Nobuo Jo

Inhibition of Experimental Abdominal Aortic Aneurysm in the Rat by Use of Decoy Oligodeoxynucleotides Suppressing Activity of Nuclear Factor κB and ets Transcription Factors

Background—Two phenomena, inflammation and matrix degradation, contribute to the progression of abdominal aortic aneurysm (AAA). Importantly, the inflammation is regulated by the transcription factor nuclear factor (NF)–&kgr;B, whereas the destruction and degradation of elastin fibers by matrix metalloproteinases (MMP) are regulated by ets. Thus, we developed a novel strategy to treat AAA by simultaneous inhibition of both NF-&kgr;B and ets by using chimeric decoy oligodeoxynucleotides (ODN). Methods and Results—AAA was induced in rats by transient aortic perfusion with elastase, whereas transfection of decoy ODN was performed by wrapping a delivery sheet containing decoy ODN around the aorta. Gel-mobility shift assay at 7 days after treatment demonstrated that both NF-&kgr;B and ets binding activity were simultaneously inhibited by chimeric decoy ODN. Transfection of chimeric decoy ODN resulted in significant inhibition of the progression of AAA such as aneurysmal dilation at 4 weeks after treatment as compared with control, accompanied by a reduction of MMP expression. Moreover, the destruction of elastin fibers was inhibited in the aorta transfected with chimeric decoy ODN. Importantly, transfection of chimeric decoy ODN demonstrated potent inhibition of aneurysmal dilatation compared with NF-&kgr;B decoy ODN alone, whereas scrambled decoy ODN had no effects. Interestingly, the migration of macrophages was significantly inhibited by chimeric decoy ODN. Conclusions—We demonstrated that inhibition of the progression of AAA was achieved by a novel strategy with chimeric decoy ODN used against NF-&kgr;B and ets in rat model. NF-&kgr;B and ets are considered to play an important role in the pathogenesis of AAA.

Pigment epithelium derived factor as a neuroprotective agent against ischemic retinal injury.

Purpose. Pigment epithelium-derived factor (PEDF) is a protein shown to have neurotrophic activity. The purpose of this study was to determine whether PEDF is neuroprotective of retinal neurons that are exposed to transient ischemia-reperfusion. Methods. Transient retinal ischemia was produced by increasing the intraocular pressure for 45 min in albino rats eyes. Immediately after reperfusion, PEDF was injected intravitreally into the experimental eyes. Injury was evaluated morphologically and by measuring the thickness of the inner retinal layers (IRL) and by counting the number of retinal ganglion cells (RGC) in epon embedded sections. Results. Morphologic and morphometric analysis of the thickness of the IRL and the counting of RGC demonstrated that PEDF injected immediately after reperfusion protected the eyes partially but significantly from the ischemic injury. Cocnlusions. Intravitreal injection of PEDF even after the ischemia can ameliorate retinal injury. PEDF may be useful in preventing neuronal degeneration in the inner retina resulting from ischemia.

In Vivo Evidence of Angiogenesis Induced by Transcription Factor Ets-1 Ets-1 Is Located Upstream of Angiogenesis Cascade

Background—A transcription factor, ets-1, regulates the transcription of metalloproteinase genes, the activity of which is necessary for matrix degradation and the migration of endothelial cells. However, no study has demonstrated that ets-1 itself has an angiogenic action in vivo. Thus, we examined (1) the effects of overexpression of the ets-1 gene on angiogenesis in a rat hindlimb ischemia model, and (2) how ets-1 induced angiogenesis. Methods and Results—In this study, we used the HVJ-liposome method, which is highly effective for transfection, to transfect the human ets-1 gene. At 4 weeks after transfection, the capillary density and blood flow were significantly increased in a hindlimb transfected with the human ets-1 gene compared with control. These data clearly demonstrated that ets-1 has the ability to stimulate angiogenesis in vivo. To elucidate the molecular mechanisms by which ets-1 induced angiogenesis, we focused especially on the expression of hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF), potent angiogenic growth factors, because the promoter regions of both genes contain ets binding sites. Interestingly, overexpression of ets-1 upregulated both tissue HGF and VEGF concentrations in rat hindlimb. More importantly, administration of neutralizing antibody against HGF and VEGF attenuated the increase in blood flow and BrdU-positive cells induced by ets-1. Upregulation of HGF and VEGF by ets-1 was also confirmed by in vitro experiments using human vascular smooth muscle cells. Conclusions—The present study demonstrated that ets-1 regulated angiogenesis through the induction of angiogenic growth factors (VEGF and HGF). Overexpression of ets may provide a new therapeutic strategy to treat peripheral arterial disease.

Upregulation of pigment epithelium-derived factor after laser photocoagulation

PURPOSE To determine the changes in the expression of pigment epithelium-derived factor in cultured human retinal pigment epithelial cells and rat retinas after laser photocoagulation. METHODS Experimental study of laser photocoagulation on human retinal pigment epithelial cells in culture and on adult rats. Reverse transcription-polymerase chain reaction and semiquantitative polymerase chain reaction analysis were used. RESULTS After photocoagulation, the mRNA expression of pigment epithelium-derived factor was upregulated in human retinal pigment epithelial cells at 6 hours and then gradually decreased. Compared with controls, significantly higher levels of pigment epithelium-derived factor were observed in rat retinas from 6 to 24 hours after laser photocoagulation (P <.005), and they were still higher than before photocoagulation at 2 weeks. CONCLUSION An upregulation of pigment epithelium-derived factor in retinal pigment epithelial cells and in the retina after photocoagulation suggests that pigment epithelium-derived factor plays a role in inhibiting neovascularization by its antiangiogenic activity.

