摘要:

The capillary beds of the eye are lined by two types of endothelia, fenestrated in the choriocapillaris and ciliary body, and continuous in the retina and iris. In this study, we wished to find a marker for each of these types of vessel beds using lectin histochemistry. Sections of glutaraldehyde fixed rat eyes embedded in epoxy resin were extracted with sodium ethoxide and rehydrated. Binding of 15 different lectins was visualized using the avidin-biotin peroxidase technique. We found WGA, WGA-s, LFA and PHA-E to strongly bind retinal vessels. In addition to the above lectins, iris vessels bound GSL-I. Choriocapillaris reacted variably only with WGA and not at all with other lectins tested. Vessels of ciliary body processes did not react with any lectin studied. The less fenestrated vessels of the base of the ciliary process bound lectins similar to the retina. We speculate that the differential lectin staining of the various vessel beds of the eye may reflect the degree of fenestration of the endothelium. This reactivity may be influenced by variations in the surrounding milieu including cells and extracellular matrix.

ISSN:0271-3683    

DOI:10.3109/02713689109013875

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Membrane preparations of cultured human retinal pigment epithelial (RPE) cells were incubated with various concentrations of [3H]arginine vasopressin (AVP) in the presence and absence of 10 uM nonradioactive AVP. Saturable, specific binding to a single site with a Kdof 6.2 nM andBmax of 111 fmol/mg protein was detected. Vaaopressin had no effect on RPE cyclic AMP levels measured by radioimmunoassay. Intracellular calcium fluxes were measured by spectrofluorometry of RPE cell suspensions preloaded with quin 2. The baseline cytosolic calcium level was 217±20 nM, and AVP caused a concentration-dependent increase in this level with a 3.5-fold maximal response at 10-−6M and an EC50of 120 nM. The production of inositol phosphates was measured in RPE preloaded with [3H]myoinositol, and AVP caused a concentration-dependent increase in their production with a 2.1-fold maximal response at 10-−5M and an EC50of 80 nM. A specific vasopressin receptor antagonist, SKF 101926, prevented the AVP-induced increase in calcium mobilization and ionistol phosphate production in RPE. These data suggest that RPE cells possess V1AVP receptors coupled to calcium mobilization and inositol phosphate metabolism.

ISSN:0271-3683    

DOI:10.3109/02713689109013876

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

The effects of griseolic acid (GA), a cyclic-AMP phosphodiesterase (PDE) inhibitor, and its 8′-pivaloyloxymethyl (POM) ester on intraocular pressure (IOP) in rabbits were investigated. When 50μ1 of 1 and 2% GA POM ester solutions were topically applied to one eye in normal rabbits, significant IOP decreases were detected at 2 hrs and at 1 to 5 hrs, respectively. Other than ocular hypotension, no other ocular effects were detected locally even after administration of 2% GA POM ester. A more marked reduction in IOP occurred after the intravitreal injection of the GA POM ester. IOP was also reduced when GA was used in an intravitreal injection but not when it was topically applied. The difference in permeability between GA and GA POM ester across the corneal epithelium may explain why GA failed to reduce IOP following topical administration. GA and the GA POM ester inhibited cAMP PDE in rabbit ciliary body at low concentrations, the I50being 0.075μM and 2.4μM, respectively, with 0.25pM cAMP as substrate. GA and the GA POM ester markedly increased cAMP levelsin vitroin iris-ciliary body specimens. Possibly, GA POM ester or its analogues may represent a new mechanistic class of ocular hypotensive agents.

ISSN:0271-3683    

DOI:10.3109/02713689109013877

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Using standardized freeze wounds in cat corneas, we tested the efficacy of human recombinant Epidermal Growth Factor (EGF) to promote endothelial healing when solubilized in either phosphate buffered saline (PBS), 1% methylcellulose (MC), or sodium hyaluronate (NaHA), in final intraocular doses ranging from 2μg to 100μg of EGF. After 6 or 7 days' healing, animals were humanely sacrificed and corneal tissues were fixed and stained for light microscopy and computation of remaining wound areas.EGF in NaHA in final intraocular doses of 2 and 10μg prompted significantly more complete healing of transcorneal freeze wounds to endothelium compared with endothelium of eyes treated with NaHA control solution alone. EGF in PBS or in MC in doses ranging from 2–100μg/eye did not promote more complete wound healing than that seen in eyes treated with their respective vehicle solutions alone. All vehicle solutions were associated with similar degrees of wound healing, implying that they have no intrinsic healing properties.

ISSN:0271-3683    

DOI:10.3109/02713689109013878

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Lenses from normal rabbits, mice, rats, cattle, guinea pigs, lambs, chicken, cats, baboons, blue acara (fish) and dogs were examined for the presence of H2O2. No previous reports exist on the presence and levels of H2O2in normal eye lenses. Freshly isolated lenses of these animals were extracted with trichloroacetic acid and the extract neutralized with Tris. H2O2was assayed in these extracts by reacting them with l-14C-alpha-ketoglutarate and measuring the14CO2produced by peroxide-dependent decarboxylation. Peroxide of the order of 10-−4M was detected in most of the lenses except in baboons wherein it exists between 10-−4and 10-−5M. Culture experiments with rat lenses demonstrated that GSH may make a major contribution to the formation of H2O2in the intact lensin vivo.

