Objective To investigate the effect of hepatocyte growth factor (HGF) on the barrier function of retinal peigment epithelium (RPE) and to detect the pathological mechanism of retinal detachment (RD) induced by over expression of HGF in RPE. Methods Sub-retina injection of E1/E3deleted adenoviral vectors encoding HGF (Ad CMV.HGF) and green fluorescent protein (Ad CMV.GFP) in adult pigmented rabbits [5times;104 plaque-forming units (pfu)/eye] to set up the model of retinal detachment. The ocular fundus and pathological changes were observed 3, 7, 14, and 28 days after injection. The expression level of HGF in retina and vitreous body was detected by immunohistochemistry and enzyme linked immunosorbent assay (ELISA). Results In the control eyes injected with AdCMV.GFP, expression of GFP only detected in RPE monolayer. The eyes injected with AdCMV.HGF had b HGF immune positive action in RPE cells at the injection site. The expression level of HGF in vitreous body reached the peak 7 days after injection and decreased to the basic level 28 days after injection. Chronic RD and chronic choroidal inflammation were found in the eyes injected with AdCMV.HGF within the time frame of HGF expression. Proliferative RPE cells were found in subretinal space in the region of RD, and multilayered cellular membranes developed in some eyes. Conclusion Over expression of HGF in RPE may induce chronic serous RD with subretinal proliferation of RPE, which suggests that HGF should be further studied as a target for therapeutic intervention in RD. (Chin J Ocul Fundus Dis, 2007, 23: 193-197)
Objective To investigate the interference effect of nerve growth factor (NGF) on apoptosis of retinal cells in experimental retinal detac hment (RD). Methods Twenty seven Sprague-Dawely rats were selected, and the left and right eyes were in the experimental control group and NGF group, respectively. After the RD model was set up by subretinal injection with sodium hyaluronate, 5mu;l NGF（1mu;g/mu;l）was injected into the vitreous body of the right eyes which were in the NGF group; 5mu;l PBS was injected into vitreous body of left eyes which were in the experimental control group. The injection was performed once every 4 days till the end of the observation period. The eye balls of the 27 rats were extrafted 1.5, 3, 6, 12 hours, 1 day, 2, 4, 8 , 16, and 32 days after the RD model was established. Another 2 rats were selected as the normal control, which underwent none of the injections but eyeball extraction at the end of the observation period. TUNEL and transmission electron microscopy were used to detect the apoptosis of the retinal cells. Cell counts and statis tical analysis were used to assess results. Results Typical apoptosis cells were observed in the early time of RD. Apoptosis was found in each retinal layers, especially in inner and outer nuclear layers. The number of apoptosis cells increased as the time of RD was prolonged（Plt;0.01）. It was also found that apoptosis cells in NGF group were less than that in the experimenta l control group（Plt;0.01）. Conclusion Intravitreous injection exogenous NGF may inhibit the apoptosis of retinal cells in experimental RD. (Chin J Ocul Fundus Dis, 2006, 22: 333-335)
ObjectiveTo observe the expression of connective tissue growth factor (CTGF) in injured model of retinal pigment epithelial (RPE) cells and the promoting effect of CTGF on migration of RPE cells.MethodsCultured monolayer-confluent human RPE cells were scraped with a trephine and a cotton stick, and set up the injured model of RPE cells with round scraped area. Immunohistochemistry and in situ hybridization(ISH) were used to detect the expression of CTGF protein and mRNA in injured RPE cells at distinct time points after injury. The number of RPE cells migrated to injured area was measured and the effect of CTGF on migration of RPE cells and the effect of dexamethasone (DEX) on the promoting process of CTGF were observed.ResultsThe results of immunohstochemistry and ISH indicated the weak positive expression of CTGF in RPE cells at the edge of scrape 6 hours after injury, and the positive expression increased gradually as time goes by after the injury. Strong positive expression of CTGF in RPE cells at the edge of scrape was found 24 and 48 hours after injury. Rebuilt human CTGF stimulated migration of RPE cells in a dose-depended manner, and DEX significantly inhabited the migration.ConclusionCTGF involves in the procedure of repair of injury of RPE cells, which may play an important role in the pathogenesis of intraocular proliferative diseases such as proliferative vitreoretinaopathy.(Chin J Ocul Fundus Dis, 2005,21:306-309)
Objective To investigate the expression of matrix metallo proteinase (MMP)-2 and MMP-9 in rats′optical nerves after extrusion wound. Methods We set up the model of rats with extrusion wound of the optical nerves, detected activity changes of MMP-2 and MMP-9 in the optical nerves by gelatin zymography, identified the attribute by Western blotting, and verified the expression of mRNA of MMP-2 and MMP-9 by reverse transcriptase-polymerase chain reaction (RT-PCR ). Results MMP-2 existed in normal optial nerves and optical nerves with extrusion wound, while MMP-9 was only detected in the latter. The expression of MMP-9 was the highest 1 day after the extrusion wound, while that of MMP-2 was the highest 7 days after the extrusion wound. Conclusions MMP-2 and MMP-9 may participate in the pathological recovery process of optical nerves after extrusion wound. The glial cells in the optical nerves may be one of the sources of MMP-2 and MMP-9. (Chin J Ocul Fundus Dis，2003，19：269-332)
