| Object: To approach significance of B-cell lymphoma factor(Bcl-2),Bcl-2 interrelated X protein(BAX)and vascular endothelial growth factor(VEGF)in the retina of rats with early stage diabetes.Methods: We intraperitoneal injected streptozotocin(STZ),and used this as the model of rats' early stage diabetic retinopathy(DR).We killed rats in 4,8,12 weeks after the model establishment,and took eyeballs to prepare paraffin section and amphiblestrodes stretched preparation.Then we used an immunohistochemical method to detect the expression of Bcl,Bax and VEGF in amphiblestrodes.We analyzed them by using the MIAS medical image analysis software.We digested retina by using pamcreatin,and at the same time,we did the ADP enzyme retinal vessel staining,and observed the morphous and the change of retinal vessel.We randomly divided 30 male Wistar rats into two groups,the diabetes model group(the DR group,n =20)and the normal control group(the BC group,n =10).We intraperitoneal injected streptozotocin(STZ)and used this as the model of rats' early stage diabetic retinopathy(DR).We fed these rats one month to prepare the DR model.We killed rats in 4,8,12 weeks after the model establishment.During the experimental period,the rats had free diet.And we detected their blood glucose and weight regularity.We took eyeballs to prepare paraffin section and amphiblestrodes stretched preparation.We stained amphiblestrodes by using a hematoxylin-eosin(HE)staining method,and detected the samples by using an immunohistochemical method.Then we operated the hematoxylineosin(HE)staining on the dehydrated and waxed samples,observed and counted the number of blood vessel endothelium cell nucleus which broke through the internal limiting membrane of amphiblestrodes under the light microscope.We took above eyeballs paraffin section to do the immunohistochemical detection.We operated the DAB staining and observed them under the light microscope.We randomly selected three visual fields under the 200 X light microscope,and analyzed them by using the MIAS medical image analysis software.After the ADP enzyme retinal vessel staining,we utilized NIH Image(Scion Image)package to calculate the total acreage of amphiblestrodes,and the amphiblestrodes acreage without blood vessel.We detected the concentration of intracellular calcium ion by using the laser scanning confocal microscope.Finally,we used the SPSS 20.0 software to do statistical analysis.Results: The expression of rat retinal Bcl-2,Bax and VEGF: Bcl-2 and Bax protein immunoreactive products and VEGF protein immunoreactive products are located in the cytoplasm with brownish yellow particles.Bcl-2 and Bax immunoreactive cells were mainly distributed in the inner nuclear layer and ganglion cell layer.The diabetic model group's Bcl-2,Bax protein VEGF,Bcl-2/Bax ratio were significantly increased compared with normal control group In rat's retina(P<0.01).Therefore,the early diabetic rat retina Bcl,Bax may be induced by high concentration of glucose,and increase the expression of VEGF.Conclusion: Bcl-2,Bax and VEGF play an important role in the development of diabetic retinopathy.In the early stage of diabetic rats,it occurred the apoptosis of RGCs,and the concentration of calcium ion in RGCs increased gradually.This study showed that,expect for our known harm on retinal vessels,hyperglycaemia also could increase the expression of VEGF by promoting the expression of HIF-1?.It took part in the formation of a new vein in the site of diabetic retinopathy.So even if in the non-ischemia period of diabetic retinopathy,there also had a risk factor of the formation of amphiblestrodes newborn blood vessels: hyperglycaemia.This provided a new thinking that the Bcl-2,Bax and VEGF may play an important part in the process that the diabetic retinal neovascularization occurrenced in diabetic retinopathy. |
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