[Melfalan in the treatment of proliferative vitreoretinopathy (experimental study)]. / Melfalan pri lechenii proliferativnoi vitreoretinopatii (eksperimental'noe issledovanie).

PURPOSE: To study of the effectiveness of the drug Melphalan as an antiproliferative agent during experimental proliferative vitreoretinopathy (PVR). MATERIAL AND METHODS: The experimental study used data from 24 eyes of 12 Chinchilla rabbits weighing 2.5-3.0 kg, which had PVR modeled in both eyes by intravitreal injection of a culture of heterogeneous activated fibroblast cells consisting of 200,000 cells in 0.1 ml. Treatment of experimental PVR was performed 1 day after the modeling process. In the first group of animals (6 eyes), 0.02 mg of Melphalan was administered intravitreally. In the second group of animals (6 eyes), 0.005 mg of Melphalan concentrated in 0.1 ml was administered intravitreally. Left eyes in both groups remained without treatment. Animals were observed for 1 month using biomicroscopy and ophthalmoscopy. 30 days after the animals were removed from the experiment, the eyes were enucleated, fixed in 10% buffered formalin and subjected to standard histological examination. The study of paraffin sections of the eyes was performed using the microscopic system «Leica¼ (Leica Microsystems, Germany) with built-in digital camera at the magnification of 200-600. RESULTS: In groups 1 and 2 of the study in the eyes of rabbits that received treatment, PVR was absent, unlike the eyes without treatment, where PVR remained. In group 1, where the dose of Melphalan was 0.02 mg in 0.1 ml, there were changes in the RPE (retinal pigment epithelium), which was regarded as a retinotoxic effect. Glial degeneration and thinning of the retina with disappearance of the photoreceptor layer (the outer nuclear and plexiform layers) resulted from the disturbance of retinal metabolism caused by RPE destruction. In group 2, structure of the retina remained more intact: isolated foci were noted with a decrease in the volume of the outer nuclear layer, shortening of rods and cones with preservation of the inner layers of the retina. CONCLUSION: A single intravitreal injection of 0.005 mg Melphalan had a positive therapeutic antiproliferative effect on the PVR model with minimal retinotoxic changes.

Development of In Vivo Model of Intraocular Retinoblastoma.

Here we describe creation of a model of intraocular retinoblastoma on 4 mature immunodeficient BALB/c nude mice of both sexes. A suspension of the primary retinoblastoma culture was injected into each of the 8 eyes of the test animals. The injections were performed under the control of an operating microscope using insulin syringes with a diameter of 29G transsclerally into the subretinal space in a volume of 0.3 ml at a rate of 10,000 cells in 25 µl physiological saline. The culture of Rb10 cells derived from a patient with retinoblastoma passed 12 passages at the time of the experiments. After 9-week follow-up, a clinical (ophthalmoscopy) and instrumental (ultrasound) study was performed with visualization of the tumor mass on the fundus. After enucleation, a morphological study was performed. The retinoblastoma tumor nodes were identified in two eyes. The proposed model can be used for further research and for testing new chemotherapeutic drugs and treatment regimens for retinoblastoma.

Morphological Changes in the Retina Under Conditions of Experimental In Vivo Regional Ischemia/Reperfusion.

Two models of retinal ischemia/reperfusion were developed in an experiment on rats and structural changes in eye tissues in the early and late postischemic periods were studied. Ischemia/reperfusion was modeled by elevation of intraocular pressure to 110 mm Hg for 30 min with air injection into the anterior chamber of the eye with a special device or subconjunctival administration of 0.2 ml 4×10-6 M endothelin-1. Morphological studies of the retina of enucleated eyes were performed in 3, 7, and 30 days after ischemia/reperfusion. In 3 days, signs of retinal ischemia were seen (retinal edema and ganglion cell damage). In the late post-ischemic period (30 days), atrophy of the outer nuclear and outer plexiform layer of the retina was observed in animals with retinal ischemia/reperfusion caused by intraocular pressure elevation and complete destruction of neuronal cells was found after administration of endothelin-1.

[The significance of fibroblasts in experimental modeling of proliferative vitreoretinopathy]. / Rol' fibroblastov v modelirovanii proliferativnoi vitreoretinopatii.

