Platelet derived growth factor and fibroblast growth factor basic levels in the vitreous of patients with vitreoretinal disorders

AIM—To determine the potential role of basic fibroblast growth factor (bFGF) and platelet derived growth factor (PDGF) in the pathogenesis of proliferative vitreoretinopathy (PVR).
METHODS—An enzyme linked immunosorbent assay technique was used to determine the quantities of bFGF and PDGF in 38 vitreous samples taken from patients undergoing trans pars plana vitrectomy (PPV) for a variety of vitreoretinal disorders.
RESULTS—bFGF levels ranged from undetectable to 318.7 pg/ml. The mean concentration was 27.57 pg/ml. PDGF levels ranged from undetectable to 160 pg/ml, the mean concentration being 40.06 pg/ml. Eight of 13 patients with clinical evidence of retinal detachment (RD) and PVR had significantly elevated bFGF concentrations, whereas only one of 11 patients with RD and no PVR had detectable bFGF; seven of eight patients with RD and PVR had elevated PDGF concentrations in the vitreous, whereas all 10 patients with RD and no PVR had no detectable vitreous PDGF. Eight patients with vitreous haemorrhage had raised PDGF concentrations, and the levels were particularly high (>120 pg/ml) in those two patients with active neovascularisation. Two out of nine patients with vitreous haemorrhage had raised bFGF levels.
CONCLUSIONS—bFGF and PDGF concentrations are elevated in PVR even in the absence of vitreous haemorrhage, and not in patients with RD uncomplicated by PVR. This observation suggests that both cytokines may be involved in the pathogenesis of PVR.

Keywords: basic fibroblast growth factor; platelet derived growth factor; proliferative vitreoretinopathy     

Macrophages in Human Epiretinal and Vitreal Membranes in Patients with Proliferative Intraocular Disorders

Purpose: Macrophages are versatile cells and have been known as a cellular component of epiretinal membranes of proliferative intraocular disorders (PID). However, the origin and functions of these cells in the membranes are not yet clear. In the present study we investigated the characterization of macrophages/monocytes in various types of human epiretinal membranes from patients with proliferative vitreortinopathy (PVR) , proliferative diabetic retinopathy (PDR) and ocular perforating injury by means of immunohistochemical techniques. Methods: A total of 49 epiretinal membranes of PID in which 24 were PDR specimens, 17 were PVR and 8 were proliferations after perforating eye injury were studied. Monoclonal antibodies against human macrophages, monocytes, HLA-DR antigen and interleukin-1 (IL-1) were used. The alkaline phosphatase anti-alkaline phosphatase (APAAP) technique was performed.Results: The results showed that 19 out of 24 PDR specimens (79%), 15 out of 17 PVR specimens (88%) and 7 out of 8 tr     

视网膜下液碱性成纤维细胞生长因子的临床意义

采用“三明治”酶链免疫分析法，检测了４３例患有视网膜脱离的视网膜下液的碱性成纤维细胞上长因子（ｂＦＧＦ）的含量，结果显示随着增殖性玻璃体视网膜病变（ＰＶＲ）严重程度的增加，样品中ｂＦＧＦ的含量也增加，高含量的ｂＦＧＦ将促进在ＰＶＲ收缩膜中占主要成分的视网膜色素上皮细胞的迁移和增殖，因此，ｂＦＧＦ有可能在ＰＶＲ在发展过程中起着重要作用。     

VEGF和bFGF在蒙古族糖尿病视网膜病变中的表达

目的：研究血管内皮生长因子( vascular endothelial growth factor,VEGF)和碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)在蒙古族糖尿病患者视网膜病变中的表达,探讨其中在糖尿病视网膜病变(diabetic retinopathy, DR)发生过程中的作用。
　　方法：对2013-02/2015-02在内蒙古自治区人民医院眼科83例蒙古族糖尿病患者根据视网膜病变情况分为：无糖尿病视网膜病变患者(NDR 组)25例、背景型糖尿病视网膜病变患者(NPDR 组)31例、增殖型糖尿病视网膜病变患者(PDR 组)27例。36例年龄匹配健康志愿者作为对照组。分别采集患者外周静脉血,采用 ELISA 法分别对血清中 VEGF 和 bFGF 的表达情况进行测定。
　　结果：蒙古族糖尿病患者血清中 VEGF 和 bFGF 的水平显著高于对照组;三组蒙古族糖尿病患者中,PDR 患者血清VEGF 和 bFGF 水平高于 NDR 和 NPDR 组,差异具有统计学意义(P<0.05),NPDR 患者血清 VEGF 和 bFGF 水平又显著高于 NDR(P<0.05)。
　　结论：高表达的 VEGF 和 bFGF 可能是蒙古族 DR 发生、发展的重要致病因素。     

