Morphological Structure Of Retina In Rhesus Monkey At Different Ages And Effects On Neural Structure Of Rhesus Monkey In Early Stage Of DR

Object The morphology of inner retina at various stages in primates during the postnatal period was not well defined and there are few studies on the early stage of diabetic retinopathy(DR).In this study,Rhesus monkeys were selected as objects to investigate structure of rhesus monkey retinas of various ages and thus contribute useful data for human eye research and selection of experimental animal’s age.Meanwhile,we will observe the changes of retinal structure in the early stage of diabetic retinopathy and provide new insights in diagnosis,intervention,prevention and treatment of DR.Methods Rhesus monkeys were divided into four groups(n=5 monkeys per group)according to age,child group,teenager group,adult group and elderly group.Animals were deeply anesthetized and the eyes were quickly enucleated,dissected,fixed and saved.The thickness of nerve fiber layer(NFL),total retina,inner plexiform layer(IPL)and outer plexiform layer(OPL)of rhesus monkey retina and the unmber of unclei with different ages were detected with Hematoxylin and eosin(H&E)staining,and quantative immunohistochemistry.The morphology and number of retina ganglion cells(RGCs),rod bipolar cells and amacrine cells in inner retina were investigated by immunoflurescence.Five rhesus monkeys with T1 DM induced by STZ,and five rhesus monkeys with T2 DM induced by high fat diet were in the early stage of DR assessed by Fundoscopy,OCT.Remaining monkeys were assigned to the T1 DM control group and T2 DM control group,and each group consisted of five age-,and refraction-matched healthy rhesus monkeys,respectively.The retinas of rhesus monkeys were stained by H&E staining and Masson staining.Then the expression of vascular endothelial growth factor(VEGF)and pigment epithelium-derived factor(PEDF)in retinas of rhesus monkeys were detected by quantitative real-time PCR and immunohistochemistry.The morphology and expression of various neurons in the inner layer and glial cell changes in the retinas of diabetic rhesus monkeys were detected by immunoflurescence.Results The retinal exhibits characteristically structure at different ages.Statistical analysis of NFL thickness reveals that there are no differences between child and teenager subjects.However,NFL thickness in the adult and the elderly is significantly(P<0.05)thicker thanthat in the young and the teenager.That is to say,the thickness of the NFL of Rhesus monkey retina is well developed in middle age.The trend of the thickness of the whole retina,IPL and OPL is the same as that of NFL.The mean density of RGCs in teenager Rhesus monkeys is significantly(P<0.05)increased compared with other periods.The density of rod bipolar cells significantly(P<0.05)increased at the teenager stage,compared with other periods,and statistically significant(P<0.05)decrease are found when the elderly compared with the adult.Quantifications have shown that the density of amacrine cells significantly(P<0.01)increased at the teenager stage,compared with the child and the elderly,and the density of amacrine cells in the elderly is significantly(P<0.01)increased than that at adult.The density of horizontal cells in teenager rhesus monkeys is significantly(P<0.05)larger when compared with the child and the elderly.There is no obvious symptom of DR in the retina of DM rhesus monkeys and their age-matched controls detected by fundus photograph.Slight exudation and effusion in the blood vessels of rhesus monkeys retina was found by OCT to filter out DM rhesus monkeys in the early stage of DR.H&E staining revealed that there is no difference in the morphology of rhesus monkeys retina in each group,but the thickness of NFL in Rhesus monkey is thinning.Masson staining show that obvious collagen fibers appear in the retina of T1 DM and T2 DM groups compared with the control group,the possible reason is an increase expression of capillary endothelial cells.VEGF m RNA and protein of T1 DM and T2 DM groups are both significantly(P<0.05)increased compared with the control group.PEDF m RNA of T1 DM and T2 DM Rhesus monkeys are both extremely significantly(P<0.05)decreased,comparing with the control groups.Histogram of mean luminance revealed a significant(P<0.05)increase in GFAP expression and Iba-1 expression in both T1 DM and T2 DM rhesus monkeys.Immunofluorescence showed that the morphology of rod bipolar cells and horizontal cells had changed in both T1 DM and T2 DM model.Compared with controls,both T1 DM and T2 DM rhesus monkeys presented a significant(P<0.01)reduction in density in rod bipolar cells.Meanwhile,the density of amacrine cells and RGCs were significantly(P<0.05)reduced in the T1 DM rhesus monkey model,but not in T2 DM rhesus monkeys.We did not find any difference in the density of horizontal cells,compared with controls.Conclusions The structure of retina in rhesus monkey is relatively immature at child,and the inner retina of rhesus monkey is mature in teenager,while NFL is the most developed in the adult group.The thickness of the each retinal layers varies in the elderly compared with the adult.These changes may influence the ability of the retina to transmission,process and integrate information.Neurodegeneration and vascular injury occurred in Rhesus monkey retina at the early stage of DR.The morphology and number of neurons in inner retina have changed and reactive gliosis occurred.Changes in the expression of VEGF and PEDF may assist in indentifying people for the early stage of DR.