摘要:

Using standardized freeze wounds in cat corneas, we tested the efficacy of basic Fibroblast Growth Factor (bFGF) solubilized in phosphate buffered saline (PBS) to promote endothelial healing when injected intraocularly at doses ranging from 0.01µg to 10µg. After 6 days, animals were humanely sacrificed and corneal tissues were fixed and stained for light microscopy and computation of remaining wound areas.A significant dose-response relationship was found between the dosages of 0.01, 0.1, and 1.0 4mUg of bFGF/eye and the stimulation of more completely healed endothelium six days after transcorneal freeze wounding. Significantly larger endothelial wounds were present six days after wounding when the eyes were treated with 10µg of bFGF/eye compared with controls treated with PBS only.

ISSN:0271-3683    

DOI:10.3109/02713689209000746

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

Purified flagella from P.aeruginosaATCC 19660 were used for active, passive, or topical immunization prior to corneal challenge with strain 19660. At 30 days post-infection, a significant number of mice actively or passively immunized with flagella and infected with the homologous bacterial strain were protected from ocular disease when compared to control animals. In topical immunization studies, premixing of 19660 flagella with the bacterial inoculum prior to ocular challenge with strain 19660, provided results similar to those of the active or passive immunization studies.A reduced lipopolysaccharide (LPS:1 E.U./mg) flagella preparation was also produced and used similarly. Again, significant protection was achieved in mice immunized by flagella regardless of the immunization route.Anin vitroadherence assay also was performed to examine quantitatively the effect of exogenously applied flagella, or an antiflagella monoclonal antibody (MAb) on bacterial adhesion. Premixing of the bacterial inoculum with flagella or the MAb prior to applying it topically to corneas in organ culture all significantly inhibited bacterial binding.These results strongly suggest that significant ocular protection is achieved with either active or passive immunization, or premixing of the bacterial inoculum with flagella from strain 19660 prior to ocular challenge with the homologous bacterial strain. They also indicate that topical application of flagella or antiflagella MAb provide protection against ocular disease by decreasing bacterial adhesion to cornea.

ISSN:0271-3683    

DOI:10.3109/02713689209000747

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

A bovine lens“native”plasma membrane fraction containing its full compliment of extrinsic proteins was prepared by sucrose density centrifugation of the water insoluble fraction. The major membrane fraction was found at the 25/45% sucrose interface. This fraction contained 73% of the total water insoluble phospholipid, 74% of the total water insoluble cholesterol and 58% of the total urea-insoluble protein. Only 9% of the total urea-soluble protein was membrane associated (extrinsic protein), most (75%) was recovered from the pellet. The major intrinsic protein (8 M-urea-insoluble) of the membrane fraction was MIP28, with lesser amounts of MP17. Extrinsic proteins (8 M-urea-soluble) were examined by SDS-PAGE, isoelectric focusing, immunoblotting and amino acid composition analysis. Approximately 70% of the total extrinsic protein appeared to beαA-crystallins and modifiedαA-crystallins. About 20% of the extrinsic protein was apparentlyβ- andγ-crystallins. The remainder contained presumed cytoskeletal proteins and perhaps other unidentified polypeptides. The native plasma membrane was found distributed throughout the lens with only minor differences in the quantitative composition of the membrane fraction.We have concluded that the native membrane fraction represents the lens plasma membrane with its extrinsic proteins which exist in vivo. These extrinsic proteins appeared to be primarily acidicα-crystallin polypeptides with minor amounts ofβ-andγ-crystallins, and presumed cytoskeletal elements. We speculate that these extrinsic proteins may serve as a nucleation site for the association of other water insoluble protein through protein-protein interactions such as those found in the non-membrane associated urea-soluble protein. Together, these interactions may form a structured cytoplasmic matrix important for the maintenance of lens transparency.

