Objective Methods Ninety male Wister rats were randomly divided into normal control group, diabetic group and FTY720 group, thirty rats in each group. Diabetes was induced by giving a single intraperitoneal injection of streptozocin. FTY720 group was administered with FTY720 at a dose of 0.3 mg/kg by oral gavage daily for 3 months after establishment of diabetes. All rats were used for experiments following intervention for 3 months in FTY720 group. Immunohistochemical staining was used to observe the expression and distribution of intercellular adhesion molecule (ICAM-1) and vascular cell adhesion molecule (VCAM-1), and the positive cells were counted. Real-time reverse transcription PCR was used to measure mRNA expression of ICAM-1 and VCAM-1. Fluorescein isothiocyanate-Concanavalin A perfusion was used to detect retinal leukocytes adhesion. Evans blue (EB) perfusion was used to analyze retinal vascular permeability. Immunofluorescence staining was used to detect retinal inflammatory cells infiltration. Results In diabetic group, both ICAM-1(t=12.81) and VCAM-1 (t=11.75) positive cells as well as their mRNA expression (t=16.14, 9.59) were increased compared with normal control group, with statistical significance (P < 0.05). In FTY720 group, both ICAM-1(t=-9.93) and VCAM-1 (t=-6.61) positive cells as well as their mRNA expression (t=-15.28, -6.10) were decreased compared with diabetic group, with statistical significance (P < 0.05). Retinal leukocytes adhesion (t=16.32) and EB permeability (t=17.83) were increased in diabetic group compared with normal control group, while they were decreased in FTY720 group compared with diabetic group(t=-9.93, -11.82),with statistical significance (P < 0.05). There were many CD45 positive leukocytes infiltration in retina of diabetic group, including CD11b positive macrophage/activated microglia, while both of them were little in FTY720 group. Conclusions FTY720 can decrease retinal leukocytes adhesion, reduce retinal vascular permeability and inflammatory cells infiltration, which is associated with down-regulation of ICAM-1 and VCAM-1.
The ocular fundus changes and the damage of visual function were various at different stages of diabetic retinopathy (DR). To get hold of timing and different therapic method correctly of early diagnosis, whole body treatment, laser photocoagulation and vitreous-retina surgery and adopting targeted interventions could help patients receiving the most reasonable and effective treatment at different stages, both of them are keys to reduce the damage of visual function. （Chin J Ocul Fundus Dis，2008，24：240-243）
ObjectiveTo observe the influence of human umbilical cord mesenchymal stem cells (hUCMSC) transplantation into vitreous cavity of diabetic rats on the retinal morphology, and the expression of glial fibrillary acidic protein (GFAP) and rhodopsin (RHO). Methods78 male Sprague-Dawley rats were used. 70 rats were injected with streptozotocin by tail vein injection at a dose of 40 mg/kg to establish the diabetes mellitus model, and another 8 rats were injected with 0.1 mol/L pH 4.0 citric acid buffer at the same dose as the normal control group. After 6 weeks of modeling, 10 rats were taken as the control group of diabetic model. hUCMSC suspension was injected into the right eye vitreous cavity of the remaining 60 rats, and the same volume of Dulbecco's modified Eagle/F12 medium was injected into the left vitreous cavity as control eyes. 1, 2 and 4 weeks after transplantation, follow-up experiments were performed. The experimental eyes were labeled as U1, U2, and U4 groups, while the control eyes were recorded as D1, D2, D4, and each group consisted of 20 eyes. After paraffin section and hematoxylin-eosin staining, the structure of the retina was observed by optical microscopy and the thickness of the outer nuclear layer and the inner nuclear layer (INL) were measured. The distribution and migration of hUCMSC in rat retina were observed by frozen section-tissue immunofluorescence assay. The mRNA and protein expression of GFAP and RHO in the retina were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot assays. ResultsThe results of optical microscope observation showed the normal structure of retina in normal control group. The retinal nerve fiber layer (NFL) was thinned and the number of retinal ganglion cells (RGC) in the control group of diabetic rats was decreased. The decreased number and disorder arrangement of RGC were observed as well in U1, D1 rats. The RGC number of U2, U4, D2, D4 rats was gradually decreased. Compared with D4 group, the thickness of INL in U4 group was significantly increased (P < 0.05). Tissue immunofluorescence assay showed that hUCMSC were distributed along the inner limiting membrane in the retina of the U1 group, while the number of hUCMSC in the U2 group was gradually decreased, mainly in the NFL and ganglion cell layers. Real-time PCR and Western blot data indicated that the relative expression of GFAP mRNA and protein in the diabetic retina was significantly increased, and the relative expression of RHO mRNA and protein decreased gradually in the diabetic model group and the D1, D2, D4 groups. Compared with D2 and D4 groups, the mRNA and protein expression of GFAP in U2 and U4 groups were decreased, and the relative expression of RHO mRNA and protein were all increased (P < 0.01). ConclusionhUCMSC could migrate and integrate into the retina, after the transplantation into the vitreous cavity of diabetic rats, which reduced the expression of GFAP, but enhanced the expression of RHO.
