Mikiko Fukushima

Vitreous fluid levels of β-amyloid(1–42) and tau in patients with retinal diseases

PurposeA decrease in β-amyloid1–42 (Aβ42) and an increase in tau in the cerebrospinal fluid are reported to be characteristic phenomena in Alzheimer’s disease patients. To test the idea that Aβ42 and tau contribute to the development of retinal diseases, we measured Aβ42 and tau concentrations in the vitreous fluid from patients with macular hole (n = 13), diabetic retinopathy (n = 15), or glaucoma concurrent with other ocular diseases (n = 8).MethodsVitreous samples were collected from patients who underwent vitrectomy, and sensitive and specific enzyme-linked immunosorbent assays were used to determine the concentrations of Aβ42 and tau.ResultsBy comparison with the levels in the control macular-hole patients (33.9 ± 7.1 pg/ml for Aβ42; 3.3 ± 3.2 pg/ml for tau), there was a significant decrease in the Aβ42 level and a significant increase in the tau level in patients with diabetic retinopathy (1.8 ± 1.9 pg/ml for Aβ42, P = 0.002; 153.7 ± 71.6 pg/ml for tau, P = 0.041) or glaucoma concurrent with other ocular diseases (2.8 ± 1.8 pg/ml for Aβ42, P = 0.006; 113.6 ± 43.1 pg/ml for tau, P = 0.023).ConclusionsOur findings indicate the possibility of a role for Aβ42 and tau in the pathogenesis of some retinal diseases.

Activation of Canonical Wnt Pathway Promotes Proliferation of Retinal Stem Cells Derived from Adult Mouse Ciliary Margin

Adult retinal stem cells represent a possible cell source for the treatment of retinal degeneration. However, only a small number of stem cells reside in the ciliary margin. The present study aimed to promote the proliferation of adult retinal stem cells via the Wnt signaling pathway. Ciliary margin cells from 8‐week‐old mice were dissociated and cultured to allow sphere colony formation. Wnt3a, a glycogen synthase kinase (GSK) 3 inhibitor, fibroblast growth factor (FGF) 2, and a FGF receptor inhibitor were then applied in the culture media. The primary spheres were dissociated to prepare either monolayer or secondary sphere cultures. Wnt3a increased the size of the primary spheres and the number of Ki‐67–positive proliferating cells in monolayer culture. The Wnt3a‐treated primary sphere cells were capable of self‐renewal and gave rise to fourfold the number of secondary spheres compared with nontreated sphere cells. These cells also retained their multilineage potential to express several retinal markers under differentiating culture conditions. The Wnt3a‐treated cells showed nuclear accumulation of β‐catenin, and a GSK3 inhibitor, SB216763, mimicked the mitogenic activity of Wnt3a. The proliferative effect of SB216763 was attenuated by an FGF receptor inhibitor but was enhanced by FGF2, with Ki‐67–positive cells reaching over 70% of the total cells. Wnt3a and SB216763 promoted the proliferation of retinal stem cells, and this was partly dependent on FGF2 signaling. A combination of Wnt and FGF signaling may provide a therapeutic strategy for in vitro expansion or in vivo activation of adult retinal stem cells.

Trabeculectomy with Mitomycin C for Neovascular Glaucoma: Prognostic Factors for Surgical Failure

PURPOSE To evaluate the prognostic factors for surgical outcomes of trabeculectomy with mitomycin C (MMC) for neovascular glaucoma (NVG). DESIGN Retrospective cohort study. METHODS We reviewed the medical records of 101 patients (101 eyes) with NVG treated at Kumamoto University Hospital. The primary endpoint was persistent intraocular pressure > or = 22 mm Hg, deterioration of visual acuity to no light perception, and additional glaucoma procedures. Multivariate analysis was performed using the Cox proportional hazards model. RESULTS The mean follow-up period was 29.3 months (range, 0.5 to 142.3 months). The probability of success 1, 2, and 5 years after trabeculectomy was 62.6%, 58.2%, and 51.7%, respectively. The multivariate model showed that younger age (relative risk [RR], 0.96/year; P = .0007) and previous vitrectomy (RR, 1.62; P = .02) were prognostic factors for surgical failure among all NVG patients. Additionally, an eye with unremoved proliferative membrane and/or unrepaired retinal detachment (RD) after vitrectomy (RR, 1.59; P = .05) was a probable prognostic factor in a subgroup of 66 eyes with previous vitrectomy, and having a fellow eye with NVG (RR, 1.73; P = .003) was a significant prognostic factor in 82 eyes with NVG attributable to diabetic retinopathy. CONCLUSIONS The prognostic factors for surgical failure of trabeculectomy with MMC for NVG were younger age and previous vitrectomy in all NVG patients, and having a fellow eye with NVG in patients with disease caused by diabetic retinopathy. Persistent proliferative membrane and/or RD after vitrectomy might contribute to poorer outcomes of trabeculectomy.

