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Purpose: To develop a non-inflammatory model of both acute and chronic angiogenesis in the rabbit cornea using a known directly angiogenic cytokine., Methods: Pellets made of the slow-release polymer Hydron (polyhydroxyethylmethacrylate) and containing sucralfate and/or basic fibroblast growth factor (basic-FGF) were implanted into rabbit corneas. The neovascular response to the implantation of pellets containing basic-FGF alone, sucralfate alone or a titration of basic-FGF in the presence of a constant amount of sucralfate was measured. The role of inflammation in the neovascular response was also investigated., Results: The addition of sucralfate to the pellets led to the sustained release of basic-FGF resulting in a predictable and aggressive neovascular response with a low dose of basic-FGF that by itself was unable to elicit neovascularisation. At a dose of 500 ng per pellet, approximately one-third of the surface area of the cornea was vascularised within eight days of implantation. Minimal or no vascularisation occurred with the same dose of basic-FGF without sucralfate. While this dose of basic-FGF induced corneal oedema, only minimal inflammation was observed and the response was unaffected by ionising radiation. A less aggressive though still robust neovascular response with no or only minimal oedema was observed when the dose was lowered to 50 ng of basic-FGF per pellet. Some induced vessels persisted for more than three months., Conclusion: This is an inexpensive in vivo model of angiogenesis with the advantages of the neovascularisation being aggressive, predictable, persistent, unassociated with an obvious inflammatory response and induced by the sustained release of an agent known to have a direct stimulatory action on endothelial cells.

Purpose: The study of angiogenesis depends on reliable and reproducible models for the stimulation of a neovascular response. The purpose of this research was to develop such a model of angiogenesis in the mouse cornea., Methods: Uniformly sized Hydron pellets containing either basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF) and sucralfate were prepared and implanted into the stroma mouse cornea adjacent to the temporal limbus., Results: Neovascularization of the corneal stroma began on day 3 and was sustained through day 8. The bFGF-induced neovascularization was consistent and dose dependent in C57B1/6, as well as in severe combined immune deficient, beige natural killer cell-deficient, and nude mouse strains. Biomicroscopic and histologic examination of bFGF- and VEGF-induced angiogenesis was notable for the absence of corneal edema or substantial inflammation., Conclusions: This noninflammatory model of corneal neovascularization is especially advantageous because it is reproducible, economical, and facilitates investigation of angiogenesis in various murine tumor models as well as in genetically defined murine strains.

Purpose: To determine whether contact lens, related corneal pannus regressed when eyes were refitted with rigid gas-permeable contact lenses and to investigate the role of hypoxia as the putative origin of this complication., Methods: Twenty-three eyes of 14 patients with corneal pannus were identified retrospectively from a referral practice population and were separated into two subgroups. One subgroup accepted refitting with rigid gas-permeable contact lenses or discontinued contact lens wear, and the other elected to continue wearing hydrogel or polymethylmethacrylate contact lenses. The response of pannus was observed by slit-lamp biomicroscopy and photographs and was compared between the two subgroups. One eye of each of six patients identified with bilateral corneal pannus secondary to daily wear of hydrogel contact lenses was selected at random. Duplicates of the hydrogel contact lenses worn by these eyes were obtained, and their peripheral oxygen transmissibility (Dk/Lp) values were compared with those of a control group of hydrogel contact lenses identical to those worn by another group of patients, matched by age, gender, and laterality to the study group, but who did not have corneal vascularization., Results: Vascularization regressed in 14 eyes of nine patients refitted with rigid gas-permeable contact lenses and also in both eyes of one patient who discontinued contact lens wear. All seven eyes of five patients who declined refitting showed persistence or increase of pannus. The mean Dk/Lp of the hydrogel contact lenses for six eyes with pannus was 3.8 X 10(-9) cm ml O2/s ml mm Hg; mean Dk/Lp was 8.6 X 10(-9) cm ml O2/s ml mm Hg for the lens of the control eyes. This difference was found to be statistically significant (P = .028)., Conclusion: Peripheral, contact lens-induced hypoxia is a risk factor for corneal pannus, and conversion from hydrogel or polymethylmethacrylate contact lenses to daily wear rigid gas-permeable contact lenses in these eyes successfully reversed this complication.

Purpose: We evaluated the effective irradiation parameters for photodynamic thrombosis of experimental corneal neovascularization enhanced by simultaneous hyperbaric oxygenation., Methods: Neovascularization was provoked in both eyes of each of 35 albino rabbit corneas using the intracorneal suture technique. The lasered animals were divided in 3 groups. Group 1 (10 rabbits) was treated under hyperbaric conditions (28 atm for 25 min.); group 2 (5 rabbits) was treated breathing pure oxygen delivered by a face mask; group 3 (10 rabbits) was treated breathing room air. The fourth group (10 rabbits) was used for control. Animals were anaesthetized, and irradiation of new corneal vessels was carried out 30 minutes after the injection of 5 mg/kg chloroaluminum sulfonated phthalocyanine. A 670 nm diode laser with a power 4 mW and a spot diameter 350 mm was used. Exposure times necessary for vascular occlusion were registered. Histological examination was carried out at the end of the follow-up time., Results: Exposure times were significantly lower in groups 1 and 2 as compared to group 3 (1.75 +/- 0.15 min., 3.1 +/- 0.4 min., and 4.75 +/- 0.15 min. respectively). Total light dose averaged 490 J/cm,2 870 J/cm,2 and 1330 J/cm,2 respectively. Histological examination revealed thrombus formation in the targeted vessels of all three investigated groups., Conclusion: Combination of PDT with hyperbaric oxygenation results in an acceleration of the photodynamic process and provides for a possibility of significant reduction of photodynamic dose.

