Search

To investigate the utility of Optos ultrawide-field fundus autofluorescence (UWF-FAF) imaging for postoperative follow-up of gas-filled eyes after vitrectomy with subretinal tissue-plasminogen activator (t-PA) injection for subretinal hemorrhage (SRH) displacement.Retrospective consecutive case series.This study included 24 eyes with SRH. Vitrectomy with subretinal t-PA injection was performed, followed by postoperative prone positioning. FAF images acquired using Optos California were examined and the SRH occupancy in the macula was calculated. The main outcome measures were displacement rate and direction of SRH for 3 days postoperatively, and postoperative best-corrected visual acuity (BCVA).The postoperative BCVA ranged from improvement (23 eyes; 95.8%) to no change (one eye; 4.2%). Analysis was done using postoperative Optos FAF images for 20 eyes (83.3%). Postoperative SRH occupancy was significantly reduced, by 27.4%, compared with the preoperative occupancy (P = 0.03). A statistically significant reduction was found between the preoperative and postoperative day (POD)1 (P = 0.04), but not between POD1 and POD2 (P = 0.7), or between POD2 and POD3 (P = 1.0).UWF-FAF imaging is useful for postoperative follow-up of gas-filled eyes after vitrectomy with subretinal t-PA injection for SRH displacement.

To describe the factors associated with epiretinal membrane (ERM) formation in eyes treated with pars plana vitrectomy (PPV) for rhegmatogenous retinal detachment (RRD).Nationwide, multicenter, clinical cohort study based on registry data.We reviewed 2239 cases treated with PPV for RRD repair registered in the Japan-Retinal Detachment Registry between February 2016 and March 2017. Associations of 13 baseline characteristics and 8 surgical procedures with ERM formation were evaluated using univariate analysis. We conducted a propensity score-matched analysis for the significantly associated clinical factor(s). The primary outcome measure was ERM formation after 6 months of vitrectomy.ERM had developed in 104 cases (4.6%) by 6 months. We found that drainage retinotomy was significantly associated with ERM after multiple testing correction (odds ratio [OR] 2.22 [95% confidence interval {CI} 1.50-3.31]; P.001). In the propensity score-matched analysis (n = 492 in each group), we confirmed a significant difference in the incidence of ERM after 6 months of vitrectomy (8.3% and 2.6% in cases with and without drainage retinotomy, respectively; OR 3.35 [95% CI 1.77-6.33]; P.001).Eyes treated with PPV combined with drainage retinotomy are more likely to develop ERM postoperatively.

Purpose: To report optical coherence tomography angiography (OCTA) findings in a case of immune choroiditis following contralateral acute retinal necrosis (ARN) with choroidal neovascularization (CNV) during anti-vascular endothelial growth factor (VEGF) therapy. Case report: A 64-year-old woman with immune choroiditis following contralateral ARN and secondary CNV presented with decreased visual acuity. Fundus examination revealed macular and peripheral yellowish lesions in the right eye. Inflammatory cells were observed in the anterior chamber and the vitreous. OCT revealed retinal exudative changes and subretinal lesions suggestive of CNV. OCTA detected an abnormal vascular net in the outer retina as well as choriocapillaris, corresponding to type 2 CNV, that reduced following intravitreous anti-VEGF therapy. Two weeks after treatment, OCTA showed re-dilated choroidal neovasculature at the outer retina despite no exudative recurrence in OCT. Six weeks after treatment, OCT detected exudative changes around the neovascular lesion. Conclusion: This case discusses the use of OCTA detection of CNV in a case of immune choroiditis following contralateral ARN. During anti-VEGF therapy for inflammatory CNV-related diseases, OCTA may be useful not only for CNV detection but also for the follow-up of CNV activity.

To investigate periostin (PN) and tenascin-C (TNC) expression in the aqueous humor and trabeculectomy specimens of patients with neovascular glaucoma (NVG) secondary to proliferative diabetic retinopathy (PDR). This study enrolled 37 eyes of 37 patients who were grouped into (1) NVG secondary to PDR (NVG; n = 8); (2) PDR without NVG (PDR; n = 9); (3) primary open-angle glaucoma (POAG; n = 11); and (4) cataract surgery patients as a control group (CG; n = 9). Aqueous humor samples were collected from the anterior chamber at the start of surgery or intravitreal injection of anti-VEGF drug. The concentrations of PN, TNC, VEGF, and TGF-β2 (transforming growth factor-beta 2) were measured by ELISA. Sclerostomy tissues containing trabecular meshwork were obtained from two NVG patients and a POAG patient who underwent trabeculectomy surgery. Immunohistochemical analyses were performed to determine the localization of PN and TNC expression in the sclerostomy tissues. PN and TNC-C levels were below detection threshold in the POAG and CG groups. The NVG group had significantly higher levels of PN and TNC compared with the PDR group (84.7 ng/ml vs 2.2 ng/ml and 18.5 ng/ml vs 4.6 ng/ml, respectively; p

ObjectiveAccurate scleral marking of retinal breaks is essential for successful scleral buckling. This study aimed to investigate the use of wide-field fundus images obtained with an Optos for preoperative estimation of the distance from the limbus to the retinal breaks.Methods and analysisThis is a retrospective review of 29 eyes from 26 patients with rhegmatogenous retinal detachment who received scleral buckling with anatomically successful repair. They underwent wide-field fundus photography with Optos California. In the pre- and postoperative fundus images, we measured distances from the macula to the retinal tears (TM), to the center of the vortex veins (VM), to the optic disc (DM), and to the posterior edge of the scleral buckle (BM).Results(BM-VM) / DM was significantly correlated with the distance from the limbus to the posterior edge of the scleral buckle that had been determined intraoperatively. (r = 0.705; pConclusionPreoperative analysis of Optos images may be useful for estimating the distance from the limbus to retinal breaks, which might aid scleral marking during scleral buckling surgery.

Purpose: To evaluate the risk factors and the preventive effects of laser panretinal photocoagulation (PRP) for neovascular glaucoma (NVG) after ophthalmic stereotactic radiotherapy. Methods: Twenty-four patients with ocular malignant tumor (such as uveal malignant melanoma, lacrimal gland cancer) who received stereotactic radiotherapy (such as gamma knife, cyber knife) were retrospectively analyzed. Patients were divided into group A without preventive PRP (n=9), and group B with preventive PRP (n=15). Survival curves were plotted using the Kaplan-Meier method and compared between the two groups with the log-rank test. Results: In group A, NVG occurred in 3 patients. In contrast, in group B, no patient encountered NVG. The significant risk factor for NVG was dose-volumes that irradiated the optic disk (p=0.045). The incidence of NVG was significantly reduced in group B compared with group A (p=0.019). Conclusions: Dose-volumes that irradiated the optic disk were risk factors for NVG. PRP is effective in the prevention of NVG.