ObjectiveTo observe the effects on the function and structure of retina in diabetic rats by intravitreal transplantation of retinal nerve stem cells (NSC) differentiated from human umbilical cord mesenchymal stem cells (hUCMSCs). MethodsFifty clean male Sprague-Dawley rats were randomly divided into normal control with 9 rats (group A) and diabetes mellitus (DM) group with 31 rats. The DM models were induced by intraperitoneal injection of streptozocin. The rats of DM group were randomly divided into four groups after 10 weeks: rats with DM only (group B), diabetic rats with saline intravitreal injection (group C), diabetic rats with NSC intravitreal injection (group D), and 9 rats for each. The rats in the group A and B received no treatment. The retinal function was examined by the flash-electroretinogram on 2, 4, 6 weeks after intervention, the latency and amplitude of a-wave, b-wave of Rod, a-wave, b-wave of Max reactions (Max-R) and the total amplitudes of OPs were recorded. The morphological changes of retina were observed by hematoxylin-eosin staining. ResultsOn 2 and 4 weeks after the intervention, the differences of latency and amplitude of b-wave of Rod, a-wave, b-wave of Max-R and the total amplitudes of OPs among group A-D were significant (P<0.05). Compared group D with group B, C, the amplitude of b-wave of Rod, Max-R and the total amplitudes of OPs were increased (P<0.05); latency of b-wave of Max-R was decreased (P<0.05). On 6 weeks after the intervention, the amplitude of b-wave of Rod and the amplitude of a-wave, b-wave of Max-R and the total amplitudes of OPs in group D were increased compared with group B and C (P<0.05), the latency of b-wave of Rod and Max-R in group D were decreased compared with group C (P<0.05). On 10 weeks after molding, each retinal layers were disordered in diabetes mellitus group. On 2 weeks after the intervention, the number of cells in the retinal layers in group B and C were reduced compared with group A, and the structure was more disorder. On 4 weeks after the intervention, the structure of each retina layer in group D arranged less disordered, and the number of retinal ganglion cells was more than group B and C. It was also found that the retinal vascular endothelial expanded and retinal blood vessels cells proliferated. ConclusionThe function of retina in diabetes mellitus rats is improved by intravitreal injection of retinal NSCs differentiated from hUCMSCs.
