Objective To investigate the protective effect of blocking the signal path of p38 mitogen activated protein kinase on blood retinal barrier (BRB) and retinal ganglion cells (RGC) in early diabetic rats.Methods A total of 60 Wistar rats were divided into the control and diabetes group, with 30 rats in each group. Diabetes was induced in rats in diabetes group by peritoneal injection of streptozotocin (STZ);the plasma glucose level of ＞16.7 mmol/L indicated that the diabetes model was set up successfully.The rats in the control group underwent peritoneal injection of equivalent sodium citrate solution. IgG leakage method was used to measure the damage of BRB function and vascular leakage. The expression and localization of caspase-3 and vascular endothelial growth factor (VEGF) in retina of diabetic rats were examined by immunohistochemistry analyses.Two weeks after the establishment of the diabtes model, the rats in diabtes group underwent intravitreal injection with SB203580, a p38 inhibitor;six weeks after the injection, the expression of caspase-3 and VEGF was detected, and the number of apoptosis RGC was counted via immunofluorescence technique.Results In the contral group, IgG staining located in the blood vessels with little leakage; while the IgG leakage was much more obvious in the diabetes group eight weeks after the establishment of the model. Six weeks after intravitreal injection with SB203580, the leakage decreased in diabtes rats. The results of semiquantitative analysis and fluorescence immunohistochemistry showed that compared with the results in diabetes rats 8 weeks after intravitreal injection (2.9 times much more than that in the control group), the fluorescence expression of VEGF decreased in diabetes rats six weeks after intravitreal injection (1.8 times much more than that in the control group).The apoptisis RGC number in rats 6 weeks after intravitreal injection of SB203580 was much less than that in rats without intravitreal injection (t＝5.731, Plt;0.01). Conclusions SB203580 can alleviate the disruption of BRB and apoptosis of RGC in early diabetes rats, which suggests that p38 MAPK pathways appear to be directly involved in the pathogenesis of early diabetic retinopathy.
Objective To quantitatively assess the damage of blood-retinal barrier (BRB) in rats with diabetic retinopathy using dynamic contrast enhanced-magnetic resonance imaging (DCE-MRI). Methods Forty 3-week-old Sprague-Dawley (SD) rats were randomly divided into experiment and control group. The rats in experiment group underwent intraperitoneal injection of streptozotocin. The rats with blood glucose over 16.65 mmol/L and ldquo;+++rdquo; of urine glucose were considered as diabetes and were further divided into four subgroups according to the course of diabetes mellitus (2, 4, 6, and 8 months).The rats in control group underwent intraperitoneal injection with the same volume of buffer and were divided into four subgroups (with 5 rats in each subgroup) according to the coordinate age of rats in experimental group.All of the eyeballs were scanned by DCE MRI and enucleation was performed after intraperitoneal injection with pentobarbitone.The data were analyzed by SPSS 12.0 statistical software.Results All the rats in experiment group became diabetic models. There was no obvious BRB permeability in control group and in 2- and 4-months experiment group.The average BRB permeability rate in 6 and 8 month experiment groups were (0.1399plusmn;0.0065) and (0.1816plusmn;0.2756) mm3/min respectively (Z=-2.121, Plt;0.05). Retinal edema and cellular disorganization appeared at 4 months and became more severe when diabetes course extended.Conclusions DCE MRI can measure the BRB permeability rate accurately and assess the extent of BRB damage quantitatively in rats with diabetic retinopathy.
