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Taiji Sakamoto

Publications and source records attributed to Taiji Sakamoto.

At least 55 records · Page 3Linked to original sources
PubMed2003

[Cell biology of hyalocytes].

There is a group of cells, called hyalocytes, in the cortical vitreous. Although hyalocytes were discovered more than a hundred years ago, the molecular and cellular biological characteristics of hyalocytes have yet to be elucidated. In this study, we investigated various aspects of hyalocytes and, also performed triamcinolone acetonide (TA)-assisted vitrectomy to remove the hyalocytes for diabetic macular edema. Immunohistochemical analysis of rat eyes showed that 90% of hyalocytes were negative for ED1 but positive for ED2, indicating that hyalocyte is a tissue macrophage. Chimeric mice were created by transplanting bone marrow from green fluorescent protein (GFP)-transgenic mice into irradiated wild-type mice, showing the origin of hyalocyte to be bone marrow cells. Bovine hyalocytes were cultured successfully. The proliferation of hyalocytes was significantly enhanced by hepatocyte growth factor (HGF), platelet-derived growth factor (PDGF), vascular endothelial growth factor (VEGF), and fibroblast growth factor (FGF-2) and inhibited by transforming growth factor(TGF)-beta. Among these, PDGF-BB stimulated the proliferation most potently through the MEK 1 pathway. Hyalocyte migration assessed by double chamber assay was also stimulated by PDGF-BB and it was mediated by the PI3K and p38 MAPK pathways. Cellular contraction of hyalocyte was significantly enhanced by PDGF-BB and TGF-beta through Rho kinase, p44/42 MAPK, and protein kinase C pathways, as measured by collagen gel contraction assay. Next, the relationship between the vitreous cavity(VC) and the immune system was studied after intravitreous inoculation with ovalbumin (OVA). Injection of OVA into the VC of C 57 BL/6 mice resulted in suppressed systemic cell-mediated immunity to OVA as determined by the ear swelling assay. This aberrant immune responsiveness following VC injection of OVA was termed VC-associated immune deviation or VCAID. The phenomenon of VCAID was mediated by intravitreous antigen-presenting cells. The histological study of chimeric mice showed these cells to be intravitreous residential cells, namely hyalocytes. VCAID was abolished by intravitreous inflammation such as experimental autoimmune uveitis. Finally, TA-assisted vitrectomy for diabetic macular edema was performed to remove cortical vitreous, because it contained many hyalocytes which could secrete inflammatory cytokines including VEGF. Although the number of treated eyes was limited, the surgical results have been favorable so far. The investigation of hyalocytes would open a new avenue for better understanding and development of treatment for various vitreo-retinal diseases.

Animals↗
PubMed2002

Functional role of Egr-1 mediating VEGF-induced tissue factor expression in the retinal capillary endothelium.

PURPOSE: To investigate the causal relationship between VEGF and tissue factor (TF) expression, and its intracellular signaling in the retinal capillary endothelium both in vitro and in vivo. METHODS: TF mRNA and protein expression in cultured bovine retinal capillary endothelial cells (BRECs) were detected by RT-PCR and western blotting. The expression and subcellular localization of Egr-1 were analyzed by immunocytochemistry and western blotting. Involvement of p44/p42 MAPK pathway in this signaling was assessed using PD98059. Electrophoretic mobility shift assay (EMSA) was performed using human TF Egr-1/Sp-1 overlapping promoter region (-85 to -70). Decoy oligonucleotide was transfected into BRECs to clarify the critical transcription factor mediating VEGF-induced TF gene expression. To evaluate the importance of GC rich region in VEGF-induced TF protein expression in rat retinas, Mithramycin was intraperitoneally administered. RESULTS: VEGF stimulated TF mRNA and protein expression in cultured BRECs, reaching maximal effect after 4 h and 10 h, respectively. VEGF activated transcription factor Egr-1 within 60 min. Inactivation of Egr-1 by PD98059 resulted in the prohibition of VEGF-induced TF gene expression. EMSA revealed the increment of Egr-1 binding with TF promoter region by displacing Sp1 after treatment with VEGF. Transfection of the Egr-1/Sp-1 overlapping decoy into BRECs inhibited VEGF-dependent TF gene expression. Mithramycin almost completely suppressed VEGF-induced TF protein expression in retinal capillary system in vivo (80%, p<0.01). CONCLUSION: Transcription factor Egr-1, which lies downstream of p44/p42 MAPK, critically mediates VEGF-dependent TF expression in the retinal capillary endothelium.