Purpose: To describe a new mouse model of corneal surface disease and neovascularization., Methods: Anatomic changes were demonstrated in corn1 and control A.By/SnJ mice from day 10 of gestation of 8 months of age by routine techniques of light microscopic and scanning electron microscopy. Corneal epithelial cell kinetics were evaluated by labeling cells in the "S" phase of the cell cycle by intraperitoneal injection of tritiated thymidine. Labeled cells were counted under 250X magnification, and the length of the corneal epithelial chord was measured by morphometric techniques. Results were expressed as labeled cells per linear millimeter of corneal epithelium. The corn1 locus was mapped using selected back-crosses., Results: Corn1 is characterized by early, irregular thickening of the corneal epithelium, development of stromal neovascularization by 20 days of age, and cataract by 48 days of age. Corneal epithelial cell kinetics demonstrated prominent labelling of corn1 mice at 30 days of age compared to the control mice. Corn1 behaves as an autosomal recessive gene and is located on mouse chromosome 2, approximately 5.2 cM from the agouti locus. Heterozygotes have no corneal disease., Conclusions: Corn1 mice, with genetically determined corneal epithelial hyperplasia and stromal neovascularization, may be particularly useful in studies of neovascularization and corneal surface proliferative disease.

Purpose: To report the danger of serious chemical ocular burn caused by common household soap., Case History: A 20 year old male developed an extensive burn of the right eye after the commonly used soft brown soap (also called floor soap) fell into his face and right eye. The burn caused conjunctival ischemia and necrosis. The cornea was oedematous and denuded of epithelium. The pH of the soap was 11.8. The patient received treatment for alkali burn of the eye. Stem cell transplantation was needed to heal the corneal surface defect. Scar formation and peripheral neovascularisation have reduced the visual acuity to counting fingers at 75 cm., Conclusion: This case of serious alkali burn caused by the common soft brown soap demonstrates the potential hazard to the eyes. The manufacturer was sought to write the pH and a warning on the exterior of the container that the "soft brown soap" may cause serious ocular injury.

The purpose of this study was to delineate the patient and surgical factors that contribute to the development of corneal neovascularization (CNV) after penetrating keratoplasty (PK). Thirty-six eyes of 36 patients with no antecedent CNV were enrolled in the study. Grafts were sutured to the host with 16 10-0 nylon sutures with the knots buried alternately in either the host or donor corneal stroma. Multiple perioperative factors were recorded for each patient, and at each postoperative visit systematic corneal drawings were used to follow the development of neovascularization. The stroma adjacent to each suture of each graft was given a neovascularization score based on the extent of vessel growth toward the wound interface. Univariate and multivariate analyses were performed, including generalized estimating equations logistic regression where each eye is considered a cluster of observations. Thirty-four patients without preoperative CNV or inflammation were followed prospectively for 6-9 (mean, 7) months after PK. Fourteen eyes (41%) developed some degree of CNV. Indication for keratoplasty, age, gender, phakic status, and size of donor button were not risk factors for CNV development. The most significant risk factor identified for any degree of CNV was placement of the suture knot in the host stroma (p = 0.00007), with the overall relative risk of CNV associated with these knots over 2 (95% confidence interval, 1.1-4.2). Furthermore, the mean recipient size in eyes with postoperative CNV was larger than eyes that did not develop neovascularization (p = 0.015), and active blepharitis was associated with a fivefold increase in the risk of developing CNV to the wound edge (p = 0.008). Embedding suture knots in the host stroma, active blepharitis, and a large recipient bed are significantly associated with postkeratoplasty CNV.

Purpose: Corneal allografts placed in human eyes at high risk often fail, and immune rejection is thought to be a major pathogenic factor. To understand the immunologic factors responsible for rejection in this instance, the authors have created "high-risk" eyes in mice by inducing corneal neovascularization. The authors then examined the fate of orthotopic corneal grafts placed in these beds and assessed the development of donor-specific delayed hypersensitivity (DH) in recipient mice., Methods: Three interrupted sutures were placed in the central cornea of recipient BALB/c mice to induce corneal neovascularization. Two weeks later, when corneal vessels occupied more than two quadrants of the cornea, mice received orthotopic corneal grafts from donor mice expressing alloantigens encoded by major and minor histocompatibility loci. Corneal allografts were evaluated by slit-lamp examination after grafting, and recipient mice were examined at the time of the rejection to determine whether they had acquired DH to alloantigens expressed on the corneal grafts., Results: Compared to grafts placed in normal eyes, a much higher incidence of rejection was observed among corneal allografts placed in neovascularized eyes (96.7% versus 46.7%). Moreover, grafts in neovascularized beds were rejected much more swiftly (2 weeks versus > 3 to 4 weeks). Rejection of corneal allografts in high-risk eyes coincided temporally with development of intense donor-specific DH, and the specificity of this immune response was directed solely at minor H antigens (not major histocompatibility complex-encoded antigens) on the graft., Conclusions: These results indicate that eyes rendered high risk by virtue of corneal neovascularization fail to provide immune privilege for orthotopic corneal allografts. In this circumstance, the grafts rapidly induce intense donor-specific DH that is readily detectable within 2 weeks of engraftment, at which time the grafts are acutely and universally rejected. The recipient DH response is directed exclusively at minor H antigens on the graft, which is consistent with the view that neovascularization creates graft beds in which recipient antigen-presenting cells infiltrate the graft and carry antigenic information by lymphatics to draining lymph nodes. In this manner, anterior chamber-associated immune deviation is avoided, and potentially allodestructive DH is promoted.

Purpose: The purpose of this study was to examine ocular surface abnormalities in aniridia., Methods: We prospectively studied the ocular surface in nine consecutive patients (18 eyes) with virtually total aniridia. Of the 18 eyes, two eyes (two patients) were excluded from the study because severe surgical damage was likely to be the main cause of their corneal abnormalities. Six volunteers whose eye examinations yielded normal results served as control subjects. Detailed slit-lamp biomicroscopic examinations were performed to assess corneal findings and confirm the presence or absence of the palisades of Vogt. Goblet cell density on the peripheral corneal epithelium and inferior conjunctival epithelium was calculated via impression cytology and was statistically compared between aniridic eyes and control eyes., Results: All 16 aniridic eyes had superficial corneal opacification and vascularization of either the peripheral or entire cornea. Palisades of Vogt were completely absent all around the limbal area. Goblet cell density on the peripheral cornea and inferior conjunctiva showed a statistically significant increase in the aniridic eyes (226 +/- 169 cells/mm2 and 406 +/- 203 cells/mm2), as compared with the control eyes (O cells/mm2 and 181 +/- 89 cells/mm2) (P = .0005 and P = .0102)., Conclusions: These findings suggest that, in the aniridic eye, conjunctival epithelium invades the cornea because of corneal epithelial stem cell deficiency.