AIM: to investigate the role of heterogeneous fibroblasts in the development of epiretinal membrane in eyes with modeled proliferative vitreoretinopathy. MATERIAL AND METHODS: The material for investigation were 6 eyes of 3 Chinchilla rabbits. Suspended fibroblasts (fibroblasts of the human skin - 200000 cells in 0.1 ml) were injected into the vitreous cavity via the pars plana. The animals were followed up for 1 month and then made out of the experiment. The eyes were enucleated and fixed in 10% neutral buffered formalin for routine histological examination. Microscopy was performed on the Leica system. RESULTS: The main clinical and morphological criteria for a rabbit model of PVR induced by intravitreal injection of heterogenic fibroblasts have been established: epiretinal membrane formation, changes in intraocular structures (the retinal pigment epithelium and retina), and inflammation (due to transplantation immunity). Particularities of the epiretinal membrane development and the role of different intraocular structures have been described. CONCLUSION: The experimental fibroblastic model of PVR reproduces the final, fibrous, stage of PVR, which is significant for efficacy evaluation of antiproliferative drugs.

[Pathological and molecular genetic characteristics in patients with extrabulbar growth of uveal melanoma]. / Patomorfologicheskie i molekulyarno-geneticheskie osobennosti u bol'nykh pri ekstrabul'barnom roste uveal'noi melanomy.

AIM: to analyze the association of extrabulbar tumor growth with pathological and molecular genetic changes in patients with uveal melanoma (UM). SUBJECTS AND METHODS: A total of 134 UM patients aged 22 to 84 years were examined and treated. The mean height of the tumor was 9.2±2.9 mm; the diameter of its base was 15.3±3.5 mm. Enucleation of the affected eye was performed in 97.8% of cases. Spindle-cell (n=61 (45.6%)), mixed cell (n=46 (34.3%)), and epithelioid cell (n=27 (20.1%)) tumors were identified according to their histological structure. Polymerase chain reaction-restriction fragment length polymorphism analysis was used to determine full and partial monosomy of chromosome 3, deletion of the short arm of chromosome 1, and RASSF1A gene methylation (n=134). The patients were divided into two groups: 1) those with extrabulbar growth (EG) (n=12) and 2) those without EG (n=122). RESULTS: There was a topographic association between the tumor invasion zone and the largest area of exit of the scleral vessels, along which the tumor invaded: the anterior and posterior segments of the eyeball. The specific features of the invasion pattern of UM were shown: there was its broader invasion in the posterior segment and thinner growing tissue interlayers in the anterior segment. Two UM types stopping the process of UM invasion through the scleral fibrous tunic of the eye were established: 1) that with nodule formation and 2) that with tumor cell dissemination within the episclera. The cellular composition of growing tumor tissue in the episclera was ascertained to differ from the main UM focus in the choroid towards its more atypization. The rate was shown to be significantly lower (20% versus 47.9% for the relatively favorable spindle cell type of UM) in the EG group. The frequency of full or partial chromosome 3 monosomy was significantly higher in the extrabulbar tumor growth group (80% versus 50.4%). CONCLUSION: The morphological features of the EG of UM were defined. The use of a statistically significant sample of patients with UM confirmed the favorable course of the tumor in its spindle cell type and the negative role of chromosome 3 monosomy, as well as the relationship to extrabulbar tumor growth.

[Viral infections and retinoblastoma in children].

In this work, the results of a comprehensive laboratory examination of 37 children with retinoblastoma were described. The presence of Igm-, IgA, - IgG- antibodies to the herpes simplex virus types 1 and 2, cytomegalovirus (СMV), epstein-Barr virus (eBV), human herpes virus (HHV) type 6, Toxoplasma gondii, mycoplasma hominis and ureaplasma urealyticum in the serum was tested using ELISA. In the polymerase chain reaction the DNA of these pathogens were detected in the blood plasma of 18 patients and tumor biopsy specimens from 10 eyes. The results showed that children with RB were predominantly infected by the herpesviruses, among which prevailed CMV. in 4 of 5 enucleated eyes the DNA of herpesvirus [CMV (2 eyes), EBV (1 eye), HHV 6 (1 eye)] and ureaplasma urealyticum (1 eye) were also present in tumor tissue. Nucleic acid of infectious microorganisms were considerably more often detected in the tumor tissue than in plasma (5 of 10, 1 of 18, respectively; p = 0.023), suggesting thereby the presence of the virus in the eye and its adverse role in the pathogenesis of the RB.

