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Biomedical subjects

Fulton Wong

Publications and source records attributed to Fulton Wong.

8 recordsLinked to original sources
PubMed2006

Targeted inactivation of synaptic HRG4 (UNC119) causes dysfunction in the distal photoreceptor and slow retinal degeneration, revealing a new function.

HRG4 (UNC119) is a photoreceptor protein predominantly localized to the photoreceptor synapses and to the inner segments to a lesser degree. A heterozygous truncation mutation in HRG4 was found in a patient with late onset cone-rod dystrophy, and a transgenic (TG) mouse expressing the identical mutant protein developed late onset retinal degeneration, confirming the pathogenic potential of HRG4. Recently, the dominant negative pathogenic mechanism in the TG model was shown to involve increased affinity of the truncated mutant HRG4 for its target, ARL2, which leads to a delayed decrease in its downstream target, mitochondrial ANT1, mitochondrial stress, synaptic degeneration, trans-synaptic degeneration, and whole photoreceptor degeneration by apoptosis. In this study, the mouse HRG4 (MRG4) gene was cloned and targeted to construct a knock-out (KO) mouse model of HRG4 in order to study the effects of completely inactivating this protein. The KO model was examined by genomic Southern blotting, Western blotting, immunofluorescence, funduscopy, LM and EM histopathology, ERG, and TUNEL analyses. The KO model developed a slowly progressive retinal degeneration, characterized by mottling in the fundus, mild thinning of the photoreceptor layer, and increase in apoptosis as early as 6 months, dramatic acceleration at approximately 17 months, and virtual obliteration of the photoreceptors by 20 months. When compared to retinal degeneration in the TG model, significant differences existed in the KO consisting of more severe and early photoreceptor death without evidence of early synaptic and trans-synaptic degeneration as seen in the TG, confirmed by LM and EM histopathology, ERG, and Western blotting of synaptic proteins. The results indicated a dysfunction in the KO outside the synapses in the distal end of photoreceptors where MRG4 is also localized. Differences in the phenotypes of retinal degeneration in the KO and TG models reflect a dysfunction in the two opposite ends of photoreceptors, i.e., the distal inner/outer segments and proximal synapses, respectively, indicating a second function of MRG4 in the distal photoreceptor and dual functionality of MRG4. Thus, inactivation of MRG4 by gene targeting resulted in a retinal degeneration phenotype quite different from that previously seen in the TG, attesting to the multiplicity of MRG4 function, in addition to the importance of this protein for normal retinal function. These models will be useful in elucidating the functions of HRG4/MRG4 and the mechanism of slow retinal degeneration.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Altered light responses of single rod photoreceptors in transgenic pigs expressing P347L or P347S rhodopsin.

PURPOSE: Numerous mutations of rhodopsin lead to rod cell death and ultimately to complete blindness, yet little is known about the alterations in the physiology of the light sensors containing the aberrant protein, the rod photoreceptors. METHODS: Suction pipettes were used to record the light responses from single rod photoreceptors isolated from the retinas of transgenic pigs of various ages and at progressive stages of retinal degeneration. RESULTS: We have observed changes in the photoresponse of transgenic porcine rods containing both wild type and mutant rhodospin. Our findings are consistent with the idea that substitutions at position proline 347 of rhodopsin interfere with the inactivation of R*. In addition the level of photoreceptor degeneration is positively correlated with an acceleration and desensitization of the photoresponse to dim flashes. CONCLUSIONS: It appears that the phototransduction cascade, even when initiated by wild type rhodopsin molecules is altered in a way that is progressive with the level of retinal degeneration. A model consistent with our observations introduces the idea of a binding site for the carboxy terminus of rhodopsin on rhodopsin kinase.

Amino Acid Substitution↗
PubMed2005

Oxidative damage is a potential cause of cone cell death in retinitis pigmentosa.