增殖性糖尿病视网膜病变基质金属蛋白酶的表达

目的:研究增殖性糖尿病视网膜病变(proliferative diabetic retinopathy,PDR)视网膜增殖膜纤维化和新生血管形成与MMP-2和MMP-9异常表达的关系.方法:采用免疫组织化学方法,检测PDR患者20例视网膜前纤维血管膜MMP-2和MMP-9的表达,并与增殖性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)10例非血管性增殖膜进行对比研究.结果:PDR增殖膜MMP-2和MMP-9染色阳性率分别为70%和85%,PVR增殖膜MMP-2和MMP-9染色阳性率分别为80%和60%.在PDR增殖膜上可见血管周围基质MMP-9染色阳性,而且与PVR增殖膜相比,PDR增殖膜MMP-9表达有显著提高.结论:PDR增殖膜MMP-2和MMP-9均有表达,在PDR新生血管形成和纤维化的病理过程中起重要作用.     

Pathomorphology of membranes appearing in proliferative vitreoretinopathies

Immunohistochemical techniques were used to investigate the cell content of epiretinal membranes occuring in proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR). Ten epiretinal membranes were obtained during surgery from eyes with PVR and five from eyes with PDR. This material was studied histopathologically and immunohistochemically. Retinal pigment epithelial cells, glial cells, macrophages, T lymphocytes and type IV collagen were identified in these membranes. The findings indicate that the cells mentioned possess a potential role in creating vitreoretinal membranes in PVR and PDR.     

Quantitative Study of Basic Fibroblast Growth Factor in Vitreous with Proliferative Vitreoretinopathy

Objective: To quantitatively study basic fibroblast growth factor (bFGF) in the vitreous of proliferative vitreoretinopathy (PVR) in orderto understand the role of bFGF in the development of PVR.Method: High sensitive sandwich enzyme immunoassay technique(ELISA) was used to measure bFGF level in vitreous of normal eyes, the eyes of PVR-C or PVR-D grade, eyes of vitreous hemorrhage and the serum levels of bFGF in PVR-D patients. Results: The levels of bFGF in the vitreous were: median 5. 20ng/L, quartile 15. 47 ng/L in 20 normal eyes; median 3. 12ng/L, quartile 10. 48 ng/L in 35 PVR-C eyes; median 46. 56 ng/L, quartile 113. 96 ng/L in 26 PVR-D eyes; median 1. 40 ng/L, quartile 6. 25 ng/L in 25 vitreous hemorrhage eyes. The vitreous bFGF level in PVR-D group was significantly higher than that in the normal group, PVR-C group and vitreous hemorrhage group ( P < 0. 01). The mean of serum-bFGF level was 18. 33 ± 3. 39 ng/ L. The vitreous bFGF level of PVR-D group was significantly higher than serum-bFGF lev     

血管内皮生长因子在增生性玻璃体视网膜病变中的作用

增生性玻璃体视网膜病变（proliferative vitreoretinopathy，PVR）发病机制尚未完全明确。大量研究证实血管内皮生长因子（vascular endothelial growth factor，VEGF）在 PVR 视网膜前增生膜组织及玻璃体中表达增高；在动物模型中，抗 VEGF 治疗可有效防止 PVR 的发生发展。VEGF 促进 PVR发展的机制可能在于 VEGF 竞争性地抑制血小板源性生长因子（platelet-derived growth factor，PDGF）结合血小板源性生长因子受体α（platelet-derived growth factor receptor α，PDGFR-α），从而促进非血小板源性生长因子（non-PDGFs）活化 PDGFR-α，激活 PI3K／Akt 信号通路，减少 p53基因的表达。（国际眼科纵览，2016，40：192-196）     

