Abstract:
:Limbal microvascular endothelial cells (L-MVEC) contribute to formation of the corneal-limbal stem cell niche and to neovascularization of diseased and injuries corneas. Nevertheless, despite these important roles in corneal health and disease, few attempts have been made to isolate L-MVEC with the view to studying their biology in vitro. We therefore explored the feasibility of generating primary cultures of L-MVEC from cadaveric human tissue. We commenced our study by evaluating growth conditions (MesenCult-XF system) that have been previously found to be associated with expression of the endothelial cell surface marker thrombomodulin/CD141, in crude cultures established from collagenase-digests of limbal stroma. The potential presence of L-MVEC in these cultures was examined by flow cytometry using a more specific marker for vascular endothelial cells, CD31/PECAM-1. These studies demonstrated that the presence of CD141 in crude cultures established using the MesenCult-XF system is unrelated to L-MVEC. Thus we subsequently explored the use of magnetic assisted cell sorting (MACS) for CD31 as a tool for generating cultures of L-MVEC, in conjunction with more traditional endothelial cell growth conditions. These conditions consisted of gelatin-coated tissue culture plastic and MCDB-131 medium supplemented with foetal bovine serum (10% v/v), D-glucose (10 mg/mL), epidermal growth factor (10 ng/mL), heparin (50 μg/mL), hydrocortisone (1 μg/mL) and basic fibroblast growth factor (10 ng/mL). Our studies revealed that use of endothelial growth conditions are insufficient to generate significant numbers of L-MVEC in primary cultures established from cadaveric corneal stroma. Nevertheless, through use of positive-MACS selection for CD31 we were able to routinely observe L-MVEC in cultures derived from collagenase-digests of limbal stroma. The presence of L-MVEC in these cultures was confirmed by immunostaining for von Willebrand factor (vWF) and by ingestion of acetylated low-density lipoprotein. Moreover, the vWF(+) cells formed aligned cell-to-cell 'trains' when grown on Geltrex™. The purity of L-MVEC cultures was found to be unrelated to tissue donor age (32-80 years) or duration in eye bank corneal preservation medium prior to use (3-10 days in Optisol) (using multiple regression test). Optimal purity of L-MVEC cultures was achieved through use of two rounds of positive-MACS selection for CD31 (mean ± s.e.m, 65.0 ± 20.8%; p < 0.05). We propose that human L-MVEC cultures generated through these techniques, in conjunction with other cell types, will provide a useful tool for exploring the mechanisms of blood vessel cell growth in vitro.
journal_name
Exp Eye Resjournal_title
Experimental eye researchauthors
Gillies PJ,Bray LJ,Richardson NA,Chirila TV,Harkin DGdoi
10.1016/j.exer.2014.12.008subject
Has Abstractpub_date
2015-02-01 00:00:00pages
20-8eissn
0014-4835issn
1096-0007pii
S0014-4835(14)00325-Xjournal_volume
131pub_type
杂志文章abstract::Crystalline lenses excised from 5-7-month-old rabbits and pigs were extracted for lipids with chloroform/methanol, 2/1 using the Folch method. The extracted crude lipids were analyzed at 202 MHz by 31P-NMR spectroscopy. Twelve membrane phospholipids were detected. Both rabbit and pig phospholipid profiles contained ph...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(90)90207-b
更新日期:1990-03-01 00:00:00
abstract::Human retinal pigment epithelial cells in culture demonstrate adenylate cyclase activity. It is membrane-bound and modulated by GTP regulatory proteins. It is effectively activated only by beta-adrenergic agonists (L-isoproterenol greater than or equal to L-epinephrine greater than L-norepinephrine) and some prostagla...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/s0014-4835(87)80158-6
更新日期:1987-04-01 00:00:00
abstract::Research over the past two decades has shown that the growth of young eyes is guided by vision. If near- or far-sightedness is artificially imposed by spectacle lenses, eyes of primates and chicks compensate by changing their rate of elongation, thereby growing back to the pre-lens optical condition. Little is known a...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1006/exer.1999.0813
更新日期:2000-04-01 00:00:00
abstract::Pericytes exert an important influence on the control of retinal blood flow; however, little is known regarding the molecular basis of retinal pericyte excitability. The purpose of this study was to elucidate the signaling pathway of how prostacyclin (PGI2), an important endogenous regulator of retinal blood flow, sti...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2006.07.011
更新日期:2006-12-01 00:00:00
abstract::Retinal degenerations, regardless of the initiating event or gene defect, often result in a loss of photoreceptors. This formal deafferentation of the neural retina eliminates the intrinsic glutamatergic drive of the sensory retina and, perhaps more importantly, removes coordinated Ca++-coupled signaling to the neural...