Purpose The purpose of this study was to evaluate the impact of intravitreal bevacizumab (IVB) on three cellular components (vascular endothelial cells, pericytes, and myofibroblasts) of the vascular microenvironment in fibrovascular membranes (FVMs) of patients with proliferative diabetic retinopathy. Methods Immunohistological studies with antibodies of CD34, αSMA, and transforming growth factor-β were performed on 20 surgical specimens obtained during a pars plana vitrectomy from 8 IVB-treated eyes, whereas 12 remained untreated. Four different indexes of vascular phenotype (vascular area, vascular major axis, CD34 endothelial area, and blood vessel density) and αSMA expression in vascular and stromal components were quantitatively analyzed. Results The intraluminal area of blood vessels, CD34 endothelial area, and the blood vessel density in IVB-treated FVMs were significantly less than in untreated FVMs. The number of CD34 blood vessels in IVB-treated FVMs was similar to that in untreated FVMs. Intravitreal bevacizumab could not affect vascular and stromal αSMA area significantly. However, the ratio of vascular αSMA area/CD34 area was significantly higher in IVB-treated FVMs than in untreated FVMs. Transforming growth factor-β expression could be observed in the IVB-treated FVM. Conclusion Intravitreal bevacizumab might primarily affect blood vessels, and the effects on pericytes and myofibroblasts might be secondary. Intravitreal bevacizumab treatment regulates vascular microenvironment by the contraction of blood vessels, the increasing pericyte ratio, and transforming growth factor-β expression in FVMs of patients with proliferative diabetic retinopathy.

Recent clinical observations have indicated that vascular endothelial growth factor (VEGF) is a key factor that stimulates the development of preretinal pathological neovascularization (NV). However, it has not been established how intraretinal physiological revascularization of hypoxic avascular areas is regulated. Our earlier study on the gene expression profile of hypoxic retinas in a mouse model of oxygen-induced retinopathy (OIR) showed that macrophage inflammatory protein-1β (MIP-1β) was the most upregulated protein. The purpose of this study was to investigate the role played by MIP-1β in recruiting bone marrow-derived monocyte lineage cells (BM-MLCs) in a mouse model of OIR. Our results showed that MIP-1β was upregulated, and its receptor, CCR5, was expressed in BM-MLCs in the hypoxic inner retina. Neutralizing Ab against MIP-1β reduced the infiltration of BM-MLCs into the OIR retinas and increased the avascular area and preretinal neovascular tufts. A very strong significant correlation was found between the area of the preretinal neovascular tufts and the avascular area, regardless of the extent of BM-MLC infiltration into the OIR retinas. Additional treatment with VEGF-A-neutralizing Ab showed that the MIP-1β-regulated pathological NV strongly depended on VEGF-A, which was probably secreted by the hypoxic avascular retinas. These results indicate that MIP-1β is involved in the recruitment of BM-MLCs, which have a significant role in the physiological revascularization of hypoxic avascular retinas. Overall, these findings indicate that the MIP-1β induction of BM-MLCs might possibly be used to promote intraretinal revascularization and thus prevent the abnormal NV in ischemic vision-threatening retinal diseases.

LYVE-1+ corneal lymphatics contribute to drainage and immunity. LYVE-1 is widely accepted as the most reliable lymphatic marker because of its continuous expression in lymphatic endothelium. LYVE-1 expression in corneal lymphatics has not been examined. In this study, we report intact CD31+ corneal lymphatic capillary endothelial cells that do not express LYVE-1. The number of LYVE-1− gaps initially increased until 8 wk of age but was significantly reduced in aged mice. C57BL/6 mice showed a notably higher number of the LYVE-1−/CD31+ lymphatic regions than BALB/c mice, which suggests a genetic predisposition for this histological feature. The LYVE-1− lymphatic gaps expressed podoplanin and VE-cadherin but not αSMA or FOXC2. Interestingly, the number of LYVE-1− gaps in FGF-2, but not VEGF-A, implanted corneas was significantly lower than in untreated corneas. Over 70% of the CD45+ leukocytes were found in the proximity of the LYVE-1− gaps. Using a novel in vivo imaging technique for visualization of leukocyte migration into and out of corneal stroma, we showed reentry of extravasated leukocytes from angiogenic vessels into newly grown corneal lymphatics. This process was inhibited by VE-cadherin blockade. To date, existence of lymphatic valves in cornea is unknown. Electron microscopy showed overlapping lymphatic endothelial ends, reminiscent of microvalves in corneal lymphatics. This work introduces a novel corneal endothelial lymphatic phenotype that lacks LYVE-1. LYVE-1− lymphatic endothelium could serve as microvalves, supporting unidirectional flow, as well as immunological hot spots that facilitate reentry of stromal macropahges.—Nakao, S., Zandi, S., Faez, S., Kohno, R., Hafezi-Moghadam, A. Discontinuous LYVE-1 expression in corneal limbal lymphatics: dual function as microvalves and immunological hot spots.

Neovascular glaucoma (NVG) is a serious complication for patients with proliferative diabetic retinopathy (PDR). Bevacizumab is a full-length humanized monoclonal antibody that binds all isoforms of vascular endothelial growth factor (VEGF). Recently, encouraging results regarding the off-label use of intravitreal bevacizumab (IVB) for the treatment of NVG have been reported. We evaluated the histology of bevacizumab-treated trabeculectomy specimens to clarify IVB’s biological effects on angle neovascularization. We retrospectively reviewed the charts of a consecutive series of 15 eyes of 13 patients who underwent trabeculectomy to treat NVG caused by PDR. In ten eyes of eight patients, 1.25 mg bevacizumab was injected intravitreally via the pars plana. Using light or electron microscopy, the surgically excised trabecular tissue was compared to that without IVB. Light microscopy revealed decreased edema, fibrin deposition, inflammation and vascular congestion in the trabecular meshwork in specimens with IVB compared to those without IVB. Electron microscopy revealed endothelial cell degeneration in the bevacizumab-treated specimens. The biological effects on angle neovascularization after IVB may involve reduced vascular permeability, decreased inflammatory reaction, loss of vascular function, and endothelial cell degeneration.

Background/aim Tumour necrosis factor-a (TNFa )i s an inflammatory cytokine that is upregulated in various vitreoretinal diseases including uveitis and diabetic retinopathy. Recently, our studies have indicated that hyalocytes contribute to the pathogenesis of these diseases. However, the impact of TNFa on the functional properties of hyalocytes is unknown. Methods Hyalocytes were isolated from bovine eyes. Cellular proliferation, migration and gel contraction in response to TNFa and the other inflammatory cytokines were analysed by thymidine uptake, Boyden’s chamber assay and collagen gel contraction assay, respectively. Furthermore, we estimated the effect of dexamethasone on these properties of hyalocytes. Results TNFa promoted proliferation, migration and gel contraction by hyalocytes. Dexamethasone inhibited TNFa-induced proliferation but not migration. Dexamethasone did not inhibit TNFa-induced gel contraction but further increased contraction. Furthermore, dexamethasone inhibited TNFa-induced extracellular signal-related kinase (ERK)1/2 phosphorylation in hyalocytes. Conclusion This study indicates that TNFa in vitreous and retina causes activation of hyalocytes, and the activated hyalocytes contribute to the pathogenesis of inflammatory vitreoretinal diseases. Steroid treatment appears to inhibit the activation of hyalocytes in the early stages of the diseases, but might have adverse effects in the late stage through membrane contraction.

