Objective To investigate the effects of genistein with different concentration on proliferation and apoptosis of cultured human retinal pigment epithelial (RPE) cells in vitro. Methods The effect of genistein with the concentration of 5，10，25，50，75，and 100 mg·L-1on the proliferation of cultured RPE was examined by tetrazolium salt (MTT) assay and AgNORs staining. The cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick-end labeling (TUNEL) methods, in the mean time, the morphologic changes of cell apoptosis were observed by light microscopy and transmission electronic microscopy, the results of which were compared with the normal RPE cells. Results Genistein with the concentration of 25, 50, 75, and 100 mg·L-1had a dose-dependent and time-dependent antiproliferative effects on RPE cells with the inhibitory rate of 12.0%-64.6% (P＜0.05). The results of AgNORs staining showed that the number of AgNORs in the nucleolus decreased when treated by genistein. In TUNEL staining, the median of percent of apoptotic RPE cells was 7.6%, 9.8%, 13.7% when treated with 50 mg·L-1genistein, 10.3%, 16.4%, 23.4% when treated with 75 mg·L-1genistein, and 15.4%, 21.2%, 35.8% when treated with 100 m g·L-1genistein respectively for 24, 48, and 72 hours. After the treatme nt with 50 mg·L-1genistein for 48 hours, the apoptosis in the nucleolus of RPE cells was found. Conclusions Genistein with different concentrations has a dose-dependent and time-dependent antiproliferative effect on RPE cells. Genistein can induce the apoptosis of RPE cells when it reaches a certain extent of concentration. (Chin J Ocul Fundus Dis，2004，20：241-244)
Objective To investigate the effect of hypericin on the activity of protein kinase C (PKC) in cultured human retinal pigment epithelium (RPE) cells in vitro.Methods RPE cells were cultured in standard medium with 10% serum concentrations containing 0.5 to 5.0 μmol/L hypericin with or without preincubation of phorbol 12-myristate 13-acetate (PMA). The activities of cytosolic PKC (c-PKC) and membranous PKC (m-PKC) were assayed by PKC kit. Results The original activities of c-PKC and m-PKC of RPE cells were (35.34±4.10) pmol·min-1·mg-1and (62.52±8.80) pmol·min-1·mg-1.The activity of c-PKC in RPE cells with PMA preincubation decreased rapidly in 5 minutes, with a subsequent slow decrease after 20 minutes and a decrease to 18% of the activity of c-PKC in RPE cells without PMA preinubation after 60 minutes. While the activity of m-PKC in RPE cells with PMA preincubation increased gradually after 5 minutes and reduced after reached the peak at 40 minutes, and then returned to baseline after 60 minutes, eventually decreased below 30% of the control group. When RPE cells were cultured with PMA for 48 hours, the activities of c-PKC and m-PKC were hardly detectable, while RPE cells were cultured with both PMA and hypericin, hypericin could counteract most of down-regulation by PMA. Conclusion Hypericin may inhibit the translocation of PKC in RPE cells,change the activity of PKC, promote the apoptosis of RPE cells likely,and then prevent proliferative vitreoretinopathy. (Chin J Ocul Fundus Dis，2003，19：55-58)
PURPOSE:To evaluate the ability Of retinoic acid(RA) in silicon oil(SiO)to inhibit the proliferation of injected intraocular fibroblast cells.
METHODS:Thirty New Zealand white rabbits (58 eyes)were divided into three groups. In control group ,only SiO(10 eyes)or BSS(10 eyes)were injected intravitreally and 5mu;g/ml (18 eyes)or 10mu;g/ml (20 eyes)RA in SiO were injected into other lwo groups respectively. Three days after gas-compression vitrectomy, 2 times;105 fibroblasts and Sio(0.5ml)or BSS(0.5ml)were injected in all eyes sequentially.The morbidity of tractional retinal detachment (TRD) were
observed by ophthalmoscope until 4 weeks.
RESULTS:After 4 weeks,in control ,5mu;g/ml RA in SiO and 10mu;g/ml RA in SiO
group,80. 00%,44.44%,and 30.00% eyes developed TRD respectively. Significant statistical differences were found between the control group and the two treated groups (P＜0.05).
CONCLUSIONS:5mu;g/ml or 10mu;g/ml RA in SiO can inhibit the occurrence of TRD effectively.
（Chin J Ocul Fundus Dis,1997,13:174-176）
Objective
To observe the expression of proinflammatory factors messenger RNA(mRNA) in periretinal membrane of proliferative vitreoretinopathy.