Elevated erythropoietin in vitreous with ischemic retinal diseases

The aim of the current study was to measure the concentrations of erythropoietin in the vitreous fluid and analyze its association with vascular endothelial growth factor (VEGF) in ischemic vitreoretinal diseases. Vitreous fluid samples were collected from patients with proliferative diabetic retinopathy, branch retinal vein occlusion and idiopathic macular hole. Concentrations of erythropoietin and VEGF in vitreous fluid were significantly elevated in patients with proliferative diabetic retinopathy and branch retinal vein occlusion as compared to patients with macular hole. There were no differences in serum concentrations of erythropoietin and VEGF among patient groups. There was significant correlation between erythropoietin and VEGF concentrations in vitreous fluid. Erythropoietin was up-regulated in ischemic disorders and may act as an endogenous neuroprotective factor against ischemic retinal disorders.

Vitreous opacities and outcome of vitreous surgery in patients with familial amyloidotic polyneuropathy

PURPOSE To report the prevalence of vitreous opacities and the outcome of vitreous surgery in patients with familial amyloidotic polyneuropathy (FAP). DESIGN Observational case series. METHODS In 37 patients with FAP and the ATTR Val30 Met mutation, vitreous opacities were present in 14 eyes of 9 patients and vitrectomy combined with phacoemulsification and intraocular lens implantation was performed in five eyes of three patients. In six patients with the ATTR Tyr114Cys mutation, vitreous opacities were present in both eyes of all six patients and vitrectomy combined with phacoemulsification and intraocular lens implantation was performed in nine eyes of six patients. The mean follow-up period after vitreous surgery was 20.9 +/- 16.8 months (range, 3 to 52 months). RESULTS The prevalence of vitreous opacities is much higher in patients with ATTR Tyr114Cys (100%) than in those with ATTR Val30 Met (24%). The mean age at the onset of vitreous opacities was significantly lower in the patients with ATTR Tyr114Cys (37.0 +/- 5.3 years) than in the nine patients with ATTR Val30 Met (52.8 +/- 9.1 years; P <.005). Visual acuity improved in all 14 eyes after vitreous surgery; however, final visual acuity decreased in one eye owing to the occurrence of a central retinal vein occlusion. Vitreous opacities mildly increased in two eyes. CONCLUSIONS Our data suggest that the ATTR Val30 Met and ATTR Tyr114Cys mutations induce different clinical features of vitreous opacities. Vitreous surgery combined with phacoemulsification and implantation of an intraocular lens is a safe and useful treatment. Careful long-term follow-up should be performed.

Combined intravitreal bevacizumab and trabeculectomy with mitomycin C versus trabeculectomy with mitomycin C alone for neovascular glaucoma.

PurposeTo evaluate the effects of intravitreal bevacizumab (IVB) before mitomycin C trabeculectomy (MMCT) for neovascular glaucoma (NVG). MethodsThe study is a retrospective, comparative, consecutive case series. The study group consisted of 57 eyes from 50 patients with NVG who underwent a first MMCT: 33 eyes were treated with MMCT alone between June 1, 2005 and May 17, 2007 (Control Group); and, 24 eyes were treated with a combination of IVB and MMCT after May 18, 2007 (IVB Group). Surgical complications, intraocular pressure (IOP), and the probability of success were compared between the 2 groups. Surgical failure was defined as IOP ≥22 mm Hg for 2 consecutive follow-up visits, a deterioration of visual acuity to no light perception, or additional glaucoma surgeries. ResultsThere were no significant differences in preoperative data between the groups. Hyphema associated with MMCT occurred significantly less often in the IVB Group (P=0.006). IOPs at 7 and 10 days after MMCT were significantly lower in the IVB Group (P=0.01 and 0.02, respectively). However, Kaplan-Meier survival-curve analysis showed the probability of success 120, 240, and 360 days after MMCT of 87.5%, 79.2%, and 65.2% in the IVB Group, and 75.0%, 71.9%, and 65.3% in the Control Group. No significant difference in survival times was found between the groups (P=0.76). ConclusionsIVB before MMCT reduced hyphema associated with MMCT for NVG. IVB provided further IOP reduction immediately after MMCT, but did not significantly improve surgical outcomes over longer periods.

