Abstract
Purpose::
Previous work from our laboratory has shown using a transforming growth factor ß (TGFß) transgenic/Smad3 knockout (KO) mouse model that the epithelial-to-mesenchymal transition of lens epithelial cells observed in anterior subcapsular cataracts (ASC) can occur in the absence of Smad3 signalling. However, the cataractous plaques that developed in these mice were reduced in size compared to wild-type littermates. Here, we investigate whether genes known to be altered in models of ASC, including the matrix metalloproteinases, MMP-2 and MMP-9, and E-cadherin, are differentially expressed in the TGFß1 transgenic/Smad3 KO mouse lenses relative to TGFß1 transgenics on the wild-type background.
Methods::
TGFß1 transgenic mice were bred with Smad3 null and heterozygote mice to generate mice with the following genotypes: TGFß1/Smad3-/- (null), TGFß1/Smad3+/-, TGFß1/Smad3+/+ and non-transgenic/Smad+/+ (controls). Lenses from mice 2-3 months old of each genotype were dissected and subjected to real time polymerase chain reaction (RT-PCR) analyses for MMP-2, MMP-9 and E-cadherin mRNA.
Results::
Lenses from TGFß1/Smad3+/+ and TGFß1/Smad3+/-, were found upon dissection, to exhibit distinct fibrotic plaques. These lenses expressed upregulated levels of both MMP-2 and MMP-9 as compared to lenses from control mice. In comparison, TGFß1/Smad3-/-, with reduced plaque formation, displayed levels of MMP-2 and MMP-9 mRNA, did not reach levels observed in transgenic mice on Smad3 heterozygote and wild-type backgrounds. Furthermore, lenses from TGFß1/Smad3+/+ and TGFß1/Smad 3+/- mice exhibited a reduction in E-cadherin mRNA compared to control mice, whereas E-cadherin mRNA levels from TGFß1/Smad3-/- lenses resembled levels in control lenses.
Conclusions::
These findings demonstrate that in the absence of Smad3 signaling, induction of MMP-2 and MMP-9 mRNA by TGFß1 is reduced. Also, E-cadherin mRNA levels are maintained at control levels in the TGFß1 transgenics on the Smad3 KO background. Further studies will be aimed at determining whether Smad3 is a direct signaling intermediate regulating expression of these three genes by TGFß.
Keywords: cataract • transgenics/knock-outs • EMT (epithelial mesenchymal transition)