ISSN:0271-3683    

DOI:10.3109/02713689209000748

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

The effect of several naturally-occurring retinoids and 13-cis-retinoic acid on the proliferation of cultured bovine retinal pigment epithelial (RPE) cells was investigated. None of the retinoids tested were toxic to the cultures and all, except retinylpalmitate, inhibited cell proliferation when given for more than 3 days. The relative potencies of the retinoids were; all-trans-retinoic acid>13-cis-retinoic acid>all-trans-retinol≅all-trans-retinaldehyde. Uptake of retinoic acid by cultured RPE cells was 10-fold less than the uptake of retinol. Although retinoic acid-treated cultures showed strong density-dependent growth inhibition, cellular proliferation was inhibited more in sparse cultures than in dense ones. Retinoic acid did not significantly inhibit the proliferation of first passage bovine or rabbit RPE cells, but partially inhibited the proliferation of first passage human RPE cells. The sensitivity of all these cultures to growth inhibition by retinoic acid increased in subsequent subcultures, yet there was no effect of passage number on retinoic acid uptake. This study demonstrates that RPE cell proliferation can be inhibited by retinoic acid but the sensitivity of these cells to the retinoid's effects are modulated by incubation time, in vitro aging, and cell density.

ISSN:0271-3683    

DOI:10.3109/02713689209000749

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

Proteoglycan synthesis was measured in chick sclera at the onset of form-deprivation myopia, as well as in the period immediately following removal of the occluder. Two day-old chicks were monocularly form vision deprived for periods from one to ten days and proteoglycan synthesis was determined after placing posterior scleral buttons in organ culture and measuring35SO4incorporation into glycosaminoglycans. Following 24 hrs of form-deprivation, proteoglycan synthesis was 33% higher in myopic eyes as compared with paired control eyes. The rate of proteoglycan synthesis further increased to levels 83% higher than controls after four days of form-deprivation and remained elevated throughout the ten day period of deprivation. Removal of the occluder after 10 days of form-deprivation resulted in a rapid drop in the rate of proteoglycan synthesis to control levels within 24 hrs. Proteoglycan synthesis was also measured in scleral chondrocytes isolated from control and myopic eyes after 10 days of form-deprivation. Proteoglycan synthesis by chondrocytes from myopic eyes did not return to control levels until 48 hrs after plating. Since the rate of proteoglycan synthesis returns to control levels more quickly during the recovery period ex vivo than when scleral chondrocytes from myopic eyes are placed in cell culture, we suggest that a mechanism is present within the eye which rapidly lowers the rate of proteoglycan synthesis in response to form vision.

ISSN:0271-3683    

DOI:10.3109/02713689209000750

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

Polyunsaturated fatty acids (PUFA), especially docosahexaenoic acid (DHA, 22:6n-3), are enriched in phospholipids of vertebrate rod outer segments (ROS). Retinal ROS can incorporate 22 carbon (C-22) PUFA from the plasma pool where C-20 PUFA are predominant. In this study, we analyzed the fatty acid composition of retinal pigment epithelium (RPE) and ROS from rats fed different fatty acid supplements to determine whether this enrichment is at the photoreceptor-RPE boundary or the RPE-choriocapillaris boundary. Long Evans rats were raised from birth for 13-14 weeks on a diet supplemented with 10% (wt/wt) hydrogenated coconut oil (COC; 0.2% 18:2n-6, no 18:3n-3), safflower oil (SAF; 73.8% 18:2n-6, 0.1% 18:3n-3), or linseed oil (LIN; 16.4% 18:2n-6, 52.2% 18:3n-3). These diets were chosen because they increased plasma levels of 20:3n-9, 20:4n-6, and 20:5n-3, respectively. These three fatty acids served as metabolic markers. Plasma levels of 22:6n-3 were reduced by the COC and SAF diets. The RPE incorporated 20:3n-9, 20:4n-6, and 20:5n-3 from the plasma. However, the levels of 20:3n-9 and 20:5n-3 were very low in ROS and 20:4n-6 was not significantly elevated in the ROS of the SAF diet group. The relative amount of total C-20 PUFA in phospholipids in RPE was similar to that found in plasma and was about 4-16 times (depending on different lipid classes) that in the ROS. In contrast, C-22 PUFA (22:6n-3 and 22:5n-6) showed a step-wise, average 3-5 fold increase in concentration from the plasma to the RPE to the ROS. Our data suggest that exclusion of C-20 PUFA from the retina occurs at the RPE-photoreceptor interface and that enrichment of C-22 PUFA occurs at both the plasma-RPE and RPE-photoreceptor interfaces.

