Basic fibroblast growth factor mRNA, bFGF peptide and FGF receptor in epiretinal membranes of intraocular proliferative disorders (PVR and PDR)

Basic fibroblast growth factor (bFGF) has been shown to be involved in epiretinal membrane formation in proliferative vitreoretinal disorders. However, up to now, little knowledge exists, as to the actual cellular source of this potent mitogen.We examined 20 epiretinal membranes from patients with proliferative diabetic retinopathy (PDR) (n = 12) and proliferative vitreoretinopathy (PVR) (n = 8) for the presence of bFGF peptide, fibroblast growth factor receptor-1 (FGFR-1) and bFGF messenger ribonucleic acid (mRNA).Using a specific antibody, we detected bFGF peptide in most (8/10) examined PDR membranes and in all (8/8) PVR membranes. Moreover, we found positive staining for the corresponding receptor.Local production of bFGF in epiretinal membranes was confirmed by nonisotopic in situ hybridisation for bFGF mRNA in some (4/7) examined PDR membranes and some (3/4) examined PVR membranes. All membranes which contained bFGF mRNA were also positive for bFGF peptide.In conclusion, bFGF is produced and stored in epiretinal membranes. Together with the corresponding receptor, bFGF may play a role in the auto- and paracrine control of the proliferative processes at the vitroretinal interface.Abbreviations aFGF acidic fibroblast growth factor - bFGF basic fibroblast growth factor - FGFR-1 fibroblast growth factor receptor-1 - mRNA messenger ribonucleic acid     

Recombinant α2(IV)NC1 domain of type IV collagen is an effective regulator of retinal capillary endothelial cell proliferation and inhibits pre-retinal neovascularisation

Background A recombinant form of the α2(IV)NC1 domain of type IV collagen has been shown to have potent anti-angiogenic activity although this peptide has not been studied in the context of proliferative retinopathies. In the current investigation we examined the potential for α2(IV)NC1 to regulate retinal microvascular endothelial cell function using a range of in vitro and in vivo assay systems. Materials and methods α2(IV)NC1 at concentrations between 0.1 and 1 μg/ml was added to retinal microvascular endothelial cells (RMECs) followed by assessment of cell attachment, proliferation and survival. This agent was also tested within a novel in vitro three-dimensional retinal angiogenesis assay and the number of angiogenic sprouts quantified. α2(IV)NC1 was also delivered intra-vitreally to mice with oxygen-induced proliferative retinopathy (OIR) and neovascularisation evaluated in comparison with vehicle-treated controls. Results RMECs treated with α2(IV)NC1 (0.1, 0.5 and 1 μg/ml) showed delayed attachment at 3 h post-seeding, although this deficit had been restored at the 6-h time point. BrdU assay of DNA replication revealed that confluent RMECs treated with α2(IV)NC1 showed no measurable response in comparison with vehicle-treated controls. By contrast, proliferation of sub-confluent RMECs was significantly reduced by α2(IV)NC1 at 0.5 μg/ml (P<0.01). α2(IV)NC1 also induced apoptosis in RMECs and inhibited angiogenesis of pre-existing retinal vascular networks in vitro (P<0.001). Intra-vitreal injection of α2(IV)NC1 in the OIR model significantly inhibited pre-retinal neovascularisation compared with vehicle-treated controls (P<0.001). Conclusion α2(IV)NC1 inhibits angiogenesis in the retinal microvasculature. This recombinant protein has potential for the treatment of neovascularisation in proliferative retinopathies. BioStratum Inc. did not sponsor this research in any way. None of the authors are paid consultants with this company.     

地塞米松缓释微粒在增生性玻璃体视网膜病变的应用

目的：观察含地塞米松（DEX）的缓释微粒治疗增生性玻璃体视网膜病变（PVR）的临床疗效。方法：9例（9只眼）视网膜脱离伴PVR患者进行常规玻璃体视网膜手术，并在玻璃体腔内植入1粒DEX微粒（含地塞米松1mg），观察术后反应及视力、PVR发展、DEX的位置及形状变化等。结果：术后炎症反应轻，7只眼视网膜复位，3只眼后极部视网膜前有增生；2只眼视网膜局限性脱离，最终3只眼玻璃腔内硅油填充。随访视力有8只眼较术前提高（P=0.015）。DEX未见对视网膜有不良影响，仅在随着处有少量色素吸附，4-6月后吸收。结论：DEX抑制PVR术后炎症反应及较远期作用是安全、有效的；对手术未完全清除的视网膜前增生有一定的抑制作用。     