Emerging evidence indicates that the tight communication between vascular endothelial cells and mural cells using platelet-derived growth factor (PDGF)-BB is essential for capillary stabilization during the angiogenic process. However, little is known about the related regulator that determines PDGF-BB expression. Using murine models of therapeutic neovascularization, we here show that a typical lymphangiogenic factor, vascular endothelial growth factor (VEGF)-C, is an essential regulator determining PDGF-BB expression for vascular stabilization via a paracrine mode of action. The blockade of VEGF type 3 receptor (VEGFR3) using neutralizing antibody AFL-4 abrogated FGF-2-mediated limb salvage and blood flow recovery in severely ischemic hindlimb. Interestingly, inhibition of VEGFR3 activity not only diminished lymphangiogenesis, but induced marked dilatation of capillary vessels, showing mural cell dissociation. In these mice, VEGF-C and PDGF-B were upregulated in the later phase after induced ischemia, on day 7, when exogenous FGF-2 expression had already declined, and blockade of VEGFR3 or PDGF-BB activities diminished PDGF-B or VEGF-C expression, respectively. These results clearly indicate that VEGF-C is a critical mediator, not only for lymphangiogenesis, but also for capillary stabilization, the essential molecular mechanism of communication between endothelial cells and mural cells during neovascularization.

Aim: To address the cellular components and the contractile mechanisms of the idiopathic epiretinal membrane (ERM). Methods: Ten surgically removed ERMs were fixed in 4% paraformaldehyde and analysed by whole-mount immunohistochemistry with anti-glial fibrillar acidic protein (GFAP) and alpha smooth-muscle actin (αSMA) antibodies. Type I collagen gel contraction assay, an established wound-healing assay in vitro, was performed using cultured bovine hyalocytes or normal human astrocytes (NHA) to evaluate the contractile property of the cells in the presence of tissue growth factor (TGF)-β2. The expression of αSMA was also analysed by western blot analysis to examine myofibroblastic transdifferentiation of the cells. Vitreous-induced collagen gel contraction was also evaluated. Results: All membranes were composed of αSMA immunopositive cells in contracted foci and GFAP immunopositive cells in the periphery. No apparent double positive cells were observed in any membranes examined. Cultured hyalocytes showed overexpression of αSMA and hypercontraction of collagen gels in response to TGF-β2, while glial cells showed marginal change. The vitreous from ERM patients also caused overexpression of αSMA and hypercontraction of the gels embedding hyalocytes, which were almost completely inhibited in the presence of anti-TGF-β2 neutralising antibody. Conclusions: Hyalocytes might be one of the critical components of ERM mediating its contractile property through the effect of TGF-β2 in the vitreous fluid.

To investigate the anatomical features of vitreoretinal interface in eyes with asteroid hyalosis (AH) with optical coherence tomography (OCT) and intravitreal triamcinolone acetonide (TA) during vitreous surgery.This study was an interventional clinical case series. Records relating to ten eyes from ten patients who underwent a TA-assisted vitrectomy for the treatment of diverse vitreoretinal diseases complicated with AH. The posterior vitreoretinal interface was examined by preoperative OCT and by intraoperative visualization of posterior vitreous cortex utilizing TA.In eight of ten AH eyes, preoperative OCT revealed abnormal vitreoretinal adhesions. In four of these eight eyes, posterior vitreoschisis could be seen on OCT. In the other four of these eight eyes, a clear no posterior vitreous detachment (PVD) pattern could be seen on OCT. Although posterior vitreous cortex could not be clearly identified with preoperative OCT in two of ten AH eyes, a complete PVD was refuted by intraoperative visualization of the posterior vitreous cortex with TA identical to the other eight eyes.These results indicate that complete PVD appears to be unlikely to occur in eyes with AH. In addition, spontaneous PVD in eyes with AH might lead to vitreoschisis or residual whole layer or posterior vitreous cortex, possibly due to anomalous vitreoretinal adhesion.

Background/aim: To examine the degree of the residual internal limiting membrane (ILM) after epiretinal membrane (ERM) peeling. Methods: Sixty-one eyes of 59 patients with ERM were enrolled. After ERM peeling, residual ILM was visualised with Brilliant Blue G (BBG). The residual ILM pattern was divided into three groups: (1) residual type (ILM mostly remained), (2) half type (approximately half of ILM remaind), (3) no residual type (ILM mostly removed with ERM). If ILM remained, residual ILM was removed in all cases and histologically examined using the flat mount method in 10 cases. The correlation between the degree of ERM evaluated by preoperative best-corrected visual acuity (BCVA) and residual ILM pattern was also examined. Results: Twenty-eight eyes (45.9%) were of the residual type. Three eyes (4.9%) were of the half type, and 30 eyes (49.2%) were of no residual type. The mean preoperative BCVA showed no significant correlation with the residual ILM pattern. Flat mount immunohistochemistry revealed many remnant cells, both glial fibrillar acidic protein positive and negative, on residual ILMs in all specimens examined. No recurrence that needed surgical treatment was observed. Conclusion: Residual ILM with remnant cells seems to be frequent after ERM removal. Intraoperative staining with BBG may be helpful in determining the extent of ILM removal.

Photoreceptor apoptosis is a critical process of retinal degeneration in retinitis pigmentosa (RP), a group of retinal degenerative diseases that result from rod and cone photoreceptor cell death and represent a major cause of adult blindness. We previously demonstrated the efficient prevention of photoreceptor apoptosis by intraocular gene transfer of pigment epithelium-derived factor (PEDF) in animal models of RP; however, the underlying mechanism of the neuroprotective activity of PEDF remains elusive. In this study, we show that an apoptosis-inducing factor (AIF)-related pathway is an essential target of PEDF-mediated neuroprotection. PEDF rescued serum starvation-induced apoptosis, which is mediated by AIF but not by caspases, of R28 cells derived from the rat retina by preventing translocation of AIF into the nucleus. Nuclear translocation of AIF was also observed in the apoptotic photoreceptors of Royal College of Surgeons rats, a well-known animal model of RP that carries a mutation of the Mertk gene. Lentivirus-mediated retinal gene transfer of PEDF prevented the nuclear translocation of AIF in vivo, resulting in the inhibition of the apoptotic loss of their photoreceptors in association with up-regulated Bcl-2 expression, which mediates the mitochondrial release of AIF. These findings clearly demonstrate that AIF is an essential executioner of photoreceptor apoptosis in inherited retinal degeneration and provide a therapeutic rationale for PEDF-mediated neuroprotective gene therapy for individuals with RP.

