UV irradiation of Type I collagen gels changed the morphology of the interconnected brain capillary endothelial cells on them.

Abstract

We cultured mouse brain capillary endothelial cell line bEnd.3 on the UV-irradiated Type I collagen gel. Morphology of bEnd.3 cells on the Type I collagen gel was drastically changed if the gel was crosslinked by UV irradiation. The interconnecting network of bEnd.3 cells which have cord-like morphology on the soft collagen gels was converted to the monolayer of the flat cells, tightly-bound each other covering the gel surface, in a confluent state. The collagen gels were mechanically stiffened by UV irradiation for 15min with UV light at 254nm showing approximately two times higher value of Young's modulus E (1.51±0.58kPa) than the control gel (3.17±1.17kPa). AFM images of the collagen fibrils were not severely changed after irradiation. Collagen subunit proteins were crosslinked and degraded simultaneously under UV irradiation proved by results of SDS-PAGE and separation by centrifugation. Expression of Integrin gene was measured by quantitative real-time PCR. Expression of the integrin α2 gene, tight junction protein 1 gene, and claudin 5 gene were down-regulated in cells on the UV irradiated collagen gel in comparison with the unirradiated one while expression of the integrin β1 gene and Integrin α1 gene did not significantly change. Thick actin filaments were more clearly observed in the cells on the UV-irradiated collagen gel than the unirradiated one by fluorescent microscopy. We conclude that UV irradiation made the collagen gel stiffened and changed the physiological state of bEnd.3 cells including their adhesion, extension, and proliferation.