[Long-term surgical results of initial trabeculotomy combined with sinusotomy performed inferiorly].

PURPOSE: To evaluate retrospectively the long-term effects of initial trabeculotomy combined with sinusotomy performed inferiorly. PATIENTS AND METHOD: Enrolled were 128 eyes of 100 patients who received initial glaucoma surgery. In 36 eyes, the removal of Schlemm's canal endothelium was also performed (removed group). The results were compared with the intact group RESULTS: In the primary open angle glaucoma (POAG), mean intraocular pressure (IOP) at 3 years after surgery was 14.6 (intact) and 15.4 mmHg (removed). Kaplan-Meier life-table analysis showed that qualified success rates for the intact group at 8 years were 62.2% and for the removed group at 5 years 45.2% defined by 20 mmHg or lower. The results in developmental glaucoma (DG) were similar to those in POAG. No statistical differences in postoperative IOP between the intact and removed groups were seen in either POAG or DG. In exfoliation glaucoma (XFG), mean IOPs for the intact group at 3 years were 17.3 mmHg and for the removed group at 2 years 15.4 mmHg. The success rates for the intact group at 3.5 years were 25.2% and for the removed group at 4.5 years 64.3%. The results in the intact group were worse than in the POAG patients. Although visual disturbance was seen in 13% of the patients, the major cause was the progression of the cataracts. CONCLUSIONS: The long-term results were the same as those of previous reports on surgery performed superiorly, including the frequency of visual disturbance. However the removal of Schlemm's canal endothelium is necessary in XFG for better IOP control.

Intraocular pressure (IOP) reduction by latanoprost in Japanese normal tension glaucoma patients over a five-year period stratified by presenting IOP.

PURPOSE: We examined the effectiveness of latanoprost for reducing intraocular pressure (IOP) in Japanese patients with normal tension glaucoma (NTG) over a 5-year period. DESIGN: Prospective interventional case series. The patients were classified into 2 groups based on mean IOP. METHODS: A total of 38 patients with NTG were studied after being classified into the high-tension (mean IOP 16 mmHg or greater, n = 27) and low-tension (mean IOP lower than 15 mmHg, n = 11) groups. IOP was measured and Humphrey Field Analyzer (HFA) examinations were conducted at 6, 12, 24, 36, 48, and 60 months after beginning a daily administration of latanoprost. RESULTS: Mean IOP before administration was 17.6 mmHg in the high-tension group, which was reduced to 13.9, 14.6, 14.4, 14.1, 13.6, and 14.6 mmHg at 6, 12, 24, 36, 48, and 60 months, respectively, after beginning administration. That in the low-tension group was 13.6 mmHg before administration, and then was reduced to 12.2, 11.4, 11.5, 12.5, 10.5, and 11.5 mmHg, respectively, after beginning administration was noted. Mean deviation (MD) values in the HFA examinations were reduced by -4.27 and -1.49 dB after 5 years in the high- and low-tension groups, respectively. CONCLUSIONS: Latanoprost administration was effective in reducing IOP over a 5-year period in a range of 3.1-4.1 and 1.3-3.6 mmHg in NTG patients with high- and low-tension levels, respectively. In addition, our results indicate that latanoprost helped to prevent a decrease in MD values in both groups, as shown by the results of HFA examinations.

Implication of the hypoxia response element of the Vegf promoter in mouse models of retinal and choroidal neovascularization, but not retinal vascular development.