Retinitis pigmentosa (RP) is a prevalent cause of blindness caused by a large number of different mutations in many different genes. The mutations result in rod photoreceptor cell death, but it is unknown why cones die. In this study, we tested the hypothesis that cones die from oxidative damage by performing immunohistochemical staining for biomarkers of oxidative damage in a transgenic pig model of RP. The presence of acrolein- and 4-hydroxynonenal-adducts on proteins is a specific indicator that lipid peroxidation has occurred, and there was strong immunofluorescent staining for both in cone inner segments (IS) of two 10-month-old transgenic pigs in which almost all rods had died, compared to faint staining in two 10-month-old control pig retinas. In 22- and 24-month-old transgenic pigs in which all rods and many cones had died, staining was strong in cone axons and some cell bodies as well as IS indicating progression in oxidative damage between 10 and 22 months. Biomarkers for oxidative damage to proteins and DNA also showed progressive oxidative damage to those macromolecules in cones during the course of RP. These data support the hypothesis that the death of rods results in decreased oxygen consumption and hyperoxia in the outer retina resulting in gradual cone cell death from oxidative damage. This hypothesis has important therapeutic implications and deserves rapid evaluation.

Acrolein↗
PubMed2004

Transplantation of full-thickness retina in the rhodopsin transgenic pig.

PURPOSE: To establish the morphology of full-thickness neuroretinal grafts transplanted to hosts with degenerative photoreceptor disease. METHODS: Twenty rhodopsin transgenic pigs received a neuroretinal sheet from a neonatal normal pig in one eye. Following vitrectomy and retinotomy with bleb formation, the grafts were positioned inside the bleb between the host neuroretina and retinal pigment epithelium. After a survival time of 4 months, eye specimens were studied by light and electron microscopy as well as with immunohistochemical markers. RESULTS: One eye developed endophthalmitis in the immediate postoperative period and was terminated. Laminated grafts with correct polarity were found in 13 of the remaining 19 eyes. In most cases, these grafts had well-developed organized photoreceptors with outer segments apposed to the host retinal pigment epithelium. The inner layers of the graft contained mostly Müller cells. Both eyes of the hosts had a reduction of photoreceptor cells in most of the retina, while inner layers remained relatively intact. CONCLUSIONS: Full-thickness neuroretinal grafts can be transplanted to a large animal host with photoreceptor degeneration. The transplantation procedure is relatively atraumatic to both graft and host tissue, and the grafts survive well for at least 4 months. The graft and host retina does not seem to form extensive neuronal contacts, and future work must be directed at stimulating such activity without disrupting the retinal neuronal organization.

Animals↗
PubMed2004

Effects of indocyanine green on the retina and retinal pigment epithelium in a porcine model of retinal hole.

PURPOSE: This study was designed to emulate human macular hole surgery and to test the effects of indocyanine green (ICG) on the retina and retinal pigment epithelium (RPE). METHODS: Yorkshire Cross pigs (n = 23) underwent vitrectomy, separation of the posterior cortical vitreous, and creation of a single retinal hole. In three study groups (n = 6, each group), air-fluid exchange was performed, following which balanced salt solution (BSS), 1.0% ICG, or 0.5% ICG was applied over the retinal hole. In one additional group (n = 5), 0.5% ICG was injected into the fluid-filled eye. At 4 weeks, the eyes were examined clinically, and fundus photographs were obtained before enucleation and light microscopic examination. RESULTS: Clinical evaluations documented a statistically significant difference between study groups (P = 0.036). There was a higher rate of moderate or severe RPE atrophy among animals where 1% or 0.5% ICG was applied in air-filled eyes (83% and 67%, respectively) compared with BSS controls (17%) and fluid-filled eyes receiving 0.5% ICG (40%). Histologic evaluation demonstrated a statistically significant difference between groups (P = 0.044), with extensive outer retinal degeneration observed in air-filled eyes receiving 1% or 0.5% ICG (66% and 60%, respectively) compared with BSS controls or fluid-filled eyes receiving 0.5% ICG (none of the eyes in either group). None of the study groups had any changes in the inner retina except at the retinal hole site. CONCLUSIONS: Retina exposed to ICG concentrations used in human vitreoretinal surgery had greater RPE atrophy and outer retinal degeneration than control eyes undergoing the same surgery without ICG. Eyes filled with infusion fluid during ICG injection had less damage to the RPE and outer retina than did air-filled eyes receiving ICG.

