RPE Cell Apoptosis by the Combination Treatment of SAHA and Lactacystin

PURPOSE: To establish a new therapeutic strategy for proliferative vitreoretinopathhy (PVR), we examined the effect of combined treatment with HDAC inhibitor SAHA and proteasome inhibitor lactacystin in human retinal pigment epithelial (RPE) cells, ARPE-19. METHODS: Viability was determined by trypan blue exclusion assay. Mitochondrial membrane potential (MMP) was measured by flow cytometry. Proteasome activity was measured by fluorophotometry. The expression and degradation of apoptosis-related proteins were assesssed by Western blotting. Subcellular location of apoptosis-related factors was monitored by confocal miscroscopy. RESULTS: A single treatment with 5 micro M SAHA or 10 micro M lactacystin did not reduce cell viability. However, combination treatment with 5 micro M SAHA and 10 micro M lactacystin substantially reduced the viability, because the mixture induced the reduction of MMP and nuclear condensation or fragmentation. Moreover, the combination treatment triggered the activation of caspase-3 and the production of PARP cleavage products. These data indicate that the combination treatment efficiently induces apoptosis in ARPE-19 cells. However, co-treatment of SAHA did not augment the proteasome inhibitory activity of lactacystin, nor did co-treatment of lactacystin augment acetylation of histones. It is notable that while p53 and CAD were observed in the mitochondria of cells treated with SAHA, they were translocated into the nucleus after the combination treatment. CONCLUSIONS: These results suggest that the combination treatment of SAHA and lactacystin effectively induced apoptosis in ARPE-19 cells. Further work is warranted to develop this combination therapy as a novel therapeutic strategy for PVR.

Comparison of 2-Octylcyanoacrylate and Suture with 8-0 Polyglactin for Conjunctival Wound Closure in Rabbits

PURPOSE: To evaluate the clinical efficacy and the histopathologic tolerance of 2-octylcyanoacrylate versus 8-0 polyglactin sutures for conjunctival wound closure in rabbits. METHODS: We performed an experimental study on 16 eyes of eight New Zealand albino rabbits. Conjunctival peritomy of 8 mm was done in the superior limbus followed by extensive subconjunctival dissection in both eyes. The eyes of the rabbits were divided into two groups. The conjunctiva was then attached to the limbus again by the proposed 2-octylcyanoacrylate adhesive (left eye, Group A) or 8-0 polyglactin suture (right eye, Group B). The clinical efficacy of the closure of the conjunctival wound, either with sutures or adhesives, and the clinical and histopathologic tolerances such as hyperemia, discharge, residual adhesive or suture, inflammatory change and fibrosis were observed at 1 day and at 1, 2, 4 and 6 weeks after surgery. Two rabbits were sacrificed at each of 1, 2, 4 and 6 weeks, and specimens of their conjunctiva were examined histologically. RESULTS: Both conjunctival surgical closure methods were found to be equally efficacious in fixing the conjunctiva to the limbus. There were no significant clinical or histopathologic tolerance differences between the two groups. The conjunctiva at 6 weeks of Group A showed nearly normalized morphology with disappeared inflammatory cells and fibroblasts, whereas those of Group B showed foreign body inflammatory reaction around the absorbing suture materials. CONCLUSIONS: 2-Octylcyanoacrylate adhesive was an efficient conjunctival closure method and was very well tolerated by the ocular surface. 2-Octylcyanoacrylate may be a possible tissue adhesive for conjunctival wound closure.