恶性和非恶性肿瘤患者正常组织细胞DNA含量参数的对比研究

目的 :探讨恶性和非恶性肿瘤患者正常组织细胞生物学特性的区别。　方法 :用流式细胞术对 2 37例恶性肿瘤患者和 148例非恶性肿瘤患者正常组织细胞的DNA含量进行了检测 ,并对二者的DNA指数 (DI)、DNA倍体类型、细胞程序性死亡水平 (Apo)和增殖活性 (SPF)做了对比分析。　结果 :2 37例恶性肿瘤患者正常组织的DI值为1.0 8± 0 .2 3,DNA异倍体检出率为 18.99% ;148例非恶性肿瘤患者正常组织的DI值为 1.0 0± 0 ,DNA倍体类型均为二倍体。恶性肿瘤患者正常组织的Apo和SPF均显著高于非恶性肿瘤患者正常组织 (P <0 .0 5 )。即使剔除DNA异倍体病例 ,其Apo和SPF仍显著高于非恶性肿瘤患者的正常组织 (P <0 .0 5和P <0 .0 1)。　结论 :恶性肿瘤患者癌旁远处组织DNA二倍体细胞的Apo和SPF显著高于非恶性肿瘤正常组织 ,不能真正代表正常组织细胞的生物学特性     

Proliferative activity of colonic mucosa at different distances from primary adenocarcinoma as determined by the presence of statin: A nonproliferation-specific nuclear protein

The field change is one hypothesis concerning the development of colorectal carcinoma. Removal of a carcinoma without its entire surrounding altered mucosa may result in the development of a recurrence. S44, a monoclonal antibody directed against statin, a nuclear protein expressed in nonproliferating cells in either a quiescent or senescent state, was used to determine the rate of cell growth in colorectal mucosa at different distances from carcinomas. The specimens of 18 patients undergoing resection of a colorectal carcinoma were immediately opened after operation, and strips of mucosa were taken at distances of 1 cm, 5 cm, and 10 cm from the carcinoma. For each location, 10 longitudinally oriented crypts were evaluated for statin-positive cells identified by the presence of a dark brown peroxidase-conjugated antibody reaction product. The average percentage of statin-positive cells per crypt was significantly lower at a 1-cm distance from the carcinoma compared with the mucosa located 5 and 10 cm from the carcinoma (20.89±4.33 at 1 cm, 32.41±5.27 at 5 cm, and 34.23±6.45 at 10 cm). None of the calculated parameters showed any significant difference between the 5-cm and 10-cm locations. The fact that the proliferation rate of the mucosal cells returns to the normal level at 5 cm from the margin of the carcinoma suggests that cells located within this distance still retain proliferative potential even though they are morphologically indistinguishable from their normal counterparts. We conclude that failure to remove this transitional, potentially proliferative mucosa may result in subsequent development of anastomotic or perianastomotic recurrences.This study was conducted with support from the Sir Mortimer B. Davis-Jewish General Hospital Foundation and the American Physician Fellowship and with grants to Eugenia Wang from the Medical Research Council of Canada and from the National Institute on Aging of the National Institutes of Health of the U.S.A.     

增生性玻璃体视网膜病变基质金属蛋白酶的定量研究

目的:研究增生性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)玻璃体中基质金属蛋白酶(matrix metalloproteinases,MMPs)的表达,探讨MMPs在PVR病理过程中的作用。方法:PVR患者采用标准三切口巩膜扁平部玻璃体切割术(pars plana vitrectomy,PPV),取未稀释的玻璃体21只眼,PPV术后复发的玻璃体腔液20只眼,意外死亡的正常人玻璃体10只眼,采用明胶酶谱分析法定量分析MMP-2和MMP-9活性水平。结果:PVR玻璃体有MMP-2活性水平增高,与正常玻璃体比较差异有显著性意义(P<0.05)。21眼PVR玻璃体中13只眼有MMP-9活性水平增高,平均(171.52±13.17)扫描单位。20眼PPV术后PVR复发的玻璃体腔液19只眼有MMP-9活性水平增高,平均(156.01±37.21)扫描单位。正常人玻璃体无MMP-9的表达。结论:PVR玻璃体有MMP-2和MMP-9活性水平增高,MMP-9活性水平增高可能与术后PVR复发有关。眼科学报2003;19:130-132。     

Numerical chromosomal aberrations in prostate cancer: correlation with morphology and cell kinetics

Eleven routinely processed radical prostatectomy specimens were studied for the presence of numerical chromosomal aberrations by means of in situ hybridization with nucleic acid probes specific for chromosomes 7, 10, 17, X, and Y. Cytogenetic information was correlated with morphology, tumour stage and volume as well as with cell kinetics, the latter being assessed by immunohistochemistry with antibodies raised against the proliferative cell nuclear antigen (PCNA) and against a formalin-resistant epitope of the Ki-67 antigen, MIB 1. In 5 of 11 cases, numerical aberrations of at least one chromosome were found. The cases with normal chromosome numbers were those with the smallest volumes of Gleason grade 4 and/or 5 tumour (mean 0.5 cm³) and represented tumours restricted to the prostate. Tumours with aberrations in the number of detected chromosomes showed advanced stages and large volumes of high-grade tumour (mean 12.5 cm³). All 4 tumours with positive surgical margins were recruited from a group with marked local heterogeneity in chromosome numbers. Immunostaining with MIB 1 and PCNA was most intense in areas of high-grade tumour and was positively correlated with the emergence of chromosomal aberrations. The data suggest that the appearance of numerical chromosomal aberrations in prostate cancer coincides with aggressive tumour behaviour and could be used as an additional prognostic marker.This work is part of E.K.'s doctoral thesis     

