SMAD4-Dependent Signaling Pathway Involves in the Pathogenesis of TGFBR2-Related CE-like Phenotype

(1) Background: Our previous data indicated that disturbance of the Transforming Growth Factor beta (TGFB) signaling pathway via its Type-2 Receptor (TGFBR2) can cause a Corneal Ectasia (CE)-like phenotype. The purpose of this study is to elucidate whether the SMAD4-dependent signaling pathway is involved in the TGFBR2-related CE-like pathogenesis. (2) Methods: Smad4 was designed to be conditionally knocked out from keratocytes. Novel triple transgenic mice, Kera(rtTA); Tet-O-Cre; Smad4(flox/flox) (Smad(4kera-cko)), were administered with doxycycline (Dox). Optical Coherence Tomography (OCT) was performed to examine Central Corneal Thickness (CCT), Corneal Radius, Anterior Chamber and CE-like phenotype and compared to the littermate Control group (Smad4(Ctrl)). (3) Results: The OCT revealed normal cornea in the Smad4(Ctrl )and a CE-like phenotype in the Smad4kera-cko cornea, in which the overall CCT in Smad4(kera-cko) was thinner than that of Smad4(Ctrl) at P42 (n = 6, p < 0.0001) and showed no significant difference when compared to that in Tgfbr2(kera-cko). Furthermore, the measurements of the Anterior Chamber and Corneal Radius indicated a substantial ectatic cornea in the Smad4kera-cko compared to Smad4(Ctrl). The H&E staining of Smad4(kera-cko) mimics the finding in the Tgfbr2(kera-cko). The positive immunostaining of cornea-specific marker K12 indicating the cell fate of cornea epithelium remained unchanged in Smad4kera-cko and the Proliferating Cell Nuclear Antigen (PCNA) immunostaining further indicated an enhanced proliferation in the Smad4(kera-cko). Both immunostainings recapitulated the finding in Tgfbr2(kera-cko). The Masson's Trichrome staining revealed decreased collagen formation in the corneal stroma from both Smad4(kera-cko) and Tgfbr2(kera-cko). The collagen type 1 (Col1a1) immunostaining further confirmed the reduction in collagen type 1 formation in Smad4kera-cko. (4) Conclusions: The aforementioned phenotypes in the Smad4(kera-cko) strain indicated that the SMAD4-dependent signaling pathway is involved in the pathogenesis of the CE-like phenotype observed in Tgfbr2(kera-cko).