ObjectiveTo investigate the clinical outcome and therapeutic efficacy of short-pulse pattern scan laser (PASCAL) photocoagulation for diabetic retinopathy (DR). MethodsForty-three DR patients (70 eyes) including 19 males (32 eyes) and 24 females (38 eyes) underwent short-pulse PASCAL pan-retinal photocoagulation (PRP). There were 24 patients (42 eyes) with proliferative diabetic retinopathy (PDR) and 19 patients (28 eyes) with severe non-proliferative diabetic retinopathy (NPDR). The best corrected visual acuity was better than or equal to 0.1 in 62 eyes, worse than 0.1 in 8 eyes. Diabetic macular edema was found in 18 eyes. Short-pulse PASCAL PRP was applied with multi-spot arrays. Macular edema was treated by PASCAL macular mode (MAC A + MAC B) and/or single spot. Visual acuity and fundus examinations were analyzed at the one-year follow-up procedure. ResultsOne year after short-pulse PASCAL treatment, the final visual acuity was improved in 10 eyes, stable in 53 eyes, decreased in 7 eyes; macular edema was relieved in 38 eyes, aggravated in 12 eyes, and stable in 20 eyes. Of 42 eyes with PDR, neovascularization were regressed in 20 eyes, uncontrolled in 11 eyes which experienced additional photocoagulation (1-2 times) during the follow-up. Among the 11 uncontrolled eyes, 3 eyes (3/11) received vitrectomy due to vitreous hemorrhage. ConclusionPASCAL might stabilize the progress of diabetic retinopathy safely and effectively.
Objective To evaluate the therapeutic effect of intravitreal injection with bevacizumab (Avastin) (IVB)combined with extra panretinal photocoagulation (E-PRP) for highrisk proliferative diabetic retinopathy (PDR).Methods A total of 57 eyes of 53 patients with highrisk PDR underwent intravitreal injection combined with E-PRP. The examinations of vision acuity, intraocular pressure, iris fluorescein angiography (IFA),fundus photos and fundus fluorescein angiography (FFA) were performed on all of the patients before and 1,2,3,and 6 months after the treatment; the results of the examinations before and after the treatment were compared and analyzed.The average follow up was 6 months.Results The mean visual acuity was (0.143plusmn;0.072) before the treatment and (0.218plusmn;0.128) 7 days after the tretment; the difference was significant (t=-7.940, Plt;0.05). The mean visual acuity 1,3,and 6 months after E-PRP (0.228plusmn;0.138, 0.223plusmn;0.125,0.220plusmn;0.134, respectively) differed much from that before IVB (Plt;0.05), but not so much from that after IVB (Pgt;0.05).The mean intraocular pressure of 21 eyes which had the neovascularization of pupil margin and iris surface before and 7 days after IVB was (26.632plusmn;2.629) and (19.316plusmn;3.092) mm Hg(1 mm Hg=0.133 kPa), respectively; the difference was significant (t=12.838, Plt;0.05) . The mean intraocular pressure 1,3,and 6 months after E-PRP was (16.947plusmn;2.345),(16.474plusmn;1.611), and (16.421plusmn;4.702) mm Hg, respectively, which differed much from that before and after IVB (Plt;0.05). Neovascularization on the disc and the retinae of 57 eyes were subsided partly, and a significant reduction or disappeared of the area of retinal neovascularization and the blood vessel leakage were observed 7 days after IVB. The neovascularization of pupil margin and iris surface of 21 eyes disappeared, and the IFA leakage decreased. The results of FFA 2 months after E-PRP showed that the one-off efficiency of E-PRP was 68.4%;12 eyes (21.1%) needed an additional laser, in which 6 eyes (10.5%) underwent vitreous surgery. Conclusion IVB combined with E-PRP as a treatment for highrisk PDR may improve the regression of retinal neovascularization and the reduction of vascular permeability,and prevent or reduce the complications and improve the therapeutic effect.
Objective To evaluate the visual prognostic factors in vitreoretinal surgery for diabetic tractional retinal detachment (DTRD). Methods 102 eyes of 86 consecutive patients with DTRD underwent vitreoretinal surgery were analyzed retrospectively. All cases diagnosed via indirect ophthalmoscope and B ultrasonic scan after mydriasis. Followup duration varied from 12 to 56 months (mean: 23 months). Best corrected visual acuity (BCVA) and anatomic success were observed postoperatively. The patients were divided into visual acuity improved group and didn't improved group. Ttest, Chisquare test and Multivariate Logistic regression analysis were performed to predict the prognosis of visual acuity. Results After primary vitreoretinal surgery, 87 eyes (85.3%) were anatomically reattached, 15 eyes (14.71%) needed reoperation because of the recurrence of retinal detachment (RD). Postoperative BCVA improved and better than 0.05 in 49 eyes (48.04%), reduced or increased but less than 0.05 in 53 eyes (51.96%). Comparing natural factors between these two groups, only combined cataract surgery and optic nerve atrophy were significant different (chi;2=5.266,9.274;P=0.022,0.002). Among post-operative complications only the RD recurrence was significant different (chi;2=12.059，P=0.000). Multivariate Logistic regression revealed recurrence of RD and optic nerve atrophy were two independent risk factors in the final BCVA (P=0.003,0.041;OR=33.518、4.079). Preoperative PRP was identified as the only protecting variable in the final BCVA(P=0.034，OR=0.270).Conclusion This study revealed recurrence of RD and optic nerve atrophy were two independent risk factors in final BCVA of DTRD patients.