Clinical Observation of Combined Trabeculotomy and Trabeculectomy on Congenital Glaucoma

PURPOSE: To investigate the result of combined trabeculotomy and trabeculectomy as an initial operative treatment of congenital glaucoma. METHODS: We retrospectively reviewed the medical records of 8 patients (14 eyes) who were operated with combined trabeculotomy and trabeculectomy as an initial operative treatment of congenital glaucoma from October 13, 1998 to February 20, 2001. The mean age of patients was 1.2 years (ranging from 0.75 to 2 years) and the mean follow-up period was 12.4 months (12.3+/-8.18). We assessed the mean age at surgery, preoperative corneal diameter, preoperative and postoperative intraocular pressure, preoperative and postoperative C/D ratio, postoperative complications. RESULTS: The mean age at first surgery was 4.4 months (4.3+/-2.45), the mean postoperative intraocular pressure (IOP) at the end of follow-up period was 13.2+/-2.01mmHg, significantly lower than the mean preoperative IOP which was 25+/-5.23mmHg (P<0.05), the mean postoperative C/D ratio at the end of follow-up period was 0.38+/-0.14, significantly lower than the mean preoperative C/D ratio which was 0.55+/-0.15 (P<0.05), C/D ratio of 3 eyes among them was decreased by more than 0.2. The overall success rate was 71.4% (10 eyes) after single procedure. All the 4 failed eyes underwent trabeculectomy again and the IOP was well controled. The complications included hyphema(5 eyes), corneal edema (3 eyes), vitreous prolapse (3 eyes), posterior synechiae (2 eyes), and they were all resolved spontaneously. CONCLUSIONS: The combined surgery of trabeculotomy and trabeculectomy as an initial single procedure of congenital glaucoma lowered the IOP without serious complications.

The Effect of Amniotic Membrane Ointment on Prevention of Adhesion in Strabismus Surgery in Rabbits

PURPOSE: The authors investigated the anti-inflammatory and the anti-fibrotic effect of amniotic membrane ointment on preventing adhesion in strabismus surgery in the rabbits. METHODS: Recession of superior rectus muscle was performed in 64 eyes of 32 rabbits and then the 32 rabbits were divided into 4 groups each 16 eyes of 8 rabbits. control group received no more treatment. 0.04% mitomycin C, base ointment, and amniotic membrane ointment were applied to the other trial groups respectively. Histopathologic studies was performed in 3 days, 2 weeks, 4 weeks, 12 weeks and evaluated the change of inflammation and fibrosis. The degree of inflammatory cell infiltration was evaluated on H and E stain and the degree of fibrosis was evaluated on Masson-trichrome stain. An immunohistochemical stain for alpha-smooth muscle actin was done for detection of myofibroblasts. RESULTS: On postoperation 3 days, 2 weeks, 4 weeks, the amniotic membrane ointment group and the mitomycin C group showed less inflammatory cell infiltration than control group(P<0.05). Masson-trichrome stain showed less fibrosis in the amniotic membrane ointment group and the mitomycin C group. The degree of immunohistochemical stain for alpha-smooth muscle actin was observed to be lower in the mitomycin C group and amniotic membrane ointment group than in the other groups during all period of observation. CONCLUSIONS: Amniotic membrane ointment might be effective adjuvant for prevention of postoperative adhesion in strabismus surgery.

The Effects of Amniotic Membrane Ointment on Corneal Alkali Burn in Rabbits

PURPOSE: The therapeutic effects of amniotic membrane transplantation (AMT) contain anti-fibrosis, anti-inflammation and facilitation of epithelialization. Thus we investigated whether applying of amniotic membrane (AM) ointment could get the same effect as AMT. METHODS: Alkali burn was induced by applying 6 mm round filter paper which was soaked with 1 N NaOH, onto the central cornea for 60 seconds on both eyes of 10 white rabbits. Then we applied AM ointment on one eye and base ointment on the other eye, four times a day for 1 week. The corneas were harvested for histopathologic studies at 1 day and 3 weeks. Corneal opacity and thickness were measured in 3 days and 1, 2, 3 weeks. RESULTS: In the AM ointment applicated group, the number of the infiltrated PMNs (polymorphonuclear cells) and apoptotic keratocytes were significantly less than that of the control group (p< 0.05). The degree of lipid peroxidation and myofibroblast differentiation were less than those of the control group. Corneal opacity and corneal edema were less in AM ointment group than control group. CONCLUSIONS: AM ointment application after alkali burn is beneficial to reduce inflammation, keratocyte apoptosis and lipid peroxidation, and is considered to suppress corneal haze by these effects. Therefore, this report may be a basic study for the AM ointment research to treat recalcitrant keratitis.