Matrix metalloproteinases and their natural inhibitors in fibrovascular membranes of proliferative diabetic retinopathy

AIM: To examine epiretinal membranes of proliferative diabetic retinopathy (PDR) for the presence of selective matrix metalloproteinases (MMPs) and their natural inhibitors (TIMPs), in order to determine whether neovascularisation and fibrosis, characteristic of this complication of diabetes mellitus, are associated with specific anomalies of MMP or TIMP expression. METHODS: The presence of selected MMPs and TIMPs was investigated in 24 fibrovascular epiretinal membranes of PDR, and the findings compared with that observed in 21 avascular epiretinal membranes of proliferative vitreoretinopathy (PVR) and five normal retinas. Specimens were examined for deposition of interstitial collagenase (MMP-1), stromelysin-1 (MMP-3), gelatinase A (MMP-2), gelatinase B (MMP-9), and three tissue inhibitors of metalloproteinases (TIMP-1, TIMP-2, and TIMP-3). RESULTS: The results showed that unlike normal retina, which constitutively expresses MMP-1 and TIMP-2, a large proportion of PDR membranes (> 62%) stained for MMP-1, MMP-2, MMP-3, MMP-9, TIMP-1, TIMP-2, and TIMP-3. There were no differences in the expression of these molecules when compared with PVR membranes. A characteristic staining for MMP-9 was observed within the perivascular matrix of PDR membranes, and there was a significant increase in TIMP-2 expression by PDR membranes (p= 0.036) when compared with PVR membranes. CONCLUSIONS: The findings that MMPs involved in degradation of fibrovascular tissue matrix, as well as TIMP-1 and TIMP-2, are found in a large proportion of PDR membranes, and that their expression does not differ from that of PVR membranes, suggest the existence of common pathways of extracellular matrix degradation in pathological processes leading to retinal neovascularisation and fibrosis.     

Expression of angiogenic and fibrogenic factors in proliferative vitreoretinal disorders

Purpose To investigate the expression of connective tissue growth factor (CTGF) in the retina of human subjects with diabetes mellitus, and CTGF, CD105, and gelatinase B in proliferative diabetic retinopathy (PDR) and proliferative vitreoretinopathy (PVR) epiretinal membranes. Methods Twelve donor eyes from six subjects with diabetes mellitus, 10 eyes from five nondiabetic subjects, 14 PDR membranes, and five PVR membranes were studied by immunohistochemical techniques. In situ zymography was used to examine gelatinolytic activity in four PDR membranes. Results In nondiabetic retinas, there was no immunoreactivity for CTGF. Diabetic retinas showed immunoreactivity for CTGF in ganglion cells and microglia. Vascular endothelial cells in PDR membranes expressed CTGF, CD105, and gelatinase B in 10 (71.4%), 11 (78.6%), and 5 (35.7%) membranes, respectively. Myofibroblasts in PDR membranes expressed CTGF, and gelatinase B in 14 (100%), and 6 (42.9%) membranes, respectively. There was a significant correlation between the number of blood vessels expressing the panendothelial marker CD34 and the number of blood vessels expressing CTGF (r = 0.7884; P = 0.0008), and CD105 (r = 0.6901; P = 0.0063), and the number of myofibroblasts expressing CTGF (r = 0.5922; P = 0.0257). There was a significant correlation between the number of myofibroblasts expressing α-smooth muscle actin and the number of myofibroblaasts expressing CTGF (r = 0.8393; P = 0.0002). In situ zymography showed the presence of gelatinolytic activity in vascular endothelial cells in PDR membranes. Myofibroblasts in PVR membranes expressed CTGF, and gelatinase B. Conclusions These results suggest a possible role of CTGF, CD105, and gelatinase B in the pathogenesis of proliferative vitreoretinal disorders.     