[Experimental realization of minimally invasive techniques of scleral collagen cross-linking]. / Eksperimental'naya realizatsiya maloinvazivnykh tekhnologii krosslinkinga sklery.

AIM: To realize two minimally invasive techniques of scleral collagen cross-linking (SXL) at the equator and posterior pole of the eye: 1) targeted irradiation of the region with ultraviolet A (UVA) and 2) sub-Tenon injection of Sklerateks. MATERIAL AND METHODS: To perform UVA-SXL, a tool was developed that includes a UV-LED light source (370 nm, 3 mW/cm2) and a polymer-coated silica multimode optical fiber located in one of the two channels of a detachable metal tip. The other channel is used to deliver riboflavin to the scleral surface. The study included 8 Chinchillas' eyes. Intact fellow eyes served as the controls. Scleral echodensity was measured in vivo with Voluson 730 Pro (Kretz) prior to the procedure and then 2 days and 1 month after. After enucleation, the elastic modulus and the degree of scleral cross-linking were established at the same time-points. A placebo-controlled study on the safety and effectiveness of sub-Tenon Sklerateks injections (solution of amino acid salts in the form of succinates) was conducted on 47 Chinchilla rabbits (94 eyes). Sklerateks or placebo (0.1 ml) was injected below the Tenon's capsule of either eye once a week for 1 month (4 injections; 1st series) or 3 months (12 injections; 2nd series). After the end of the course, 22 eyes were studied morphologically. In 72 eyes, scleral samples were obtained in order to evaluate the elastic modulus (Autograph AGS-H tester, SHIMADZU, Japan) and the rate of cross-linking (judging from the denaturation temperature) by differential scanning calorimetry (Phoenix DSC 204 calorimeter, Netzsch, Germany). RESULTS: After UVA irradiation, the scleral echodensity increased from 86.7±5.1 to 98±4.9 dB. The elastic modulus appeared 1.5 times higher than that of the control samples. The denaturation temperature also increased indicating the rate of scleral cross-linking as high as 15-18%. Weekly Sklerateks for 1-3 months has been shown to induce neither clinical, nor morphological signs of local irritative, damaging, or toxic effect. The findings also include: a 1.8 times higher rate of scleral cross-linking, activation of cellular elements, neoformation of connective tissue on the scleral surface, and vascular growth, which together indicate a pronounced metabolic and strengthening effect of Sklerateks on the sclera. CONCLUSION: Experimental results on minimally invasive techniques of SXL allow to recommend them for further clinical investigation as a promising treatment of progressive myopia.

Experimental Modeling of Proliferative Vitreoretinopathy. An Experimental Morphological Study.

A model of proliferative vitreoretinopathy induced by simultaneous intravitreal injection of recombinant IL-1ß and platelet concentrate is created and its main morphological manifestations are studied on Chinchilla rabbits. The model reflects pathogenesis of proliferative vitreoretinopathy: epiretinal membrane with the formation of retinal plication, traction detachment of the retina; moderate inflammatory reaction in the uveal tract, in the optic nerve infundibulum, in the vitreous body; intact structural elements of the retina, dissociation of the retinal pigmented epithelium cells with their subsequent migration. The model is adequate to the clinical picture of proliferative vitreoretinopathy in humans, which recommends it for experimental studies of the efficiency of drug therapy and prevention of this disease.

[Morphogenesis of complications after long-term intraocular silicon oil filling (clinical histopathological study)].