The Effects of Amniotic Membrane for Prevention of Adhesion in Strabismus Surgery in Rabbits

PURPOSE: The formation of adhesion following strabismus surgery resulted in restrictive ocular motility dysfunction, recurrence, and failure of adjustable strabismus surgery. From these results, authors used amniotic membrane and evaluated its effect for prevention of adhesion in strabismus surgery in rabbit eyes. METHODS: Recession of superior rectus muscle was performed in 48 eyes of 32 rabbits. In 16 rabbits, both eyes were operated. Sixteen left eyes of 16 rabbits, control group, received no treatment after operation. In 16 right eyes of 16 rabbits, a sponge soaked in mitomycin C was applied after recession of superior rectus muscle. In 16 right eyes of other 16 rabbits, amniotic membrane transplantation was performed after procedure. After operation, 4 rabbits were sacrified in 24 hours, 2 weeks, 4 weeks, 3 months and evaluated the change of inflammation & fibrosis. The degree of inflammatory cell infiltration was evaluated on H & E stain and Masson-trichrome stain was performed to evaluate the degree of fibrosis. An immunohistochemical stain for alpha-smooth muscle actin was done for detection of myofibroblasts. RESULTS: On post-operation 24 hours and 2 weeks, the amniotic membrane-used group and the mitomycin C-used group showed less inflammatory cell infiltration than control group. On post-operation 2 weeks, 4weeks, and 3 months, amniotic membrane-used group and the mitomycin C-used group showed less fibrosis and fibroblast proliferation than control group (P<0.05). CONCLUSIONS: From these findings, authors conclude that the use of amniotic membrane transplantation after strabismus surgery is an effective method for prevention of postoperative adhesion.

The Effect of ECMs on MHC Class II Molecule Expression in Retinal Pigment Epithelium

Expression of MHC class II molecule by retinal pigment epithelial cells and the interaction of the cell with extracellular matrix molecules involve in the pathogenesis of proliferative vitreoretinopathy and chorioretinitis. In this study interferon-gammainduced the expression of MHC class II molecules on RPEJ cells. Extracellular matrix molecules increased the interferon-gamma-i n d u c e d expression of MHC class II molecules. TGF-beta2 inhibited the interferon-gammainduced expression of MHC class II molecules. However, there was no significant effect on such inhibitory function according to the types of extracellular matrix molecules. Blocking the autocrine effect of TGF-beta2 by the specific antisense oligonucleotides decreased its inhibitory function. PLC-gamma1-specific antisense oligonucleotide inhibited the effect of TGF-beta2, which suggests that PLC-gamma1 involves in the signal transduction of TGF-beta2 on the expression of MHC class II molecules. In conclusion, the present study provides further understandings to the previous knowledge of pathogenesis of immunologic retinal disorders.

Comparison of Histologic Changes According to Treatment Method in Experimentally Perforated Rabbit Eyes

Perforation of the choroid and retina during strabismus surgery is fairly common and optimal management of global perforation is contoversial. The authorsinvestigated the histological changes of the rabbit eyes to compare the treatment effects of diode laser photocoagulation, cryotreatment, and nontreatment in experimentally perforated rabbit eyes.We perforated the globe of 6 pigmented rabbits (12 eyes)and treated four eyes with diode laser and four with cryotherapy and did not treat the remaining four.Proliferation of stromal cells and chorioretinal adhesion were seen in untreated eyes. The cryotherapy eyes revealed retinal destruction and pigment-laden cells in all retinal layers.The eyes treated by diode laser showed firm chorioretinal adhesion and relatively well preserved inner layers of retina.We concluded that directed diode laser photocoagulation was a safe and effective method of retinopexy to prevent the retinal detachment after globe perforation durng strabismus surgery.