Objective To observe the expression of erythropoietin (EPO) and its receptor (EPOR) mRNA and protein levels in retinae of mice with oxygen-induced retinopathy, and to evaluate the effect of EPO and EPOR in retinal vascular develo pment and in the occurrence and development of oxygen-induced retinopathy. Methods One hundred and thirty-two 7-day-old C57BL/6J mice were divided into two g rou ps: normal control group (control group) and oxygen-induced retinopathy group (experimental group). The proliferative neovascular response was estimated by obse rving the vascular pattern in adenosine diphosphatease (ADPase) stained retina flat-mounts by executing 6 mice in each group at the 12th, 15th, and 17th day, respectively. The expression of EPO, EPOR mRNA was determined by reverse transcription-polym erase chain reaction (RT-PCR), and the protein levels of EPO and E PO R were determined by immunohistochemistry. RT-PCR and immunohistochemistry were done every other day from the 7th to the 21st day. Results In the control group, retinal vascularization was found. In the experimental group, the large vesse ls were constricted straight, the branches decreased, and alarge nonperfusion area was observed at the 12th day; the large vessels were dilated and tortuous and neovascularization occurred at the 15th day; a mass of neovascularization was found and the vascular net structure of the deep and shallow layer was destroye d at the 17th day. The expression of EPO mRNA decreased from the 7th day and kee p decreasing in the whole oxygen-breathing duration in the experimental group. A fter the mice were returned to room air, the expression increased obviously from the 15th day and kept the high level until the 21st day. The expression of EPO mRNA increased at the 7th day and reached the peak at the 11th day, and kept the high level until the 21st day. The changes of protein levels of these three fac tors were later than that of their mRNA, but had the same trend. The difference of the expression between the two groups at the different time point was signifi cant except for the 7thday point (Plt;0.05). Conclusion It 's suggested that EPO and EPOR played important roles on the development of normal retina vascularizati on and the pathogenesis of ROP, which may provide new conception and method for the prevention and treatment of the oxygen-induced retinopathy.
ObjectiveTo investigate the expression and relation of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in rats with diabetic retinopathy.MethodFifty-five Wistar rats were randomly divided into the control group (10 rats), and 1, 3, and 5-month-diabetes group (15 rats in each diabetes group), and the diabetic models were set up. The expressions of VEGF and bFGF were detected by situ hybridation and immunohistochemistry on retinal paraffin sections.ResultsThe results of situ hybridation showed that expression of bFGF was found in 3-month-deatbtes group with the percentage of 77.8%, and 88.9% in 5-month-deatbtes group; the positive expression of VEGF was not found in 3-month-deatbtes group but in 5-month-deatbtes group with the percentage of 66.7%. Immunohistochemistry indicated that the positive expression of bFGF started in 3-month-deatbtes group with the percentage of 55.6%, and 88.9% in 5-month-deatbtes group; the percentage of the expression of VEGF was 33.3% in 3-month-deatbtes group and 88.9% in 5-month-deatbtes group.ConclusionThe expression of VEGF occurs after the expression of bFGF in rats with DR.(Chin J Ocul Fundus Dis, 2005,21:37-40)
Objective To determine whether kringle 4-5 could inhibit choroidal neovascularization (CNV) in mice induced by argon laser photocoagulat ion. Methods Fundus laser photocoagulation was performed on C57BL/6J mice to induce CNV. In treatment group, 20 μg (low dosage group) and 50 μg (high dosage group) kringle 4-5 were injected retrobulbarly after photocoagulation. In control group, equilibrium liquid was injected retrobulbarly. Choroidal neovascularization was evaluated on the 7th and 14th day after photocoagulation by fundus fluorescein ang iography. The mice were killed on the 14th day after photocoagulation, the lesions were evaluated histologically and immunohistochemically, and the expression of CD105 was detected. The Expression of VEGF and bFGF was detected by immunohist ochemistry on the 4th day after photocoagulation.Results The incidence of CNV was 64.3% in control group, 51.2%（P＜0.05）in low dosage group, and 44. 1% (P＜0.01) in high dosage group. The CNV lesions were smaller in kringle 4-5 injected eyes in a dose-dependent manner and the number of proliferative vascular endothelial cells in the subretinal membrane of the treated eyes was smaller than that of the control eyes. There was no significant difference of the expression of VEGF and bFGF between the mice in control and treatment group.Conclus ions Kringle 4-5 could inhibit the development of choroidal neovascularization in the experimental mice model.(Chin J Ocul Fundus Dis，2003，19：201-268)