The question of necessity and time of silicon oil (SO) removal after tamponade for traumatic retinal detachment (TRD) is still controversial. So the study of changes in the eye filled with SO for a long time is of great interest. Histopathologic analysis of 9 enucleated eyes filled with SO for TRD during long-time period (from 6 months till 30 years) was performed. The characteristic change was inflammatory reaction with fibrin membranes surrounding silicon drops and development of proliferative vitreoretinopathy and tractional retinal detachment in the following. In 2 cases the inflammatory process around SO was associated with bone formation. The most severe complication was the imbibition of retina with SO in a vacuole-like manner associated with total atrophy of neural structures. Obtained data show that time for SO removal should be less than 6 months after surgery.

[Congenital corneal staphylomas in children].

Clinical manifestation, results and outcomes of surgical treatment were studied in children with very rare anomaly of cornea and anterior segment. Symptoms and signs, histological findings are described in details, results of ophthalmoscopy, ultrasound biomicroscopy and maternal ophthalmotrophic infections are studied in children with corneal staphylomas.

[Immunomorphology in the diagnosis of orbital rhabdomyosarcoma].

Immunohistochemistry (IHC) using monoclonal antibodies is presently the most reliable method to verify tumor tissue belonging. The histological diagnosis of rhabdomyosarcoma (RMS) was immunohistochemically confirmed in 7 of 10 orbital neoplasms in children. In 3 cases the diagnosis of RMS was established after IHC study that revealed the atypical tumor cell expression of the muscular antigens--myoglobin, protein Myo D1, actin HHF-35, which is indicative of the rhabdomyogenic genesis of a tumor. IHC data indicated that, no matter what the histological pattern, orbital RMS is characterized by the expression of cytospecific proteins--myoglobin, Myo D1, actin HHF-35, and tissue-specific (intermediate filaments)--vimentin and desmin. The use of appropriate antibodies offers a means of making a histogenetic diagnosis even in cases of the atypical morphological pattern.

Mitochondria-targeted plastoquinone derivatives as tools to interrupt execution of the aging program. 4. Age-related eye disease. SkQ1 returns vision to blind animals.

Mitochondria-targeted cationic plastoquinone derivative SkQ1 (10-(6'-plastoquinonyl) decyltriphenylphosphonium) has been investigated as a potential tool for treating a number of ROS-related ocular diseases. In OXYS rats suffering from a ROS-induced progeria, very small amounts of SkQ1 (50 nmol/kg per day) added to food were found to prevent development of age-induced cataract and retinopathies of the eye, lipid peroxidation and protein carbonylation in skeletal muscles, as well as a decrease in bone mineralization. Instillation of drops of 250 nM SkQ1 reversed cataract and retinopathies in 3-12-month-old (but not in 24-month-old) OXYS rats. In rabbits, experimental uveitis and glaucoma were induced by immunization with arrestin and injections of hydroxypropyl methyl cellulose to the eye anterior sector, respectively. Uveitis was found to be prevented or reversed by instillation of 250 nM SkQ1 drops (four drops per day). Development of glaucoma was retarded by drops of 5 microM SkQ1 (one drop daily). SkQ1 was tested in veterinarian practice. A totally of 271 animals (dogs, cats, and horses) suffering from retinopathies, uveitis, conjunctivitis, and cornea diseases were treated with drops of 250 nM SkQ1. In 242 cases, positive therapeutic effect was obvious. Among animals suffering from retinopathies, 89 were blind. In 67 cases, vision returned after SkQ1 treatment. In ex vivo studies of cultivated posterior retina sector, it was found that 20 nM SkQ1 strongly decreased macrophagal transformation of the retinal pigmented epithelial cells, an effect which might explain some of the above SkQ1 activities. It is concluded that low concentrations of SkQ1 are promising in treating retinopathies, cataract, uveitis, glaucoma, and some other ocular diseases.

[Topography and functional changes of the rabbit retina with a single injection of triamcinolone].

Intravitreal injections of 6 and 8 mg of triamcinolone (kenalog) were studied for impact on the time course of electroretinographic changes and the retinal structure of healthy rabbits. Histopathological study failed to reveal the signs of toxic damage to the retina. The electroretinographic findings suggest that kenalog primarily affect the function of photoreceptors and Muller's glia. The fact that the alteration of retinal functional activity may be linked with the early cytotoxic effects of the agent cannot be ruled out. However, the changes found in retinal electrogenesis are most likely to reflect the mechanisms of kenalog's beneficial therapeutic action that is primarily mediated through its effect on the function of photoreceptors and Muller's cells.