Levels of vascular endothelial growth factor and pigment epithelium-derived factor in eyes before and after intravitreal injection of bevacizumab.

PurposeBevacizumab is a human monoclonal IgG1 antibody that blocks the action of vascular endothelial growth factor (VEGF). The purpose of this study was to determine the level of VEGF and pigment epithelium-derived factor (PEDF) in eyes with proliferative diabetic retinopathy (PDR) before and after an intravitreal injection of bevacizumab.MethodsEleven eyes of ten patients were studied. Patients were included if they had neovascular glaucoma, rubeosis of the iris with PDR, or aggressive PDR. Samples of aqueous humor were collected just before the injection of bevacizumab and the vitrectomy. The concentrations of VEGF and PEDF in the aqueous humor were measured by enzyme-linked immunosorbent assay, and the effects of bevacizumab on PDR were evaluated.ResultsThe free VEGF concentration before the injection was 676.5 ± 186.7 pg/ml (mean ± SEM, n = 11). Seven days later, it was significantly reduced to 7.1 ± 7.1 pg/ml (P < 0.005, n = 9). The PEDF concentration before the injection was 2.32 ± 0.49 μg/ml (n = 11), and 7 days later, it was 3.23 ± 0.76 μg/ml (P = 0.33). During the vitrectomy, patients had less intraoperative bleeding when the neovascular tissues were cut.ConclusionsAn intravitreal injection of bevacizumab significantly decreased the free VEGF in the aqueous humor by 7 days, indicating that the clinical effects of bevacizumab appear rapidly. However, intravitreal bevacizumab did not affect the level of intraocular PEDF.

Pars Plana Vitrectomy with Removal of Posterior Hyaloid Face in Treatment of Refractory Diabetic Macular Edema Resistant to Triamcinolone Acetonide

BackgroundTriamcinolone acetonide (TA) has recently been used to treat diabetic macular edema (DME) but its effectiveness is limited.CasesThree patients (three eyes) with unresolved diffuse DME who did not respond to a posterior sub-Tenons injection of TA underwent vitrectomy.ObservationsIntraoperatively, it was found that all of the eyes had a posterior hyaloid face that was adherent to a large area of the posterior pole retina, although this had not been detected by slit-lamp biomicroscopy or optical coherence tomography. After vitrectomy and removal of the posterior hyaloid face, there was a significant reduction in the central macular thickness of all three eyes and an improvement in the visual acuity of the patients.ConclusionsWhen TA treatment is not effective for DME, vitrectomy with the complete removal of the posterior hyaloid face, including removal of the internal limiting membrane, should be considered.

Effective transfection of a cis element "decoy" of the nuclear factor-κB binding site into the experimental choroidal neovascularization

Purpose. To evaluate the efficacy of the gene transfer of a double-stranded phosphorothioate oligonucleotides (ODNs), called a “decoy”, against the NF-?B binding site into cells of an experimentally-induced choroidal neovascularization. Methods. FITC-labeled decoy was injected into the subretinal space of rat eyes by the HVJ-liposome delivery system, and 3 days later, choroidal neovascularization was induced by laser photocoagulation. The eyes were removed and the transfected cells were detected by fluorescence microscopy and also detected by immunohistochemistry. The degree of neovascularization was evaluated by fluorescein angiography. Results. The decoy was transfected into the retinal pigment epithelial (RPE) cells, inner and outer segment of the photoreceptors at 3 days after the injection. When choroidal neo-vascularization was induced, highly effective transfection of the decoy was observed 3 to 14 days after photocoagulation, after which the level decreased. Decoys were transfected into the RPE cells and macrophages in the choroidal neovascularization. The eyes transfected with NF-?B decoy showed a weaker leakage in fluorescein angiograms than that of the control eyes transfected with scrambled decoy. Conclusions. A decoy can be transfected into retinal cells and cells within a choroidal neovascularization by the HVJ-liposome method. The transferred NF-?B decoy reduced the degree of choroidal neovascularization. Decoy targeted against NF NF-?B may be considered as a potential therapy for neovascularization.

Intraocular Pressure (IOP) Reduction by Latanoprost in Japanese Normal Tension Glaucoma Patients Over a Five-Year Period Stratified by Presenting IOP

PURPOSE We examined the effectiveness of latanoprost for reducing intraocular pressure (IOP) in Japanese patients with normal tension glaucoma (NTG) over a 5-year period. DESIGN Prospective interventional case series. The patients were classified into 2 groups based on mean IOP. METHODS A total of 38 patients with NTG were studied after being classified into the high-tension (mean IOP 16 mmHg or greater, n = 27) and low-tension (mean IOP lower than 15 mmHg, n = 11) groups. IOP was measured and Humphrey Field Analyzer (HFA) examinations were conducted at 6, 12, 24, 36, 48, and 60 months after beginning a daily administration of latanoprost. RESULTS Mean IOP before administration was 17.6 mmHg in the high-tension group, which was reduced to 13.9, 14.6, 14.4, 14.1, 13.6, and 14.6 mmHg at 6, 12, 24, 36, 48, and 60 months, respectively, after beginning administration. That in the low-tension group was 13.6 mmHg before administration, and then was reduced to 12.2, 11.4, 11.5, 12.5, 10.5, and 11.5 mmHg, respectively, after beginning administration was noted. Mean deviation (MD) values in the HFA examinations were reduced by -4.27 and -1.49 dB after 5 years in the high- and low-tension groups, respectively. CONCLUSIONS Latanoprost administration was effective in reducing IOP over a 5-year period in a range of 3.1-4.1 and 1.3-3.6 mmHg in NTG patients with high- and low-tension levels, respectively. In addition, our results indicate that latanoprost helped to prevent a decrease in MD values in both groups, as shown by the results of HFA examinations.