Phosphorylation of alphaB-crystallin in epiretinal membrane of human proliferative diabetic retinopathy

AIM: To examine phosphorylation of alphaB-crystallin (p-?BC), a vascular endothelial growth factor (VEGF) chaperone, and immunohistochemically investigate relationship between p-?BC, VEGF and phosphorylated p38-mitogen-activated protein kinase (p-p38 MAPK) in the epiretinal membrane of human proliferative diabetic retinopathy (PDR). METHODS: Eleven epiretinal membranes of PDR surgically excised were included in this study. Two normal retinas were also collected from enucleation tissues due to choroidal melanoma. Paraformaldehyde-fixed, paraffin-embedded tissue sections were processed for immunohistochemistry with anti-p-?BC, VEGF, CD31, and p-p38 MAPK antibodies. RESULTS: Immunoreactivity for p-?BC was observed in all of the epiretinal membranes examined, where phosphorylation on serine (Ser) 59 showed strongest immunoreactivity in over 70% of the membranes. The immunolocalization of p-?BC was detected in the CD31-positive endothelial cells, and co-localized with VEGF and p-p38 MAPK in PDR membranes. Immunoreactivity for p-?BC, however, was undetectable in endothelial cells of the normal retinas, where p-p38 MAPK immunoreactivity was less marked than PDR membranes. CONCLUSION: Phosphorylation of ?BC, in particular, phosphorylation on Ser59 by p-p38 MAPK may play a potential role as a molecular chaperon for VEGF in the pathogenesis of epiretinal membranes in PDR.     

Proliferative activity in epiretinal membranes. The use of the monoclonal antibody Ki-67 in proliferative vitreoretinal diseases.

Epiretinal membranes occur in a number of pathogenetically different diseases, such as proliferative vitreoretinopathy (PVR), proliferative diabetic retinopathy (PDR), macular pucker, and ischemic, inflammatory, and degenerative vitreoretinal disorders. The formation of epiretinal membranes is characterized by the proliferation of histogenetically different cell types. Knowledge of the proliferating potential of surgically excised epiretinal membrane tissue might be clinically relevant to determine which patients face a high risk of recurrences. The monoclonal antibody Ki-67 specifically stains a cell-cycle associated nuclear antigen that is only expressed by cells in the G1, S, and G2/M phases of the cell cycle. With this antibody, the actively proliferating growth fraction can be stained in frozen tissue samples of surgically removed epiretinal membranes by the indirect immunoperoxidase method. In this study, the monoclonal antibody Ki-67 was used to screen 37 epiretinal membranes in PVR, PDR, macular pucker, and in recurrences after intraocular silicone oil tamponade for the presence of actively proliferating cells. Additionally, the number of Ki-67 positive cells in a section of the membrane was quantitatively set in relation to the total cell number of this section. Thus a proliferative index (PI) was ascribed to each membrane as a decimal quotient. The proliferative indices can be graded into four subgroups for missing or very low (PI less than 0.1), low (PI 0.1-0.3), moderate (PI 0.3-0.6), and high (PI greater than 0.6) proliferative activities. High proliferative activities were found in 4 of 5 PVR membranes, in 9 of 14 PDR membranes, in 6 of 11 recurrent membranes after intraocular silicone oil tamponade, and in 2 of 6 macular pucker membranes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Expression of HLA-DR antigen in epiretinal and vitreous membranes

The pathogenic mechanisms of epiretinal membranes are not clearly understood nowadays in ophthalmology. Trying to elucidate it from another aspect, we examined the presence of HLA-DR antigen in 41 epiretinal membrane specimens from patients with proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR) by using immunohistochemical technique (APAAP). The results showed that 82.93% of the membranes (34 out of 41) were HLA-DR antigen positive. HLA-DR antigen was considered to be expressed by macrophages in epiretinal membranes. The findings here reveal that the formation and development of epiretinal membranes are probably correlated with immune responses.     

