| Objective: To observe the effect of safflower yellow pigment to the expression of platelet derived growth factor(PDGF) and vascular endothelial growth factor(VEGF) in the retina of diabetic rats, and to explore mechanism of safflower yellow pigment to the treatment of diabetic retinopathy. To provide more experimental evidence of clinical application of safflower yellow pigment, and provide a new treatment for diabetic retinopathy.Method: 50 clean and health male SD rats, weight about 200 ± 20 g.10 rats were randomly selected as a control group, and the remaining 40 rats were intraperitoneally injected with streptozotocin(STZ)(at a dose of 50 mg / kg) to create diabetic rat model. Then the animals were randomly divided into diabetic group(positive control group) of 20 and safflower yellow treatment group of 20. The rats in treatment group were intraperitoneal injection safflower yellow pigment as a dose 40mg/kg daily, while the diabetic group and control group injected with NS over the same period. Drawn bilateral eye of the rats after consecutive injection of 2, 4, 6, 8th week. Prepared of retinal tissue biopsies, and observed morphological changes of retinal ganglion cells(RGCs) in light microscopy and calculated the number of surviving RGCs.Observed expression of VEGF and PDGF by immunohistochemical, and used computer image analysis technique semi-quantitative detection values of the average optical density(AOD) of VEGF and PDGF in retinal tissue.All data in this experiment were data of normally distributed, results are expressed as mean±standard deviation, the experimental results used SPSS13.0 statistical software for statistical analysis. Data using one-way ANOVA analysis at different time points in the same group(One-way ANOVA), data of different groups in the same time point used independent-samples t test. Inspection standards for the α=0.05, P<0.05 was considered statistically significant.Results: 1. General Growth of ratsDuring the experiment, all rats were survived. In the beginning of the experiment the weight of rats in three groups has no significant difference,(P>0.05). With the extension of the observation time, weight of rats in the diabetic group and treatment group were significantly lower than the normal group(P<0.05) while increasing food intake, water intake and urine output than normal rats. 2 HE staining of retinal tissue and results of light microscopy 2.1 Morphological Changes of RGCsIn the control group, the structure of rat retinal from the inside to out was: the ganglion cell layer, bipolar cell layer and photoreceptor cell layer. Cells in the ganglion cell layer were monolayer, orderly, the cell density and cell nuclear clear, bipolar cell layer and photoreceptor cell layer were closely arranged. In the diabetic group, some cells appeared edema and cavitation model change in early stage of DR, along with the extension of time, the cell nucleus of RGCs were changed sparse, disordered, cavitation degree enhanced, appeared necrotic cells, bipolar cell layer and photoreceptor cell layer were different level to reduce. The morphological changes of RGCs in treatment group were damaged lighter than control group in each time point. 2.2 Number change of RGCsThe average number of RGCs was not obvious difference in normal group, every ×400 visual field were: 24.297±0.090 、 24.315±0.381 、24.293±0.426、24.581±0.548. The average number of RGCs in diabetic group at each time were 16.778±1.683、12.172±0.527、8.064±0.761、3.512±0.719. And treatment group were 21.469±0.737、14.678±1.740、9.918±1.073、6.524±0.921. Our experimental results show that the number of RGCs in diabetic group and treatment group were tend to decline after DR, the number of RGCs in treatment group was higher than diabetic group at each time point, and all data had statistically significant(P<0.01). In the every time the number of RGCs in diabetic group and treatment group were lower than the normal group, and all data had statistically significant(P<0.01). 3 Immunohistochemical of VEGF and PDGF in retina and results of semi-quantitative test 3.1 Expression of VEGF in the retinaThe results of immunohistochemical showed that VEGF in the normal group had slight expression, and no significant different at each time point. AOD in diabetic group at each time pointwere0.262±0.017、0.341±0.018、0.382±0.017、0.422±0.025. AOD in treatment group were 0.241±0.019、0.309±0.019、0.357±0.023、0.375±0.026. Thus, the diabetic group showed the positive expression of VEGF in 2week, and increased gradually by time. At the same time, the positive expression of VEGF in treatment group was lower than diabetic group, the differences has statistically significant(P<0.05). 3.2 Expression of PDGF in the retinaThe expression of PDGF in the normal group had slight expression, and no significant different at each time point. AOD in diabetic group at each time pointwere0.116±0.016、0.194±0.015、0.244±0.025、0.264±0.019. AOD in treatment group were 0.089±0.015、0.168±0.019、0.211±0.019、0.234±0.019. Thus, the diabetic group showed the positive expression of PDGF in 2week, and increased gradually by time. At the same time, the positive expression of PDGF in treatment group was lower than diabetic group, the differences has statistically significant(P<0.05).Conclusion: The expression of VEGF increased in the retina of diabetic rats, and the treatment of safflower yellow pigment can make expression of VEGF become lower. The expression of PDGF increased in the retina of diabetic rats, and the treatment of safflower yellow pigment can make expression of PDGF become lower. It is suggested that safflower yellow pigment reduced VEGF and PDGF in retina of diabetic rat by some mechanism, thereby inhibiting the formation of new blood vessels, further delaying the development of diabetic retinopathy. |
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