Background Recombinant Sendai virus vectors (rSeV) constitute a new class of cytoplasmic RNA vectors that have shown efficient gene transfer in various organs, including retinal tissue; however, the related immune responses remain to be overcome in view of clinical applications. We recently developed a novel rSeV from which all envelope-related genes were deleted (rSeV/dFdMdHN) and, in the present study, assess host immune responses following retinal gene transfer. Methods rSeV/dFdMdHN or conventional F-gene deleted rSeV (rSeV/dF) was injected into subretinal space of adult Wistar rats or C57BL/6 mice. The transgene expression and histopathological findings were assessed at various time points. Immunological assessments, including the expression of proinflammatory cytokines, natural killer (NK)-cell activity, as well as SeV-specific cytotoxic T lymphocytes (CTLs) and antibodies, were performed following vector injection. Results rSeV/dFdMdHN showed high gene transfer efficiency into the retinal pigment epithelium at an equivalent level to that seen with rSeV/dF. In the early phase, the upregulation of proinflammatory cytokines, local inflammatory cell infiltration and tissue damage that were all prominently seen in rSeV/dF injection were dramatically diminished using rSeV/dFdMdHN. NK cell activity was also decreased, indicating a reduction of the innate immune response. In the later phase, on the other hand, CTL activity and anti-SeV antibodies were similarly induced, even using rSeV/dFdMdHN, and resulted in transient transgene expression in both vector types. Conclusions Deletion of envelope-related genes of rSeV dramatically reduces the vector-induced retinal damage and may extend the utility for ocular gene transfer; however, further studies regulating the acquired immune response are required to achieve long-term transgene expression of rSeV. Copyright © 2007 John Wiley & Sons, Ltd.

Vessels of the lymphatic system are highly permeable and specialized for the uptake of fluid and macromolecules from the interstitium and their return to venous circulation (2, 21). Embryonic development of the lymphatic vessels starts when a subset of endothelial cells in the cardinal vein commits to the lymphatic lineage and sprouts to form the primary lymph sacs (2, 21, 22). Recent studies have identified specific transcription factors and growth factors required to regulate the development of lymphatic vessels. In mice, the lymphatic vasculature starts to develop at embryonic day 10.5 (E10.5), when the cardiovascular system is already functioning. After the formation of the initial lymph sacs, the peripheral lymphatics are generated by centrifugal sprouting. Several genes have been shown to play essential roles in embryonic lymphatic vessel development. The Prox1 homeobox transcription factor, vascular endothelial growth factor C (VEGF-C), and its receptor, vascular endothelial growth factor receptor 3 (VEGFR-3), are essential for the generation of lymphatics (2, 21). Hematopoietic intracellular signaling proteins, Syk, SLP-76, and PLCγ2, are important for the separation of lymphatic vessels from blood vessels (1, 30). Furthermore, the final patterning and maturation of lymphatic vasculature require angiopoietin 2, neuropilin 2, and podoplanin/T1 (2, 21). However, regulatory genes for lymphatic vessel development have not been identified yet. Recently, Sprouty/Spred family proteins were identified as negative regulators for growth factor- and cytokine-induced ERK activation (5, 9, 14, 29). In mammals, four Sprouty homologues and three Spred (for Sprouty-related Ena/VASP homology 1 domain-containing proteins) homologues have been identified (6, 14, 29). Spred-1 has been implicated in hematopoiesis, since bone marrow-derived mast cells and eosinophils from Spred-1−/− mice were more sensitive to interleukin 3 and interleukin 5, respectively, than those from wild-type (WT) mice (11, 20). In Spred-2−/− mice, embryonic hematopoiesis was enhanced in the aorta-gonad-mesonephros region compared with WT mice (19). However, except for a shortened face and slight growth retardation, neither Spred-1−/− nor Spred-2−/− mice showed strong developmental abnormalities in most organs (11, 19). To define overlapping functions of Spred-1 and Spred-2, we generated Spred-1/Spred-2 double-knockout (DKO) mice. We found that Spreds are key regulators of embryonic lymphangiogenesis and that they can specifically regulate VEGF-C signaling by suppressing VEGFR-3-mediated ERK and Akt activation.

Sprouty genes encode cytoplasmic membrane-associated proteins that inhibit receptor tyrosine kinase signaling. Four orthologs of Drosophila Sprouty (dSpry) (Sprouty1-4) have been identified in mammals. Physiological function of Sprouty1 and Sprouty2 has been investigated using gene targeting approaches, however to date detailed examination of Sprouty4 knockout (KO) mice has not been reported. In this study, Sprouty4 KO mice were generated and characterized. Although a significant fraction of Sprouty4 KO mice died shortly after birth due to mandible defects, the remainder were viable and fertile. Growth retardation was observed for most Sprouty4-deficient mice, with nearly all Sprouty4 KO mice having polysyndactyly. ERK activation was sustained in Sprouty4 KO mouse embryonic fibroblasts (MEFs) in response to FGF, but not to EGF. Sprouty2 and Sprouty4 double KO (DKO) mice were embryonic lethal and showed severe defects in craniofacial, limb, and lung morphogenesis. These findings suggest both redundant and non-redundant functions for Sprouty2 and Sprouty4 on embryonic development and FGF signaling.

Objective. The aim of this study is to evaluate the feasibility of extraperitoneal laparoscopic para-aortic and common iliac lymphadenectomy for cervical and endometrial carcinoma. Methods. Seventy-six patients underwent extraperitoneal laparoscopic para-aortic and common iliac lymphadenectomy between February 1999 and September 2005. The lymph nodes dissected with the laparoscopic procedure included the inframesenteric para-aortic lymph nodes, the sacral lymph nodes, and the bilateral common iliac lymph nodes. The extraperitoneal laparoscopic operation was performed with pelvic open surgery using Lap Disc to ensure the safety of patients. Results. The number of patients with cervical and endometrial carcinoma was 36 and 40, respectively. The median age of patients was 51 years (range 24–75 years). Conversion to open surgery was necessary in 8 patients. These include 3 patients who encountered blood loss of 400, 136 and 128 ml; 2 extremely obese women; and 3 patients who had peritoneal tears causing CO 2 gas leakage. Among the remaining 68 patients, the median operating time for extraperitoneal laparoscopic para-aortic and common iliac lymphadenectomy was 75 min (range 45–145 min), and the median estimated blood loss was 5 ml (range 5–138 ml). The median total number of resected nodes was 14 (range 2–31), and 4 patients had lymph node metastasis. No patient encountered postoperative complications attributable to extraperitoneal laparoscopic para-aortic and common iliac lymphadenectomy. Conclusions. Extraperitoneal laparoscopic para-aortic and common iliac lymphadenectomy with pelvic open surgery using Lap Disc is a feasible procedure, particularly in the surgeons learning phase.

Nagao S, Fujiwara K, Ishikawa H, Oda T, Tanaka K, Aotani E, Kohno I. Hormonal function after ovarian transposition to the abdominal subcutaneous fat tissue. Int J Gynecol Cancer 2006;16:121–124. We previously reported a new technique for ovarian transposition to the abdominal subcutaneous fat tissue (OTAFT) following hysterectomy. The purpose of this study is to assess the hormonal function after OTAFT. From 1993 to 2000, OTAFT was performed in 27 patients (group A). Forty-two women underwent hysterectomy and retained ovaries without transposition (group B). In 19 cases, bilateral oophorectomy with hysterectomy was performed, and they received a hormone replacement therapy (HRT) (group C). Serum follicle-stimulating hormone (FSH) level of patients was monitored every 2–12 months, and the time of menopause (defined as FSH >40 mIU/mL two times consecutively) was determined in groups A and B. After a median follow-up of 65 months, cumulative ovarian survival did not show significant difference between group A and group B (HR = 0.52, 95% CI = 0.17–1.16; P= 0.10). In patients who were 40 years old or younger, ovarian function declined significantly in group A compared to group B (HR = 0.29, 95% CI = 0.02–0.91; P= 0.04). However, FSH level of postmenopausal patients in group A was not different from FSH level of patients in group C, but FSH level of postmenopausal patients in group B was significantly higher than FSH level of patients in group C (P= 0.002). Although the procedure of OTAFT may somewhat affect the ovarian function, the transposed ovary in postmenopausal women presumably still secrete a small amount of estrogen which is equivalent to an estrogen level by HRT.