Objective To investigate the effect of intravitreal injection with dexamethasone on leukocyte accumulation, vascular permeability, and the expression of intercellular adhension molecule (ICAM-1) in rats with diabetes. Methods Seventy-two BN rats were divided into 4 groups: control group, diabetes group, diabetes+ physiologic saline group, and diabetes+ dexamethasone group, with 18 rats in each group. Streptozotocin was injected into the rats to set up the diabetic model. Accumulated leukocytes were counted in vivo by acridine orange leukocyte fluorography, and retinal vascular permeability was measured by Evans blue assay. The expression of mRNA and protein level of ICAM-1 were evaluated by real-time quantitative polymerase chain reaction analysis and enzymelinked immunosorbent assay. Results In the diabetes+ dexamethasone group, accumulated leukocytes were reduced, retinal vascular permeability decreased, and the expression of ICAM-1 decreased. The expression of ICAM-1 mRNA and protein levels in control group, diabetes group, diabetes+ physiologic saline group, and diabetes+ dexamethasone group were 0.43plusmn;0.07，0.76plusmn;0.21，0.74plusmn;0.18，and 0.55plusmn;0.13; (37.90plusmn;4.56), (76.74plusmn;6.68), (74.32plusmn;7.11), and (39.61plusmn;4.47) pg/mg respectively. Conclusions Dexamethasone can reduce accumulated leukocytes and retinal vascular permeability, which may be caused by inhibiting the expression of ICAM-1. (Chin J Ocul Fundus Dis,2007,23:273-276)
Obiective lt;brgt;To investigate the change of the activity of proliferation in cultivated Muuml;ller cells treated by advanced glycation endoproducts (AGEs), and the effect of these changes on expression of occludin in bovine retinal vascular endothelial cells (BREC). lt;brgt;Methods lt;brgt;The cultivated Muuml;ller cells were devided into normal growth group and cultured with AGEs group. The cultured BREC were devided into 4 groups:group 1, without any medium; group 2, with normal growth Muuml;ller cell medium (MCM); group 3,MCM treated by AGEs; group 4, without cell as the control. Enzyme-linked immuno sorbent assay was used to detect the content of occludin in the medium in the 4 groups. lt;brgt;Results lt;brgt;The content of expression of occludin was the most in group 2, less in group 1, and the least in group 3. lt;brgt;Conclusion lt;brgt;AGEs may promote the abnormal proliferation of Muuml;ller cells and inhibit the expression of occludin in BREC. lt;brgt;(Chin J Ocul Fundus Dis, 2006, 22: 28-30)
Objective To investigate the effect of batroxobin on the blood-retinal barrier (BRB) and vascular endothelial growth factors (VEGF) in diabetic rats. Methods Sixty Sprague-Dawley rats were used to establish diabetic models by intraperitoneal injecting with streptozotocin (60 mg/kg), and were divided into 3 groups: diabetic group (n=20), batroxobin (40 mg/kg) group (n=20) and batroxobin (20 mg/kg) group (n=20). Twenty-five else rats were in control group. All of the rats were executed 7 days later. The function of BRB was observed by Evans blue method. Results concentration of VEGF protein was detected by enzyme-linked immunoabsorbert assay (ELISA). The results of each group were compared. Results The content of BRB leaked into retina was obvious lower in the control group than which in the other 3 diabetic groups（Plt;0.01）. There was no significant difference of the content of Evans blue between the two groups with different dosage of batroxobin (P＞0.05). The content of Evans blue was lower in the 2 diabetic groups with different dosage of batroxobin than which in the control group (Plt;0.05). The content of VEGF in retina was obviously lower in control group and 2 diabetic groups with different dosage of batroxobin than which in the diabetic group (Plt;0.01), and obviously lower in batroxobin (40 mg/kg) diabetic group than which in the control group (P＝0.01). The content of VEGF in control group and batroxobin (20 mg/kg) diabetic group (P＝0.06) didnprime;t differ much, which occurred similarly in batroxobin diabetic groups with different dosage (P＝0.78). Conclusions Batroxobin may alleviate the damage of function of BRB in diabetic rats and reduce the expression of VEGF, which suggests that batroxobin can protect the function of BRB to a certain extent. (Chin J Ocul Fundus Dis, 2006, 22: 16-19)
Objective To investigate the relationship between expression of retinal intercellular adhesion molecule-1 (ICAM-1) and blood-retinal barrier (BRB) rupture and therapeutic effect of intravitreous injection with triamcinolone acetonide (TA) on blood-retinal barrier rupture in rats with diabetes mellitus (DM). Methods Diabetic model of Wistar rats was induced and were divided into normal control group, DM-4-month group and DM-6-month group. Each group was subdivided into immunohistochemcial staining and BRB measurement groups. BRB measurement group was further divided into non-TA treatment group, 1-week-TA treatment group, and 2-week-TA treatment group. The rats were intravitreously injected with 5 mu;l TA. The digested retinal preparation was stained by immunohistochemcial method to observe the expression of retinal ICAM-1 and morphological changes. The mean optic density (A) value of endothelial cells was measured by image-analyzing software to quantify the expression of ICAM-1. BRB changes were measured by content test of retinal Evans blue (EB). Results In the immunohistochemcial staining groups, there was no significant positive expression of ICAM-1 in retinal capillary in control group. Compared with the control, there was significant positive expression of ICAM-1 in DM-4-month group (P＜0.001) with some morphological changes such as irregular width of capillary caliber, and there was enhanced positive expression of ICAM-1 in DM-6-month group (P＜0.001) with aggravated morphological changes and even acellular capillary. In the BRB measurement