| Objectives: Diabetic retinopathy (DR) is the main complication of diabetes mellitus (DM), and also is one of the most serious diseases in ophthalmology. It remains a leading cause of vision loss and new-onset blindness. Due to the lack of safe and effective medicine, the management of patients with DR has always been a tough job in ophthalmology. With the onset of diabetes, cellular and biochemical alterations within the retina are set into motion. Recent studies show that cytokines may play important roles in the occurrence of DR. Transforming Growth Factor-p (TGF-p) is a multifunctional cytokine, whose numerous cell and tissue activities include cell-cycle control, the regulation of early development, differentiation, proliferation , migration , extracellular matrix formation, hematopoesis, angiogenesis, chemotaxis, immune functions, and the induction of apoptosis. In this study we choose Transforming Growth Factor-pi (TGF-p 1) and Transforming Growth Factor-p2 (TGF-p2) genes as the target genes, to quantitatively detect the expression level of TGF-P 1 and TGF-P2 genes in the retina of normal rats in order to determine the expression difference of TGF-p 1 and TGF-P2 in retina, which will be able to provide the technical and substantial foundation for the study of gene in retina . We also detected the expression of TGF-P2 in different stage of diabetic rats' retina to observe and analyze the effect of TGF-P2 on the retina of rat diabetic animal model. This study finally confirmed the effect of TGF-P2 in DR, and provided experiment data and experience for the further clinic study.Methods: 1. Sprague-Dawley (SD) rats ( male , approximately 200 g ) were used. The animals were housed in stainless steel cages and fed standard rat chow and tapwater .They were held in a room on a 12-hour light-dark cycle with an ambient temperature of 22 +1 . The animals were killed by an ether overdose and decapitated at the respective date. The eye was then enucleated; the animals' retinas were dissected. The total RNA was isolated from which the first strand of cDNA was prepared. 2. The expressions of TGF-pl and TGF-J32 gene mRNA were demonstrated quantitatively by Real Time Fluorescence Quantitative reverse transcription PCR (QRT-PCR). 3. Diabetes mellitus was induced by a single intraperitoneal injection of 60mg/kg streptozotocin (STZ) and the animals were held without insulin treatment until sacrifice. Age-matched rats treated with saline were used as controls. Tail vein blood glucose was measured after 2 days and rats were considered hyperglycemic with a blood glucose reading of > 16.7 mmol/L. Animals with blood glucose < 16.7 mmol/L were excluded from the study. The animals were killed for 4weeks, Sweeks, 12weeks, 16weeks, 20weeks and 24weeks stages. The retinas were dissected as the first. 4. The expressions of TGF-P2 gene mRNA were demonstrated by reverse transcription PCR (RT-PCR).Results: 1. The RNA of rat retina was integrative enough to be used to further QRT-PCR analysis. 2. The levels of TGF-p2/(3-actin were 0.0378 ?.009 . The levels of TGF-pl/p-actin were 0.0008 ?0.0003. The mean ration of TGF-P2 /TGF-pl were 55.00+26.61 (P<0.01) . The mRNA level of TGF-p2 was about 55 times as that of TGF-pl in normal rat retina .3. Compared with control groups, the expressions of TGF-P2 mRNA in diabetic rats' retinas were up-regulated at 4 weeks, but there was no statistic difference (p=0.201) ;down-regulated at Sweeks, there was statistic difference (p=0.041) ;also down-regulated at 12weeks, there was statistic difference(p=0.029); at 16weeks there was no statistic difference(p=0.361); up-regulated at 20 weeks, but there was no statistic difference (p=0.136) ;continue up-regulated at 24 weeks, there was statistic difference (p=0.047) .Conclusions: Our data implicated: 1. QRT-PCR could specifically and accurately detect gene expression level in rat retina. The method is practical and feasible. 2. Innormal retina the TGF-(i2 gene was expressed more abundantly than TGF-pl. It was suggested that TGF-p2 played an important rol...
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