Background: The presence of vessels has a negative influence on corneal transplant survival. Closure of such vessels prior to transplantation may improve the transplant results, and this might be achieved by irradiating the vessels with argon laser light after intravenous administration of a photosensitizer, e.g. bacteriochlorin a (BCA). A suture-induced corneal neovascularization model in rats was set up to test this hypothesis., Methods: Suture-induced vessels in the cornea of male Wistar rats were irradiated with argon laser light after intravenous administration of BCA. We applied irradiation of varying energy levels and duration and assessed the changes in the vessels by slit-lamp examination, fluorescein angiography and histology., Results: Suture-induced corneal vessels in the rat could be used effectively to study photothrombosis therapy. Intravenous administration of BCA prior to irradiation (lambda = 514.5 nm) of the corneal vessels led to vessel closure at lower energy levels and of longer duration than occurred with laser treatment alone., Conclusions: Suture-induced corneal neovascularization in the rat can be used as a model to study the efficacy of photothrombosis therapy. BCA can be used to enhance the rate and duration of vessel closure.

Human amniotic membrane isolated from the placenta contained basement membrane components such as type IV collagen, laminin, and 6 and 4 integrins, all of which remained detectable while preserved in glycerin for one week. One month after the n-heptanol removal of the total corneal epithelium and the limbal lamellar keratectomy, all rabbit eyes carried features of limbal deficiency, including conjunctival epithelial ingrowth, vascularization and chronic inflammation. Ten control eyes then received a total keratectomy, and 13 experimental eyes received an additional amniotic membrane transplantation. Three-month follow-ups revealed that all control corneas were revascularized to the center with granuloma and retained a conjunctival phenotype. In contrast, in the experimental groups, 5 corneas became clear with either minimal or no vascularization; the rest had either mild peripheral (5) or total (3) vascularization and more cloudy stroma. Using monoclonal antibodies for epithelial markers and matrix components, we concluded that the success correlated with the return of a cornea-like epithelial phenotype and the preservation of the amniotic membrane, whereas the failure maintained a conjunctival epithelial phenotype and the amniotic membrane was either partially degraded or covered by host fibrovascular stroma. Measures taken to facilitate the former might prove this procedure clinically useful for ocular surface reconstruction.

Background: Sclerocornea is a congenital, uni- or bilateral, non-progressive, non-inflammatory malformation characterized by histological changes of the cornea resembling scleral tissue., Case Report: We report the case of a 2-month-old child with bilateral dysplasia of the anterior segment including: 1. extreme corneal ectasia; 2. vascularized, opacified microcornea; 3. presence of rudimental limbus; 4. anterior scleral dysplasia. The axial length measured by means of ultrasonography (A-scan) was about 24 mm in both eyes. A penetrating keratoplasty was first performed in the right eye and one week later in the left eye. Intraoperatively an advanced iris atrophy with extensive anterior synechiae, as well as a localized anterior subcapsular cataract were seen bilaterally. A pale optic disk with a maximally enlarged optic cup could be seen only in the left eye., Conclusion: Although the histologic examination was compatible with the diagnosis of sclerocornea, the presence of atypical clinical findings previously unreported does not allow a definite classification of the case reported herein.

Various pathological conditions may cause vascularisation of the normally avascular and transparent cornea. The purpose of this study was to investigate the fractal geometry of corneal vascularisation and relate it to different growth models which may have implications for the understanding of fundamental processes involved in vasculogenesis. Photographs of 12 cases of severe superficial corneal vascularisation were obtained from accidentally alkali burned eyes long after the acute inflammatory reaction was over. The fractal dimension of these vessel patterns was calculated by means of the density-density-correlation function method to be D = 1.893 +/- 0.044 (m +/- sd) (n = 12). It is in excellent agreement with percolation models. The results suggest the importance of the surface characteristics of the invading cells as well as that of the extracellular matrix and the related interaction between them.

Dipivefrin hydrochloride is a lipophilic prodrug for epinephrine hydrochloride, allowing lower concentration of the drug to achieve the same intraocular pressure lowering effect and having also less harmful effects than epinephrine hydrochloride. However, harmful effects have been associated also with the use of dipivefrin and here we report of a case of corneal vascularization during long-term use of dipivefrin.

We evaluated the efficacy of topical cyclodextrin-encapsulated FK-506 in the prevention of experimental corneal allograft rejection. Two weeks after inducing corneal inflammation and neovascularization with 8-0 silk sutures, 23 albino rabbits received a unilateral 8-mm diameter central penetrating corneal allograft from pigmented donors. Rabbits were randomly assigned to no treatment (eight eyes), topical cyclodextrin four times daily for 28 days (seven eyes), or topical FK-506 0.3 mg/ml in a cyclodextrin suspension (eight eyes) four times daily for 28 days. Grafts were examined daily for degree of inflammation, neovascularization, edema, and signs of rejection for up to 100 days. Seven of eight (88%) untreated grafts and five of seven (71%) cyclodextrin-treated grafts rejected at a median of 3 weeks after transplantation, whereas only two (25%) of eight FK-506-treated grafts rejected and did so at a significantly longer interval (p < 0.005). Topical FK-506 prevents or delays corneal allograft rejection after experimental corneal transplantation.

Corneal neovascularization (CNV) was induced by alkali burn injuries in 20 rabbit corneas. The process of CNV was observed with biomicroscopy in vivo and histologically by light and electron microscopy. At 8 hours after injury, an obvious acute inflammatory response as evidenced by infiltration of neutrophils had already occurred in the pericorneal limbal area. Many neutrophils in the extravascular space contained abundant intracytoplasmic phagolysosomes. The vascular sprouts appeared 2 days after injury. The infiltration of neutrophils and their debris were visible around newly formed blood vessels in the cornea. The results suggest that there be the participation and an inductive action of neutrophils in the pathogenesis of CNV.

