TY - JOUR
T1 - Characterization of an in vitro model of elastic fiber assembly
AU - Robb, Bruce W.
AU - Wachi, Hiroshi
AU - Schaub, Theresa
AU - Mecham, Robert P.
AU - Davis, Elaine C.
PY - 1999/11
Y1 - 1999/11
N2 - Elastic fibers consist of two morphologically distinct components: elastin and 10-nm fibrillin-containing micro fibrils. During development, the micro fibrils form bundles that appear to act as a scaffold for the deposition, orientation, and assembly of tropoelastin monomers into an insoluble elastic fiber. Although microfibrils can assemble independent of elastin, tropoelastin monomers do not assemble without the presence of microfibrils. In the present study, immortalized ciliary body pigmented epithelial (PE) cells were investigated for their potential to serve as a cell culture model for elastic fiber assembly. Northern analysis showed that the PE cells express microfibril proteins but do not express tropoelastin. Immunofluorescence staining and electron microscopy confirmed that the microfibril proteins produced by the PE cells assemble into intact microfibrils. When the PE cells were transfected with a mammalian expression vector containing a bovine tropoelastin cDNA, the cells were found to express and secrete tropoelastin. Immunofluorescence and electron microscopic examination of the transfected PE cells showed the presence of elastic fibers in the matrix. Biochemical analysis of this matrix showed the presence of cross-links that are unique to mature insoluble elastin. Together, these results indicate that the PE cells provide a unique, stable in vitro system in which to study elastic fiber assembly.
AB - Elastic fibers consist of two morphologically distinct components: elastin and 10-nm fibrillin-containing micro fibrils. During development, the micro fibrils form bundles that appear to act as a scaffold for the deposition, orientation, and assembly of tropoelastin monomers into an insoluble elastic fiber. Although microfibrils can assemble independent of elastin, tropoelastin monomers do not assemble without the presence of microfibrils. In the present study, immortalized ciliary body pigmented epithelial (PE) cells were investigated for their potential to serve as a cell culture model for elastic fiber assembly. Northern analysis showed that the PE cells express microfibril proteins but do not express tropoelastin. Immunofluorescence staining and electron microscopy confirmed that the microfibril proteins produced by the PE cells assemble into intact microfibrils. When the PE cells were transfected with a mammalian expression vector containing a bovine tropoelastin cDNA, the cells were found to express and secrete tropoelastin. Immunofluorescence and electron microscopic examination of the transfected PE cells showed the presence of elastic fibers in the matrix. Biochemical analysis of this matrix showed the presence of cross-links that are unique to mature insoluble elastin. Together, these results indicate that the PE cells provide a unique, stable in vitro system in which to study elastic fiber assembly.
UR - http://www.scopus.com/inward/record.url?scp=0032694642&partnerID=8YFLogxK
U2 - 10.1091/mbc.10.11.3595
DO - 10.1091/mbc.10.11.3595
M3 - Article
C2 - 10564258
AN - SCOPUS:0032694642
VL - 10
SP - 3595
EP - 3605
JO - Molecular Biology of the Cell
JF - Molecular Biology of the Cell
SN - 1059-1524
IS - 11
ER -