Objective. To clarify the pharmacological advantage of carboplatin-based intraperitoneal chemotherapy using the three-compartment mathematical model. Methods. Eleven consecutive patients in one institution underwent intraperitoneal administration of carboplatin, and 11 consecutive patients in another institution received intravenous administration. Carboplatin (AUC = 6 mg × min/ml) was diluted in 500 ml 5% glucose and administered either as an intraperitoneal bolus infusion or intravenous drip infusion during 1 h. Patients undergoing intravenous injection also received an infusion of 500 ml 5% glucose to obtain intraperitoneal samples. Intraperitoneal fluid and blood samples were obtained, immediately and 1, 2, 4, 8, 12, and 24 h after administration. The mathematical model consisting of a three-compartment model was applied to analyze the pharmacokinetics. The model was created with simultaneous differential equations and was solved by the Runge–Kutta method. Results. The rate constants of platinum diffusion from the peritoneal cavity to serum, serum to peritoneal cavity, serum to peripheral space, peripheral space to serum, and elimination were 0.94 ± 0.79 (mean ± SD), 1.28 ± 2.50, 16.50 ± 9.26, 0.99 ± 0.62, and 4.14 ± 1.45 (h −1 ), respectively. When the theoretical pharmacological concentration of platinum was calculated using this mathematical model, 24-h platinum AUC in the serum was exactly the same regardless of intraperitoneal or intravenous administration of carboplatin. However, the 24-h platinum AUC in the peritoneal cavity was approximately 17 times higher when carboplatin was administered by the intraperitoneal route. Conclusion. The present pharmacological analysis suggests that intraperitoneal infusion of carboplatin is feasible not only as an intraperitoneal regional therapy but also as a more reasonable route for systemic chemotherapy.

Objective. Although the Calvert formula is the standard method to calculate the dose of carboplatin, there is no consensus how to determine the glomerular filtration rate (GFR) without using [ 51 Cr]-ethylenediamine tetraacetic acid ( 51 Cr-EDTA). Creatinine clearance (Ccr), calculated using the Cockroft–Gault, Jelliffe, Modified-Jelliffe or Wright formulae, has been used as a substitute for the GFR. In addition to these four formulae, the Chatelut formula has been proposed as a way to calculate carboplatin clearance. Among these formulae, Jelliffe formula does not include body surface area (BSA) or body weight to adjust the body size and thus may have greater bias than the other four formulae in estimating carboplatin clearance. The purpose of this study is to evaluate if these five formulae could equally estimate the carboplatin clearance. Methods. Carboplatin clearance was estimated in 253 patients with gynecologic cancer who received carboplatin-based chemotherapy between January 1996 and August 2004. Ccr was estimated using the Cockroft–Gault, Jelliffe, Modified-Jelliffe or Wright formulae. Carboplatin clearance was also calculated directly by using the Chatelut formula. The five estimations of carboplatin clearance were compared with each other using the post-hoc Wilcoxon signed rank test. The median percent error (MPE) and the median absolute percent error (MAPE) were evaluated by comparing carboplatin clearance. The relationships between BSA and ratios of estimated carboplatin clearance (Jelliffe/Cockroft–Gault, Jelliffe/Modified-Jelliffe, Jelliffe/Wright, Jelliffe/Chatelut) were evaluated by using simple regression. Results. The estimated carboplatin clearances were: Cockroft–Gault formula, 109.8 ± 28.4; Jelliffe formula, 128.5 ± 28.2; Modified-Jelliffe formula, 110.8 ± 25.7; Wright formula, 112.2 ± 24.3; Chatelut formula, 114.1 ± 33.0. A statistically significant difference was observed between carboplatin clearance calculated using Jelliffe formula and that given by each of the other four formulae. Comparing the results of the Cockroft–Gault formula with the Jellife, Modified-Jelliffe, Wright and Chatelut formulae yielded MPEs of +19%, +2%, +3% and +3% and MAPEs of 27%, 5%, 6% and 6%, respectively. There was a significant correlation between BSA and ratio of estimated carboplatin clearance (Jelliffe/Cockroft–Gault, Jelliffe/Modified-Jelliffe, Jelliffe/Wright, Jelliffe/Chatelut), with Pearson correlation coefficients of 0.928, 0.847, 0.965 and 0.839 respectively. As the BSA of patient became smaller, the differences between the carboplatin clearance calculated by the Jelliffe formula from other four formulas became larger. Conclusions. Estimates of carboplatin clearance calculated by the Jelliffe formula tend to have greater positive bias compared to the other four formulae, particularly when the BSA of the patient is small. In order to conduct collaborative international studies, it may be necessary to standardize the formula used to estimate carboplatin clearance to perform international collaboration studies.

Objective A recent study demonstrated that docetaxel (DTX) was an effective agent for second line chemotherapy against ovarian cancer. Weekly administration of taxane compounds had been more effective compared with a 3 week interval administration in ovarian cancer. The role of biweekly administration of DTX has been unknown. We conducted a dose determination and feasibility study of biweekly DTX administration in patients with ovarian cancer. Methods Patients with histologically confirmed epithelial ovarian cancer who received one or more regimens of prior chemotherapy with more than 4 weeks of treatment-free interval were eligible. DTX was administered as 1-h intravenous infusion every two weeks for at least four courses. The starting dose was 40 mg/m 2 (level 1) and the dose was escalated to 50 mg/m 2 (level 2) and 60 mg/m 2 (level 3) in consequent patient cohorts. Results Nine patients were examined in this study. The treatments were completely performed in all cohorts. Mean treatment delay ranged from 0 to 2.0 days. Dose level did not affect treatment delay. At the first dose level, no patients experienced grade 3/4 neutropenia. Two patients in level 2 experienced grade 3/4 neutropenia. In level 3, all patients had grade 4 neutropenia. Nonhematologic toxicities were tolerable. Of eight patients with measurable disease, all patients in level 1 showed progressive disease, and all patients in level 2 were no-change. There were two responders showing complete response and partial response and one case was no-change in level 3. Conclusion The present study showed that biweekly administration of 60 mg/m 2 DTX was feasible for recurrent ovarian cancer.

