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Sandeep Kumar, Alex Jones, Zach Berriochoa, Shixian Wang Wang, Yingbin Fu; The proteolytic activity of HTRA1 is responsible for its pathological role in polypoidal choroidal vasculopathy. Invest. Ophthalmol. Vis. Sci. 2013;54(15):4991.
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© ARVO (1962-2015); The Authors (2016-present)
We recently reported the generation of the first PCV model by transgenically expressing human HTRA1, a multi-functional serine protease, in mouse retinal pigment epithelium (RPE). We showed that increased HTRA1 induced characteristic features of PCV, including branching networks of choroidal vessels and polypoidal lesions. Transgenic hHTRA1+ mice also developed occult CNV. To test the hypothesis that the proteolytic activity of HTRA1 is responsible for its pathological role in PCV, we have generated transgenic mice expressing the protease inactive mutant of HTRA1 (S328A) in mouse RPE. In addition, we generated an improved transgenic HTRA1 mouse model Tg44, with more consistent phenotypes.
The expression of HTRA1 and HTRA1-S328A was determined by western blotting and immunohistochemistry. The phenotypes of various HTRA1 mice were examined by fluorescein angiography (FA), indocyanine green angiography (ICGA), fundus imaging, spectral-domain optical coherence tomography (SD-OCT), histology and electron microscopy (EM).
We have generated two S328A mouse lines (Tg26 and Tg33) and one HTRA1 mouse line Tg44. The protein level of HTRA1-S328 is ~ 2-3 times that of HTRA1 in Tg44. The protein level of HTRA1-S328 in Tg33 is similar to that of HTRA1 in Tg44. All transgenic proteins were expressed in mouse RPE. On ICGA, Tg44 mice exhibited characteristic features of PCV: 1) late geographic hyperfluorescence and hyperfluorescent plaque; 2) polypoidal lesions (polyp and branching vascular network). SD-OCT depicted RPE atrophy in lesion areas. Color fundus photograph showed hemorrhagic pigment epithelial detachment and scar formation at polypoidal lesions. Histology staining revealed serous exudation, thin-walled choroidal vessels, and regional loss of choriocapillaris. EM analysis revealed deteriorative changes in the elastic lamina of Bruch’s membrane, RPE, photoreceptors and choroid vasculature. In contrast to Tg44 mice, neither Tg26 nor Tg33 mice expressing HTRA1-S328A show any PCV phenotypes.
Our results provide strong evidence that the proteolytic activity of HTRA1 is responsible for its pathological role in PCV. The transgenic HTRA1 mice exhibited cardinal features of human PCV, which will be invaluable for further studies on the mechanisms and treatment strategies of PCV.
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