Effect of Experimentally Induced Anisometropia on Binocular Vision

We investigated the effect of experimentally induced anisometropia on binocular function. Unilateral myopia, hyperopia, or astigmatism(Axis 90degrees) was induced in eighteen adults with normal binocular function using softcontact lenses. Sensory status was assessed by measuring stereoacuity, Worth four dot test, and Bagolini lens test. Binocular function of all subjects decreased with increasing levels of anisometropia. In Worth four dot test, foveal suppression increased in proportion to the levels of anisometropia. Stereoacuity was similarly degraded by the induced anisometropia. Bagolinilens responses were normal in almost all subjects. Our study suggest that experimentally induced anisometropia cause significant abnormalities in binocular function in adults.

Effects of Topical 2% Dorzolamide(Trusopt(R)) in Q-switched Nd:YAG Laser Induced Glaucoma Rabbits

The purpose of this study is to evaluate the effects of 2% dorzolamide(Trusopt(R)) on the ocular blood flow and retinal microcirculation. To creat an experimental glaucoma model in rabbits and to study the effects of elevated intraocular pressure(IOP), ocular blood flow, retinal effects of elevated intraocular pressure(IOp), ocular blood flow, retinal microcirculation on rabbits eyes, we treated trabecular meshwork of 6 adult pigmented rabbits with Q-switched Nd;YAG laser. And then we investigated the IOP lowering effect, ocular blood flow, and the microcirculation on retina of 2% dorzolamide(Trusopt(R)) in experimental glaucoma model. The IOP, ocular blood flow and the microcirculation were measured with applanation pneumotonography(Alcon, Texas), pneumotonometric probe linked to Langham ocular blood foow system(OBF, Blue mountain) and Heidelberg Retina Flowmeter(HRF). During sustained IOP elevation, 2% dorzolamide(Trusopt(R)) was instilled in one eye and normal saline in the fellow eye. The IOP and ocular blood flow were measured 1, 2, 4 and 8 hours after instillation. The retinal microcirculation was measured 2 and 8 hours after instillation. The retinal microcirculation was measured 2 and 8 hours after instillation at 200micrometer apart from the superior optic disc margin. There were statistically significant reductions in IOP in both 2% dorzolamide(Trusopt(R)) and normal saline-treated eyes(P0/1). From the above results, we concluded that 2% dorzolamide(Trusopt(R)) reduced the IOP but did not alter ocular blood flow and peripapillary retinal microcirculation.

Clinical Significance of Nasal Step Index in Early Glaucoma

Nasal step index(NSI) is an index of light sensitivity of the nasal step area, the early sign of visual field defects in glaucoma. It represents the asymmetry between the upper-nasal and lower-nasal quadrant within 30 degree field. We applied this index to 66 eyes with normal visual field and 204 eyes with mild-to-moderate glaucomatous visual field defect (mean defect(MD)or=1.04), respectively. AUC values related to the validity of this study were NSI-1:0.38, NSI-2;0.44 and MD;0.83 in early glaucoma patients. But AUC values in high NFBI patients were NSI -1;0.50, NSI -2;0.50, MD;0.82. Therefore, NSI is shown to have lesser diagnostic value than MD in detection of early glaucoma in this study.

The Effect of Endothelin-1 on IOP and Aqueous Outflow Facility in the Rabbit Eye

Endothelin(ET) is known as a family of potent hydrophobic, vasoactive peptide. We investigated the effect according to the concentrations of ET on intraocular pressure(IOP), aqueous outflow facility, pupillary diameter and light reflex and iris vessels. Twenty-four hours after injection of 2.5 microgram and 10 microgram of ET-1 into the anteriorvitreous of rabbit eyes, the IOP was reduced by 69% and 80%, respectively and did not return to the level of prefreatment until at least 14 days and 20 days, respectively. But the decrease of IOP was not due to the increased aqueous outflow. The pupillary diameter of ET-1 treated eyes was 1 to 2mm larger than the pretreatment. The time course of the pupillary effects generally ran paralled with the reduction of IOP. The iridial and conjunctival hyperemia was detectable during the pupillary dilatation. Endothelins are therefore potential participants in the local regulation of IOP, ocular blood vessel tone, and iris smooth muscle tone.

