TGF-β1 mediates pathologic changes of secondary lymphedema by promoting fibrosis and inflammation

Abstract

Background: Secondary lymphedema is a common complication of cancer treatment, and previous studies have shown that the expression of transforming growth factor-beta 1 (TGF-β1), a pro-fibrotic and anti-lymphangiogenic growth factor, is increased in this disease. Inhibition of TGF-β1 decreases the severity of the disease in mouse models; however, the mechanisms that regulate this improvement remain unknown.

Methods: Expression of TGF-β1 and extracellular matrix molecules (ECM) was assessed in biopsy specimens from patients with unilateral breast cancer-related lymphedema (BCRL). The effects of TGF-β1 inhibition using neutralizing antibodies or a topical formulation of pirfenidone (PFD) were analyzed in mouse models of lymphedema. We also assessed the direct effects of TGF-β1 on lymphatic endothelial cells (LECs) using transgenic mice that expressed a dominant-negative TGF-β receptor selectively on LECs (LEC^DN-RII ).

Results: The expression of TGF-β1 and ECM molecules is significantly increased in BCRL skin biopsies. Inhibition of TGF-β1 in mouse models of lymphedema using neutralizing antibodies or with topical PFD decreased ECM deposition, increased the formation of collateral lymphatics, and inhibited infiltration of T cells. In vitro studies showed that TGF-β1 in lymphedematous tissues increases fibroblast, lymphatic endothelial cell (LEC), and lymphatic smooth muscle cell stiffness. Knockdown of TGF-β1 responsiveness in LEC^DN-RII resulted in increased lymphangiogenesis and collateral lymphatic formation; however, ECM deposition and fibrosis persisted, and the severity of lymphedema was indistinguishable from controls.

Conclusions: Our results show that TGF-β1 is an essential regulator of ECM deposition in secondary lymphedema and that inhibition of this response is a promising means of treating lymphedema.

Keywords: TGF-β; fibrosis; inflammation; pirfenidone.