Role of inhibitors of isoprenylation in proliferation, phenotype and apoptosis of human retinal pigment epithelium.

BACKGROUND: Isoprenoid biosynthesis is known to be essential for diverse cellular functions, including cell proliferation. The aim of this work was to study the effects caused by the addition of different inhibitors of isoprenylation (lovastatin, manumycin A, farnesyltransferase inhibitor III and N-acetyl-S-farnesyl-L-cysteine) to human retinal pigment epithelium (RPE) in culture, as potential coadjunctive-to-surgery treatments applicable to proliferative vitreoretinopathy. METHODS: Human RPE cell cultures were established from adult corneal donors. Proliferation levels were evaluated using the incorporation of 5-bromo-2'-deoxyuridine into the DNA. Cell viability was measured by tetrazolium bromide transformation. Apoptosis was determined by DNA fragmentation assay, TdT-mediated d-UTP-X nick-end labeling (TUNEL) and phosphatidylserine exposure assessment. Changes in cell morphology and actin cytoskeleton were evaluated using a phase-contrast microscope and by fluorescent staining of actin cables with TRITC-phalloidin. RESULTS: We found that lovastatin showed an important antiproliferative effect on human RPE cells in culture. This effect was clearly dose-dependent, and adding mevalonate could reverse it. We also found that lovastatin induced changes in the distribution of actin cytoskeleton and, finally, that it also induced RPE apoptosis. Manumycin A, farnesyltransferase inhibitor III and N-acetyl-S-farnesyl-L-cysteine also showed antiproliferative effects in RPE. However, they do not have any effect on cell morphology or induction of apoptosis. DISCUSSION: We identified various effects of lovastatin on human RPE cultures: inhibition of cell proliferation, modifications of the phenotype and induction of apoptosis. Interestingly, the addition of different inhibitors of protein isoprenylation only affected the proliferation of the cells. There was no evidence that isoprenylated proteins inhibition is related to lovastatin-induced RPE apoptosis.

Retinitis pigmentaria / Retinitis pigmentosa

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Retinosis pigmentaria: resultados del análisis del gen de la rodopsina en la población gallega / Retinitis pigmentosa: results of rhodopsin gene analysis in the galician population

Objetivo: La retinosis pigmentaria es la degeneración retiniana hereditaria más frecuente. Las manifestaciones clínicas son variables en lo que se refiere a severidad, edad de inicio y progresión. Esta heterogeneidad clínica es paralela a la heterogeneidad genética (hasta el momento actual se han descrito más de 20 loci diferentes). Los objetivos de este trabajo fueron identificar mutaciones puntuales en el gen de la rodopsina y determinar las frecuencias de los diferentes tipos genéticos de retinosis pigmentaria en la población gallega. Métodos: Hemos estudiado a 47 pacientes, previamente diagnosticados de retinosis pigmentaria, y a sus familiares. Las diferentes formas genéticas se establecieron según los datos recogidos de la historia familiar y los derivados de la exploración clínica. Se realizó un estudio de screening, para la búsqueda de mutaciones puntuales en el gen de la rodopsina, mediante amplificación por reacción en cadena de la polimerasa, análisis por SSCPs y secuenciación directa en 36 pacientes no emparentados con RP no sindrómica. Resultados: Presentamos la distribución por frecuencias de las diferentes formas genéticas de la RP. En el análisis mediante SSCPs del gen de la rodopsina, encontramos diferentes patrones de movilidad: 1 variante en la región 5' no codificante del gen y 1 variante en el tercer intrón. La secuenciación directa demostró dos transiciones: A269®G y C3982®T, respectivamente. Ademas, observamos un cambio de base en el codón 160 (C®A) de este gen. Conclusiones: Los polimorfismos son hallazgos comunes en los exones 1 y 3 del gen de la rodopsina. Son variaciones neutras y no conllevan cambio en la proteína. No hemos encontrado diferencias significativas en la frecuencia de los polimorfismos A269®G y C3982®T entre los tres grupos de pacientes y los individuos normales. No hubo desviación significativa del equilibrio Hardy-Weinberg en ninguno de los grupos (AU)

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Changes in anterior chamber depth and angle width after filtration surgery: a quantitative study using ultrasound biomicroscopy.

