This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.This progress report has been prepared on the basis of the work that has been done during the first six months of the current budget period.
The specific aim for the first year was to characterize retinal and endothelial (EC) cells, maintained alone or in co-culture, in the horizontally rotating bioreactor (HRB). In most in-vitro models of neovascularization, the formation of new blood vessel-like structures is slow and takes 2-6 weeks and is very difficult to examine during the early stages of development. HRB co-cultured retinal and EC generate a tissue-like 3-D assembly, and formed capillary-like structures within 3 days. Cells cultured in the HRB show changes in the expression of several factors, which play a role in the retinal neovascularization. The rationale for our approach is that several different cell types, including retinal, retinal pigmented epithelial cells (RPE), endothelial (EC) cells, participate in the development of retinopathies, and associated neovascularization. However, most in vitro models have looked at the impact of only one cell type at a time, or their secreted factors, on neovascularization. The strength of the HRB culture system is that it allows for co-spatial localization of different cell types, thus approximating the in vivo tissue structure more closely. The HRB permits cell lines to display phenotypic characteristics that are not normally seen for these cells in monolayer culture. Confluent retinal and EC were harvested; cells mixed with previously laminin-coated microcarrier beads and incubated for 4 hr to allow the cells attach. The cells and beads were maintained in the HRB for 1-5 days. The morphological appearance and expression of several pro-angiogenic factors was assessed.
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