Fatty Acids In Tea (Camellia Sinensis (L.) O.Kuntze) And Their Effects On The Growth Of Choroids-Retinal RF/6A Endothelial Cell Lines

There has been much focus recently on the long chain fatty acids, which were considered as a major determinant of growth, visual and neural development. Epidemiological studies and clinical data suggested that ω-3 series polyunsaturated fatty acids may do goods to retinal development. On the other hand, elevated accumulation of fatty acids was often reported as a cause for obesity and type 2 diabetes mellitus. Dyslipidemia was supported to be a critical factor in the development of diabetic retinopathy, and fatty acids were represented as the link between diabetes, dyslipidemia, and diabetic retinopathy.Though the fatty acids in tea shoots were related with the tea quality, it has been unclear that whether they have any effects on human healthy. Tea was considered to be with a relationship with facility of sight in Chinese traditional medicine. It's also confirmed by modern medicine that the tea leaf components like vitamine A, B1, B2 and C were essential in maintaining visual function. Thus, here we investigated the fatty acid compostion of tea shoots with their characteristics in order to examine their effect on retianl endothelial cells and study the possible mechanisms.In this dissertation, the rhesus macaque choroids-retinal endothelial cells, RF/6A, was used to investigate the biological activity of three long chain fatty acids detected in tea shoots. The composition and contents of fatty acids in tea shoots were described by using the technique of gas chromatography, and their effects on cell growth were investigated in vitro. Retinal RF/6A endothelial cells viability employed in the experiment was determined by MTT assay. DNA fragment was analysed by gel electrophoresis, and apoptotic characteristics of RF/6A cells were examined by flow cytometry. Electron spin resonance (ESR) was used to detecte oxygen consumption and free radical scavenging ability of unsaturated linoleic acid.By using gas chromatography, the content of lipids in tea shoots was described as the most abundant long chain fatty acids were palmitic (C16:0), linoleic (C18:2n-6) and linolenic acid (C18:3n-3). Results showed that there was an appreciable amount of essential fatty acids in tea shoots, which accounted for 49.0~65.2% of total fattyacids and achieved 3.16 mg/g in the 2nd tea (C. sinensis cv. zhuyeqi) fresh leaf, comprising 1.25 mg/g linoleic acid and 1.91 mg/g linolenic acid.MTT assay demonstrated the viability of RF/6A cells by exposure to various representative long chain fatty acids existed in tea shoots with pathologically relevant concentrations (0-500 u M). It seemed that those fatty acids with different structure had various effects on the cell proliferation at their relative low concentrations. Saturated palmitic acid (40-400 u M) did not affect the cell viability, n-3 polyunsaturated linolenic acid (40-200 u M) had a significant proliferative effect on the growth of RF/6A cells within 24 h and n-6 polyunsaturated linoleic acid (80-160 u M) also activated their proliferation. These findings implied that the long chain fatty acids in tea shoots might be relative to the proliferative diabetic retinopathy. Meanwhile, the cell growth was significantly inhibited by a high dose of the essential fatty acids. 500 u M palmitic acid, 400 u M linoleic acid and 300 u M linolenic acid inhibited the cell proliferation respectively.From DNA fragment analysis and flow cytometry, the different effects of these fatty acids on the endothelial cells were evaluated and their biological activities were clarifed. Results showed that the growth inhibitory effects of high dose of fatty acids were neither through apoptic-inducedc pathway nor cell necrosis. Moreover, the microscopic images of RF/6A cells treated with varied concentrations of fatty acids weere described, which showed that there were floating cells in the linoleic and linolenic treated midia.ESR results showed that linoleic acid may directly react with DPPH radical in DPPC ethanol and liposome suspensions. Furthermore, it suggested that the oxygen consumption rate in Egg PC containing 0.05 M linoleic acid system was lower than Egg PC alone. Thus, linoleic acid expressed antioxidant activity in the present study, which is contrary to inducing lipid peroxidation in retina.