Background: Corneal neovascularization is generally undesirable because it can lead to corneal scarring, lipid deposits, and corneal graft failure. To eliminate these vessels, several techniques are available including laser photocoagulation., Methods: This prospective study was designed to compare the effectiveness of three laser wavelengths (continuous wave Nd:YAG, 1064 nm; argon green 514 nm; and yellow dye, 570 nm) to obliterate experimentally induced corneal neovascularization in the rabbit. Corneal vascularization was created in 12 rabbits by placing 7-0 silk sutures through two quadrants of the cornea. Once neovascularization was complete, the suture was removed and one of the three lasers was applied to occlude vessels at one of the neovascular sites. The other site was used as a control., Results: The yellow wavelength, when compared with the green required fewer exposures to occlude corneal vessels. At no time during observation was any laser more effective than the control. In the continuous wave Nd:YAG group, tissue necrosis was needed to achieve closure of vessels., Conclusions: Yellow and green laser light are equally effective in eliminating the corneal vessels. Continuous wave Nd:YAG, as used here, appears to be a poor choice.

Corneal vascularization is an important clinical event that requires careful management. Vessel penetration can serve to heal a threatening inflammatory process, cause opacification and loss of vision, induce graft rejection, or serve as a warning of corneal distress. The latter condition is particularly common in contact lens wearers. Practitioners should be aware of the subtle signs of early neovascularization, and work toward its remission. Acute vessel penetration is a difficult clinical situation to manage. Quiescent vessels in the cornea must be explained by history and managed with either monitoring or intervention.

The effect of rabbit vitreous with endotoxin-induced uveitis was investigated for its role in the modulation of rat corneal neovascularization (NV). Elvax pellets implanted into at cornea contained either uveitic vitreous, intact vitreous or Elvax alone. The latter two kinds of pellets did not elicit NV. Uveitic vitreous pellets caused, in 100% of eyes, a persistent NV with maximal growth on the 10th day post implantation. Prostaglandin E2 and leukotriene B4 levels in the uveitic rabbit vitreous at 36 h postendotoxin injection were 7- and 5-fold higher than baseline, respectively. Histopathologically, the neovascularized cornea showed a significant inflammatory reaction attributable to the uveitic vitreous extract.

The antiangiogenic effect of tecogalan sodium on corneal neovascularization was investigated. Tecogalan sodium, a sulfated polysaccharide peptidoglycan complex isolated from an Arthrobacter species, has been reported to inhibit angiogenesis induced by basic fibroblast growth factor (bFGF) as well as tumor angiogenesis related to Kaposi's sarcoma. Corneal neovascularization induced by bFGF was inhibited by tecogalan sodium in a dose-dependent manner. Since bFGF is known to have a promoting effect on corneal neovascularization, tecogalan sodium may be possible therapeutic agent for corneal neovascularization, which can cause severe visual disturbances.

We studied the histotoxic effects of mitomycin C and 5-fluorouracil (5-FU) in the eyes of New Zealand white rabbits using light microscopy. Twenty-four eyes of 24 rabbits received subconjunctival injections of mitomycin C (0.025, 0.05, 0.10, 0.20, 0.40 or 0.80 mg/mL) or 5-FU (10.0 mg/mL), either alone (0.25 mL) or in combination with anterior chamber injections (0.05 mL), once daily for 4 consecutive days. Two eyes of two rabbits received subconjunctival and anterior chamber injections of unpreserved sterile saline. The eyes were examined regularly for external signs of toxicity and were enucleated 4 weeks after the last injection. Mitomycin C produced considerable tissue damage in the anterior segment, the severity being related to both the concentration and the route of administration. Eyes that received both subconjunctival and anterior chamber injections showed more damage than those that received subconjunctival injections alone. At the highest concentration of mitomycin C the cornea was inflamed, with stromal necrosis and marked endothelial loss. Hemorrhagic iris necrosis was also seen. In contrast, the 5-FU-treated eyes showed no microscopic evidence of toxicity. We conclude that there is greater risk of toxic anterior segment effects when mitomycin C is used as adjunctive therapy following filtration or pterygium surgery.

Background: Corneal neovascularization is a common clinical entity. Although visual acuity is usually impaired and corneal function compromised, there has been only limited success in the clinical management of this condition. We evaluated the efficacy of laser photocoagulation of neovascularization in the rabbit cornea., Methods: New vessel formation was provoked by the placement of sutures in the corneas. Rose bengal was injected intravenously and new vessels in the upper part of the corneas were treated with an argon laser. The lower halves were used as controls. Eighteen rabbits were divided into 2 groups. In group A neovascularization was treated 28 days after suture removal, when corneal inflammation had regressed. In group B treatment was performed 3 days after suture removal, when the cornea still exhibited marked inflammation. Postoperatively, the corneas were studied by slit-lamp microscopy, fluorescein angiography, and light, as well as electron microscopy., Results: In group A, treatment led to the immediate occlusion of the vessels and to their gradual disappearance during the course of 3 months. In group B, no occlusion was seen during the 3-month follow-up period. The main histologic findings in the occluded vessels were endothelial cell disruption and degeneration, and the formation of clots., Conclusion: Our results suggest that argon laser photocoagulation using rose bengal is an effective method of occluding corneal new vessels, providing there is no corneal inflammation at the time of treatment.

Objective: To investigate the immunosuppressive effect of rapamycin in prolonging allograft survival in the rat model of orthotopic allogeneic penetrating keratoplasty., Design: Thirty inbred Lewis rats received corneal allografts from Brown Norway donors. Animals were divided into two rapamycin treatment groups and one allogeneic control group., Results: By the second week after surgery, all of the control animals had experienced allograft failure due to allograft rejection. However, allografts in seven of 10 animals in the low-dose treatment group and allografts in seven of nine animals in the high-dose treatment group remained clear. In addition, corneal neovascularization was markedly reduced in the treated animals., Conclusions: The systemic administration of rapamycin prolongs corneal allograft survival and significantly inhibits the neovascular component of rejection in the rat model of orthotopic allogeneic penetrating keratoplasty.