Objective To evaluate the effect of vascular endothelial cell growth factor (VEGF) antisense oligodeoxynucleotides (ASODNs) on the expression of VEGF in rats with oxygen-induced retinopathy. Methods Thirty newborn Sprague-Dawley (SD) rats were randomly divided into 3 groups:normal control group, disposal group and non-disposed group, The animal models with oxygen-induced proliferative retinopathy were established by raising the rats in hyperoxic environment. Retrobulbar injection was performed with VEGF ASODNs or normal saline on the rats in 3 groups respectively. The intraocular tissues (all the tissues except the cornea, sclera, and lens) and serum were collected, and the expressions of VEGF were determined by using competitive enzyme immunoassay.Results The expressions of VEGF in intraocular tissues of rats in disposal group were significantly lower than those in non-disposed group (P＜0.05), and there was no statistical difference between the disposal and normal control group (P＞0.05). There was no significant difference of the expressions of VEGF in serum of rats between the disposal and non-disposed group (P＞0.05), which were both lower than those in the normal control group (P＜0.05). Conclusion VEGF ASODNs could significantly inhibit the expression of VEGF in intraocular tissues. (Chin J Ocul Fundus Dis，2003，19：172-174)
Objective To determine the effect of methimazole (MMI) on retinal vascular development in neonatal rats, and to investigate the relationship between the concentration of insulin-like growth factor-I (IGF-I) in serum and the development of normal blood vessels and between the concentration of IGF-I and the formation of abnormal blood vessels. Methods There were 75 neonatal SpragueDawley rats in experimental group whose mothers were raised with water with 0.1% MMI at the first day of parturition. Another 50 neonatal rats were in the control group whose mothers were raised with normal water. The rats in the two groups were sub-divided into 4day and 10day subgroup, respectively. The retinal flatmount of the right eyes were stained with adenosine diphosphatase (ADPase); with the paraffin section of the left eyes, the number of nucleolus breaking through retinal inner limiting membrane was counted and the retinal blood vessels were evaluated. Serum IGF-I levels were detected by radioimmunoassay, and the weight of the neonatal rats in each group were observed and recorded. Results The incidence of retinal neovascularization in 10 day MMI group was 27%, and 0% in 4-day MMI group and control group. The serum IGF-I level in 4-day and 10-day MMI group (73.07 ng/ml, 175.13 ng/ml) was obviously lower than which in the 4-day and 10-day control group (168.73 ng/ml,306.38 ng/ml) （P=0.00）. Obvious slow growth of the neonatal rats was found in MMI group compared with which in the control group. Conculsions MMI may inhibit the normal growth of retinal blood vessels and lead neovascularization, which may relate to the initial decrease of the serum IGF-I level. (Chin J Ocul Fundus Dis, 2007, 23: 198-201)
Objective To investigate the effects of intravitreal injection of matrix metalloproteinase-3 (MMP-3) on the vitreoretinal adhesion and the vitreous gelatin. Methods Twenty-four pigmented rabbits were randomly divided into 3 experimental groups（group A, B, and C）and one control group with 6 rabbits (12 eyes) in each. Different concentrations of 0.1 ml MMP-3 (5,10, 20 ng in group A, B, and C, respectively) and equivalent dose of balanced salt solution were intravitreally injected to the rabbits, respectively. Clinical examinations (such as gross observation, slit-lamp biomicroscopy, indirect fundus ophthalmoscopy ), electroretinography (ERG) and fundus fluorecein angiography (FFA) were taken before and after injection. Results One week after injection, posterior vitreous detachment (PVD) and focal vitreous liquefaction were recognized clinically for the first time in 1 eye in group B. By the end of this study, clinically detected PVD developed in 1 eye in group A, 3 eyes in group B, but the synchisis developed slowly, and no liquefaction or PVD occurred in control group. As for the histological examination, partial PVD was observed in 1 eye in group A and 3 eyes in group B 60 minutes after injection. All of the eyes in group A and B showed partial PVD 1 week after injection, and the area of PVD enlarged in contrast with before. Complete PVD were recognized in 1 eye in group A and 3 eyes in group B 15 weeks after injection, and the cleavage was narrow and limited. In group C, inflammatory cell infiltration in the inner layer of retina, destruction of retinal structure, and fluorescein leakage at late phase was found in the early period after injection. Conclusions MMP-3 is effective in disrupting the adhesion of the posterior hyaloid to the inner limiting membrane leading to PVD, and helpful to some extent in producing vitreous liquefaction. The dose of 10 ng MMP-3 is safe and effective for the rabbits eyes, which may be used as an promising assistant of vitreous surgery. (Chin J Ocul Fundus Dis，2004，20：67-132)