Animals↗
PubMed2002

Triamcinolone-assisted pars plana vitrectomy improves the surgical procedures and decreases the postoperative blood-ocular barrier breakdown.

PURPOSE: To determine the effect of a triamcinolone-assisted pars plana vitrectomy (PPV) on the visibility of hyaloid during surgery and the postoperative clinical outcome. METHODS: Thirty-one patients with proliferative retinal disease [8 with diabetic macular edema (DME), 10 with proliferative diabetic retinopathy (PDR), 13 with proliferative vitreoretinopathy (PVR)] underwent PPV, where the vitreous body was visualized by the intravitreal injection of triamcinolone solution during the operation. The visual acuity, intraocular pressure (IOP), tamponade, corneal pathology, after-cataract, vitreous hemorrhage, and necessity of reoperation, were thereafter examined for at least 3 months after surgery. The anterior chamber laser flare cell meter was used on postoperative day 8 in DME eyes with triamcinolone-assisted PPV and with routine PPV to evaluate the breakdown of the blood-ocular barrier. RESULTS: The vitreous body was clearly seen by means of triamcinolone during surgery, which greatly helped us to perform a posterior hyaloid resection safely and thoroughly. Six of 8 DME eyes, 8 of 10 PDR eyes, and 5 of 13 PVR eyes showed an improvement in their vision postoperatively. No eye except one experienced IOP elevation above 21 mmHg for 7 days. Six eyes had vitreous hemorrhage. The DME eyes which received triamcinolone-assisted PPV showed significantly less breakdown of the blood-ocular barrier than those with routine PPV (Mann-Whitney U-test, P<0.01). CONCLUSION: Triamcinolone improved the visibility of the hyaloid and the safety of the surgical procedures during PPV and also inhibited the postoperative breakdown of the blood-ocular barrier. Although the long-term effects are still unknown, this method appears potentially useful as an improved treatment for proliferative retinal diseases.

Blood-Aqueous Barrier↗
PubMed2002

Morphological and functional damage of the retina caused by intravitreous indocyanine green in rat eyes.

BACKGROUND: This study was designed to investigate the influence of intravitreal indocyanine green (ICG) on retinal morphology and function. METHODS: Brown Norway rats eyes ( n=24) were vitrectomized by the injection of 0.05 ml of 100% SF(6) gas. Two weeks later, ICG solution was injected into the vitreous cavity of vitrectomized eyes at a dose of 25 mg/ml, 2.5 mg/ml, 0.25 mg/ml or 0.025 mg/ml (0.05 ml/eye). Retinal toxicity was histologically assessed by light microscopy on day 10. The retinal function was also evaluated by electroretinography (ERG) in the low-dose groups (0.25 mg/ml and 0.025 mg/ml) after 10 days and again after 2 months,. Sham-operated eyes (SF(6) injected followed by 0.05 ml of BSS plus, n=6) were used as controls. RESULTS: In the high-dose group (25 mg/ml ICG), the retinal structure was severely deformed and the retinal pigment epithelium partly disappeared. In eyes with 2.5 mg/ml ICG, the retinal structure was also affected but less strongly so than with 25 mg/ml. No apparent pathologic change was observed in the low-dose groups (0.25 mg/ml or 0.025 mg/ml) by light microscopy. In contrast, 10 days later the amplitude of dark-adapted a- and b-waves of ERGs in the eyes of low-dose group rats were found to have decreased. In addition the light-adapted b-waves did not change significantly. These changes remained for 2 months. CONCLUSION: Even at a low dose (0.025 mg/ml), intravitreous ICG induced functional damage of the retina without any apparent morphological damage. This information should be taken into account when clinically administering ICG into the vitreous cavity.

Animals↗
PubMed2002

Targeted gene transfer to corneal stroma in vivo by electric pulses.

This study was conducted to develop a method of targeted gene transfer by electric pulses to a selected area of corneal stroma in vivo. Plasmid DNA with a green fluorescent protein (GFP) gene under a cytomegalovirus promoter was injected through the corneal pocket into the corneal stroma of the adult Brown Norway rat, and various intensities of electric pulses ranging from 10 to 30 V were delivered to the corneal epithelial side with an electric probe. Direct stereomicroscopy of the fluorescent using real-time imaging was used to determine in vivo gene expression on days 1, 2, 4, 6, 8, 10, 15, and 20 after gene transfer. Transgene expression was detected in the corneal stroma as early as day 1 and until day 15. The most intense expression was noted on days 4 and 6. Gene transfer was most effective using eight electric pulses of 20 V for 50 msec. Histologic study disclosed GFP expression in keratocytes within the targeted area. There was no apparent cell damage in the gene transferred cells. No apparent inflammation was found in the anterior chamber or trabecular cells when electric pulses less than 30 V were used. In summary, the present technique transferred the gene of interest to a highly selected area of corneal stroma with no apparent damage. This method will likely be useful not only for developing gene therapy for corneal diseases but also for corneal research in general.