As mononuclear cell infiltration and growth of pannus critically depend on synovial neovascularization in rheumatoid arthritis (RA), inhibition of the synovial blood vessels would have the potential to reduce rheumatoid inflammation. In this investigation, we studied the effect of gold sodium thiomalate (GST) and auranofin (AUR) on neovascularization in vivo by using a micropocket technique. Both GST and AUR suppressed rabbit corneal neovascularization in a dose-dependent fashion. Significant inhibition was observed by 3 mg/kg GST and 1 mg/kg AUR injected intravenously every other day. These injections maintained serum gold concentrations at the level of 2-5 micrograms/ml and less than 2 micrograms/ml in GST- and AUR-injected rabbits, respectively. These are concentrations attained in the serum or synovium of rheumatoid patients treated by gold compounds. Similar inhibition was observed by both intramuscular administration of GST and oral administration of AUR. In contrast, no inhibition was observed when non-steroidal anti-inflammatory drugs (NSAIDs; 20 mg/kg acetylsalicylic acid, 10 mg/kg ibuprofen and 10 mg/kg indomethacin) were injected intravenously on a daily basis. These results suggested that gold compounds have an antiangiogenic effect in vivo. The inhibition of neovascularization by gold compounds suggested that they may suppress rheumatoid synovitis by reducing the number of small blood vessels required for mononuclear cell infiltration and synovial tissue proliferation.

The aim of this study was to evaluate the possible influence of phthalocyanine dose and of time interval between phthalocyanine injection and irradiation commencement on the rate of experimental corneal neovascularization photodynamic thrombosis in albino rabbits. New corneal vessels were irradiated with a diode laser (670 nm, 2 mW) after the intravenous injection of chloroaluminum sulfonated phthalocyanine. Different animals were irradiated either 5 min after the injection of different phthalocyanine doses (3, 6, 8, 12, or 14 mg/kg), or at different times (5 min, 24 h, or 58 h) after a standard phthalocyanine dose (3 mg/kg) injection. Irradiation time necessary for vascular occlusion was recorded. Decrease of phthalocyanine dose as well as delay of irradiation onset resulted in a statistically significant increase of irradiation time. Electron and light histological examination revealed platelet thrombi inside irradiated corneal new vessels. Damage in the vascular endothelial cell membrane and in intercellular contact structure was noted, leading to disorganization of the endothelial cells layer and death of most endothelial cells. These results indicate that both early commencement of irradiation after phthalocyanine injection and phthalocyanine dose increase accelerate the rate of phthalocyanine mediated corneal neovascularization photodynamic thrombosis. Thrombosis seems to result from photochemically induced vascular endothelial cell damage.

Ten corneal specimens obtained at the time of penetrating keratoplasty for long-standing refractory Acanthamoeba keratitis were reviewed. Histopathologic findings included epithelial denudation and variable degrees of necrosis, inflammation, and cysts or trophozoites of acanthamoeba in the stroma. No blood vessels were found in the stroma despite the presence of long-standing corneal inflammation. The absence of corneal stromal neovascularization on biomicroscopic and histopathologic examination is a sign of Acanthamoeba keratitis.

Purpose: To reexamine the possible effect of human thrombospondin on in vivo angiogenesis., Methods: In vivo angiogenesis in the rabbit cornea was induced by implants of Elvax-40 sequestering human recombinant basic fibroblast growth factor (bFGF) or bacterial endotoxin lipopolysaccharide (LPS). Implants sequestering various concentrations of thrombospondin were examined for their ability to induce angiogenesis and also for their possible influence on the angiogenic potential of bFGF- or LPS-sequestering implants., Results: Constant and reproducible angiogenic stimuli were obtained with implants sequestering 250 ng or more of bFGF or 100 ng or more of LPS. Implants sequestering 500 ng of thrombospondin induce very little clinical change but larger concentrations induce infiltration of leukocytes and a mild angiogenic stimulus. Combination of thrombospondin implants with bFGF or LPS implants enhanced the angiogenic response to either of these factors. The thrombospondin enhancing effect was more prominent when LPS was the stimulating factor. Histologic examination of the tested corneas disclosed that the LPS angiogenic stimulus follows the influx of leukocytes. Conversely, the bFGF angiogenic stimulus appears to be associated with the proliferation of stromal keratocytes and corneal epithelial cells. The thrombospondin angiogenic enhancing effect on both the LPS and bFGF stimuli was correlated with an increased infiltration of polymorphonuclear cells., Conclusion: In this system, thrombospondin enhanced the in vivo angiogenic process induced by bFGF or LPS. This enhancement appears to be associated with an in vivo activation and chemotactic effect on the polymorphonuclear cells.

Corneal neovascularization complicates many anterior segment diseases. Corneal laser photocoagulation using yellow light (577 nm) has been shown to reduce corneal neovascularization. No histopathologic studies of the effects of this treatment on the eye have been reported, however. Target (cornea) and nontarget (iris, lens, retina, and choroid) ocular tissue were studied 1, 24, and 48 h and 5 days after yellow dye corneal laser photocoagulation in a rabbit model of corneal neovascularization. Biomicroscopic examination of the corneas revealed intracorneal hemorrhage in five of 24 (21%) eyes of nonpigmented rabbits. Faint lenticular opacities were observed in two eyes of pigmented rabbits 24 h after laser treatment. Histopathologic examination revealed increased cellularity (neutrophils) (p < 0.005) in the cornea, increasing from 1 h after treatment, peaking 24 h later, and persisting past 5 days. Distortion of the corneal lamellae by red blood cells occurred in eyes in which intracorneal hemorrhage developed. These results indicate that corneal laser photocoagulation using yellow light is a relatively safe procedure for reducing corneal neovascularization.

Purpose: The present study considered the hypothesis that spontaneous neovascularization of the cornea of nude mice results from a local imbalance between angiogenic and anti-angiogenic factors., Methods: The presence of angiogenic and anti-angiogenic factors was revealed by exchanging orthotopic corneal allografts between nude BALB/c mice and normal (hirsute) euthymic BALB/c mice and observing the presence, intensity, and degree of corneal neovascularization before and after grafting., Results: Avascular corneal grafts from normal BALB/c donors resisted neovascularization after grafting to spontaneously vascularized graft beds in nude mice. In contrast, spontaneously vascularized corneal grafts from nude mice remained vascularized over 2 mo after grafting to similar nude recipients. Although corneal grafts from nude donors stimulated neovascularization in normal BALB/c recipients, most of the vessels regressed by the 6th week post transplantation., Conclusions: The results confirm that there is an imbalance between angiogenic and anti-angiogenic factors in the nude mouse cornea. The cornea of the nude mouse displays more angiogenic activity and less anti-angiogenic activity than that of the normal mouse. Most angiogenic activity of the nude mouse cornea appears to reside in the epithelium.