Retinal neovascularization (NV) and macular edema, resulting from blood-retinal barrier (BRB) breakdown, are major causes of visual loss in ischemic retinopathies. Choroidal NV (CNV) occurs in diseases of the retinal pigmented epithelium/Bruch's membrane complex and is another extremely prevalent cause of visual loss. We used mice in which the hypoxia response element (HRE) is deleted from the vascular endothelial growth factor (vegf) promoter (Vegf(delta/delta) mice) to explore the role of induction of VEGF through the HRE in these disease processes. Compared to wild type (Vegf+/+) mice with oxygen-induced ischemic retinopathy (OIR) in which vegf mRNA levels were increased and prominent retinal NV and BRB breakdown occurred, Vegf(delta/delta) littermates with OIR failed to increase vegf mRNA levels in the retina and had significantly less retinal NV and BRB breakdown, but showed prominent dilation of some superficial retinal vessels. Vegf(+/delta) littermates with ischemic retinopathy developed comparable retinal NV to Vegf+/+ mice, exhibited intermediate levels of BRB breakdown, and did not show vasodilation. In a mouse model of CNV, due to laser-induced rupture of Bruch's membrane, the area of CNV at Bruch's membrane rupture sites was more than tenfold greater in Vegf+/+ mice than in Vegf(delta/delta) littermates. In contrast to these dramatic differences in pathologic ocular NV, Vegf(delta/delta) mice showed subtle differences in retinal vascular development compared to Vegf+/+ mice; it was slightly delayed, but otherwise normal. These data suggest that induction of VEGF through the HRE in its promoter is critical for retinal and CNV, but not for retinal vascular development.

Different effects of angiopoietin-2 in different vascular beds: new vessels are most sensitive.

In this study, we used double transgenic mice with inducible expression of angiopoietin-2 (Ang2) to investigate the role of Ang2 in the retinal and choroidal circulations and in three models of ocular neovascularization (NV). Mice with induced expression of Ang2 ubiquitously, or specifically in the retina, survived and appeared grossly normal. They also had normal-appearing retinal and choroidal circulations, demonstrating that high levels of Ang2 did not induce regression of mature retinal or choroidal vessels. When Ang2 expression was induced soon after birth, there was increased density of the deep capillary bed on postnatal day (P) 11 that returned to normal by P18, the time that retinal vascular development is usually completed. In mice with ischemic retinopathy, induction of Ang2 during the ischemic period resulted in a significant increase in retinal NV, but induction of Ang2 at a later time point when ischemia (and vascular endothelial growth factor [VEGF]) was less, hastened regression of NV. In triple transgenic mice that coexpressed VEGF and Ang2, the increased expression of Ang2 inhibited VEGF-induced NV in the retina. Increased expression of Ang2 also resulted in regression of choroidal neovascularization. These data suggest that ocular neovascularization, but not mature retinal or choroidal vessels, is sensitive to Ang2; a high Ang2/VEGF ratio promotes regression, while high Ang2 in the setting of hypoxia and/or concomitantly high Ang2 and VEGF stimulate neovascularization.

Angiopoietin 1 prevents retinal detachment in an aggressive model of proliferative retinopathy, but has no effect on established neovascularization.

Vascular endothelial growth factor (VEGF) plays a central role in vasoproliferative diseases in the retina, however, other gene products modulate its effects. The angiopoietins are particularly important in this regard. Angiopoietin 2 (Ang2) collaborates with VEGF to stimulate neovascularization (NV) in some situations, but in other situations causes regression of NV. Ang2 also causes a transient increase in vascular density during retinal vascular development. In this study, we sought to determine if Ang1 has similar activities. The effects of Ang1 were tested in double transgenic mice with inducible expression of Ang1. Increased expression of Ang1 in the retina during retinal vascular development did not cause a detectable alteration in vascular density. Also, unlike Ang2, increased expression of Ang1 had no effect on established retinal or choroidal NV. However, when Ang1 expression was initiated simultaneously with that of VEGF, it strongly suppressed VEGF-induced NV and prevented retinal detachment. These data indicate that the timing of Ang1 expression is a critical determinate of its effects on VEGF-induced NV in the retina; it effectively blocks the initiation and progression of NV, but cannot reverse established NV or reduce leakage from NV. These data suggest that increased expression of Ang1 may be a good strategy for prophylaxis of retinal NV, but is unlikely to be effective as monotherapy of established NV.

Increased expression of VEGF in retinal pigmented epithelial cells is not sufficient to cause choroidal neovascularization.