This multicenter collaborative study prospectively evaluated the effect of omentoplasty and omentopexy on the prevention of complications after pelvic lymphadenectomy. Sixty-four consecutive patients (42 cervical and 22 endometrial cancer) were enrolled and examined periodically for 12 months. All patients underwent simple, semiradical, or Okabayashi's radical hysterectomy and complete pelvic lymphadenectomy. The infracolic omentum was longitudinally divided in half and omentoplasty was performed so that bilateral omental flaps would reach the pelvic floor. The omental flaps were inserted into the retroperitoneal space and the edges of the flaps were sutured to the psoas muscle. The omental flap was then covered by the peritoneum. Incidence of lymphocele, lymphedema, and severe complications associated with lymphocele, such as infection or urinary stenosis, was evaluated at intervals for at least one year after surgery.Among the 64 patients, 12 patients received pelvic radiation because of occult lymph node metastasis. Planned omentoplasty was not possible in one patient because her omentum was too small; therefore, only unilateral omentopexy was performed. Asymptomatic lymphoceles only were detected by ultrasonogram in 12 patients (18.8%). Three patients (4.7%) had a symptomatic but pressure-only lymphocele. Hydronephrosis and bladder compression probably due to lymphocele were observed in one patient, respectively (3.1%), but resolved within 6 months. Lymphedema was observed in seven patients (10.9%) and persisted for more than 6 months in two patients (3.1%). We conclude that this simple technique of omentoplasty and omentopexy appeared to be effective in reducing the incidence of complications after pelvic lymphadenectomy.

Carboplatin has proven to be effective in the treatment of patients with recurrent ovarian cancer who have responded to platinum-based first-line chemotherapy. However, thrombocytopenia is a problem in heavily pretreated patients. Paclitaxel has a so-called platelet-sparing effect, but until now it was unknown whether this effect could be obtained in heavily pretreated patients. We treated four patients with recurrent epithelial ovarian cancers who had previously undergone heavy chemotherapy. Because these patients had responded to platinum-based first-line therapy, single carboplatin administration was employed. Although satisfactory tumor marker responses were obtained, thrombocytopenia occurred. When paclitaxel was administered before the carboplatin infusion in the subsequent treatments, the thrombocytopenia became milder. It was concluded that a platelet-sparing effect was also observed in patients who had undergone heavy prior chemotherapy.

Fujiwara K, Suzuki S, Yoden E, Ishikawa H, Imajo Y, Kohno I. Local radiation therapy for localized relapsed or refractory ovarian cancer patients with or without symptoms after chemotherapy.The purpose of this paper is to prospectively evaluate the effects of local radiation therapy upon localized ovarian cancer following chemotherapy. Patients with objective relapses or refractory disease but with localized epithelial ovarian cancers and who had undergone at least one regimen of chemotherapy were enrolled in this study. External irradiation was performed on all patients. Twenty patients, with a mean age of 53.8 ± 10.3 y, were enrolled in this study. The median number of previous chemotherapies was 2. The interval between previous chemotherapy and radiation therapy was 4.5 months. The maximum diameter of the lesions was 3.6 ± 1.8 cm. The irradiation dose was 52.3 ± 8.3 Gy. Neither hematologic nor intestinal toxicity >grade 3 was observed. Forty-four disease sites, including the lymph nodes, vaginal cuff, pelvis, abdomen, subcutaneous regions, and the brain were irradiated. Thirty of these sites were symptom-free before irradiation. In patients with symptoms, the symptomatic relief was obtained in approximately 50% of patients. Smaller lesions (P = 0.024) and lymph nodes (P = 0.042) demonstrated better responses than larger lesions or other sites, respectively. Regression rates correlated with longer survivals (P = 0.0195) after radiation therapy. Survival was significantly better when radiation therapy was given before patients had symptoms (P = 0.001). Survival was also better in patients with lymph node disease only (P = 0.0069). We conclude that local radiation therapy may be one of the treatment options for relapsed or refractory but localized ovarian cancer, particularly when the tumor is small and/or located in the lymph nodes, even when patients had no symptoms.

The choroid is an area of the eye commonly affected both by metastatic tumors and by primary malignant melanomas [1]. Although the accuracy of the clinical diagnosis of uveal melanoma has improved [2], atypical ocular manifestations may lead to misdiagnosis in some cases. In the case presented here, we observed choroidal metastasis with melanocytic proliferation that mimicked malignant choroidal melanoma.

Indoleamine 2,3-dioxygenase (IDO)-initiated tryptophan degradation in the placenta has been implicated in the prevention of the allogeneic fetus rejection [Munn, Zhou, Attwood, Bondarev, Conway, Marshall, Brown, and Mellor (1998) Science 281, 1191-1193]. To determine how IDO is associated with the development of the fetus and placenta, the time course of IDO expression (tryptophan-degrading activity, IDO protein and IDO mRNA) in the embryonic and extra-embryonic tissues as well as maternal tissues of mice was examined. A high tryptophan-degrading activity was detected in early concepti on days 6.5 and 7.5, whereas IDO protein and its mRNA were not expressed during early gestation, but appeared 2-3 days later, lasted for about 3 days and declined rapidly thereafter. The expression of IDO basically coincided with the formation of the placenta. On the contrary, the early tryptophan-degrading activity was due to gene expression of tryptophan 2,3-dioxygenase (TDO), as shown by Northern and Western analysis. These findings indicate that IDO is transiently expressed in the placenta but that the expression does not last until birth, and that the IDO expression is preceded by expression of another tryptophan-degrading enzyme, TDO, in the maternal and/or embryonic tissues in early concepti.

A case of florid endocervical glandular hyperplasia with pyloric gland metaplasia is described. MR images showed multiple and conjugated cysts in the uterine cervix, which suggested adenoma malignum. A postoperative examination revealed florid but definitely benign endocervical glandular hyperplasia. Hitherto described radiologic features considered suggestive of adenoma malignum are not necessarily specific and can be a diagnostic pitfall.

Surgical repair of a radiation-induced vesico-vaginal fistula by a simple closure technique is extremely difficult. Here, we report a patient with a radiation-induced vesico-vaginal fistula that was successfully reconstructed using a gracilis myocutaneous flap. The vesico-vaginal fistula developed in an 83-year-old Japanese woman who had undergone a radical hysterectomy for cervical cancer of the uterus at age 63, and had pelvic radiation therapy for recurrent disease 17 years after the surgery. The fistula was repaired by a reconstruction technique that employed a gracilis myocutaneous flap. Two-thirds of the vaginal mucosa on the side where the fistula was present was completely removed to obtain a wide mucosal defect. Irradiated fibrous tissue around the fistula was removed, and the bladder mucosa was mobilized, and then closed with absorbable sutures. An island gracilis myocutaneous flap was designed on the medial aspect of the patient's left thigh. The first incision was made through the proximal outline of the flap, and its vascular pedicle, the medial circumflex femoral vessels, was dissected. After a complete incision through the flap, the skin flap, including the entire gracilis muscle, was elevated with the pedicle vessel. The distal portion of the myocutaneous flap was then turned to overlay the mucosal defects on the vagina. Postoperatively, there was no flap necrosis and no infection in the vaginal cavity. Indwelling catheterization was required for 8 weeks. Retrograde cystography confirmed complete closure of the fistula.