Methods
Fourteen specimens of periretinal membrane were collected during vitrectomy, and they were made into paraffin sections.The presence of mRNA coding for IL-1,IL-6,IL-8 and TNF alpha was observed by in situ hybridization(ISH) with biotin labeled oligonuclotide probes respectively.The eyeball after corneal grafting was made as normal control.
Results
No expression of proinflammatory factors mRNA was found in normal human retina.Positive staining was present in 5 specimens.In these specimens, IL-1βmRNA was found in 3 specimens and TNFαmRNA was found in 3 specimens,there is 1 specimen expressed IL-8 mRNA and 3 specimens expressed IL-6 mRNA.In these positive specimens, one contained cells expressing mRNA for IL-1βbeta and IL-6, and one exhibited cells expressing mRNA for IL-1β、IL-8 and TNFα,two membranes possessed positive cells for IL-6 and TNFαmRNA, one membrane contained IL-1βmRNA positive cells only.
Conclusion
These findings suggest that these cytokines may be locally produced by cells infiltrating epiretinal membranes. The expression of IL-1β, IL-6, IL-8 and TNFαmRNA within retinal membranes provides further evidence of a pathogenic role of these cytokines in proliferative vitreoretinopathy.
(Chin J Ocul Fundus Dis, 2002, 18: 286-288)
Objective
To investigate the therapeutic effects of subconjunctival verapamil on outcome in an experimental model of traumatic proliferative vitreoretinopathy.
Methods
An experimental model of traumatic proliferative vitreoretinopathy was induced in pigme nt rabbits,which then were selected randomly to receive either subconjunctival verapamil injection treatment or a placebo injection(control)daily for 3 weeks.Animals were examined by indirect ophthalmoscopy at weekly intervals for 5 weeks. Eyes were enucleated for light microscopy 5 weeks later.
Results
Fifty-six percent(18 of 32)of the rabbits receiving subconjunctival verapamil injection had developed tractional retinal detachment,whereas eighty-one percent(26 of 32)of control animals had developed tractional retinal detachment(chi;2=4.655,P=0.031).The results of clinical examination and light microscopy didn't show evidence of toxicity between the verapamil treated animals and control animals.
Conclusion
Subconjunctival verapamil decreased the incidence of tractional retinal detachment due to traumatic proliferative vetreoretinopathy in this rabbit model.Verapamil at the dose used in this model has no evident toxicity on rabbit eyes.Further studies are needed to determine the doseresponse and efficacy of the drug.
(Chin J Ocul Fundus Dis,1999,15:69-71)
PURPOSE:To measure the concentration changes of tumor necrosis factor a (TNF-alpha;)in vitreous during the development of experimental PVR induced by macrophages and explore the initial and mediated factors which stimulate the
cellular proliferation.
METHODS:Rabbit PVR model was induced by macrophages and the vitreous was taken at the 7th,14th,21st and 28th day and 4 eyes in each group. The TNF-alpha; levels in vivreous of the above examinated and control eyes were
measured with an ELISA kit.
RESULTS:The TNF-alpha; level in the vitreous reached its peak 434mu;g/ml at 21st day in the mod-el, then rappidly decreased to 122mu;g/ml at 28th day. CONCLUSION:The changes of TNF-a in the vitreous of the PVR model were parallel to the natrual phases of the development of PVR,indicating TNF-alpha; may play an important role in initiating and mediating the inflammation and cellular proliferation in the vitreous.
（Chin J Ocul Fundus Dis,1997,13: 231-233）
Objective
To study the expression of the fibronectin (FN) and beta;1 integrin (beta;1) in epiretinal membranes(ERM) of eyes with proliferative vitreoretinopathy(PVR).
Methods
wenty epiretinal membranes were obtained from eyes undergone vitrectomy for retinal detachment complicated with PVR and observed by immunohistochemical methods.
Results
Overexpression of FN and beta;1 were observed in 18 and 16 membranes respectively．
Conclusion
The synergism of FN and beta;1 in their action mignt be one of the important roles in the development of PVR.
(Chin J Ocul Fundus Dis, 2001,17:119-121)
Objective
To detect the variation rule of different cellular components, extracellular matrix, matrix-metalloproteinases (MMPs), and tissue inhibitors of metalloproteinases（TIMPs）in proliferative membranes in proliferative vitreoretinopathy (PVR) with different courses of disease, and to investigate the remodeling mechanism of PVR.