Identification of two distinct CD5- B cell subsets from human tonsils with different responses to CD40 monoclonal antibody

This study investigated the response of different CD5^? B cell subsets to CD40 monoclonal antibody (mAb) in various combinations with interleukin (IL)-4 or rabbit anti-human μ chain antibody (a-μ-Ab). The different CD5 B cell subsets were isolated from tonsillar B cell suspensions depleted of CD5⁺ B cells and subsequently fractionated on Percoll density gradients. While resting CD5⁺ B cells proliferated and produced IgM molecules in response to a-μ-Ab, IL-4 and CD40 mAb as well as to Staphylococcus aureus Cowan strain I (SAC) and IL-2, resting CD5^? B cells, which were co-purified in the same 60% Percoll fractions, consistently failed to respond. These cells were, however, activated by the stimuli employed, as demonstrated by their capacity to express the surface activation markers CD69, CD25 and CD71. Resting CD5⁺ B cells had the typical phenotype of mantle zone B cells (IgM⁺ IgD⁺ CD39+ CD38^? CD10^? CDw75^dim), whereas resting CD5^? B cells were CD38^? CD39^? CD 10^? CDw75 intermediate and expressed surface IgM but relatively little surface IgD and could not be classified as mantle zone or germinal center cells. The finding that purified germinal center cells (CD38⁺ CD10⁺ CD39^? CDw75^bright, IgG⁺) responded to CD40 mAb and IL-4 and also to SAC plus IL-2 further underlined the differences to resting CD5^? B cells. However, some of the data collected suggest possible relationships between CD5^? B cells and germinal center cells. The CD5^? B cells isolated from the 50 % Percoll fraction proliferated in response to a-μ-Ab, CD40 mAb and IL-4 as well as to SAC and IL-2. These cells had the same mantle zone B cell phenotype as the CD5⁺ B cells, but their capacity to respond to the stimuli in vitro was unrelated to a possible contamination with CD5⁺ B cells, as documented by the appropriate controls. Furthermore, upon exposure to SAC or phorbol esters, the large majority of CD5^? B cells from the 50 % Percoll fraction did not express surface CD5 and there was very little if any accumulation of CD5 mRNA. Finally, most of the cycling cells in the stimulated CD5^? B cells did not express CD5. The CD5^? B cells from the 50 % Percoll fraction were comprised of a consistent proportion of cells that expressed surface activation markers. The removal of these cells abrogated the capacity of the suspensions to respond to the stimuli in vitro, possibly suggesting that these cells additional activation signals in vivo which were essential to acquire the capacity to respond and that could not be reproduced in vitro. The present study underlines the phenotypic and functional heterogeneity of CD5^? B cells and contributes to the identification of two subsets of these cells which differ in phenotype, tissue distribution and in vitro responses to different stimuli.     

A novel isoform of the smooth muscle cell differentiation marker smoothelin

Studies on smooth muscle cell differentiation and those on vascular development in mouse and humans have long been hampered by the lack of suitable markers. Here we describe a novel, large isoform of smoothelin, a structural protein of differentiated, contractile smooth muscle cells. The protein, which is highly conserved in mouse and humans, shows homology with other cytoskeleton-associated smooth muscle cell proteins and contains an actinin-type actin-binding domain. Northern blot analysis from various mouse organs identified short and long smoothelin mRNA forms, which exhibit distinct tissue expression patterns. The short form is highly expressed in visceral muscle tissues such as intestine and stomach and is not detectable in brain, while the long mRNA form is expressed in all vascularized organs. These results may provide new tools and approaches to study both smooth muscle cell differentiation and proliferative vascular disease. Received: 25 August 1998 / Accepted: 19 October 1998     

前部增生性玻璃体视网膜病变引起慢性低眼压房水生成率的测定

目的：探讨前部增生性玻璃体视网膜病变（anterior prolieferative vitreoretinopathy,aPVR)低眼压状态下房水生成率变化,从房水动力学的角度揭示aPVR引起慢性低眼压的发病机制。方法：利用培养的同种兔皮肤成纤维细胞制作aPVR引起慢性低眼的动物模型。于术前及术后不同时间点分别观测眼压、术后14d,28d,56d分别以高效液相色谱仪测量房水荧光素清除率,进一步计算房水生成率。结果：术后2周、4周、8周实验组平均眼压、房水生成率明显低于对照组（P＜0．05或P＜0．01）。结论：在aPVR病理状态下,低眼压的形成与房水生成率下降有关。