[Expression of mouse telomerase reverse transcription in a mouse model of oxygen-induced retinopathy].
摘要
OBJECTIVE: To establish oxygen-induced retinal neovascularization in mice and to detect the expression of mTERT in mice. METHODS: It was an experimental study Establishment of oxygen-induced retinal neovascularization in mice. Thirty-two 7-day-old C57BL/6J mice were divided into oxygen-induced retinopathy group and control group without restriction of gender. In oxygen-induced retinopathy group, 16 mice were exposed to 75% +/- 2% oxygen for 5 days and then to room air; In control group, 16 mice were raised in room air. Observation of the retinal neovascularization. On the postnatal day 19, The mice's vena caudalis were perfused with 2% Evens blue solution. Eyeballs were enucleated and fixed in 4% paraformaldehyde for half an hour. Then the retina was separated and flat-mounted on the slide. The morphologic changes of retinal vessel were observed and captured under fluorescence microscope. Histological observation and vascular endothelial cells counting. The eyeballs were enucleated and then fixed. After paraffin imbedding, 4 microm serial slices, hematoxylin-eosin staining, select one section every 60 microm to count the endothelial cell nucleus that break through the inner limiting membrane. Expression of mTERT mRNA were confirmed by reverse-transcription polymerase chain reaction (RT-PCR). In the each group, the retina were all carefully dissected on the postnatal day 19. The total RNA was isolated and cDNA was synthesized before RT-PCR was performed. The PCR products were separated by 2% agarose gel electrophoresis and photographed. Expression of mTERT mRNA were confirmed by Real-time PCR The total RNA was isolated and cDNA was synthesized (The same procedure as RT-PCR). Fluorescent real-time quantitative polymerase chain reaction system (total 20 microl) was made. The Fluorescent signals were detected at 60 degrees C. The expression of mTERT were confirmed by immunohistochemistry. At P19, 4 microm cross sections were made in the hyperoxia-exposed and normal reti