Methods
Sixteen surgically excised specimens of proliferative membranes from patients with rhegmatogenous retinal detachment combined with PVR with the course of disease of 2 months to 8 years were selected. The different cellular component of retinal pigment epithelial (RPE) cells and glial cells, component of extracellular matrix including fibronectin, laminin，and collagen types Ⅰ to Ⅳ, and matrix metalloproteinases (MMP2, MMP9) and TIMP1 in proliferative membrane were labeled by immunohistochemical method. The variati on of those labeled components in proliferative membrane in PVR duration and the correlation between these components and the course of PVR were analyzed.
Results
As the duration of PVR increased,the expression of RPE cells, fibronectin and MMP2 decreased (Plt;0.05）,while glial cells,collagen type Ⅰ and Ⅲ increased (Plt;0.05）.The positive staining of laminin and collagen type Ⅱ and Ⅳ were found, but the association with PVR duration was not detected. A negative correlation between PVR duration and RPE cells, MMP2, and fibronectin respectively and a positive correlation between PVR duration and glial cells, collagen Ⅰand Ⅲ respectively were detected. MMP2 positively related with variation of fibronect in. Positive staining of MMP9 and TIMP1 was recorded but did not change with the variation of the disease course.
Conclusion
During the formation and development of proliferative membrane in PVR, RPE cells, glial cells, fibronectin, collagen type Ⅰand Ⅲ and MMP2 take part in the remodeling of proliferative membrane.
(Chin J Ocul Fungdus Dis, 2006, 22:308-312)
Objective
To observe the expression of vascular endothelial growth factor (VEGF) in aqueous humor and vitreous body in eyes with proliferative vitreo-retinal diseases, and to investigate the role of VEGF plays in the pathoge nesis of proliferative vitreo-retinal diseases.
Methods
The concentration of VEGF in aqueous humor and vitreous body in eyes with proliferative vitreoretinopathy (PVR), retinal vein occlusion (RVO), proliferative diabetic retinopathy (PDR), and neovascular glaucoma (NVG) were measured by double antibodies sandwich enzyme-linked immunosorbent assay (ELISA).
Results
The concentration of VEGF in aqueous humor and vitreous body in eyes with PVR, RVO, PDR and NVG were obviously higher than that in the control group (Plt;0.05), respectively. Among all of the diseases, the concentration of VEGF in aqueous humor and vitreous body decreased orderly in NVG, PDR, RVO and PVR (Plt;0.05). The concentration of VEGF in vitreous body in eyes with PVR, RVO, PDR and in the control group were much higher than that in aqueous humor in corresponding groups (Plt;0.05). There was a negative correlation between the disease history and content of VEGF in aqueous humor and vitreous body in patients with PVR (r=－0.819, －0.823；Plt;0.05). The disease history positi vely correlated with the concentration of VEGF in aqueous humor and vitreous body in patients with RVO (r=0.913, 0.929；Plt;0.05), and the time of vitreous hemorrhage positively correlated with the concentration of VEGF in aqueous humor and vitreous body in patients with PDR (r=0.905, 0.920；Plt;0.05).
Conclusion
The concentration of VEGF in aqueous humor and vitreous body in patients with proliferative vitreo-retinal diseases significantly increases, and VEGF may play an important role in the pathoge nesis of proliferative vitreo-retinal diseases.
(Chin J Ocul Fundus Dis, 2006, 22: 313-316)
Objective
To investigate the occurrence, progress and conversion of hypotony in anterior proliferative vitreoretinopathy (aPVR), and to provide knowledge about how to prevent and treat it．
Methods
Animal models of chronic hypotony by aPVR were made with cultured ho mologous dermal fibroblasts on pigmented rabbits．The intraocular pressure (IOP) and ultrasound biomicroscopy(UBM) examination were taken preoperatively and on days 7,14, 28 and 56 postoperatively．Rabbits were killed on days 14, 28 or 56 postoperatively, prepared for histology and ultrastructure examination．
Results
The average IOP of experimental group was lower than that of control group on days 7,14,28 and 56 significantly (Plt;0．01)．UBM demonstrated that trip like echo emerged in front of ciliary body four weeks postoperatively, and tractional retinal detachment was found four weeks and eight weeks postoperatively in experimental group． Microscopic examination showed atrophy orabsence of the non-pigmented ciliary epithelium on days 28 and 56 postoperatively in experimental group．Electronic microscopy showed that the amount of mitochondrions decreased and there were many vacuoles in the non-pigmented ciliary epithelium in experimental group four and eight weeks postoperatively．
Conclusions
Atrophic change of the non-pigmented epithelium due to dragging effect of the ciliary body from the epiciliary membrane in aPVR might be the main cause of hypotony．
(Chin J Ocul Fundus Dis, 2001,17:216-220)