血清中VEGF和bFGF水平与糖尿病视网膜病变关系的临床研究

目的:探讨糖尿病视网膜病变患者血清中血管内皮细胞生长因子(VEGF)、碱性成纤维细胞生长因子(bFGF)水平与糖尿病视网膜病变的关系。方法:收集患有糖尿病视网膜病变白内障患者的血清样本,并以无糖尿病白内障患者的血清样本作为对照,用ELISA方法测定血清中bFGF,VEGF的含量。结果:对照组12例血清样本bFGF含量为(34.95±9.62)ng/L,VEGF含量为(150±20)μg/L;单纯型DR组中10例bFGF含量为(52.6±7.54)ng/L,VEGF含量为(330±50)μg/L;增生型DR组7例bFGF含量为(64.929±11.917)ng/L,VEGF含量为(580±50)μg/L。和对照组相比糖尿病患者血清中bFGF,VEGF的含量均显著增加(P<0.01),并且随着DR病情的进展而增高(P<0.05);血清中VEGF与bFGF之间呈正相关(r=0.419,P<0.01)。结论:VEGF,bFGF参与了DR的发生发展,并与DR后期新生血管形成关系密切。     

Detection of eicosanoids in epiretinal membranes of patients suffering from proliferative vitreoretinal diseases

AIM—Arachidonic acid is metabolised via lipoxygenase to 15-HETE (15-hydroxyeicosatetraenoic acid) and 15-HPETE (15-hydroperoxyeicosatetraenoic acid), which are believed to influence proliferation in tissue culture. 15-HETE is the reduction product of 15-HPETE. Cell proliferation is believed to be decreased by 15-HPETE and increased by 15-HETE. The aim of this study was to investigate epiretinal membranes for the presence of these lipoxygenase products and to compare membranes from different disease processes.
METHODS—Epiretinal membranes of 15 patients suffering from proliferative vitreoretinopathy (PVR, n=7) and proliferative diabetic retinopathy (PDR; n=8) were removed during vitrectomy and analysed by means of thin layer chromatography. The plates were evaluated by digital image analysis.
RESULTS—Both 15-HETE and 15-HPETE were identified in membranes from eyes of patients with PVR and PDR with HETE values significantly higher (p<0.05) than HPETE values (HETE/HPETE ratio = 5.2).
CONCLUSION—This study demonstrates that eicosanoids are present in the epiretinal membrane tissue of patients with PVR and PDR. Considering that HETE increases cell proliferation while HPETE inhibits it, it is conceivable that eicosanoids are an additional factor contributing to the regulation of membrane growth in proliferative retinal disorders. Thus, inhibition of lipoxygenase could be a therapeutic approach in these diseases.

     

糖尿病患者血浆碱性成纤维细胞生长因子的变化

目的:讨论糖尿病患者血浆碱性成纤维细胞生长因子(basic fibroblast growth factor,bFGF)变化的临床意义,研究bFGF在糖尿病视网膜病变(diabetic retinopathy,DR)发病机制中的作用。方法:收集糖尿病不伴有眼底改变与患有糖尿病视网膜病变白内障患者的血浆样本,并以无糖尿病白内障患者的血浆样本作为对照,用ELISA方法测定了血浆中bFGF的含量。结果:对照组12例血浆样本bFGF含量为(38.9±9.6)ng/L。糖尿病患者无眼底改变组中,15例bFGF含量为(49.12±6.01)ng/L;单纯性DR组中10例bFGF含量为(50.6±9.5)ng/L;增生性DR组7例bFGF含量为(61.9±12.9)ng/L。和对照相比糖尿病患者血浆中bFGF的含量均显著增加(P<0.01),并且随着DR病情的进展而增高(P<0.05)。结论:bFGF积极参与了DR的发生发展,并与DR后期新生血管形成关系密切。     

碱性成纤维细胞生长因子对牛眼晶体上皮细胞的促增殖作用

目的观察碱性成纤维细胞生长因子（ｂａｓｉｃｆｉｂｒｏｂｌａｓｔｇｒｏｗｔｈｆａｃｔｏｒ，ｂＦＧＦ）对牛眼晶体上皮细胞的作用。方法首先进行牛眼晶体上皮细胞原代与传代培养；在第２代培养细胞中添加ｂＦＧＦ，浓度１０－２～１０２ｎｇ／ｍｌ，借助甲基噻唑基四唑（ｍｅｔｈｙｌｔｈｉａｚｏｌｙｌｔｅｔｒａｚｏｌｉｕｍ，ＭＴＴ）测定方法观察ｂＦＧＦ对晶体上皮细胞增殖的影响。结果牛眼晶体上皮细胞体外具有易于贴附、细胞融合较快以及生长迅速等特点。添加ｂＦＧＦ后，可促进晶体上皮细胞的增殖，尤其是１０２ｎｇ／ｍｌ显示出明显的促增殖作用。结论ｂＦＧＦ在促进白内障术后晶体上皮细胞的增殖及后囊混浊上起了重要作用。     