Suppression of Choroidal Neovascularization by N-Acetyl-Cysteine in Mice

Purpose: N-acetyl-cysteine (NAC) is a potent antioxidant known to be a precursor of glutathione. The purpose of this study was to investigate the role of NAC in the development of choroidal neovascularization (CNV). Methods: CNV was induced in C57BL/6 mice by laser photocoagulation of the ocular fundus. Mice were injected intraperitoneally with NAC or vehicle alone. The levels of 4-hydoroxy-2-nonenal (4-HNE)-modified protein and nucleus factor (NF)-κB were determined by wester blotting. The recruitment of macrophages and neutrophils after laser injury was analyzed immunohistochemically and in myeloperoxidase (MPO) assays. Enzyme-linked immunosorbent assays (ELISA) were used to measure monocyte chemotactic protein (MCP)-1, CXCL1, vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR)-1, and VEGFR-2. The extent of CNV was evaluated 7 d after laser injury by lectin staining. Results: In NAC-treated mice with laser-induced injuries, the induction of 4-HNE-modified protein after 3 hr and the activation of NF-κB in nuclear extracts after 6 hr were markedly suppressed compared to vehicle-treated mice. Macrophage and neutrophil recruitment were inhibited and the levels of MCP-1, CXCL1, VEGF, and VEGFR-1 were also lower in NAC-treated mice compared to vehicle-treated mice. Furthermore, the extent of CNV induced was significantly lower in NAC-treated compared to vehicle-treated mice (p = 0.027). Conclusions: Our results clearly showed that NAC inhibited indicators of oxidative stress and the activation of NF-κB induced by laser injury, and, consequently, suppressed macrophage and neutrophil infiltration and the development of CNV. This suggests novel preventative and interventional therapeutic strategies for age-related macular degeneration.

Developmental and denervation changes in c-ret proto-oncogene expression in chick motoneurons

Receptor tyrosine kinases (RTKs) play important roles in cellular differentiation, survival, and proliferation. To search for genes involved in the survival of motoneurons, we isolated the RTKs specifically expressed on them. We used RT-PCR, by which RNA was obtained from purified embryonic day 5 (E5) chick motoneurons, and screened by in situ hybridization. Of 17 RTK cDNA clones, c-ret expression gradually increased in the motoneurons of the spinal cord during development. Compared with the expression of the neurotrophin receptor, TrkC, that of c-ret was very high in motoneurons from around E17 to adulthood. The level of c-ret expression on the motoneurons was unaffected by deafferentation at E2, but changed after denervation at post-hatching day 2. The in situ hybridization signal for c-ret mRNA increased slightly at day 1, then decreased progressively up to day 8, and increased again 2 weeks after sciatic nerve denervation in the motoneurons of the lumbar spinal cord. There were also changes in the signal of the lesioned sciatic nerve. In the most distal part from the lesioned site, the signal for c-ret mRNA significantly increased from day 3 on after denervation. These results suggested that the c-ret gene may encode the receptor for a factor involved in motoneuron differentiation and the promotion of regeneration of injured peripheral nerves.

Involvement of the Hipk family in regulation of eyeball size, lens formation and retinal morphogenesis

Members of the homeodomain‐interacting protein kinase (HIPK) family are involved in various intracellular regulatory mechanisms. The present study focused on clarifying the functions of HIPK family members in ocular organization during late embryogenesis. HIPK1 and HIPK2 were expressed in the inner retina during late embryogenesis. Hipk1 +/− Hipk2 −/− mice had a greater frequency of small eyes with a lens deficiency and abnormally laminated and thickened retinas than did wild‐type littermates. These data indicate that Hipk1 and Hipk2 are involved in regulation of eye size, lens formation and retinal lamination during late embryogenesis.

Pitavastatin: Protection against Neuronal Retinal Damage Induced by Ischemia-Reperfusion Injury in Rats

Purpose: To evaluate the neuroprotective effects of pitavastatin against neuronal retinal damage induced by ischemia-reperfusion injury in rats. Methods and Results: Ischemia-reperfusion injury was induced in Sprague-Dawley rats using ocular hypertension. Pitavastatin (0.1, 0.5, or 1.0 mg/kg) was given intravenously 12 hr or 5 min before, or 12 or 24 hr after the induction of ischemia-reperfusion injury. Morphometric and retrograde labeling analyses revealed neuroprotective effects when pitavastatin (0.5 mg/kg) was administered 5 min before—even 12 and 24 hr—after induction of ischemia-reperfusion injury. These effects depended on dose; protection was noted at pitavastatin concentrations of 0.5 and 1 mg/kg but not 0.1 mg/kg. Furthermore, preadministration of pitavastatin (0.5 mg/kg) reduced expression of P-selectin and intercellular adhesion molecule-1 at 12 and 24 hr after induction of ischemia-reperfusion injury. Conclusions: As pitavastatin was efficacious in preventing retinal neuronal death, it may be a novel therapeutic modality for ischemic retinal diseases.