ISSN:0271-3683    

DOI:10.3109/02713689109013879

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Lens fiber cells are coupled by communicating junctions that comprise over 50% of their appositional surfaces. The main intrinsic protein (MIP26) of lens fibers is a 23.2 kDa protein that forms large gap junction-like channels in reconstituted systems. Previously, we have shown that Ca++-activated calmodulin (CaM) regulates the permeability of reconstituted MIP26 channels and changes the conformation of MIP26, as measured by intrinsic fluorescence and circular dichroism spectroscopy. Examination of the MIP26 amino acid sequence has revealed a basic amphiphilic a-helical segment (Pep C) on the C-terminus with residue distribution similar to that found in other CaM binding proteins. To test the interaction between the amphiphilic segment and CaM, both a 20-mer peptide and trp-substituted fluorescent analog have been synthesized and purified by HPLC. Evidence from spectrofluorometric titration shows that the Pep C binds with CaM in 1:1 stoichiometry and with a Kdof-10 nM. Neither Ca++nor H+alone affects the conformation of the Pep C. However, when mixed with CaM the Pep C undergoes both a dramatic blue-shift in tryptophan fluorescence emission, indicative of strong hydrophobic interaction, and an increase in circular dichroism absorption in theα-helical region. Additional fluorescence blue-shift and a-helical content occur when Ca++is added to the CaM:Pep C complex.

ISSN:0271-3683    

DOI:10.3109/02713689109013880

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Three fundamentalin vitroexperiments have been done in the present report: 1) comparison of three different nutrient media on their abilities to culture and passage the human corneal epithelial cells; 2) evaluation of the ability of extracellular matrix material to promote the growth of cultured human corneal epithelium on collagen corneal shields; and 3) determination of the feasibility of the shield to serve as a carrier for the transfer of cultured cells to allogeneic, denuded corneal surfacein vitro.Primary cultures of human corneal epithelium were established from explants which were obtained from limbal and peripheral corneal tissue by three different nutrient media respectively: KGM (Keratinocyte Growth Medium), SHEM (Supplemental Hormonal Epithelial Medium), and one combination of the two media (KGM/SHEM). We found the KGM/SHEM combination to be more favorable because morphology was better preserved, the proliferation rate increased five-fold over the 14 days observed time course, and we were able to subculture the tissue for at least three passages. With this combined medium, a suspension of cultured corneal epithelial cells (5×K5/ml) was seeded onto either the concave surface of collagen corneal shields or onto shields which had been coated with extracellular matrix materials (Matrigel™or type IV collagen). The cells attached readily to all the coated shields (20/20) but to only a few of the uncoated shields (3/10), and formed a stratified tissue (2 to 3 layers) within seven days once the cells attached. However, the cells on the shields coated with Matrigel™failed to become confluent under these conditions. The stratified tissue on type IV collagen coated shields could then be subsequently transferred to denuded human corneal stroma in organ culture by placing them together and incubating for 2–7 days. After that, histologic examinations showed that the epithelial cells had attached tightly to the recipient stromal surface, even after the removal of the collagen shield.

ISSN:0271-3683    

DOI:10.3109/02713689109013881

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

The Major Intrinsic Polypeptide (MP26) of lens membranes contains an-asn-gly-sequence, which has been shown In other proteins to be particularly susceptible to spontaneous deamidation. To determine If the asparagine residue of this sequence undergoes age-dependent deamidationin vivo, antiserum to a synthetic peptide containing the sequence was used to monitor purification of a tryptic peptide containing this sequence from fetal versus mature bovine lenses. The peptide from fetal lenses contained the-asn-gly-sequence, while the peptide from mature lenses contained an-asp-gly-sequence, demonstrating that age-dependent deamidation of this asparagine residue was occurring in the lens.

ISSN:0271-3683    

DOI:10.3109/02713689109013882

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

The bovine lens multicatalytic proteinase complex (MFC) (MW 700 kDa) comprises at least twelve subunits in the molecular mass range 22–35 JcDa. Three of the subunits, LI (27 kDa), L2 (24 kDa) and L3 (29 kDa), were purified by reverse phase HPLC. Their amino acid composition and N-terminal sequences indicate that they are not identical. LI and L2 subunits show very high (>90%) sequence homology with specific subunits of rat liver and human reticulocyte MPC and these are considered to be homologous components of the MPC which are highly conserved in evolution.

ISSN:0271-3683    

DOI:10.3109/02713689109013883

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor

摘要:

Quantitative studies of collagen density and fibril size distribution as well as elastin density were carried out in the optic nerve head and sclera of human and experimental monkey glaucoma eyes. The collagen fibrils of the normal lamina cribrosa are smaller and more uniform in size than those of the sclera. This feature may be an adaptation to maximize either elasticity or resistance to mechanical stress. In glaucomatous nerve heads, there is a major disruption of the structure of the lamina cribrosa beam structure, including a decrease in collagen density. The peripapillary sclera undergoes similar collagen density changes to those in the nerve head in human glaucoma eyes. Elastin fiber density is unchanged in the glaucomatous nerve heads that we studied.

ISSN:0271-3683    

DOI:10.3109/02713689109013884

出版商:Taylor&Francis    

年代:1991

数据来源: Taylor