journal_title:Experimental eye research
pub_type: 杂志文章,评审
doi:10.1016/j.exer.2005.03.006
更新日期:2005-08-01 00:00:00
abstract::The polarity of retinal pigmented epithelia (RPE) from chicken embryos was studied in primary cell culture. Since cultured RPE approximates the morphological polarity of RPE in vivo, we investigated whether this polarity extends to the distribution of plasma membrane proteins that are peculiar to RPE. In contrast to o...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(90)90156-o
更新日期:1990-10-01 00:00:00
abstract::The photopigment of avian pineal which mediates light sensitivity was sought via its chromophore. Chick pineal cells in primary cultures were incubated overnight in the dark with [3H]retinol. Reduction of Schiff's bases with cyanoborohydride prior to SDS-PAGE revealed radioactivity bound to a 30 kDa component in pinea...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/s0014-4835(88)80042-3
更新日期:1988-06-01 00:00:00
abstract::Transport of circulating reduced glutathione (GSH) was studied at the basolateral side of the lens epithelium by using an in situ vascular eye perfusion technique in guinea-pigs with rapid sampling to ensure detecting initial uptakes. The unidirectional transport rates of [35S]-GSH (4 nM) from plasma and aqueous into ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1006/exer.1996.0004
更新日期:1996-01-01 00:00:00
abstract::The aim of the study was to investigate the molecular genetics of uveal melanoma (UM) metastases and correlate it with disease progression. Twelve pathologically confirmed UM metastases from 11 patients were included. Molecular genetic alterations in chromosomes 3 (including the BAP1 region), 8q, 6p, and 1p were inves...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2012.04.010
更新日期:2012-07-01 00:00:00
abstract::A method for the isolation and purification of human ocular mucin from the brief saline extract of human ocular mucus is reported. Initial purification of ocular mucin was achieved by sequential chromatography of the saline-soluble mucus extract from an individual donor's mucus pool on columns of Sephadex G-50 and Sep...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(88)90002-4
更新日期:1988-08-01 00:00:00
abstract::Choroidal neovascularization (CNV) is one of the severe pathological consequences of the end-stage of age-related macular degeneration (AMD). Several lines of evidence implicate increased levels of vascular endothelial growth factor (VEGF) in the retinas of AMD patients. Current available agents for the inhibition of ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2010.06.019
更新日期:2010-09-01 00:00:00
abstract::The routine cultivation of human corneal endothelial cells, with the view to treating patients with endothelial dysfunction, remains a challenging task. While progress in this field has been buoyed by the proposed existence of progenitor cells for the corneal endothelium at the corneal limbus, strategies for exploitin...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2014.07.002
更新日期:2014-10-01 00:00:00
abstract::(O-acyl) ω-hydroxy fatty acids (OAHFAs) are a recently found group of polar lipids in meibum. Since these lipids can potentially serve as a surfactant in the tear film lipid layer, the surface properties of a molecule of this lipid class was investigated and compared with a structurally related wax ester and a fatty a...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2013.06.016
更新日期:2013-10-01 00:00:00
abstract::Rat lenses incubated in tissue culture medium (M 199) maintain their transparency for a long period of time. The soluble corticosteroid, solumedrol (methyl prednisolone sodium succinate) was added to the medium, at concentrations including the range expected during rejection episodes following organ transplantation (3...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(83)90150-1
更新日期:1983-07-01 00:00:00
abstract::The binding of eight fluorescence-labeled lectins to the photoreceptors of the monkey retina was investigated using a post-embedding staining method. Concanavalin A (specific for mannosyl and glucosyl residues) bound to the outer and inner segments of both rods and cones, while the degree of staining was more intense ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(83)90094-5
更新日期:1983-01-01 00:00:00
abstract::Electrical stimulation of the preganglionic cervical sympathetic nerve produced ocular vasoconstrictor responses as measured from the anterior choroid of anesthetized cats using laser-Doppler flowmetry. Ipsilateral nictitating membrane contractions were simultaneously measured as an established index of neural sympath...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1006/exer.1993.1146
更新日期:1993-10-01 00:00:00
abstract::This study was performed to determine whether cells in the posterior stroma undergo apoptosis in response to endothelial cell injury and to determine whether basement membrane component nidogen-1 was present in the cornea. New Zealand White rabbits had an olive tip cannula inserted into the anterior chamber to mechani...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2018.03.025
更新日期:2018-07-01 00:00:00