Background: Conventional treatment of corneal stromal vascularization is often inadequate. The authors developed a rabbit model of corneal stromal vascularization, treated it with laser photocoagulation, and then studied the histopathology., Methods: A reproducible model of corneal stromal vascularization was developed in albino rabbits by injecting sodium hydroxide into the corneal stroma. Corneal stromal vascularization was produced in both eyes of 13 rabbits, and treated after stabilization at 5 weeks with 577-nm yellow dye laser in 1 eye of each rabbit. Seven rabbits were followed for 6 months with corneal angiography and photography, and the corneal stromal vascularization quantified with a grid. The other 6 rabbits were killed at 1, 4, 8, 24, 48 hours, and 6 days after laser photocoagulation and examined by light and transmission electron microscopy., Results: Stable corneal stromal vascularization was observed in the anterior and midstroma for at least 6 months in the model. Laser photocoagulation reduced corneal stromal vascularization significantly compared with the controls (P < or = 0.05), resulting in 40.7% +/- 5.0%, 45.3% +/- 3.3%, and 34.9% +/- 5.2% (mean +/- standard error of the mean) reduction at 2, 4, and 6 months, respectively. Maximum inflammatory cell infiltrates were detected at 8 hours after laser photocoagulation, which diminished markedly at 6 days. The stroma of unlasered eyes showed no inflammatory cells and considerably more patent blood vessels than the lasered eyes. In the lasered eyes, transmission electron microscopy showed damaged vascular endothelial cells, extravasated erythrocytes, haphazardly arranged collagen fibrils, thrombus formation, and ghost vessels in the stroma. No damage was observed in the deep corneal stroma or endothelium in the lasered eyes., Conclusion: Laser photocoagulation is effective in reducing corneal stromal vascularization in this model for at least 6 months. It does not damage the deeper stroma or endothelium.

We used chloroaluminum sulfonated phthalocyanine as a photo-sensitizer and a diode laser as a light source for induction of photothrombosis of corneal neovascularization. Corneal neovascularization was induced in 1 eye of each of 10 New Zealand white rabbits using intrastromal 6.0 silk sutures. After the intravenous injection of phthalocyanine in a dose of 4 mg per kg of body weight, photothrombosis was carried out using a diode laser emitting at 675 nm. The animals were followed up by both fluorescein angiography and slit-lamp photography for up to three months. At the end of the follow-up period the eyes were prepared for histology. After photothrombosis, thrombus formation was induced within new vessels demonstrating histological characteristics of both arterioles and venules. Most of the vessels disappeared or remained closed during the follow-up period. Recanalization of some of the thrombosed vessels occurred within the first 20 days after the operation. The combination of phthalocyanine and a 675 nm diode laser is effective for the induction of photothrombosis of corneal neovascularization.

The first results of a prospective study on 50 patients who were followed up 2, 4, 6, and 12 months after keratoplasty are reported. The patients received a human cornea transplant, which had been cultured in a modified MEM medium for up to 28 days. We observed that the loss of epithelium during organ culture leads to delayed regeneration after keratoplasty. This is especially important for patients who have had this problem almost before transplantation. The slower increase of visual acuity after keratoplasty with organ-cultured material is due to increased folding of Descemet's membrane. Nevertheless the best vision was usually reached within 6 to 12 months. We saw seven immune reactions by which five transplants were lost. A comparatively low rejection rate was observed especially in the group of high-risk patients (27.2%). Organ culture may therefore have some additional positive effects for these patients.

This report describes an in vitro system that could be useful for investigating corneal neovascularization. When isolated bovine capillary endothelial cells (CEC) were cocultivated with rabbit corneal keratocytes, capillary-like cords extended actively from the endothelial cells in the multilayers of the keratocytes. The developing cords continued to elongate, branch out, and anastomose with each other, forming a capillary-like network by the 12th day of coculturing. Electron microscopic observation revealed that the cords were tubular structures composed of three to seven endothelial cells in the multilayers of keratocytes, and that a basal lamina-like matrix was situated at the abluminal face of the endothelial cells. The growth of cellular cords from the cloned CEC also occurred when CEC were seeded onto a cell-free extracellular matrix that had been synthesized by keratocytes. Keratocyte-conditioned medium alone did not stimulate proliferation of CEC. These observations clearly indicate that the formation of capillary-like structures by CEC depended upon the presence of an extracellular matrix produced by keratocytes, rather than upon the keratocytes themselves or any other keratocyte byproduct. This simple in vitro experimental system is proposed as a useful tool for studying corneal neovascularization.

Perfluorocarbon liquids are fully fluorinated, synthetic, transparent compounds with a high specific gravity. These compounds are being increasingly used as an intraoperative tool for repair of complicated retinal detachments. A potential complication of their use, however, is liquid entering the anterior chamber in aphakic patients. In the study described herein we evaluated the effects of two of these liquids when placed in the anterior chamber of the rabbit eye. Sixteen rabbit eyes underwent anterior chamber injection of 0.05 ml of either perfluoroctane, perfluoropolyether, or balanced salt solution (control eyes). Animals were monitored clinically by biomicroscopy and external photography for up to 14 days, after which they were sacrificed and the corneas processed for light and for scanning electron microscopy. The animals injected with perfluoropolyether showed more intense stromal inflammation and corneal vascularization (p less than 0.02) than did those that received perfluoroctane. However, the perfluoroctane group showed more of the "fish-egging" phenomenon (p less than 0.03). Loss of endothelial cells was similar in the two groups, as determined by light and scanning electron microscopy. These results suggest that the corneal toxicity of these two perfluorocarbon liquids is such that their use as vitreous substitutes should be limited to short-term replacement.

Endothelin, a potent vasoconstrictor, was found to be ineffective in the treatment of experimentally induced corneal neovascularization. Endothelin was administered topically, subconjunctivally, and intraluminally in serial concentrations ranging from 0.0005 to 5.0 micrograms/mL in New Zealand white rabbits without effect. Electron microscopy of the neovascular cornea revealed the vessels consisted only of endothelin and pericytes. Hence, the vessels were not responsive to endothelin because they lacked contractile smooth muscle.