Objective To compare the therapeutic effects of 577 nm laser and 532 nm laser panretinal photocoagulation (PRP) in the treatment of non-proliferative diabetic retinopathy (NPDR). Methods This is a prospective controlled study. A total of 23 patients (41 eyes) with clinically diagnosed severe NPDR were randomly divided into two groups including 577 nm group (11 patients, 20 eyes) and the 532 nm group (12 patients, 21 eyes). 577 nm group and 532 nm group received 3 - 4 times PRP with single-point mode. The laser energy and the number of laser spots were compared, and the laser energy density was calculated. Before treatment and 1 day, 1, 3, 6 and 12 months after treatment, the changes of best corrected visual acuity (BCVA), average threshold sensitivity, a/b-wave amplitude of flash ERG (F-ERG) in the 30° - 60° visual field, and fundus fluorescein angiography (FFA) were compared between two groups. Results The response rate was 85.0% and 23.8%, respectively in the 577 nm and 532 nm group, the difference was statistically significant (χ2=15.43，P < 0.05).Compare to the pre-treatment measurement, the average threshold sensitivity, a/b wave amplitude of F-ERG and the 30° - 60°visual field were reduced at 1 day after treatment both in the 577 nm and 532 nm group, the difference were statistically significant (F=8.68, 7.57, 4.52; P < 0.05). The average threshold sensitivity (t=2.41, 3.48, 1.23), a/b wave amplitude (a wave: t=5.82, 4.45, 7.83；b wave: t=5.40, 3.23, 4.67) of F-ERG were different between 577 nm and 532 nm group at 3 , 6 and 12 months after treatment (P < 0.05). There was no retinal neovascularization and non-perfusion region in two groups at 6 months after treatment. The average laser power were (436.25±54.65) and (446.43±35.61) mW, number of laser spots were (1952.95±299.09) and (2119.05±302.69) spots, energy density were (7.60±1.30) and (7.60±3.00) mW×ms/μm2 in the 577 nm group and 532 nm group, respectively. There was no difference in average laser power (t=1.35), number of laser spots (t=2.85) and energy density (t=1.99) between two groups (P > 0.05). Conclusion Compared with the 532 nm laser, 577 nm laser treatment has better visual outcomes for NPDR patients.