Animals↗
PubMed2002

Expression of matrix metalloproteinases and their inhibitors in experimental retinal ischemia-reperfusion injury in rats.

Matrix metalloproteinases (MMPs) are endopeptidases that degrade the extracellular matrix (ECM) and are involved in the pathogenesis of retinal degeneration along with tissue inhibitors of metalloproteinases (TIMPs). The present study examined the expression and activation of two specific members of MMPs (MMP-2 and MMP-9) and their related inhibitors (TIMP-1 and TIMP-2) in an experimental retinal ischemia-reperfusion injury. Retinal ischemia-reperfusion injury (RIRI) was induced in adult rats with a ligation method. After one hour of ischemia and a varied reperfusion time (0, 3, 6, 12, 24, 48 and 76 hr), the rat eyes were enucleated. Retinal extracts underwent zymographic analysis to measure the activity of MMP-2/9. The activity of TIMP-1 and TIMP-2 was measured by reverse zymography. The protein level was examined by Western blot. Immunohistochemistry analysis was undertaken to assess the anatomical distribution of MMP-9 in the retina after RIRI. The gelatinolytic activity of ProMMP-2 (72 kDa) was increased markedly at 6 hr after RIRI. ProMMP-9 (92 kDa) was not detected in the control specimens, while it appeared at 3 hr, increased markedly at 6 hr, and reached maximal levels at 24 hr after RIRI. The gelatinolytic activity found ian retinal extracts was shown to be inhibited by 10 m M EDTA and activated in vitro by a known metalloproteinase activator (4-aminophenylmercuric acetate (APMA)), indicating that these enzymes were of the metalloproteinase class. By western blot, MMP-2/9 levels increased parallel to protein activity level in zymography. No corresponding increase in TIMP-1 and TIMP-2 protein activity and protein level was detected by reverse zymography and western blot. Elevated levels of MMP-9 and its distribution in retina were confirmed by immunohistochemistry. Expression of MMP-9 was detected in the inner and outer segments of rat retina, and the level becomes stronger at 24 hr after RIRI. In this study, ProMMP-2 and ProMMP-9 were expressed and increased significantly, but their inhibitors (TIMP-1 and TIMP-2) remained relatively unaltered in ischemic retina after RIRI in rats. These results suggest that MMP-2 and MMP-9 may play an important role in the pathomechanism of retinal ischemic injury.

Animals↗
PubMed2002

Recombinant Sendai virus-mediated gene transfer into adult rat retinal tissue: efficient gene transfer by brief exposure.

To determine the usefulness of recombinant Sendai virus (SeV) for ocular gene transfer, the authors characterized SeV-mediated gene transfer to the retinal tissue of adult rats via subretinal injection. Recombinant SeV encoding the lacZ gene achieved frequent transgene expression in the retinal pigment epithelium (RPE) (mean=38.76%), while gene transfer to other retinal cells was rare. These findings are similar to those of previous reports using adenoviruses. Peak reporter gene expression of SeV in cultured RPE cells was similar to that of adenovirus at the same titer; however, SeV achieved high levels of expression after a brief vector-cell contact time, while adenovirus required over 3hr for efficient gene transfer. This finding was also observed in vivo following a brief SeV filling in the subretinal space, and may therefore provide a clinical advantage in avoiding retinal damage due to prolonged detachment. The observed SeV-mediated gene expression in the rat retina was transient. The initial phase of the decrease in luciferase activity could be prevented by daily eye drops of dexamethasone, suggesting that the corticosteroid-sensitive host reaction may affect early clearance of the virus. The late decline of transgene expression (2 weeks) was inhibited by the immunosuppressant, cyclosporin A, in a dose-dependent manner, suggesting that the cytotoxic T-lymphocyte response may be important in this phase. This work represents the first report of SeV-mediated gene transfer to ocular tissue, and identifies recombinant SeV as a new tool for studies of retinal gene transfer and gene therapy.

Adenoviridae↗
PubMed2002

Neovascularization in the anterior segment of the rabbit eye by experimental anterior ischemia.