Increased expression of vascular endothelial cell growth factor (VEGF) in the retina is sufficient to stimulate sprouting of neovascularization from the deep capillary bed of the retina, but not the superficial retinal capillaries or the choriocapillaris. Coexpression of VEGF and angiopoietin 2 (Ang2) results in sprouting of neovascularization from superficial and deep retinal capillaries, but not the choriocapillaris. However, retina-derived VEGF and Ang2 may not reach the choriocapillaris, because of tight junctions between retinal pigmented epithelial (RPE) cells. To eliminate this possible confounding factor, we used the human vitelliform macular dystrophy 2 (VMD2) promoter, an RPE-specific promoter, combined with the tetracycline-inducible promoter system, to generate double transgenic mice with inducible expression of VEGF in RPE cells. Adult mice with increased expression of VEGF in RPE cells had normal retinas and choroids with no choroidal neovascularization (CNV), but when increased expression of VEGF in RPE cells was combined with subretinal injection of a gutless adenoviral vector containing an expression construct for Ang2 (AGVAng2), CNV consistently occurred. In contrast, triple transgenic mice with induced expression of Ang2 and VEGF in RPE cells, did not develop CNV. These data suggest that increased expression of VEGF and/or Ang2 in RPE cells is not sufficient to cause CNV unless it is combined with a subretinal injection of a gutless adenoviral vector, which is likely to perturb RPE cells. These data also suggest that the effects of angiogenic proteins may vary among vascular beds, even those that are closely related, and, therefore, generalizations should be avoided.

Angiopoietin-2 enhances retinal vessel sensitivity to vascular endothelial growth factor.

Increased expression of vascular endothelial growth factor (VEGF) in the retina starting after postnatal day (P)7 results in neovascularization originating from deep retinal capillaries, but not those in the superficial capillary bed. Doxycycline was administered starting P0 to double transgenic mice with inducible expression of VEGF in the retina. These mice showed proliferation and dilation of superficial retinal capillaries, indicating that at this stage of development, the superficial capillaries are sensitive to the effects of VEGF. Angiopoietin-2 (Ang2) is expressed along the surface of the retina for several days after birth, but by P7 and later, Ang2 is only expressed in the region of the deep capillary bed. In mice with ubiquitous doxycycline-inducible expression of Ang2, in the absence of doxycycline, intravitreous injection of a gutless adenoviral vector expressing VEGF (AGV.VEGF) resulted in neovascularization of the cornea and iris, but no retinal neovascularization. After treatment with doxycycline to induce Ang2 expression, intravitreous injection of AGV.VEGF caused retinal neovascularization in addition to corneal and iris neovascularization. The retinal neovascularization originated from both the superficial and deep capillary beds. These data suggest that Ang2 promotes sensitivity to the angiogenic effects of VEGF in retinal vessels.

RPE cells modulate subretinal neovascularization, but do not cause regression in mice with sustained expression of VEGF.