Objective To compare the clinical results of yellow micro-pulse laser and traditional laser grid (MLG) photocoagulation for diabetic macular edema (DME). Methods Seventy-eight patients (106 eyes) with DME diagnosed by fundus fluorescein angiography (FFA) and optical coherence tomography (OCT) were enrolled in this study. The patients were divided into micro-pulse group (39 patients, 51 eyes) and MLG group (39 patients, 55 eyes). The patients of micropulse group underwent 577 nm yellow micro-pulse laser therapy, while the patients of MLG group underwent continuous wavelength laser photocoagulation with a 561 nm yellow green laser. All the patients were examined documenting corrected visual acuity, macular retinal thickness (CMT) and mean sensitivity within macular 10 deg; examination before and after treatment. Six months after treatment was considered as the judgment time for the therapeutic effects. The mean corrected visual acuity, CMT and MS were comparatively analyzed. Results Six months after treatment, the mean corrected visual acuity of micropulse group and MLG group were 0.45plusmn;0.20 and 0.42plusmn;0.20, which increased significantly compared to those before treatment (t=3.404,2.316; P＜0.05). The difference of mean corrected visual acuity between before and after treatment of micro-pulse group and MLG group were 0.08plusmn;0.02 and 0.06plusmn;0.03, the difference was statistically significant between two groups (t=0.532, P＞0.05). The mean CMT of micropulse group and MLG group were (323.94plusmn;68.30) and (355.85plusmn;115.88) mu;m, which decreased significantly compared to those before treatment (t=4.028, 2.039; P＜0.05). The difference of mean CMT between before and after treatment of micro-pulse group and MLG group were (55.12plusmn;13.68) and (22.25plusmn;10.92) mu;m. The difference was not statistically significant between two groups (t=1.891,P＞0.05). The mean MS of micro-pulse group and MLG group were (6.63plusmn;2.65) and (4.53plusmn;1.81) dB. The mean MS of micro-pulse group increased significantly compared to that before treatment(t=3.335,P＜0.05). The mean MS of MLG group decreased significantly compared to that before treatment (t=3.589,P＜0.05). The difference of mean MS between before and after treatment of micro-pulse group and MLG group were (1.10plusmn;0.33) and (-0.91plusmn;0.25) dB.The difference was statistically significant between groups (t=4.872，P＜0.05). Conclusions In the treatment of DME, yellow micro-pulse laser therapy and MLG can improve visual acuity, and reduce CMT. In addition, yellow micro-pulse laser therapy can improve the MS, but MLG reduces MS.
Objective To observe the effects of dual targets intervention on the expression of vascular endothelial growth factor (VEGF) and connective tissue growth factor (CTGF) in diabetic rat retina. Methods Forty-eight Sprague -Dawley rats were randomly divided into control group (CON1 group) and diabetes mellitus group (DM group). The rats of DM group were induced with streptozotocin injection creating a diabetic model. Retinas were obtained at eight, 10, 12 weeks after DM induction from both groups. CTGF and VEGF mRNA levels were examined by realtime reverse transcriptionpolymerase chain reaction (RT-PCR). Based on the results of above experiments, 60 rats with same conditions were selected. Fifty rats were induced with streptozotocin injection creating a diabetic model, and 10 rats comprised the control group (CON2 group). Then the 50 diabetic rats were randomly divided into ranibizumab and CTGF shRNA dual targets intervention group, ranibizumab singletarget intervention group, CTGF shRNA singletarget intervention group and nonintervention group. Retinas were obtained at one week after intervention from all the groups. CTGF and VEGF mRNA levels were examined by RT-PCR. Results The levels of CTGF mRNA were significantly higher in DM group than that in CON1 group at the 8th weeks after DM induction, and this upregulation was maintained through the 12th week (t=-2.49, -2.67, -2.42;P＜0.05). There was no difference on VEGF mRNA levels between DM group and CON1 group at the 8th weeks after DM induction(t=-0.443,P=0.669). VEGF mRNA levels of DM group started to be significantly elevated over those in the CON1 group at the 10th week, and remained to be higher at the 12th week (t=-2.35, -2.57;P＜0.05). The VEGF mRNA of ranibizumab single-target intervention group was significantly lower than that in non-intervention group (t=-3.44,P=0.014), which was similar to CON2 group (t=-1.37,P＞0.05); however, the CTGF mRNA level was significantly increased as compared to the nonintervention group (t=2.48,P＜0.05). In the CTGF shRNA single-target intervention group, the levels of CTGF and VEGF mRNA were decreased as compared to the non-intervention group (t=0.23, -2.92;P＜0.05). In the ranibizumab and CTGF shRNA dual targets intervention group, the levels of CTGF and VEGF mRNA were decreased as compared to the non-intervention group (t=-6.09, -5.11;P＜0.001), which was similar to CON2 group (t=-1.16, 1.139; P＞0.05). Conclusions Both CTGF and VEGF gene expression are up-regulated in early diabetic rat retina, and the level of CTGF increased earlier than VEGF. Ranibizumab combined with CTGF shRNA could simultaneously reduce the level of CTGF and VEGF mRNA in diabetic rat retina.