The Effects of Mitomycin C on Trabeculectomy

Mitomycin C (MMC) is an antimetabolite, antibiotic which prevents the proliferation of fibroblast and fibrosis by inhibition of DNA-dependent RNA synthesis. It has been used to increase the success rate of filtering surgery. We performed trabeculectomy and peripheral iridectomy with 0.4 mg/ml MMC application for 3 minutes in poor prognostic glaucoma patient, 26 eyes of 24 patients from February 1992 to December 1994. The postoperative change of intraocular pressure (IOP) and complications were followed up from 3 to 30 months (mean 12.4 months). The preoperative and postoperative mean IOP after 24 months were 32.3 +/- 11.4 mmHg and 12.7 +/- 3.1 mmHg, respectively. The decrease rate of IOP was 60.7% and the success rate was 71.7%. The postoperative complications were 15 eyes including 5 eyes of shallow anterior chamber, 5 eyes of hyphema, 2 eyes of conjunctival wound leakage, one eye of bleb rupture, one eye of choroidal detachment, and one eye of cataract formation.

The Cycloplegic Effect of Atropine

Atropine is known as one of the strongest cycloplegics,But its side-effects often limits its use only to the patient of strabismus and to the children below five. The purpose of this study is to find a way to apply atropine with reduced side effects. We examined twenty children below 6 years of age who are commonly considered to have strong hyperopic refractive capability or squints. Their eyes were examined 60 minutes and 90 minutes after administring a drop of atropine twice with five minutes' interval. For three days afterwards, Atropine was applied three times a day. And the results was examined with Cannon Autorefractometer. Refractive error after the single dose application(90 minutes) showed more hyperopia by +0.2 diopter than conventional three day application(P>0.05), and it has no statistical significance between the measurement after 60 and 90 minutet(P>0.15). It was also found that twenty nine eyes(72.5%) showed less than 0.5 diopter difference between both methods. No central nervous toxicity was found in both applications, while peripheral toxicity was found less in the single-dose application(10%) than conventional three-day application(30%). So for the practical purpose the refractive data evaluated after 90 minutes atropinization can be used as a basis for the prescription of glasses.

Study for Reversibility on Myopathy of Extraocular Muscle after Subconjunctival Injection of Gentamicin(R) in Rabbit

Subconjunctival injection of gentamicin(R)(GM) can cause an acute myopathy of the extraocular muscles. The purpose of this study is to demonstrate the effect of various dosage of GM and reversibility of tissue changes after subconjunctival injection of GM with 10m/ml, 20mg/ml and 40mg/ml each rabbit respectively. We obtained the superior rectus at 1 day, 1 week, 4 weeks and 12 weeks after injection and examined the specimens by light and electron microscopes. On 1 day, light microscopy showed inflammatory change with polymorphonuclear leukocytes and monocytes on 1 week. The higher the concentration of GM was, the more marked inflammation and degeneration of the muscle fibers were. The control group was normal. On 4 weeks there were markedly regenerated muscle fibers with focal degeneration. On 12 weeks there were nearly normal finding of the muscle with some degeneration. On 1 day and 1 week, electron microscopy showed various degrees of degeneration of the muscle fibers. On 4 and 12 weeks the muscle fiber showed satellite nucleus, deposit of abundant Golgi networks, increased numbers and enlargement of mitochondria with some degeneration. On early stage of injecton with normal saline, some vacuoles were seen in the control group. Our results suggest that gentamicin-induced myopathy is dose-related and reversible in histological aspect.