PURPOSE: To evaluate the anterior chamber configuration by means of ultrasound biomicroscopy in patients with glaucoma and control subjects, and to determine quantitative changes in this configuration after glaucoma filtration surgery (trabeculectomy) and combined cataract and filtration surgery. METHODS: The study included 33 eyes of 33 patients with glaucoma (diagnosed with primary open-angle or exfoliative glaucoma) in which filtration surgery (n = 17) or combined cataract and filtration surgery (n = 16) was performed, and 25 eyes of 25 age-matched control subjects. Ultrasound biomicroscopy was used to measure anterior chamber depth and the angle width at 500 microm from the scleral spur in all eyes. The patients with glaucoma were examined 2 days before surgery and 3 and 6 months after surgery. RESULTS: There were no significant differences in anterior chamber depth and angle width between patients with glaucoma before surgery and control subjects. Postoperative values for anterior chamber depth were significantly greater in patients with glaucoma who underwent combined surgery, but no significant changes were observed in those who underwent filtration surgery alone. In contrast, angle width was significantly greater after surgery both in patients who underwent combined surgery and in those who underwent filtration surgery alone. CONCLUSION: On ultrasound biomicroscopic evaluation, there were no differences in anterior chamber depth and angle width between patients with glaucoma and control subjects. Trabeculectomy alone widens the angle but does not affect the anterior chamber depth; however, combined surgery both deepens the anterior chamber depth and widens the angle.

[Retinitis pigmentosa: results of rhodopsin gene analysis in the Galician population]. / Retinosis pigmentaria: resultados del análisis del gen de la rodopsina en la población gallega.

PURPOSE: Retinitis pigmentosa (RP) is the most prevalent inherited degeneration in the retina. The clinical manifestations are variable in terms of severity, age of onset and progression. The clinical variation is paralleled by genetic heterogeneity (more than 20 different loci have been described to date). The aim of this work was to identify mutations in rhodopsin gene and to determine the frequencies of the different genetic forms of RP in the Galician population. METHODS: 47 previously diagnosed RP patients and their relatives were studied. Genetic forms of RP were identified by recording full family history and clinical examination. DNA samples from patients with RP and control individuals were screened for point mutations in the rhodopsin gene by using PCR SSCPs (Single Strand Conformation Polymorphisms) and direct sequencing in 36 unrelated nonsyndromic RP patients. RESULTS: We report the frequency distribution of the different genetic RP forms. In the SSCPs analysis of rhodopsin gene we found different mobility shifts: one variant in the 5'-untranslated region of the gen and one variant in the third intron. Direct sequencing revealed an A269-->G and an C3982-->T transitions, respectively. Additionally, we observed a single base change in codon 160 (C-->A) of this gene. CONCLUSIONS: Polymorphisms are commom findings in the exon 1 and 3 of rhodopsin gene. They are neutral variations and do not represent a change in the protein. No significant differences in the frecuencies of A269-->G and C3982-->T polymorphisms among the three groups of RP patients (ADRP, ARRP, Esporadic RP) and normal individuals were found. There was no significant deviation from Hardy-Weinberg's equilibrium in each genotype in any group.

Ultrasound biomicroscopy in the diagnosis of supraciliochoroidal fluid after trabeculectomy.

PURPOSE: To report supraciliochoroidal effusion after trabeculectomy with the use of ultrasound biomicroscopy. METHODS: In a prospective study, 28 eyes of 19 patients with primary open-angle glaucoma that underwent trabeculectomy were evaluated preoperatively and postoperatively by ultrasound biomicroscopy. RESULTS: Four eyes showed a hypoechogenic suprachoroidal space that remained stable for 6 months postoperatively. These four eyes had intraocular pressures of 11 mm Hg or less on no antiglaucoma medications and without signs of choroidal detachment. CONCLUSION: Ultrasound biomicroscopy proved to be a useful method of detecting, after trabeculectomy, supraciliochoroidal fluid without clinically detectable choroidal detachment. This fluid may signify an iatrogenic cyclodialysis during surgery or, less likely, subclinical ciliochoroidal detachment.

Repair of scleral perforation with preserved scleral and amniotic membrane in Marfan's syndrome.

To describe the surgical technique used in the repair of a large scleral perforation in a patient with Marfan's syndrome and a past history of various surgical interventions in both eyes. Scleral homograft and amniotic membrane transplant were used to reconstruct the large scleral defect present in his left eye. One month after surgical intervention, the patient showed excellent restoration of the scleral perforation without signs of inflammation or infection. The combination of scleral homograft and amniotic membrane transplant constitute an effective alternative to autologous scleral and conjunctival grafts when these cannot be used.

Dorzolamide effect on ocular blood flow.