ISSN:0271-3683    

DOI:10.3109/02713689209000751

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

The glycerolipid and fatty acid compositions of frog retinal pigment epithelium (RPE) were determined and compared with rod outer segments (ROS), retina, plasma, and red blood cells (RBC). The glycerolipid class composition of RPE was similar to RBC and ROS or retina, with phosphatidylcholine and phosphatidylethanolamine being the major components. The fatty acid composition of RPE differed substantially from that of plasma or RBC; the former contained much higher levels of C-20 and C-22 polyunsaturated fatty acids (PUFAs), such as 20:4n-6 and 22:6n-3, but less C-18 mono-, dienoic, and trienoic acids. The difference between RPE and ROS or retina with respect to fatty acid profile was also dramatic; RPE had relatively less 22:6n-3, but more 20:4n-6 and 18:2n-6, than ROS or retina. These results suggest that frog RPE cells may selectively take up C-20 and C-22 PUFAs from the circulation, but preferentially deliver 22:6n-3 to the ROS and retina. Fatty acid analyses show that 20:4n-6 and 22:6n-3 were unevenly distributed among RPE glycerolipids; phosphatidic acid, diglyceride, triglyceride, and phosphatidylserine are relatively more enriched in 22:6n-3 compared with 20:4n-6. This information might imply that these two PUFAs are metabolized differently inside the frog RPE cells.

ISSN:0271-3683    

DOI:10.3109/02713689209000752

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

Plasma membrane with its associated extrinsic proteins was isolated from normal and cataractous rat lenses by centrlfugation of the total water insoluble fraction from homogenized lenses on a discontinuous sucrose gradient. Membrane, which we call“native”membrane, was recovered mainly from the 25/45% sucrose interface. Development of the experimental U18666A cataract resulted in plasma membrane shifting to higher density (the 50/55% sucrose fraction) and great increases in the urea soluble protein content of the lens. At early stages of cataract development, most of the increased urea soluble protein was membrane associated, presumably as extrinsic protein. With advancing cataract, most of the urea soluble protein appeared in an essentially membrane-free pellet fraction. The urea soluble protein associated with the cataract membrane was shown by combined IEF, SDS-PAGE, Western blotting, amino acid compositional analysis and protein sequence determinations to be mainly composed of modified alpha- and beta-crystallins. Alpha A-crystallin truncated by not more than 27 residues from the carboxyl terminus plusβb1 crystallin truncated by 49 residues from the amino terminus were conclusively identified. In addition toβbl, a population of six alpha-crystallin derived polypeptides were specifically enriched in the cataract membrane fraction. Four of these six alpha-crystallins appear to be truncated from their carboxyl terminus, a modification which should have increased their hydrophobicity. The pellet fraction, which accumulated in the lens nucleus as the cataract advanced, was enriched in urea soluble gamma-crystallin derived polypeptides. We suggest that protein insolubilization in this experimental cataract involves the selective and tight association of principally modified alpha-crystallins to the fiber cell plasma membrane.

ISSN:0271-3683    

DOI:10.3109/02713689209000753

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

Alpha crystallin was prepared from newborn and aged bovine lenses. SDS-PAGE and tryptic peptide napping demonstrated that both preparations contained only the alpha-A and alpha-B chains, with no significant contamination of other crystallins. Compared with alpha crystallin from the aged lens, alpha crystallin from the newborn lens was much more effective in the inhibition of betaLcrystallin denaturetion and precipitation inducedin vitroby heat. Together, these results demonstrate that during the aging process, the alpha crystallins lose their ability to protect against protein denaturation, consistent with the hypothesis that the alpha crystallins play an important role in the maintenance of protein native structure in the intact lens.

ISSN:0271-3683    

DOI:10.3109/02713689209000754

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor

摘要:

RETINOPATHY OF PREMATURITY, A CLINICIAN'S GUIDE, edited by J.T. Flynn and W. Tasman, published by Springer-Verlag GmbH&Co., Germany, 1992.MANUAL OF OCULOTOXICITY TESTING OF DRUGS, edited by O. Hockwin, K. Green and L.F. Rubin, published by Gustav Fischer Verlag, Stuttgart, 1992

ISSN:0271-3683    

DOI:10.3109/02713689209000755

出版商:Taylor&Francis    

年代:1992

数据来源: Taylor