The Effect of 0.1% Topical Bunazosin(R) on the lOP of Normal Subjeets

The effects of topically applied bunazosin hydrochloride, Which has been developed as a highly selective alpha-1 adrenergic antagonist, on intraocular pressure(IOP), pupillary diameter, conjunctival injection, ptosis, and blood pressure were investigated in 14 normal volunteers. It is known that the mechanism of lOP reduction appears to be an increase in uveoscleral outflow and/or a decrease in episcleral venous pressure. The purpose of this experiment is to observe the effects of 0.1 % Bunazosin(R) on the lOP, pupil size, lid, conjunctival change and blood pressure in the normal human subject. The effects of 0.1% topical bunazosin(R) have been observed with placebo-controlled, double-blind single-dose study. The drug reduced lOP for 10 hours, and the decrease was statistically significant(p value <0.05). The lOP was decreased maximally at 3 hours after the instillation of the drug and the amount of the lOP reduction was 4.2mmHg. There were no significant changes of pupil size, lid configuration, conjunctival injection, and blood pressure in this experiment.

Effect of Topical Betoptic(R) and Mikelan(R) on the Perfusion Pressure

The effects of topical Betoptic(R) 0.5% and Mikelan(R) 2% on the ophthalmic artery blood pressure and perfusion pressure were investigated in 30 healthy subjects using ophthalmodynamometer and Goldmann applanation tonometer. The subjects were divided into two groups and each has 15 healthy subjects. In a randomized, double-blind study, one eye of each subject received one drop of Betoptic(R) 0.5% or Mikelan(R) 2%, and fellow eye was received a drop of placebo. Diastolic ophthalmic artery blood pressure (OABPd), systolic artery blood pressure (OPBPs), and intraocular pressure (lOP) were measured just before and 2 hours after the instillation of the drugs. Mean ophthalmic arery blood pressure (OABPm) was calculated as OABPm = OABPd + 1/3 (OABPs - OABPd) and the perfusion pressure (PP) was calculated as PP = OABPm - lOP. The perfusion pressure (pp) was Betoptic(R)-treated group was increased by 9%, and that of Mikelan(R)-treated group was increased by 13%, these increasement were statistically significant (Both group P<0.01).

Infiltrating Orbital Granular Cell Tumor: A case report

Granular cell tumor is an uncommon neoplasm with uncertain histogenesis that rarely occurs in the orbit. Although it is regarded as a benign lesion, one case of malignancy has been recorded. The authors present a case of recurred, infiltrating orbital granular cell tumor in a 39-year-old woman. On light microscopic examination, the tumor cells showed the characteristic findings of granular cell tumor, but there were hyperchromatic and pleomorphic nuclei with unusual mitotic figures, suggesting a malignant nature.

Effect of the Viscoelastic Substance on Surgery of the Extraocular Muscles in Rabbit: 1. Histopathologic Study by a Light Microscope

We evaluated histopathologically the effect of the viscoelastic substance on surgery of the extraocular muscle in the 26 rabbits with 1. 6-2. 5kg of weight. The viscoelastic substances were 1% sodium hyaluronate(Healon) and 2% hydroxypropyl methylcellulose(Methocel), which were introduced between the extraocular muscle and its adjacent tissue on surgery. The rabbits were categorized by group 1(the rabbits with Healon), group 2(with Methocel), and group 3(with normal saline as control). These rabbits were enuclated at 3rd, 7th, 14th, 28th and 56th day following operations, and the eyes were fixed in 2.5% glutaraldehyde solution. The tissue was stained with hematoxylin-eosin and Masson's trichrome, and examined under a light microscope. The results were as follows: 1. In the first group(Healon), the tissue adhesion and degeneration of the muscle were milder than those of control(saline) group. 2. The changes in second group(Methocel) was milder than in the first group and more severe than in control group. 3. It is concluded that 2% Methocel can be used clinically in the surgery of the extraocular muscle.