PURPOSE: To evaluate the effect of dorzolamide on ocular blood flow in normal and glaucomatous eyes. METHODS: Twenty-six eyes with documented open-angle glaucoma of 26 patients and 13 normal control eyes of 8 age-matched subjects were included in this study. All eyes underwent color Doppler imaging for measuring peak-systolic velocity, end-diastolic velocity, and resistance index in the ophthalmic and central retinal arteries and the maximal and minimal velocities in the central retinal vein. Eyes were grouped in control and initial and advanced glaucoma categories. Measurements were made in all groups before and after application of topical dorzolamide. Intragroup comparisons between baseline and dorzolamide conditions were made using paired Student's t-test. Intergroup comparisons under baseline conditions between normal and glaucomatous eyes were made by using the one-way ANOVA test. Statistical significance was set at P < 0.05. RESULTS: The peak-systolic velocity of the central retinal artery in glaucomatous eyes and the end-diastolic velocity of the ophthalmic and central retinal arteries in all groups were significantly higher after application of dorzolamide. The minimal velocity of the central retinal vein showed significantly higher values after dorzolamide, whereas the maximal velocity remained unchanged. The peak-systolic velocity of the ophthalmic artery in all groups and the peak-systolic velocity of the central retinal artery in normal eyes also remained unchanged. The resistance index was significantly lower in the ophthalmic and central retinal arteries in all groups after dorzolamide. The intraocular pressure was significantly reduced in all groups after dorzolamide. Under baseline conditions normal control eyes and glaucomatous eyes showed differences in various measurements. Peak-systolic velocity was significantly lower in glaucomatous eyes than in normal control eyes with the exception of the ophthalmic artery in the initial glaucoma group. End-diastolic velocity was lower in glaucomatous eyes than in control eyes in both arteries. Maximal and minimal velocities of the central retinal vein were lower in glaucomatous eyes than in normal control eyes. Resistance index was higher in glaucomatous eyes than in normal control eyes in the ophthalmic artery but not in the central retinal artery. CONCLUSIONS: Most hemodynamic parameters of intraocular and periocular vessels improve after application of topical dorzolamide in both normal control and glaucomatous eyes. Dorzolamide should be regarded as a useful drug for treatment of glaucoma not only because it reduces intraocular pressure but also because it improves the ocular blood supply.

C-C chemokines in the vitreous of patients with proliferative vitreoretinopathy and proliferative diabetic retinopathy.

PURPOSE: To investigate the presence of three C-C chemokines (monocyte chemotactic protein-1 [MCP-1], macrophage inflammatory protein-1alpha [MIP-1alpha], and MIP-1beta) in vitreous samples from eyes with proliferative vitreoretinopathy (PVR), proliferative diabetic retinopathy (PDR), or retinal detachment (RD). SUBJECTS AND METHODS: Vitreous samples were obtained and assayed by solid phase enzyme-linked immunosorbent assay for the chemokines MCP-1, MIP-1alpha, and MIP-1beta in a prospective study of 43 consecutive patients. Eighteen samples from cadaveric control eyes were also assayed. RESULTS: Monocyte chemotactic protein-1 was detected in all samples. The vitreous of eyes with vitreoretinal disorders showed significantly higher levels than the vitreous of cadaveric control eyes (P < or = 0.05). Median level (5th-95th percentile) in the PVR cases (n = 20) was 890.18 pg/mL (286.04-1806.20); in RD (n = 8), 296.69 pg/mL (171.44-1310.02); and in PDR (n = 15), 434.60 pg/mL (124.56-1092.94). In the cadaveric control eyes (n = 18), median level was 83.97 pg/mL (26.09-208.38). Macrophage inflammatory protein-1alpha and MIP-1beta were not detected in any samples. CONCLUSION: Monocyte chemotactic protein-1 might be involved in the recruitment of macrophages and monocytes into the vitreous of eyes with proliferative vitreoretinal disorders.

Linkage analysis in a large Spanish family with X-linked retinitis pigmentosa: phenotype-genotype correlation.

X-linked retinitis pigmentosa (XLRP) accounts for 10-25% of RP families and causes the most severe form of the disease in terms of onset and progression. Although three different loci (RP3, RP2 and RP15) have been proposed on the short arm of the X-chromosome by linkage analysis, RP3 represents the disease locus in the majority of XLRP families. The identification of female carriers of X-linked RP is important for genetic counselling. The presence of fundus and electroretinogram (ERG) abnormalities have been reported to be as high as 87 and 90%, respectively. However, in clinical practice it has not always been possible to know the carrier state of females at risk. Thirty-five members of a Spanish family with X-linked RP were evaluated by linkage analysis using nine polymorphic markers (CYBB, DXS1110, M6, DXS6679, DXS1068, DXS1058, MAOA, MAOB and DXS6849) that map to the X-chromosome region Xp21.1 to Xp11.3, in an attempt to determine the carrier state of these females at risk. It was possible to establish that a RP3 mutation is, most likely, segregating in this family.