Development of normal and injury-induced gene expression of aFGF, bFGF, CNTF, BDNF, GFAP and IGF-I in the rat retina

Focal mechanical injury to the retina substantially increases basic fibroblast growth factor (bFGF) and ciliary neurotrophic factor (CNTF) mRNA expression, accompanied by a transient increase in FGFR-1 mRNA, and this response is thought to protect photoreceptors near the injury site from inherited and light-induced retinal degenerations. We have now examined retinal gene expression of the principal survival factors involved in the response to injury in normal rats as a function of postnatal age both in normal and injured retinas. Sprague-Dawley rats were injured in one eye by needle incision through the retina at postnatal day (P) 10, 22, 35, 60, 90, 120 and 180. The other eye was uninjured and served as the control. Retinas were taken 1 day post-injury. Northern blot analysis was performed to determine the mRNA expression of the following factors and receptors: bFGF and acidic fibroblast growth factors (aFGF) and FGF receptor-1 (FGFR-1); CNTF and CNTF receptor alpha (CNTFR-alpha); brain-derived neurotrophic factor (BDNF) and its receptor trk B; and insulin-like growth factor-I (IGF-I) and IGF-I receptor (IGF-IR); glial fibrillary acidic protein (GFAP) and opsin. In the uninjured control eyes, mRNA expression of most of the factors increased with postnatal age, with little expression at P10 and maximal expression levels reached at P22 (opsin), P35 (aFGF), P60 (BDNF) or P90 (bFGF, FGFR-1, CNTF and GFAP). In contrast, IGF-1 mRNA rapidly decreased from a high level of expression at P10 to about 55% of that level by P22, reaching a stable 45-50% of the P10 level at P35 and thereafter. The response to injury of bFGF, FGFR-1, CNTF and GFAP mRNAs increased with postnatal age. Unexpectedly, only minimal increases in bFGF, FGFR-1, CNTF and GFAP over those seen in the control eyes were observed before P35. Thereafter, the increase of bFGF mRNA after injury reached a maximum of three-fold at P60, maintained this level to P120, and slightly decreased to 2.5-fold by P180. Expression of FGFR-1 mRNA showed a maximum increase of 2.6-fold at P90. Expression of CNTF and GFAP mRNAs followed a time course similar to that of bFGF. Mechanical injury did not alter the mRNA levels of aFGF, BDNF, IGF-I, and receptors, CNTFR-alpha, trk B and IGF-IR. These data show that the response to injury is minimal at early postnatal ages but increases with age and peaks at P60-90 for most potential survival factors.     

三种视网膜病视网膜前膜中基质金属蛋白酶及其天然抑制分子的表达

目的 研究增殖性糖尿病视网膜病变(proliferative diabetic retinopathy,PDR)、增殖性玻璃体视网膜疾病(proliferative vitreoretinopathy,PVR)和急性视网膜坏死(acute retinalnecrosis,ARN)患者视网膜前膜中基质金属蛋白酶(matrixmetalloproteinases:MMPs)及其天然抑制物(tissueinhibitorsofmetalloproteinages,TIMPs)的表达情况.方法 玻璃体手术中剥取PVR、PDR和ARN患者的视网膜前膜,同供体眼视网膜作为正常对照,冰冻切片后进行免疫组织化学染色,包括:MMP-1,MMP-2,MMP-3,MMP-7,MMP-9,TIMP-1和TIMP-2.结果 正常视网膜中能够观察到MMP-1,MMP-3,TIMP-1和TIMP-2的表达,在PVR、PDR和ARN患者标本中各种分子的表达都增强,尤以MMP-2,MMP-3和MMP-7明显.结论 正常视网膜中存在MMPs和TIMPs分子维持着细胞外基质动态的平衡,在PVR,PDR和ARN患者中MMP-2,MMP-3和MMP-7等MMPs分子表达增强,在其病变过程中可能起重要作用.