PURPOSE: Previous studies using models of choroidal neovascularization (CNV) in which the angiogenic stimulus is not sustained, have concluded that the retinal pigmented epithelium (RPE) causes regression of neovascularization (NV). However, the withdrawal of angiogenic stimuli may actually be the major modulator of NV, and RPE cells may simply be responding to withdrawal of the angiogenic stimuli or something released by NV because of the withdrawal. In this study, the long-term course of NV and the behavior of the RPE in rhodopsin/VEGF transgenic mice, in which there is a sustained angiogenic stimulus, was investigated. METHODS: Hemizygous mice from the V-6 line were killed at 0.75, 1, 3, 6, and 12 months after birth, and at each time point mRNA for VEGF, VEGF-R1, and VEGF-R2 was measured by RT-PCR. Some mice were perfused with fluorescein-labeled dextran and retinal flatmounts were examined by fluorescence microscopy. Light and electron microscopy was performed on Epon-embedded eyes. RESULTS: The mRNA levels for VEGF, VEGF-R1, and VEGF-R2 remained constant from the earliest to the latest time point. Retinal flatmounts showed numerous small areas of subretinal NV at 3 weeks and at 1 month, and there were a similar number of larger lesions. By 6 months, many of the individual NV lesions had grown together to form large networks of new vessels. At 12 months, NV networks were similar to those at 6 months, but some of the vessels were not perfused. Light microscopy showed serous retinal detachments overlying NV lesions in mice up to 3 months of age, but at 6 and 12 months, the RPE completely surrounded new vessels and formed tight junctions to reestablish the outer blood-retinal barrier, and there were no serous detachments. Electron microscopy showed that compared with more acute NV lesions, chronic lesions contained thinner endothelial cells, similar to those of the choriocapillaris in that they had scant cytoplasm and numerous fenestrations, or pinocytotic vesicles with thick basement membrane surrounded by extracellular matrix (ECM). Bruch's membrane remained intact. CONCLUSIONS: Despite persistent high expression of VEGF and its receptors, NV stopped growing and reached a plateau in older V-6 mice. RPE cells modulated the NV by surrounding it and reestablishing the blood-retinal barrier, but did not cause regression, although some vessels in chronic lesions were not perfused. These data do not support the conclusion of several previously reported studies, that RPE cells cause regression of CNV.

Combretastatin A-4 phosphate suppresses development and induces regression of choroidal neovascularization.

PURPOSE: Combretastatin A-4 (CA-4) is a naturally occurring agent that binds tubulin and causes necrosis and shrinkage of tumors by damaging their blood vessels. In this study the effect of a CA-4 prodrug, combretastatin A-4-phosphate (CA-4-P), was tested in two models of ocular neovascularization. METHODS: The effect of CA-4-P was quantitatively assessed in transgenic mice with overexpression of vascular endothelial growth factor in the retina (rho/VEGF mice) and mice with choroidal neovascularization (CNV) due to laser-induced rupture of Bruch's membrane. RESULTS: In rho/VEGF mice, daily intraperitoneal injections of 4.0 mg/kg CA-4-P starting at postnatal day (P)7, the time of onset of transgene expression, resulted in a significant reduction in the number of neovascular lesions and total area of neovascularization per retina at P21, compared with vehicle-injected mice. In mice with laser-induced rupture of Bruch's membrane, daily intraperitoneal injections of 75 or 100 mg/kg CA-4-P resulted in a significant reduction in the area of CNV at rupture sites compared with vehicle-injected mice. In mice with established CNV, daily intraperitoneal injections of 100 mg/kg CA-4-P for 1 week resulted in a significant reduction in CNV area at rupture sites compared with the baseline area before treatment or the area of CNV in vehicle-treated mice. CONCLUSIONS: These data indicate that CA-4-P suppresses the development of VEGF-induced neovascularization in the retina and both blocks development and promotes regression of CNV. Therefore, CA-4-P shows potential for both prevention and treatment of ocular neovascularization.

A comparison of optic disc topographic parameters in patients with primary open angle glaucoma, normal tension glaucoma, and ocular hypertension.

BACKGROUND: Heidelberg Retina Tomograph (HRT) findings have been employed to quantitatively assess the topography of optic discs. We measured topographic parameters of optic discs in patients with primary open-angle glaucoma (POAG), normal-tension glaucoma (NTG), and ocular hypertension (OH) using an HRT in order to determine whether HRT topographic parameters can be used to differentiate those conditions. METHODS: Seventeen eyes in 17 patients with POAG, 23 eyes in 23 patients with NTG, and 15 eyes in 15 patients with OH were examined using an HRT, and the results were analyzed by age, refractive error, and topographic parameters. RESULTS: Among the HRT parameters, the mean values for rim area, rim volume, cup disk area ratio, and classification showed significant differences among POAG, NTG, and OH eyes. The mean values for cup area, cup volume, mean RNFL thickness, and RNFL cross section area showed significant differences between POAG and NTG eyes, and NTG and OH eyes, however, not between POAG and OH eyes. Cup shape measure showed significant differences between POAG and OH, and NTG and OH eyes, but not between POAG and NTG eyes. CONCLUSIONS: Our results suggest that POAG is distinguishable from NTG and OH based on evaluations of rim area and rim volume. Patients with NTG tend to have larger cupping, smaller rims, and thinner retinal nerve fiber layers as compared to POAG and OH patients. Thus, HRT topographic parameters are useful to differentiate patients with POAG, NTG, and OH.