When isolated bovine capillary endothelial cells were co-cultivated with rabbit corneal keratocytes, capillary-like cords extended actively from the capillary endothelial cells in multilayers of keratocytes. The developing cords continued to elongate, branch out and anastomose with each other, forming a capillary-like network by the 12th day of co-culturing. Electron microscopic observation revealed that the cords had tubular structures, which were composed of 3-8 endothelial cells in the multilayers of keratocytes, and that a basal-like matrix was situated on the outer surface of the endothelial cells. The roles of keratocytes in the cord formation by capillary endothelial cells were discussed. This simple in vitro experimental system is proposed as a useful tool in the study of corneal neovascularization.

We sought to determine if cyclosporine, which has been shown to suppress corneal allograft rejection, could also suppress corneal neovascularization induced by interleukin 2. Thirty A/J mice were treated with daily intramuscular injections of cyclosporine (25 mg/kg in olive oil) for 3 days before and 2 weeks following the intrastromal injection of 0.5 microL (5 IU) of recombinant mouse interleukin 2. Controls received intramuscular injections of olive oil. The mean area of corneal neovascularization 4, 8, and 12 weeks after injection was 9.2, 9.1, and 9.2 mm2, respectively, in controls, and 5.0, 5.2, and 5.2 mm2 in cyclosporine-treated mice (P less than .02; Student's t test). Cyclosporine causes a significant reduction in interleukin 2-induced corneal neovascularization that may, in part, account for its ability to prolong corneal allograft survival in high-risk cases.

A model for keratoepithelioplasty (KEP) was developed using the Lewis rat, and histological studies were performed using this model. The entire corneal epithelium was removed mechanically and a 1.5-mm width of the conjunctiva including the limbus was excised. An oval corneal lamellar graft (3 x 1.5 mm) with an intact epithelium taken from another Lewis rat was transplanted on the denuded limbus. Biomicroscopic observations showed much less vascular invasion in the part of the cornea adjacent to the lenticule than in other parts of the cornea, and the cornea remained clear adjacent to the lenticule. Histologically, a few vessels were observed in the corneal stroma under the lenticule. Epithelial cells on the lenticule specimens showed histological characteristics of the corneal epithelium. These findings indicate that one of the functions of KEP is to block neovascularization in the newly developing corneal epithelium by transplanting the lenticule between the corneal epithelium and conjunctival vessels. The present study also confirmed that this model is useful in the research of the pathophysiological mechanism of KEP.

In nine rhesus monkeys, human corneal grafts were used for penetrating keratoplasty in one eye of each. The data were divided into three groups according to the size of grafts. Group 1 (n = 3) received 5.5-mm human corneal grafts in 5.0-mm recipient beds; group 2 (n = 3) received 6.2-mm human corneal grafts in 6.0-mm beds; and group 3 (n = 3) received 7.5-mm grafts in 7.0-mm beds. Five of nine eyes in groups 1 and 2 (55%) maintained graft clarity for more than 1 year, but only two of the three in group 1 maintained clear human corneal grafts for 32 and 36 months, respectively. However, the three grafts in the third group maintained transparency for only 4-7 months. Our findings suggest that xenogeneic corneal grafts may have a greater success rate in humans than previously believed.

The rabbit corneal angiogenesis assay was modified to allow quantification of the neovascular response induced by growth factors. Basic Fibroblast Growth Factor (bFGF) was used for this experiment. bFGF was diluted in phosphate buffer solution (PBS) in such a way that 2 microliter of solution contained concentrations ranging from 17 to 1,200 ng. Each drop was absorbed by a 1 x 2 mm particle of a dried 70% hydratable hydrogel and implanted into a mid-stroma corneal pocket, 2 mm from the limbus. Iodinated bFGF was used to measure growth factor diffusion in the cornea, which was found to be isotropic. Autoradiography showed that bFGF was stored in the cornea at both epithelial and endothelial level. Corneal neovascularization occurred on the second day after implantation and was maximal on the 7th day. At this time the neovascular surface was measured by planimetry on corneal photographs and compared with controls. This method allows precise definition of the dose of any substance to be implanted into the cornea and induces a rapid neovascular response, thus allowing quantitative evaluation of neovascularization within one week. A neovascular response was detectable for doses as low as 35 ng of bFGF and increased proportionally to the dose of bFGF implanted.

Deep corneal neovascularization is a recently described complication of contact lens wear. Visual loss associated with deep corneal neovascularization has been associated with stromal lipid leakage. Deep intracorneal hemorrhage is rare and has been documented infrequently with the use of contact lenses. The authors present five cases of deep stromal hemorrhage associated with contact lens-related deep corneal neovascularization. These patients had no other discernible cause for their vascularization and subsequent hemorrhage. One patient required a penetrating keratoplasty for visual rehabilitation. Contact lenses of all types may cause neovascularization. The deep stromal neovascularization develops insidiously and may progress in the absence of acute symptoms. The presence of deep stromal neovascularization must be watched carefully and managed as a potentially vision-threatening complication of contact lens use.

During wound healing, embryological development, and solid tumor growth, the established vasculature gives rise to large numbers of new blood vessels. This neovascular response occurs at the level of the capillary bed, where endothelial cells divide rapidly, locally remodel the surrounding stroma, and migrate away from existing vessels to form capillary sprouts. In order to examine the environment of these newly forming vessels, actively growing blood vessels in neovascularized rabbit and guinea pig corneas were examined immunohistochemically using antibodies against laminin, type IV collagen, heparan sulfate proteoglycans, entactin, and factor VIII-related antigen. Sequential serial 5-microns sections taken from the unfixed frozen corneas in a plane perpendicular to the direction of vessel growth were stained with these antibodies. It was possible to follow well-defined lumenized vessels out through sequential sections to the point where they became single factor VII-R positive cells in the region of the capillary sprout. Examination of these stained sections has shown the presence of four important basement membrane components--laminin, type IV collagen, heparan sulfate proteoglycan, and entactin--associated with actively migrating and invading capillary sprouts. These results suggest that the extracellular matrix of the actively invading capillary sprouts does not differ qualitatively from that of the established vasculature.