BACKGROUND: Although cytologic findings of clear cell adenocarcinoma of the female genital tract have been reported sporadically, the background on the smear has received little attention. CASE: A 16-year-old female had a large, necrotic mass in the cervix. As cervical brushing cytology and a punch biopsy of the mass could not make a definitive diagnosis, fine needle aspiration cytology was performed from the mass. The smears revealed loose, three-dimensional clusters and sheet arrangements of atypical cells. Dispersed atypical cells were also seen. The atypical cells were large and had abundant, weakly stained cytoplasm and round or oval nuclei with large nucleoli. There were a few tumor cells with clear cytoplasm and distinct cell borders in Papanicolaou-stained smears. The background in Diff-Quik-stained smears revealed a tigroid background and basement membrane material. CONCLUSION: The malignant tumor, revealing both a tigroid background and basement membrane material, seems to have been clear cell adenocarcinoma. Both features are diagnostic clues to clear cell adenocarcinoma in the female genital tract. Our case indicates that aspiration cytology is also an effective method of diagnosing a cervical tumor when the tumor is polypoid and the surface is extensively necrotic.

We describe three cases of florid endocervical glandular hyperplasia with intestinal and pyloric gland metaplasia, which can be a benign mimic of adenoma malignum. In two cases, adenoma malignum was seriously considered preoperatively because of watery vaginal discharge and the results of imaging studies. The three cases shared common histopathological features, i.e., (i) proliferating endocervical glands surrounded by clusters of smaller glands, resembling the pyloric glands of the stomach; (ii) occasional intestinal metaplasia; (iii) bland nuclear features; and (iv) predominantly PAS-positive neutral mucin in the glandular epithelium. In two cases, glands were densely and irregularly arranged in some areas. Immunohistochemistry disclosed that the intracytoplasmic mucin of the metaplastic epithelium was positive for M-GGMC-1 (HIK1083), which reacts with mucin of pyloric glands. Monoclonal CEA was negative in all cases. This pseudoneoplastic benign condition should be recognized by both gynecologists and pathologists, although it might be difficult to establish a definite diagnosis preoperatively even with deep cone biopsy.

Background. There is no standard formula to estimate doses for the intraperitoneal (IP) administration of carboplatin. We evaluated a combination of the Cockcroft and the Calvert (Cockcroft-Calvert) formula to determine the area under the curve (AUC) for IP carboplatin co-administered with intravenous (IV) cyclophosphamide (CPM). We also evaluated the correlation of carboplatin clearance determined by the Chatelut formula with carboplatin clearance determined by the Cockcroft-Calvert formula. Methods. We performed a retrospective study of the records of 149 treatments in 30 patients who received IP carboplatin and IV CPM for ovarian carcinoma. The glomerular filtration rate was calculated with the Cockcroft formula. Carboplatin doses were determined based on the body surface area. The Cockcroft-Calvert formula was used to calculate the AUC. The Chatelut formula was also used to calculate the clearance of carboplatin and the AUC. Results. The AUC calculated with the Cockcroft-Calvert formula was well correlated to the AUC calculated with the Chatelut formula (r 2 = 0.965). During the first four courses of IP carboplatin combined with IV CPM (300–500mg/m2), the correlation between the percent decrease in platelet count and the calculated carboplatin AUC varied among methods: Cockcroft-Calvert formula AUC:r 2 = 0.460; Chatelut formula AUC:r 2 = 0.431; body surface area dose:r 2 = 0.204; total dose:r 2 = 0.143. Conclusion. To decrease patient platelet count by 67%, the optimal target AUC following IP administration of carboplatin in combination with 300–500mgCPM/m2IV was calculated as 6.5, using the Cockcroft-Calvert formula, and as 7.5, using the Chatelut formula. Considerable modification of the IP carboplatin dose is required after the fourth course. A prospective study is ongoing to confirm these results.

A case of papillary squamous cell carcinoma (PSCC) of the uterine cervix is reported. The patient was a 73-year-old Japanese woman with acute renal failure and bilateral hydronephrosis. A cauliflower-like mass was found in the uterine cervix. A uterine cervical biopsy specimen revealed PSCC in situ, while clinically it was an invasive carcinoma. Uterine cervical biopsy was performed a second time to confirm its stromal invasion. However, only small fragments were obtained because of heavy bleeding from the tumor and they showed PSCC in situ again. Following this, computed tomography of the pelvis revealed a 5 cm mass in the uterine cervix, invading the vagina and urinary bladder. Though deep-wedge biopsy, loop electrosurgical excision, or cone biopsy is recommended to evaluate PSCC, it may be impossible to perform any of these procedures because of bleeding such as that seen in our case. In these circumstances, good communication between pathologists and clinicians is important since lack of communication may cause PSCC to be microscopically misinterpreted as in situ carcinoma rather than invasive carcinoma.

The aim of this study was to retrospectively evaluate the effect of the combination of preoperative donation and acute normovolemic hemodilution of the autologous blood in gynecologic operations. The records of 42 patients undergoing extensive gynecologic operations with a blood loss of 800 ml and who received either no autologous transfusion (n = 7), hemodilution alone (n = 19) or the combination of preoperative donation and hemodilution (n = 16) were reviewed. The incidence and volume of allogeneic blood transfusion and postoperative anemia were analyzed. The incidence of allogeneic blood transfusions was 85.7%, 26.3% (P = 0.02), and 20% (P

Purpose Inflammation and immune cells regulate choroidal neovascularization (CNV) and could become therapeutic targets in age-related macular degeneration (AMD). Lymphangiogenesis is a key component of various inflammatory diseases. Whether lymphangiogenesis and lymph node-mediated immunity are involved in the pathogenesis of AMD is not understood. Methods To investigate lymphangiogenesis in CNV, we generated CNV in animals by laser and studied surgically removed CNV membranes from uveitis and AMD patients. Immunohistochemistry was performed with lymphatic vessel endothelial hyaluronate receptor 1 (LYVE-1) and podoplanin antibodies. VEGF-C and VEGFR-3 expressions were examined with immunohistochemistry and Western blotting. To examine the role of lymph node in CNV, we lasered lymphotoxin alpha-deficient mice (LTα-/-) and measured the CNV volume. Results Immunohistochemistry showed that LYVE-1(+) macrophages infiltrated in acutely induced CNV, although lymphatic tubes did not form. CNV membranes from patients did not show LYVE-1(+)podoplanin(+) vessels, suggesting the lack of lymphangiogenesis in AMD and uveitis. Western blots and immunostaining revealed VEGF-C and VEGFR-3 expression in CNV lesions, mainly in macrophages and angiogenic endothelial cells. Using fluorescent microsphere tracers, we show a path for cellular migration from the eye to the cervical lymph nodes (LNs) during CNV. However, CNV injury did not cause LN swelling. CNV volume did not differ between wild-type and LN-deficient mice, suggesting that LN is not a key component of early CNV formation. Conclusions Laser-induced CNV is not primarily dependent on acquired immunity, nor does the fundus injury affect peripheral LNs. Our results reveal a previously unknown cellular connection between the ocular fundus and the cervical LNs. This connection that in function resembles lymphatics is actively utilized in CNV.