abstract::Transforming growth factor β (TGFβ) signaling is one of the most important signaling pathways regulating cell behavior in ocular tissues. Its functions are mainly linked to tissue fibrosis and inflammatory responses in ophthalmology. In epithelial cells, however, the growth inhibitory activity of TGFβ was reported in ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2019.03.014
更新日期:2019-05-01 00:00:00
abstract::There is increasing evidence that the macular pigment carotenoids, lutein and zeaxanthin, may play an important role in the prevention of age-related macular degeneration, cataract, and other blinding disorders. Although it is well known that the retina and lens are enriched in these carotenoids, relatively little is ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1006/exer.2000.0954
更新日期:2001-03-01 00:00:00
abstract::We designed an experiment to study the role of light history and polyunsaturated fatty acids (PUFA) on the susceptibility of the albino rat retina to light damage. Albino rats were born to dams that had been kept in either 1 lx or 250 lx cyclic light for 0-4 days prior to delivery and fed one of three diets containing...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/s0014-4835(05)80112-5
更新日期:1995-03-01 00:00:00
abstract::Herpes simplex virus type-1 (HSV-1) infection leads to impaired corneal sensation and, in severe cases, to corneal ulceration, melting and perforation. Here, we explore the potential therapeutic action of pigment epithelial-derived factor (PEDF) plus docosahexaenoic acid (DHA) on corneal inflammation and nerve regener...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2017.06.015
更新日期:2017-08-01 00:00:00
abstract::Contact inhibition is an important mechanism for maintaining corneal endothelium in a non-replicative state. Protein tyrosine phosphatases (PTPs) play a role in regulating the integrity of cell-cell contacts, differentiation, and growth. In this study, we aimed to evaluate whether phosphatases are involved in the main...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2005.03.015
更新日期:2005-11-01 00:00:00
abstract::The objective of this study were to establish a cell line derived from porcine hyalocytes and to investigate the regulation of hyaluronan (HA) synthesis in response to cytokines. After 50 passages of the cells derived from porcine vitreous tissue, a cell line was generated. The immortalized cells showed fibroblastic m...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2007.07.006
更新日期:2007-10-01 00:00:00
abstract:PURPOSE:The feasibility of corneal reconstruction with cultured adult human corneal endothelial cells (HCEC) was examined in a nude rat model. METHODS:Endothelial cells were removed from the corneas of Lewis rats using a sterile cotton swab. Cultured adult HCEC labelled with a fluorescent marker chloromethyl-benzamido...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2004.05.001
更新日期:2004-08-01 00:00:00
abstract::Photoreceptor outer segments (OS) in the vertebrate retina undergo a process of continual renewal involving shedding of disc membranes that are cleared by phagocytic uptake into the retinal pigment epithelium (RPE). In dystrophic Royal College of Surgeons (RCS) rats, OS phagocytosis is blocked by a mutation in the gen...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2015.08.006
更新日期:2015-11-01 00:00:00
abstract::Simvastatin is a cholesterol-lowering drug which exhibits numerous pleiotropic effects including anti-cancer activity. Yet, the anti-cancer effects in choroidal melanoma remain poorly characterized. Therefore, in this study, we investigated the effects of simvastatin on OCM-1 cells growth, apoptosis and cycle. Simvast...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2013.05.013
更新日期:2013-08-01 00:00:00
abstract::We have investigated: (a) Phospholipid composition and phosphoinositide-PO4 turnover in rabbit cornea tissues; and (b) the effects of adrenergic and serotonergic agonists on breakdown of phosphoinositides in the rabbit cornea. The data obtained from these studies can be summarized as follows: (1) in the cornea phospha...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/s0014-4835(87)80047-7
更新日期:1987-06-01 00:00:00
abstract::Lack of an optimal in vitro model of human corneal epithelial (HCE) cells is a major limitation in studying normal functions and gene regulations in HCE. Moreover, availability of a multi-layered HCE culture can reduce the usage of animals in the toxicity testing of consumer products. We have developed tetracycline-re...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/s0014-4835(03)00175-1
更新日期:2003-10-01 00:00:00
abstract::The levels of alpha 1-proteinase inhibitor (alpha 1-antitrypsin) in keratoconus, normal human, and other diseased corneas were examined. Using an immunoperoxidase technique, the presence of this inhibitor was demonstrated in the epithelium, stroma and endothelium of all corneal sections. Compared with normal human con...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/0014-4835(90)90044-u
更新日期:1990-05-01 00:00:00
abstract::RPE65-related Leber's congenital amaurosis (LCA) is a rod-cone dystrophy whose clinical outcome is mainly attributed to the loss of rod photoreceptors followed by cone degeneration. Pathogenesis in Rpe65(-/-) mice is characterized by a slow and progressive degeneration of rods dependent on the constitutive activation ...
journal_title:Experimental eye research
pub_type: 杂志文章
doi:10.1016/j.exer.2011.12.019
更新日期:2012-03-01 00:00:00