[Histological studies of carbon felt (carbotextim M) in late periods after its orbital implantation].

The authors performed histological studies of a grown carbon felt implant and its adjacent tissues in late postoperative studies. During planned operations (cavity or stump correction), pieces of a grown implant with its adjacent tissues were taken for examination from 25 patients in whom carbon had been earlier implanted during enucleation and delayed stump plastic repair. Morphological studies indicated that mature connective tissue had grown between the fibers of the implant, that a connective tissue capsule had formed around the whole implant, and that there was no inflammatory reaction in both the implant itself and its adjacent tissues. By taking into account the fact that there is a good tissue basis and vascularity, the authors present the results of additional simulation of an implant in late postoperative periods and those of additional correction of the implant's volume.

[Sympathetic ophthalmia. Clinical and morphological study in 6 cases after penetrating injury and subsequent repeated vitreoretinal operations].

The paper outlines the clinical and morphological picture of 6 sympathizing eyes enucleated in 1996 to 2003 after vitrectomy and other endovitreal interventions into early injured or operated eyes. Resurgery was attempted early after wounds or a primary operation on the average following 20 days. Sympathetic ophthalmia (SO) generally occurred in the posterior uveal tract as panuveitis or posterior uveitis and diagnosed in the late periods. Specific granulomatous inflammation in the uveal tract was observed in all eyes and the morphological feature of sympathizing eyes was the spread of an inflammatory process to the retina with the development of the latter's epithelioid-cell granulomas and adhesive chorioretinitis. The authors also discuss the possibility of additional antigenic stimulation accompanied by a progressive autoimmune process in the eye and by the generalization of a granulomatous inflammation, as well as the role of retinal minor lesions in this process as a source of additional autoantigens during repeated vitreoretinal operations. The latter, as the authors believe, are a risk factor of SO. In this connection, when repeated vutreoretinal interventions should be made in the presence of an uncompleted wound process in the eye and when they should be conducted under the conditions of active immunosuppressive therapy gain in great importance.

[Acute enterovirus uveitis in infants].

Enterovirus uveitis (EU) is a new infant eye disease that was first detected and identified in Russia in 1980-1981. Three subtypes of human echoviruses (EV19K, EV11A, and EV11/B) caused 5 nosocomial outbreaks of EU in different Siberian cities and towns in 1980-1989, by affecting more than 750 children mainly below one year of age. Sporadic and focal EU cases (more than 200) were also retrospectively diagnosed in other regions of Russia and in different countries of the former Soviet Union. There were following clinical manifestations: common symptoms of the infection; acute uveitis (rapid focal iridic destruction, pupillary deformities, formation of membranes in the anterior chamber of the eye); and in 15-30% of cases severe complications, cataract, glaucoma, vision impairments. Uveitis strains EV19 and EV11 caused significant uveitis in primates after inoculation into the anterior chamber of the eye, as well as sepsis-like fatal disease with liver necrosis after venous infection. The uveitis strains are phylogenetically and pathogenetically close for primates to strains EV19 and EV11 isolated from young children with sepsis-like disease. The contents of this review have been published in the Reviews in Medical Virology, 2004, vol. 14, p. 241-254.

[New modification of autoconjunctival plastic surgery in urgent corneal surgery]. / Novaia modifikatsiia autokon''iunktival'noi plastiki i neotlozhnoi khirurgii rogovitsy.

A new modification of plastic repair of deep defects of the cornea was performed: autoconjunctival plasty of the cornea with a flap on a pedicle with its rigid fixation in the corneal bed. The operation was carried out in 30 patients with infectious, trophic, and autoimmune involvement of the cornea, 24 of these with perforation of the cornea. The patients were observed for 1 year after the operation. The conjunctival-corneal complex formed in 24 cases, dislocation of the flap without its necrosis occurred in 2 cases, and necrosis of the flap in 4 cases. This modification of conjunctival plasty of the cornea can serve as the operation of choice in urgent ophthalmic surgery in cases when transplantation material is unavailable.