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Y. Maruyama, S. Mikawa, T. Sasaki, Y. Hotta, K. Sato; BMP4 Expression in the Developing Rat Retina. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1232.
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Bone morphogenetic protein-4 (BMP4) is a member of the transforming growth factor ß superfamily. BMP4 signaling has been shown to play an important role in the development of central nervous system. Also in the early stages of the developing eye, BMP4 has been reported to play pivotal roles, including the lens formation, topographic retinotectal projection, and apoptotic cell death. However, little information is available for BMP4 expression in the late embryonic period and postnatal period. In the present study, we thus investigated the expression of BMP4 in the developing retina.
Male wistar rats at various ages (at embryonic day 15, 19 and postnatal day 1, 7, 14, 21, 49; n=3 at each time point) were used. For immunochemistry, the embryos were fixed in 4 % paraformaldehyde in 0.1 M phosphate buffer (PB, pH 7.4) at 4 °C overnight. The postanal rats were perfused transcardially under deep diethylether anesthesia with saline followed by 0.1 M PB, containing 4 % paraformaldehyde. The eyes of the postnatal rats were removed rapidly, postfixed in the same fixative for 2 hrs at 4 °C. All samples were immersed in 30 % bufferd sucrose at 4 °C overnight. Frozen sections (12 µm) in thickness were cut on a cryostat. An Avidin-Biotin-peroxidase method was applied for immunohistochemistry. The sections were incubated with primary antibodies against mouse monoclonal BMP4, and then were incubated with secondary antibody, biotin conjugated against goat IgG, and horse-radish peroxidase conjugated streptavidin. These antibodies were purchased from Novocastra, Inc., UK. Staining was visualized using 3-3'-diaminobenzidin.
At E19, we found very intense BMP4-immunoreactivity (IR) in the nerve fiber layer, suggesting that ganglion cell axons express abundant BMP4 protein. At P1, the inner plexiform layer, where ganglion neurons, amacrine cells, and bipolar cells make synapses at that time, exhibited very strong BMP4-IR. Thereafter, abundant BMP4 expression was kept to the adult period.
BMP4 is abundantly and differentially expressed throughout the development of the retina. These results suggest that BMP4 plays pivotal roles in the retina not only in the early embryonic period but also in the late embryonic and postnatal periods, and even in the adult.
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