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V. E. Trinkaus-Randall, C. Mayo, R. Ren, C. Rich; The P2X7 Receptor Modulates Stromal Collagen Organization. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4043.
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P2X receptors are nucleotide gated ion channels that are expressed in the cornea. Previously we demonstrated that the P2X7 KO reduces epithelial migration and alters the expression of cadherin. As the wound margin was fragile we investigated the epithelial mesenchymal transition and the stroma of KO and WT. Our goal was to examine what role the receptor had in modulating the structure of the corneal stroma.
Corneas from P2X7 KO and WT mice were were fixed and embedded in paraffin for light microscopic and immunohistochemical analysis. Tissues were fixed and analyzed also at the electron microscopic level. Fibril diameter and interfibrillar spacing were measured in micrographs from non-overlapping regions of the anterior, mid and posterior stroma. Cuprolinic blue staining was performed on fixed tissue and sulfated glycosaminoglycans depicted as electron dense filaments were measured.
P2X7 KO mice cornea were transparent but displayed a thin central cornea. The area underlying the basal lamina lacked the orderly organization of collagen fibrils present in the WT. In addition the lamellae thickness was 1.5 fold greater than WT and extensive swirling of collagen fibrils was detected in the posterior and middle stroma. The collagen fibrils had a reduced diameter with a larger interfibrillar distance (1.4 fold greater). In contrast to WT there was no change in fibril diameter over the depth of the cornea. Cuprolinic blue staining revealed 33% fewer sulfated GAGs along fibrils in the KO stroma.
These studies show for the first time that the P2X7 receptor is necessary for the proper organization of stromal collagen. Knock down of the P2X7 receptor lead to morphological changes in the stroma including smaller collagen fibrils with fewer GAGs per fibril than in the WT.
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