PURPOSE. Anti–VEGF-A antibody (Ab) (e.g., bevacizumab, ranibizumab) is widely used as a treatment against retinal angiogenesis and edema. The purpose of this study was to evaluate whether intravitreal anti-VEGF Ab injection modulates inflammatory cells in retinal angiogenesis. METHODS. To investigate whether intravitreal bevacizumab injections affect the number of inflammatory cells in proliferative diabetic retinopathy (PDR) membranes in patients, immunohistochemical staining with CD45 Ab (pan-leukocyte marker) was performed using the surgically obtained membranes in pars plana vitrectomy with or without pretreatment with bevacizumab. To check whether anti–VEGF-A Ab affects leukocytes going in and out of blood vessels during retinal angiogenesis, the authors performed their novel leukocyte transmigration assay and CD45 immunostaining in a mouse model of oxygen-induced retinopathy (OIR). RESULTS. The authors’ new imaging approach revealed that intravitreal injection of anti–VEGF-A Ab blocks leukocyte infiltration as well as angiogenesis. The Ab injection inhibited leukocyte transmigration before affecting the angiogenenic area. CD45 staining showed no significant difference in the leukocyte number in the angiogenic retina or the human PDR membranes between the anti–VEGF-A Ab injected group and the control group. Furthermore, VEGF-A inhibition also affected leukocytes going out from the retina. CONCLUSIONS. Intravitreal injection of anti-VEGF-A Ab could inhibit leukocyte trafficking in the retina, suggesting that antiVEGF-A therapy could serve as a treatment in retinal inflammation. (Invest Ophthalmol Vis Sci. 2012;53:4323– 4328) DOI:10.1167/iovs.11-9119

Asteroid hyalosis (AH) is a condition in which cream-colored or white spherical particles are suspended in the vitreous body. Asteroid hyalosis is considered not to cause decreased vision or any other visual symptoms except in rare cases. There have been a few reports of AH in patients with retinitis pigmentosa (RP).To assess the prevalence of AH in patients with RP, 320 patients with typical forms of RP were studied. One patient was offered a standard three-port vitrectomy, and the spherical particles obtained from her vitrectomy sample were analyzed using an energy-dispersive x-ray spectrometer.Ten patients (two men and eight women) developed AH. Among them, four had bilateral AH and two had rapidly increasing vitreous opacity that led to decreased vision. One patient was a 48-year-old woman with progressive AH in the left eye. After treatment with a vitrectomy, her vision improved from 0.4 to 0.8. The spherical particles were composed of mainly calcium and phosphorus.The prevalence of AH in RP was higher than in previous reports, and we encountered two rare cases of progressive AH with decreased vision. We conclude that AH might lead to decreased vision in patients with RP.

While cytoplasmic features of ovarian surface epithelial tumors are well-known, the nuclear findings have received little attention. We reviewed imprint cytology materials of the ovary which were collected at the Kawasaki Medical School Hospital between January 1989-July 1999, and identified 15 mucinous cystadenomas, 3 borderline mucinous tumors, 4 mucinous cystadenocarcinomas, 4 serous cystadenomas, 4 borderline serous tumors, 7 serous cystadenocarcinomas, 6 endometrioid carcinomas, and 2 clear-cell adenocarcinomas. We microscopically observed nuclear findings of the 45 cases. Coffee-bean nuclei were observed in 15.0%, 15.8%, and 10.1% of the tumor cells in mucinous adenomas, borderline mucinous tumors, and borderline serous tumors, respectively. The frequencies of the coffee-bean nuclei in the three tumors were higher than in the remaining tumors (P < 0.001). Intranuclear cytoplasmic inclusions were observed in 2.1% of the tumor cells in mucinous cystadenoma, and their frequency was significantly higher than that in cases of other surface epithelial ovarian tumors (P < 0.001). Semilunar-shaped nuclei were seen in all cases of mucinous cystadenomas and borderline mucinous tumors, and in 3 of 4 mucinous adenocarcinomas. The remaining surface epithelial tumors did not reveal the semilunar-shaped nuclei. In the cytology of the ovary, the semilunar nuclei are characteristic of mucinous tumors, and the intranuclear cytoplasmic inclusion may be a diagnostic clue to mucinous cystadenoma, when it is conspicuous. The coffee-bean nuclei can be seen in mucinous cystadenoma, borderline mucinous tumors, and borderline serous tumors.

Purpose To examine the histopathologic effect of a single intravitreal injection of bevacizumab on newly formed vessels in eyes with proliferative diabetic retinopathy (PDR). Design Interventional case series and laboratory investigation. Methods Two days after intravitreal injection of bevacizumab (1.25 mg/eye), pars plana vitrectomy or trabeculectomy was performed for the treatment of PDR or neovascular glaucoma (NVG) associated with PDR. Ten surgically removed preretinal proliferative tissues and 6 deep scleral flaps containing trabecular meshwork were fixed in 2% glutaraldehyde or 4% paraformaldehyde and were subjected to transmission electron microscopic analysis, immunohistochemical analysis, and terminal deoxyuridiine triphosphate (dUTP) nick-end labeling staining. Two surgically removed preretinal proliferative tissues and 2 deep scleral flaps from patients with PDR and NVG, but without preoperative intravitreal injection of bevacizumab (IVB), served as controls. Results In control tissues, vascular endothelial cells possessed many fenestrations and were accompanied by pericytes. Apoptotic vascular endothelial cells frequently were observed in tissue after intravitreal injection of bevacizumab, whereas they were not observed in control tissues. Additionally, no apparent fenestration was observed in newly formed vessels from either proliferative tissue or trabecular meshwork after intravitreal injection of bevacizumab. In both PDR and NVG tissues after intravitreal injection of bevacizumab, overexpression of smooth muscle actin was observed in newly formed vessels, suggesting that the treatment may have increased pericytes on the vasculature as compared with control tissue. Conclusions Intravitreal injection of bevacizumab may induce changes in immature, newly formed vessels of PDR or NVG tissue, leading to endothelial apoptosis with vascular regression, while inducing normalization of premature vessels by increasing pericyte coverage and reducing vessel fenestration.

While statins have an anti-angiogenic property, their underlying mechanisms are not fully understood. We investigated intracellular mechanisms of simvastatin-mediated reduction in VEGF-induced signalings.The effects of simvastatin on cell proliferation and viability were evaluated by [(3)H]-thymidine incorporation in retinal endothelial cells (RECs) and cell counting. The impact of simvastatin on VEGF-induced phosphorylation of p44/42 mitogen-activated protein (MAP) kinase, myosin light chain (MLC), and VEGF-receptor (VEGFR) 2 were examined by Western blotting. Involvement of the mevalonate pathway in VEGF-induced signaling was also examined.Simvastatin (1 and 10 microM) suppressed VEGF-induced RECs proliferation in a concentration-dependent manner, without affecting cell viability. Simvastatin significantly inhibited VEGF-induced phosphorylation of VEGFR2 and its downstream mediators, p44/42 MAP kinase and MLC. Mevalonate completely reversed VEGF-induced VEGFR2 phosphorylation, but only partially reversed the phosphorylation of p44/42 MAP kinase and MLC.These data indicate that simvastatin exerts its anti-angiogenic effects through the reduction of VEGFR2 phosphorylation in RECs at least in part. However, there seems to be both mevalonate-dependent and independent pathway in simvastatin's anti-angiogenic property.