Increased expression of brain-derived neurotrophic factor preserves retinal function and slows cell death from rhodopsin mutation or oxidative damage.

There are no effective treatments for inherited retinal degenerations, which are prevalent causes of visual disability. Several proteins promote the survival of various types of neurons, and increasing expression of one or more of these survival factors is a promising strategy for a new treatment. Studies examining the effects of intravitreous injections of brain-derived neurotrophic factor (BDNF) in models of inherited retinal degenerations have suggested that BDNF has little survival-promoting activity for photoreceptors. In this study, we generated double transgenic mice with doxycycline-inducible expression of BDNF in the retina. In a model of primary rod photoreceptor degeneration, expression of BDNF resulted in significant delay in photoreceptor cell death and maintenance of retinal function assessed by electroretinogram recordings. Expression of BDNF also caused strong protection of photoreceptors from oxidative damage-induced cell death. These data suggest that continuous expression of BDNF, unlike intravitreous injections, results in morphologic and functional benefit in animal models of inherited retinal degeneration. Double transgenic mice with inducible expression of survival factors provide valuable tools for selection of survival factor candidates for gene therapy.

Topical nepafenac inhibits ocular neovascularization.

PURPOSE: Topical nepafenac readily penetrates the cornea and is metabolized to amfenac, a potent cyclooxygenase (COX)-1 and COX-2 inhibitor. In this study, we tested the effect of topical nepafenac in three murine models of ocular neovascularization (NV). METHODS: A masked trial was performed to compare the topical effects of vehicle with one of several concentrations of nepafenac (0.01%, 0.03%, 0.1%, or 0.5%), 0.1% diclofenac, or 0.5% ketorolac tromethamine in mice with oxygen-induced ischemic retinopathy, mice with choroidal NV (CNV) due to laser-induced rupture of Bruch's membrane, or transgenic mice with increased expression of vascular endothelial growth factor (VEGF) in photoreceptors (rho/VEGF transgenic mice). RESULTS: Mice treated with 0.1% or 0.5% nepafenac had significantly less CNV and significant less ischemia-induced retinal NV than did vehicle-treated mice. Nepafenac also blunted the increase in VEGF mRNA in the retina induced by ischemia. In rho/VEGF transgenic mice, nepafenac failed to inhibit neovascularization. In additional studies, compared with vehicle-treated mice, mice treated with 0.1% or 0.03% nepafenac had significantly less CNV, whereas eyes treated with 0.1% diclofenac showed no significant difference. Mice treated with 0.5% ketorolac tromethamine for 14 days had high mortality, but when evaluated after 7 days of treatment showed no difference from mice treated with vehicle for 7 days. CONCLUSIONS: Topical nepafenac inhibits CNV and ischemia-induced retinal neovascularization by decreasing production of VEGF. The absence of effect in rho/VEGF transgenic mice is consistent with this mechanism. Topical nepafenac may provide an effective new treatment for ocular neovascularization. The excellent corneal penetration of nepafenac certainly plays an important role in this effect. It is possible that other antiangiogenic agents are also amenable to topical application after formulations are identified that maximize their corneal penetration. Because of the many advantages of the topical route of delivery, this is a possible topic for exploration.

Blockade of nitric-oxide synthase reduces choroidal neovascularization.