PURPOSE: To investigate the effects of anterior ischemia accompanied by neither retinal nor choroidal ischemia on the anterior segment of the eye. METHODS: Both long posterior ciliary arteries in the right eye of 14 rabbits were directly cauterized with an electric coagulator. The eyes were enucleated 1, 2, 4, 7, 9 or 14 days after cauterization, then fixed with 4% paraformaldehyde. Semi-thin sections were studied by light microscopy. Several sections were stained with Griffonia simplicifolia lectin, which bound specifically to mammalian vascular endothelium. Other specimens were examined immunohistochemically for vascular endothelial growth factor (VEGF) protein. The tissue specimens of the first postoperative day were studied for expression of VEGF mRNA by in situ hybridization. RESULTS: Atrophy of the iris and ciliary body was seen after the second postoperative day. Corneal neovascularization appeared after 7 days. Neovascularization on the anterior surface of the iris and in the trabecular meshwork was detected after the ninth postoperative day. The proliferative tissues with newly formed vessels obstructed the iridocorneal angle 14 days after the treatment. There was no histological change in either the retina or choroid. Immunohistochemically, VEGF protein was detected in the epithelial and vascular cells of the iris on the first and fourth postoperative day. Expression of VEGF mRNA was detected in the epithelial cells of the ciliary body on the day following the treatment. CONCLUSIONS: Anterior segment ischemia, when unaccompanied by retinal ischemia, causes neovascularization in the cornea, iris and trabecular tissue.

Animals↗
PubMed2002

Electron microscopic study of monkey retina after photodynamic treatment.

The purpose of this histological study was to determine the effects of photodynamic treatment, using a hematoporphyrin derivative and argon laser, on normal retinas of monkeys. Ten cynomolgus monkeys were treated with a hematoporphyrin derivative, given intravenously at a dose of 2.5 mg/kg. Forty minutes or 1 or 3 days after the injection, argon laser photoradiation was given over a 2.0-mm-diameter with a 10-min exposure and at an intensity of 40, 100, or 200 mW. The eyes were enucleated 1, 3, 4, 15, 18, 21, 35, or 38 days after the photoradiation and tissue samples were observed under a transmission electron microscope. The most fragile regions in the retina were the retinal nerve fibers, the outer segments of the visual cells, and the retinal pigment epithelium. Vascular endothelial cells were also fragile. The retinal capillary was easily obstructed, and the choriocapillaris was also occluded in an animal with severe retinal damage. The Mueller cells had the highest tolerance to the photodynamic treatment. Thus, exposing the normal part of the retina to light during photodynamic therapy should be avoided.

Animals↗
PubMed2002

Critical role of photoreceptor apoptosis in functional damage after retinal detachment.

PURPOSE: Although apoptosis is assumed to play a pivotal role in retinal function loss, its mechanism and real influence on retinal function are still unclear. To investigate the relation between retinal function and apoptosis, we studied photoreceptor apoptosis in experimental retinal detachment (RD). METHODS: We induced RD by subretinal injection of sodium hyaluronate in Brown Norway rats. Apoptotic photoreceptors were detected by TdT-dUTP Terminal Nick-End Labeling (TUNEL). To evaluate the function of the detached retina, electroretinograms (ERGs) were taken on day 1, 3 with corneal electrodes and full-field stimulation. RESULTS: Apoptotic DNA fragmentation appeared 12 hours after RD, was most prominent on day 3, and decreased thereafter. The ERGs showed that the amplitudes of dark-adapted a-waves and light adapted 2 Hz b-waves decreased immediately after RD and continued to decrease over time. The administration of Fas/Fc chimera recombinant protein or a caspase inhibitor, Z-VAD.fmk, failed to prevent either photoreceptor apoptosis or retinal functional damage. In contrast, brain derived neurotrophic factor (BDNF) and basic fibroblast growth factor (bFGF) significantly impeded both apoptosis and dysfunction. The ERGs recognized the functional changes sensitively, and these ERG changes correlated well to the amount of photoreceptor apoptosis. Immunohistochemical study showed that apoptosis-inducing factor (AIF), a novel caspase-independent apoptotic factor, was relocalized from mitochondria to the nucleus in this process. CONCLUSIONS: The present results showed that apoptosis was a key phenomenon in the retinal dysfunction in RD and that this process was transmitted mainly by mitochondria-dependent pathways rather than Fas/Fas-L or downstream caspase dependent pathways.

