| Diabetes mellitus(DM) is a metabolic disorder disease, which eventually causes brain, kidney, eye and peripheral nerve system damage and dysfunction of diabetic retinopathy.Diabetic retinopathy(DR) is the most serious and common in the diabetic microvascular complications.The persistent hyperglycemia status acts on diacylglycerol(DAG),activates the DAG/protein kinase C(PKC) pathway, causes retinal blood flow dynamic change, induces retinal blood microcirculation disorder.In addition, the activation of the DAG/PKC pathway directly impacts on the downstream signal factor-vascular endothelial growth factor(VEGF),up-regulates the expression of VEGF.As a classic factor of inducding retinal neovascularization,VEGF is one of the important factors involved in the formation of DR pathologic changes.At present, the study about pathogenesis of DR has become a key link for the treatment of DR.There are various western medicine towards DR. Researches about traditional Chinese medicine for DR have made progress,which preliminary show advantages of traditional Chinese medicine in the prevention and treatment of DR.Sericine is a kind of water-soluble protein, exists in silkworm cocoons,accounts for 30% of total silk.In recent years, study have found that sericine has a certain impact on prevention and treatment of diabetic rats and its target organs such as kidney, testis complications, but the specific mechanism has not yet been elucidated.In this study, we used high-sugar and high-fat diet combined with low-dose streptozotocin(STZ) intraperitoneal injection to build type 2 diabetes model rats,investigated the protective effects of sericine on diabetic retinopathy of type 2 diabetes model rats,laid theoretical foundation for further study of sericine to prevent and treat diabetes complications.Objective:By observing the influence of sericine on the factor VEGF and PKC expressions of type 2 diabetic rats retina to explore the protective effects of sericine on diabetic retinopathy.Methods:1 SPF level male SD rats(170g-190g) were divided into 3 groups in accordance with the random number table:Normal control group,diabetes model group and sericine treatment group.Normal control group was fed by standard diet;diabetes model group and sericine treatment group were fed by high-sugar and high-fat diets,and then injected with 2% STZ(25mg·kg-1·d-1,3d)intraperitoneally to build type 2 diabetes model, blood glucose?16.7mmol/L meant modeling successfully.Sericine treatment group was given gavage of sericine(2.4g·kg-1·d-1);at the same time,the normal control group and diabetes model group were given the same does of saline.2 Fasting blood glucose meter was used to detect the blood glucose of all groups and living state was observed at regular intervals.3 After administration,all rats received intraperitoneal anesthesia, removed the eye balls for text specimen processing.4 HE staining was used to observe the pathologic changes of retinal tissue.5 SP immunohistochemical staining was used to observe the PKC and VEGF protein expression of retinal tissue.6 Western blotting was used to detect the PKC and VEGF protein expression and quantitative analysis.7 RT- PCR was used to detect the PKC and VEGF mRNA expression and quantitative analysis.Results:Living state of each group:before the sericin treatment group rats received treated,the rats were irritable with noise and vibration,less active,with dry hair and shedding,more drinking and urinating, lean;After treated with sericine lavage, the hair of rats were more shining, gained weight, less drinking and urinating.As the develpoment of DM,diabetes model group rats were more irritable, with poor quality of hair, shedding, lost weight,drinking and urinating more than before. During the whole process, the normal control group rats has no obvious change.Blood glucose monitoring result showed that the blood glucose of rats in diabetes model group increased obviously compared with that in normal control group(P<0.01);the blood glucose of rats in sericin treatment group decreased obviously compared with that in diabetes model group(P<0.01).The HE staining changes showed as follows.Normal control group :the retinal structure was complete and stratified clearly, cells morphology were in good condition and the inner limiting membrane was smooth, no changes of swelling and thickening.diabetes model group: inner limiting membrane was markedly swolling and thickening, the retinal layers owed clear, some cells vacuolar degeneration and karyopyknosis, cell distribution disorder and sparse. Some fragmentation of inner limiting membrane causing membrane surface rough, hyperplasia of capillaries endothelial cells out from the membrane.Sericine treatment group:stratified clear, inner limiting membrane mild swelling and thickening.Compared with diabetes model group,the appearance of decreasing in the cell number and cell disorder were less.Immunohistochemical staining in combination with Western blotting and RT-PCR results showed: compared with diabetes model group, the expression of VEGF 、 PKC protein and mRNA decreased significantly in sericine treatment group(P<0.01);compared with normal control group, the expression of VEGF、PKC protein and mRNA increased in diabetes model group(P<0.01).Conclusion:The expression of PKC and VEGF protein and mRNA increase significantly in diabetic rats retina,sericine can reduce the expression of PKC and VEGF to exert a protective effect toward diabetic rats retina. |
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