Air↗
PubMed2003

Lensectomy and vitrectomy decrease the rate of photoreceptor loss in rhodopsin P347L transgenic pigs.

BACKGROUND: Photoreceptor degeneration in retinitis pigmentosa (RP) runs an inevitable, gradually progressive course. A wide variety of growth factors of different origins have been shown to slow the rate of degeneration in some rodent models of RP. Recently, lens-derived neurotrophic factors have been shown to rescue degenerating ganglion cells in crush models of the optic nerve. Our objective was to evaluate the potential rescue effect of lensectomy and vitrectomy (L&V) on photoreceptor degeneration in a large-animal model, the rhodopsin P347L transgenic pig. METHODS: We operated on one eye of each of 49 3-week-old pigs--15 vitrectomies and 34 L&V, 6 of which received steroids. Retinal paraffin sections were prepared for all eyes, in addition to immunohistochemistry in four eyes, 8 weeks after L&V. RESULTS: At eight weeks after L&V, operated eyes showed significantly more nuclei in the outer nuclear layer (ONL) than the unoperated fellow eyes. The better preservation of the ONL persisted but was less prominent by 20 weeks after surgery. Steroid treatment did not markedly reduce the better preservation of the ONL seen at 8, 10, and 12 weeks after surgery. The significant difference in cell count between operated and unoperated eyes in the L&V group at 8 weeks was due to the difference in the number of rods, not the cones. CONCLUSION: Lensectomy and vitrectomy delay photoreceptor degeneration in rhodopsin P347L transgenic pigs. Lens-related rescue effect is a probable reason for the delayed degeneration.

Animals↗
PubMed2002

RGD peptide-assisted vitrectomy to facilitate induction of a posterior vitreous detachment: a new principle in pharmacological vitreolysis.

PURPOSE: To evaluate a new concept in pharmacological vitreolysis by studying the efficacy of intravitreal RGD peptide-assisted vitrectomy in facilitating the separation of the posterior cortical vitreous from the retinal surface in an animal model. METHODS: Eight rabbits (16 eyes) received an intravitreal injection of 1 or 5 mg of RGD peptide in one eye and either RGE peptide (inactive control) or phosphate buffered saline in the fellow eye. After 24 hours, a pars plana vitrectomy with low aspiration (< or =30 mmHg) was performed in an attempt to create a detachment of the posterior cortical vitreous. A masked observer performed pre- and postoperative indirect ophthalmoscopy and B-scan ultrasonography. Postoperative scanning electron microscopy evaluated the vitreoretinal surface in selected eyes. Two additional rabbits received intravitreal injections of RGD peptide in one eye (1 mg and 5 mg) and 1 mg of RGE peptide in the fellow eye to examine apoptosis of the retinal cells by TUNEL assay. RESULTS: Based on postoperative ultrasound findings, six of the eight rabbits had a greater degree of posterior vitreous detachment in the RGD eye compared to the fellow eye (p = 0.03). The total number and the average number of detached quadrants in the group of RGD peptide eyes was twenty-three and 2.85 respectively compared to seven and 0.85 for the control fellow eyes (p = 0.02). Scanning electron microscopy confirmed the presence of postoperative posterior vitreous detachment. There was no evidence of retinal cell apoptosis in RGD injected eyes. CONCLUSION: RGD peptide-assisted vitrectomy facilitated posterior vitreous detachment in rabbit eyes, suggesting that RGD-containing peptides may prove to be effective adjuncts in producing posterior vitreous separation during vitreous surgery.

Animals↗