groups, there was no significant difference of EB content(P＞0.05) among control groups. The EB content in two DM groups significantly increased compared with that in the controls (Plt;0.001), and higher in DM-6-month group than that in DM-4-month group (Plt;0.01). In TA treatment groups, the EB content in all the DM groups significantly decreased (Plt;0.001) but with no significant difference among the groups(P＞0.05). EB content in DM-4-month group after 2-week treatment almost reached to normal value (P＞0.05) while was higher in the rest of TA treatment groups than that in the control group (Plt;0.05). Rectilinear correlation between A value of endothelial cells and the retinal EB content(r=－0.959)was found. Conclusion There is a positive relation between the expression of ICAM-1 and BRB rupture in retina of DM rats, and intravitreous injection with TA can effectively alleviate BRB rupture. (Chin J Ocul Fundus Dis, 2006, 22: 24-27)
Objective To investigate the bloodretinal barrier（BRB）function of porcine retinal pigment epithelial（RPE）cells cultured in vitro. Methods Primary porcine RPE cells were cultured，and the third generation were inoculated in a microporous filter with the filter membrane of polyvinylpyrrolidone（PVP）-free polycarbonate membrane.After 1，2，3 and 4 weeks of culture，the surface of filter membrane was observed by light microscope，and after 2 weeks of culture，the section of filter membrane was observed by light microscope and transmission electron microscope.Transepithelial electrical resistance（TER）was detected and the permeability was measured with fluorescein sodium（FS）and horseradish peroxidase（HRP）. Results Primary porcine RPE cells were cultured successfully.RPE cells converged1week after inoculation; 2 and 3 weeks after inoculation，the density of RPE cells did not changed obviously; 4 weeks after inoculation，the density of RPE cells decreased.The characteristics of polarized growth of monolayer were found in RPE cells on the surface of filter membrane; 2 weeks after inoculation，the TER of RPE cells was（97.44plusmn;11.36）Omega;/cm2 which maintained till the 3rd week after inocubation.After incubated for 30 minutes，only 0.27% of FS and 0.17% HRP reached the inferior filter membrane，and the permeability rate of SF with low molecular weight was higher than which of HRP with high molecular weight. Conclusions The filter with PVPfree polycarbonate membrane may be used to set up the model of RPE cells with polarized growth of monolayer and investigate the barrier function of RPE cells. (Chin J Ocul Fundus Dis, 2006, 22: 188-191)
ObjectiveTo investigate the effect of nerve growth factor (NGF) on recuperate of optic nerve after contusion by clamping in adult rabbits. MethodsSixteen adult rabbits were randomly divided into NGF and the control group with 8 rabbits in each group. After the optic nerve of the right eyes was clamped，tissue engineering nerve containing 0.06 ml NGF（concentration: 5×10-4 g/L, NGF group） and 0.06 ml of PBS (control group) was immediately transplanted into the injured eyes respectively, and 0.02 ml NGF（concentration: 5×10-4 g/L, NGF group）and 0.02 ml of PBS (control group) were injected into the vitreous of right eyes respectively. Flash visual evoked potential (FVEP) test was performed on the eyes 1 day, 2 weeks and 8 weeks after the injury. The number of retinal ganglion cells (RGCs) and changes of optic nerves were observed by light microscopy and electron microscopy at the 8th week after contusion,and a computer-image-analysis system was used to count the optic nerve axons.ResultsThe ratio of amplitude of FVEP of the injured and healthy eyes was 0.765±0.150 in NGF group and 0.494±0.108 in the control at the 2th week after injury with a significant difference between the two groups (Plt;0.05); and was 0.581±0.138 and 0.409±0.119 respectively at the 8th week after contusion with statistical difference between the two groups (Plt;0.05). The results of light microscopy and electron microscopy showed that degeneration of RGCs and optic nerves in the NGF group was lighter than that in the control group 8 weeks after injury, while the amount of optic nerve axons was (10 955±608.7) axons/ mm2 in the NGF group and (7 898±608.8) axons/mm2 in the control with statistical difference between the two groups (Plt;0.05). ConclusionNGF may redound to the survival of RGCs and regeneration of the axons in some degree, which can promote the recuperation of optic nerve and visual function. (Chin J Ocul Fundus Dis, 2005,21:253-257)
Objective To investigate effects of neural retina on development of the structure of outer blood retinal barrier in embryogenesis. Methods The retinal neural epithelium (RNE) and pigment epithelium (RPE) layers of 150, 120 and 90 embryonic chicken eyes incubated for 7,10, and 14 days were peeled off. RNE was used to prepare the culture medium with different conditions (7drcSF3, 10drcSF3, 14drcSF3). RPE cells of 7- and 14-incubated chicken embryos were cultured on laminin-coated transwell filter. The SF3, 7drcSF3, 10drcSF3 , 14drcSF3 medium were used respectively in the apical chamber and SF2 was used in basolateral chamber. After the formation of monolayer, the transepithelial electrical resistance of the RPE was detected. After the fixation of RPE cells, the condition of the tight junction among the cells was observed by immunohis tochemistry and transmission electron microscopy. Results For the RPE cells of 7-and 14-day incubated embryonic eyes, the difference of TER in various medium of SF3/SF2, 7drcSF3/SF2, 10drcSF3/SF2, 14drcSF3/SF2 was statistically significant (P＜0.01). The polarity of RPE cells was induced and the netlike tight junctional strands was urged in the retina-conditioned medium. Conclusion The neural retina may actively promote the formation of the structure of outer blood retinal barrier. (Chin J Ocul Fundus Dis，2004，20：237-240)