The rabbit corneal pocket assay is one of the most frequently used systems for the study of angiogenesis. This model particularly is useful to identify stimulators of new blood vessel formation. More recently, however, interest in inhibitors of angiogenesis has grown, and several antiangiogenic agents have been identified. Investigations of angiogenesis inhibitors require a reliable model for the stimulation of neovascularization. One method was modified to produce corneal neovascularization by implanting into the rabbit cornea a sustained-release polymer containing endotoxin (Escherichia coli lipopolysaccharide). The implant was prepared by mixing weighed quantities of endotoxin with ethylene vinyl acetate copolymer (Elvax) and forming 1-mm3 pellets containing 1%, 7.5%, 15%, 20%, 30%, and 40% (w/w) of endotoxin. Pure Elvax pellets were implanted as controls. Intrastromal corneal pockets were created in 92 eyes of male, albino New Zealand rabbits (n = 80), and sterilized endotoxin-copolymer implants were introduced. The growth rate of new vessels was measured by slit-lamp biomicroscopy. Endotoxin loads of 15% (n = 40) produced a strong neovascularization response with minimal stromal edema, with a mean growth rate of 0.21 +/- 0.12 mm/day. Loads of 1%, 7.5%, and 20% yielded 0.1 +/- 0.03 mm/day, 0.27 +/- 0.05 mm/day, 0.30 +/- 0.06 mm/day, respectively (n = 8, each group). Higher loads (30% and 40%; n = 8, each group) produced intense neovascularization, accompanied by severe corneal edema that obscured accurate measurement of the vessels. Corneal pockets that did not contain polymer implants were avascular. When endotoxin-Elvax pellets were removed, the new vessels regressed within 2 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)

It is known that hydrocortisone can be converted to a potent angiogenesis inhibitor by coadministration with heparin or with a sulfated cyclodextrin. The activity of tetrahydrocortisol-S, a purely angiostatic corticosteroid, can be potentiated by beta-cyclodextrin tetradecasulfate as shown in this study. This drug "pair" and other pairs of corticosteroids and beta-cyclodextrin tetradecasulfate can be applied topically to inhibit corneal neovascularization. Endotoxin-induced corneal neovascularization in rabbits was treated with beta-cyclodextrin tetradecasulfate coadministered with either: hydrocortisone, tetrahydrocortisol-S, or 6-alpha-fluoro-17,21-dihydroxy-16-beta-methyl-pregna-4,9(11),diene,3, 20-dione. When optimal ratios of steroid and cyclodextrin were used, neovascularization was reduced to 13%, 26%, and 28% of untreated controls for the three steroids, respectively. Hydrocortisone-cyclodextrin drug pairs suppressed virtually all inflammatory cell infiltration (induced by endotoxin), whereas tetrahydrocortisol-cyclodextrin pairs only partially reduced inflammation. These results demonstrate that corneal neovascularization and corneal inflammation are separable processes and that the neovascularization may be treated specifically using angiostatic steroids without inflammatory activity.

We have previously described corneal neovascularization (CNV) induced by the intrastromal injection of interleukin-2 (IL-2) in inbred mice. Photodynamic therapy (PDT), administered by a deeply penetrating 630 nm fiberoptic laser, can destroy neoplasms and their associated neovascularization with some selectivity, but can damage neighboring tissues when used for CNV. We performed PDT with a 514 nm ophthalmic argon laser in an attempt to induce regression of CNV and reduce the associated toxicity. Eight weeks following IL-2 injection, mice with CNV were injected i.v. with dihematoporphyrin ether (DHE). Seventy-two hours later, 11 eyes (group I) were irradiated with eight 800 mW, 1000 mu, 2 s spots. Controls included 11 vascularized corneas from mice that received DHE but no laser (group II), 11 that received laser but no DHE (group III), and 35 untreated vascularized corneas (group IV). Comparison of the mean areas of CNV in groups I through IV pretreatment (6.0, 6.5, 6.7, and 7.6 mm2) and 12 weeks posttreatment (4.3, 6.3, 5.6, and 7.5 mm2) revealed that a significant decrease was seen in group I only (p less than 0.04, ANOVA). Complications in group I included blepharitis (9%) and iris damage (18%). Histologic studies revealed no evidence of posterior segment damage. PDT with the 514 nm laser is safe and efficacious for the treatment of IL-2-induced CNV in this model.

The process of corneal neovascularization induced by alkali burns was periodically observed with a newly developed macroscope. The central corneas were burned using filter discs measuring 6 mm in diameter that had been immersed in 1 N NaOH. At 0, 1, 3 and 7 days and at 2, 3 and 4 weeks after injury, the corneas were observed with the macroscope and then examined histologically. At 1 day post-burn, the limbal vascular plexus was engorged but no new vessel formation was detected. By 3 days, many vascular sprouts had arisen from the limbal vascular arcade. At 7 days, the vascular sprouts grew and became fine new vessels. At 2 weeks, the new vessels lengthened further to the central cornea. At 3 weeks, trunk vessels extended and branched like a vascular tree. Blood in the trunk vessels appeared to flow slowly to and fro. The ends of the vessels swelled in a fusiform shape on the application of slight pressure of the macroscope probe. Histological examination revealed that the ends of the vessels consisted of single vascular endothelial cells and the trunk vessels were covered by pericytes. By 4 weeks, the branch vessels around the burned lesion had degenerated and collapsed. Thus, our in vivo study using the new macroscope not only clarified the process of corneal neovascularization from the early to the regressive phases but also provided some valuable new information.

Antagonists of leukotriene D4 (LTD4), 3-[7-(2-n-propyl-3-hydroxy-4-acetyl- phenoxy)-propoxy-2-methyl-3,4-dihydro-4-8-n-propyl-2H-1-benzopyran- 2-yl]-propionic acid (SC-39070) and 7-[3-(4-acetyl-3-hydroxy-2-propylphenoxy)-2-hydroxy- propoxy]-4-methyl- 2-oxo-2H-1-benzopyran-3-acetic acid (FPL-60319) were locally administered to eyes of rabbits with immunogenic keratitis. The period of circular leukocyte infiltration (Wessely's ring) in the cornea was markedly decreased. Edema and neovascularization of the cornea were not influenced.