160Thr mutation in the rhodopsin gene associated with retinitis pigmentosa.

Mutations in the rhodopsin gene were studied in 23 unrelated Spanish patients with sporadic retinitis pigmentosa (RP). A codon 160 Thr C-->A transition was found in 4 of the 23 patients vs. none of the 159 controls (p < 0.001) suggesting that this mutation may be an informative marker in RP.

Comparative study of incomplete posterior vitreous detachment as a risk factor for proliferative vitreoretinopathy.

BACKGROUND: Abnormal vitreoretinal relationships have recently been implicated in many vitreoretinal disorders. Sites of abnormal vitreoretinal adherences are likely to exist in eyes predisposed to rhegmatogenous retinal detachment (RD), causing either retinal tears or incomplete posterior vitreous detachment (PVD). The present study was designed in two parts to identify the risk for preoperative and postoperative proliferative vitreoretinopathy (PVR) due to incomplete PVD. METHODS: We prospectively evaluated the vitreoretinal relationships using high-resolution kinetic echography in 102 consecutive eyes of 100 patients with rhegmatogenous RD. In the first part, a case-control study was conducted to compare the vitreous status in patients with preoperative PVR (cases) with that in patients with non-PVR-complicated RD (controls). During the second part, patients with noncomplicated RD (65 eyes) who were operated on by a simple retinal attachment procedure were followed up for a mean period of 6.6 months to compare the recurrence of RD due to postoperative PVR according to their vitreous status. RESULTS: Patients with PVR on study entry had a higher prevalence of partial PVD (28 of 32 eyes, 87%) than did controls (25 of 70 eyes, 35%). The statistical significance of this difference was independent of all other variables studied. After a mean follow-up period of 6.6 months, the incidence of recurrence of RD associated with postoperative PVR was 33% in the eyes with incomplete PVD, compared with 4.9% in the eyes without incomplete PVD. CONCLUSIONS: Our results support the notion that the occurrence of incomplete PVD in RD is a significant risk factor for preoperative and postoperative PVR.

A c-myc antisense oligonucleotide inhibits human retinal pigment epithelial cell proliferation.

The purpose of this work was to investigate if MYC-dependent intracellular mitogenic pathway is active in cultures of human retinal pigment epithelial (hRPE) cells and whether myc antisense phosphorotioate oligonucleotides (c-myc-AS-ODN) are useful tools for inhibiting the proliferation of hRPE cells. Cultures of hRPE cells were established from adult human corneal donors. These cells were positively stained for cytokeratins and vimentin. Myc mRNA expression was determined by Northern blot analysis and it was determined by means of immunofluorescence if MYC was expressed. C-myc-AS-ODN effect on cell proliferation was estimated by evaluating the incorporation of 5-bromo-2'-deoxy-uridine into cellular DNA. Cell number was estimated by using a tetrazolium bromide based colorimetric method. Human RPE cells in culture expressed MYC and myc mRNA as well as prothymosin alpha mRNA--a gene whose transcription is under MYC control--indicating that MYC-dependent intracellular mitogenic pathway is active in these cells. In accordance with this, we found that blocking the expression of myc by the addition of c-myc-AS-ODN to the culture medium inhibited hRPE cell proliferation. The effect of the c-myc-AS-ODN was found to be sequence specific (the use of a control oligonucleotide with the same sequence but in an opposite direction had no effect) and dose-dependent (4 microM was the lowest effective dose tested). By using RT-PCR we found that the c-myc-AS-ODN inhibition of cell proliferation was related to a diminution in c-myc mRNA expression, and by immunofluorescence we detected a diminution in c-MYC protein staining in RPE cells after 48 hr of treatment with c-myc-AS-ODN. Furthermore, growth inhibition remained for at least 5 days after addition of a single dose of the c-myc-AS-ODN to the culture. We conclude that hRPE cell proliferation is under MYC control. Blocking the expression of myc by c-myc-AS-ODN inhibited hRPE cell proliferation. These findings establish a rationale for investigating the potential use of a c-myc-AS-ODN as a novel therapeutical tool in the treatment of Proliferative Vitreoretinopathy.

