβ signaling may be activated to some extent in the Marfan vessel wall, or alternatively the mere suppression of its baseline activity may be sufficient to normalize the vessel wall thickening that is initiated by increased TGFβ activity. Consistent with an important role of PDGFRβ in vascular thickening is a recent report that showed transphosphorylation of a truncated PDGFRβ induced by the Angiotensin II type 1 receptor (AT1) 
, which can also activate Smad2-dependent signals independent of TGFβ 
. AT1 is inhibited by Losartan, a drug that also reduced wall thickening in a mouse model of Marfan syndrome 
. Taken together, these findings indicate a critical role for Smad2-and AT1-dependent signaling mechanisms in Marfan and Marfan-like syndromes and identify PDGFRβ as another important component of the pathogenic mechanism. Our findings also suggest that rosiglitazone, a clinically approved antidiabetic drug, may be useful for the treatment of vasculopathies that are caused by increased TGFβ signaling, e.g. Marfan syndrome, as well as for the supportive treatment of certain malignancies, such as malignant glioblastomas