Animals↗
PubMed2002

Photocoagulation-induced retinal gliosis is inhibited by systemically expressed soluble TGF-beta receptor type II via adenovirus mediated gene transfer.

Retinal gliosis is one of the major causes of visual dysfunction due to the loss of the retinal regular structure and function in various diseases, including diabetic retinopathy, retinal detachment, and glaucoma. Transforming growth factor-beta (TGF-beta) is assumed to play an important role in this disease process. In the present study, we determined whether the systemically expressed extracellular domain of the TGF-beta type II receptor by adenovirus-mediated gene delivery could inhibit experimental retinal gliosis both in vitro and in vivo. Cultured bovine retinal glial cells, Müller cells, were stimulated by recombinant TGF-beta and the expression of the glial marker, glial fibrillary acidic protein (GFAP), was evaluated by immunohistochemistry, semiquantitative RT-PCR, and Western blotting. In cultured Müller cells, TGF-beta stimulated the GFAP expression in a dose-dependent fashion, and the conditioned medium from 293 cells transfected with adenovirus encoding for a soluble form TGF-beta type II receptor (AdT beta-ExR) inhibited the expression of GFAP stimulated by exogenous TGF-beta (p < 0.05). In this process, Smad4 protein, which plays a key role in intracellular signaling after cell surface receptors, actually translocated from cytosol to nucleus with TGF-beta stimulation. The conditioned medium from AdT beta-ExR also inhibited the cytosol-nuclear translocation of Smad4. For in vivo studies, AdT beta-ExR was injected into the femoral muscles of Brown Norway rats and retinal photocoagulation was subsequently carried out. Immunohistochemical studies revealed that GFAP was strongly expressed around the photocoagulation spots after 12 days and these phenomena were inhibited by AdT beta-ExR. Western blotting of total retinal extract demonstrated the same results as those observed after immunohistochemistry. Our results suggest that TGF-beta plays a pivotal role in the pathologic processes in retinal gliosis, and that the systemically expressed soluble TGF receptor by gene delivery may thus have a potential therapeutic value by inhibiting excessive retinal gliosis in various ocular diseases.

Adenoviridae↗
PubMedDate not supplied

Prefoveolar membrane in macular hole opercula formation.

PURPOSE: To better understand the process of macular hole opercula formation by both optical coherence tomography and intraoperative observations. METHODS: Seventy-nine eyes of 71 consecutive patients with stages 1 to 3 idiopathic macular holes were studied using optical coherence tomography (OCT). In eyes with stage 1 or 2 holes undergoing vitrectomy, meticulous observation of the posterior hyaloid and the macular hole was carried out before and after peeling of the posterior hyaloid. RESULTS: In 6 of 12 eyes with stage 1 holes, OCT showed tiny steps on the anterior wall of the foveal cyst, connecting to the detached posterior hyaloid face. In eyes with stage 2 holes, opercula were incompletely detached and connected to the hole edge. In eyes with stage 1 holes that were operated on, a small semitransparent opacity was noted at the posterior hyaloid face after peeling of the posterior hyaloid in the absence of defects of the anterior wall of the cyst. In 10 of 12 eyes with stage 2 holes undergoing vitrectomy, the size of the foveal opening remained unchanged after peeling of the posterior hyaloid, and a semitransparent opacity was observed at the detached hyaloid face. All opercula in stage 3 holes that were clearly imaged by OCT were positioned above the plane of the posterior hyaloid face. CONCLUSIONS: These findings suggest that the anterior wall of an evolving macular hole is composed of two layers: a prefoveolar membrane and the inner retinal layer. The prefoveolar membrane may play an important role in both persistent vitreofoveal adhesion and macular hole opercula formation.

Adult↗
PubMedDate not supplied

A nontraumatic macular hole in a 10-year-old girl.

BACKGROUND: Full-thickness macular holes usually develop in the elderly population. To the best of our knowledge, there has been no written report of a nontraumatic macular hole in a pediatric patient. CASE: A 10-year-old girl noticed decreased central vision in her left eye without any history of trauma. OBSERVATIONS: Fundus examination of the left eye revealed a full-thickness macular hole and a thin fibrous membrane on the superior peripapillary retina. She underwent standard macular hole surgery with stripping of the membrane, resulting in closure of the hole. CONCLUSIONS: A full-thickness macular hole may develop in pediatric patients. Although the etiology of the macular hole in the present patient is unclear, tangential traction induced by contraction of the peripapillary membrane, presumed to be an incomplete regression of the Bergmeister papilla, might have been responsible for the formation of the macular hole.

Child↗