Iris metastasis from endometrial carcinoma.

PURPOSE: To report a case of metastasis to the iris from endometrial carcinoma. METHOD: Case report. A 67-year-old woman with a history of endometrial carcinoma and local recurrence after surgery presented 11 months later with two yellow-pink nodules on the iris of the right eye. RESULTS: Systemic medical evaluation demonstrated no other metastases. The iris tumors were removed surgically, and histology demonstrated adenocarcinoma consistent with endometrial carcinoma. CONCLUSIONS: Endometrial carcinoma can metastasize to the iris. This possibility should be considered because the frequency of endometrial carcinoma is increasing.

Corneal graft after drug-induced toxic epidermal necrolysis (Lyell's disease).

Toxic epidermal necrolysis (TEN) is a clinical syndrome characterized by extensive epidermal loss similar to that found in scalding. Drug sensitization is generally considered to be the mechanism leading to this condition. Severe ocular manifestations are present in nearly all patients. We report a case of TEN with severe ocular involvement, on whom two penetrating keratoplasties were performed in the same eye. Although the prognosis of keratoplasty is reported to be poor when cicatricial changes are present, after the second graft, our patient's symptoms of pain decreased and her visual acuity from light perception improved to 20/40. The graft remains transparent under immunosuppressive therapy twenty-three months after surgery.

Ocular echography in the prognosis of vitreous haemorrhage in type II diabetes mellitus.

PURPOSE: In this study, we have attempted to demonstrate the presence of various echographic parameters which could be associated with a non-spontaneous resorption of vitreous haemorrhage in type II diabetes mellitus and correlate these parameters with clinical outcome. SUBJECTS AND METHODS: We studied 297 eyes of 257 patients with diabetic retinopathy and vitreous haemorrhage without tractional macular retinal detachment ophthalmoscopically and echographically. Of the total eyes studied, a 3-month follow-up visit (including ultrasound) was available in 208 eyes. We retrospectively reviewed the medical records of each patient. RESULTS: The echographic parameters associated with non-resorption of the vitreous haemorrhage were: extramacular tractional retinal detachment, fibrovascular membranes and location of the haemorrhage within the subhyaloidal space (in contrast to within the intragel space). In addition, the duration of the vitreous haemorrhage and the presence of panretinal laser photocoagulation at the time of presentation with a vitreous haemorrhage influenced the resolution of the vitreous haemorrhage. We were also able to construct a logarithmic function that could be used to predict the prognosis of a vitreous haemorrhage in type II diabetes mellitus. CONCLUSIONS: When employed to evaluate vitreous haemorrhages in non-insulin-dependent diabetes mellitus, ocular ultrasound can provide useful prognostic information regarding the lack of resorption of vitreous haemorrhages in type II diabetics.

Ocular blood flow velocity reduction after buckling surgery.

BACKGROUND: It has been reported that scleral buckling reduces the blood flow velocity in retinal vessels. Blood flow changes may also appear in other ocular and extraocular vessels. This study describes the blood flow velocity changes in the ophthalmic artery (OA) after performing this procedure. METHODS: The study was carried out in 12 patients (12 eyes) with rhegmatogenous retinal detachment. Color Doppler imaging was used to measure the peak and average blood flow velocity in the OA. Measurements were taken 1 day before and 2 days after scleral buckling surgery was performed. Intraocular pressure (IOP) was measured prior to each ultrasound study. RESULTS: We found that statistically significant reductions in the peak flow velocity (33%) and average flow velocity (31%) occur in the OA after scleral buckling. All patients showed an increase in IOP after surgery. CONCLUSION: Buckling surgery reduces the blood flow velocity in the OA. Since the OA is the origin of the arterial branches that supply blood to the eye, our results suggest that scleral buckling may decrease not only retinal but also choroidal blood perfusion. Some extraocular structures might also be affected.

Anteroposterior shift in rigid and soft implants supported by the intraocular capsular bag.

We compared the depth of the anterior chamber and the optimal distance refraction in a group of patients with soft and rigid intraocular implants under pilocarpine (maximal ciliary contraction) and cyclopentolate (maximal ciliary relaxation) in order to determine if lens movement might account for the apparent accommodation phenomenon. Lens shifts ranging from 1.5 to 0.02 mm and refractive variations up to 1 D were found. However, the discrepancies between amount of shift and refractive variations suggest that lens movement does not play a relevant role in this phenomenon.