Nitric oxide (NO) promotes retinal and choroidal neovascularization, although different isoforms of nitric-oxide synthetase (NOS) are critical in each. Deficiency of endothelial NOS (eNOS) suppresses retinal but not choroidal neovascularization, whereas deficiency of neuronal NOS (nNOS) or inducible NOS (iNOS) suppresses choroidal, but not retinal neovascularization. In this study, we investigated the effect of N(G)-monomethyl-L-arginine (L-NMMA), a nonspecific NOS inhibitor, in three models of ocular neovascularization. Oral administration of L-NMMA caused significant inhibition of choroidal neovascularization in mice with laser-induced rupture of Bruch's membrane and significantly inhibited subretinal neovascularization in transgenic mice with expression of vascular endothelial growth factor (VEGF) in photoreceptors (rho/VEGF mice) but did not inhibit retinal neovascularization in mice with ischemic retinopathy. By extensive mating among mice deficient in NOS isoforms, triple homozygous mutant mice deficient in all three NOS isoforms were produced. These mice had marked suppression of choroidal neovascularization at sites of rupture of Bruch's membrane and near-complete suppression of subretinal neovascularization in rho/VEGF mice but showed no difference in ischemia-induced retinal neovascularization compared with wild-type mice. These data indicate that NO is an important stimulator of choroidal neovascularization and that reduction of NO by pharmacologic or genetic means is a good treatment strategy. However, the situation is more complex for ischemia-induced retinal neovascularization for which NO produced in endothelial cells by eNOS is stimulatory, but NO produced in other retinal cells by iNOS and/or nNOS is inhibitory. Selective inhibitors of eNOS may be needed for treatment of retinal neovascularization.

Caspase-3 inhibitor transiently delays inherited retinal degeneration in C3H mice carrying the rd gene.

BACKGROUND: The effect of a caspase-3 inhibitor on retinal degeneration in C3H mice carrying the rd gene, a mutation of a rod-specific phosphodiesterase, was investigated. METHODS: A quantity of 2 mg/kg of Ac-DEVD-CHO, as inhibitor, was injected intraperitoneally every other day from 8 days of age, and retinal damage was compared with that in saline-treated C3H mice at 13 days (1 day after the third treatment) and 17 days of age (1 day after the fifth treatment). Retina of ICR mice not carrying rd gene was also evaluated under the same protocol. The efficacy of Ac-DEVD-CHO was evaluated based on total retinal thickness and outer retinal thickness (thickness of outer nuclear layer and photoreceptor layer). An apoptotic index and a cell proliferation index for the photoreceptor cells, at 13 days of age, were calculated based on terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling (TUNEL) and proliferating cell nuclear antigen (PCNA) labeling, respectively. RESULTS: At 13 days of age, total and outer retinal thickness in saline-treated C3H mice were 140.3 microm and 37.5 microm, compared with 160.4 microm and 49.5 microm, respectively, in Ac-DEVD-CHO-treated C3H mice ( P<0.01, respectively). In ICR mice, total and outer retinal thickness were 182.1 microm and 90.9 microm, respectively, in saline-treated mice and 183.8 microm and 89.6 microm in Ac-DEVD-CHO-treated mice (not significant). At this time, the TUNEL index was 23.52 cells/10(4) microm (2) of outer nuclear layer in saline-treated C3H mice; Ac-DEVD-CHO treatment significantly reduced this value to 18.73 cells/10(4) microm(2) ( P<0.05). The TUNEL index in saline- and Ac-DEVD-CHO-treated ICR mice was 0.59 cells/10(4) microm(2) and 0.80 cells/10(4) microm(2), respectively (not significant); Ac-DEVD-CHO treatment had no influence on normally developing retina. The PCNA index was not affected by Ac-DEVD-CHO-treatment. However, at 17 days of age, Ac-DEVD-CHO treatment did not ameliorate retinal degeneration. CONCLUSIONS: The caspase-3 inhibitor was transiently effective in delaying retinal degeneration through inhibition of the apoptosis of photoreceptor cells in rd gene-carrying mice. The use of caspase-3 inhibitors may have therapeutic applications in the treatment of human retinal degeneration.