Wilms' tumor gene WT1 promotes homologous recombination-mediated DNA damage repair.

The Wilms' tumor gene WT1 is overexpressed in leukemia and various types of solid tumors and plays an oncogenic role in these malignancies. Alternative splicing at two sites yields four major isoforms, 17AA(+)KTS(+), 17AA(+)KTS(-), 17AA(-)KTS(+), and 17AA(-)KTS(-), and all the isoforms are expressed in the malignancies. However, among the four isoforms, function of WT1[17AA(-)KTS(+)] isoform still remains undetermined. In the present study, we showed that forced expression of WT1[17AA(-)KTS(+)] isoform significantly inhibited apoptosis by DNA-damaging agents such as Doxorubicin, Mitomycin, Camptothesisn, and Bleomycin in immortalized fibroblast MRC5SV and cervical cancer HeLa cells. Knockdown of Rad51, an essential factor for homologous recombination (HR)-mediated DNA repair canceled the resistance to Doxorubicin induced by WT1[17AA(-)KTS(+)] isoform. GFP recombination assay showed that WT1[17AA(-)KTS(+)] isoform alone promoted HR, but that three other WT1 isoforms did not. WT1[17AA(-)KTS(+)] isoform significantly upregulated the expression of HR genes, XRCC2, Rad51D, and Rad54. Knockdown of XRCC2, Rad51D, and Rad54 inhibited the HR activity and canceled resistance to Doxorubicin in MRC5SV cells with forced expression of WT1[17AA(-)KTS(+)] isoform. Furthermore, chromatin immunoprecipitation (ChIP) assay showed the binding of WT1[17AA(-)KTS(+)] isoform protein to promoters of XRCC2 and Rad51D. Immunohistochemical study showed that Rad54 and XRCC2 proteins were highly expressed in the majority of non-small-cell lung cancer (NSCLC) and gastric cancer, and that expression of these two proteins was significantly correlated with that of WT1 protein in NSCLCs. Our results presented here showed that WT1[17AA(-)KTS(+)] isoform had a function to promote HR-mediated DNA repair.

Block suture for trabeculectomy using only 2 anchors.

We examined both a parallel and a cross technique in comparison with operations not using block suture. Of the 44 eyes that underwent surgery without block suture, aqueous leakage occurred in 4 eyes (9.0%). Of 51 eyes that underwent the parallel technique, 1 eye (2.0%) leaked from the limbus incision. In contrast, leaks were not detected in any of the 58 eyes that underwent the cross technique.

Indocyanine green angiography in a case of punctate inner choroidopathy associated with acute zonal occult outer retinopathy.

BACKGROUND: The etiology of punctate inner choroidopathy (PIC) and acute zonal occult outer retinopathy (AZOOR) are currently unknown, although both diseases are hypothesized to be part of the spectrum of a single disorder. CASE: We report indocyanine green (ICG) angiographic findings in a 24-year-old woman in whom PIC was associated with AZOOR. OBSERVATIONS: The patient had been diagnosed with bilateral PIC. She noticed a visual field defect in her right eye 8 months later, and a defect in her left eye 25 months later. Perimetry showed an arcuate scotoma (right eye) and a central scotoma (left eye), which could not be explained by funduscopy or late-phase fluorescein angiography. Multifocal electroretinography led to a diagnosis of bilateral AZOOR. ICG angiography demonstrated late multiple patchy hypofluorescent spots throughout the posterior pole in the right eye and geographic hypofluorescence in the macular region from initial to late phase in the left eye. However, these findings were not evident at the onset of PIC. CONCLUSION: These angiographic results suggest that at the onset of AZOOR, choriocapillaris injuries had occurred in the areas corresponding to the scotomas.

Expression of erythropoietin receptor in human epiretinal membrane of proliferative diabetic retinopathy.

PURPOSE: It is widely accepted that intravitreous levels of erythropoietin (Epo) are elevated in patients with ischaemic retinal diseases such as proliferative diabetic retinopathy (PDR). The aim of this study was to examine the expression of Epo and the Epo receptor (EpoR) in epiretinal membranes with and without diabetes. METHODS: Eighteen epiretinal membranes (PDR (n = 10), idiopathic epiretinal membranes (IERMs) without diabetes (n = 4) and inner limiting membranes (ILMs) (n = 4)) were obtained during pars plana vitrectomy. Formalin-fixed and paraffin-embedded tissues were examined by immunohistochemistry with anti-Epo and EpoR antibodies. RESULTS: The histopathological findings demonstrated that PDR membranes consisted of a variety of endothelial cells forming a microvascular cavity with red blood cells and non-vascular stromal mononuclear cells. Membranous and cytoplasmic immunoreactivity for EpoR was strongly detected in endothelial cells and stromal cells in all PDR patients. Although microvessels were not observed in IERMs and ILMs, immunoreactivity for EpoR was noted in the cellular component of IERMs, and was weakly detected in ILMs. Epo was not expressed in any membrane. CONCLUSION: EpoR was strongly expressed in microvessels of all PDR membranes. The in vivo evidence in this study suggests that Epo in the vitreous binds to EpoR in PDR membranes, which subsequently leads to the proliferation of new retinal vessels. EpoR immunoreactivity in non-vascular stromal cells in PDR membranes, and IERMs and ILMs might be indirectly correlated with ischaemia.

Positive association of pigment epithelium-derived factor with total antioxidant capacity in the vitreous fluid of patients with proliferative diabetic retinopathy.

BACKGROUND: Pigment epithelium-derived factor (PEDF), a glycoprotein with potent neuronal differentiating activity, was recently found to inhibit advanced glycation end product (AGE)-induced retinal hyperpermeability and angiogenesis through its antioxidative properties, suggesting that it may exert beneficial effects on diabetic retinopathy by acting as an endogenous antioxidant. However, the inter-relationship between PEDF and total antioxidant capacity in the eye remains to be elucidated. AIMS: To determine vitreous PEDF and total antioxidant levels in patients with proliferative diabetic retinopathy (PDR), and to investigate the relationship between them. METHODS: Vitreous levels of PEDF and total antioxidant capacity were measured by an ELISA in 39 eyes of 36 patients with diabetes and PDR and in 29 eyes of 29 controls without diabetes. RESULTS: Vitreous levels of total antioxidant capacity were significantly lower in patients with diabetes and PDR than in controls (mean (SD) 0.16 (0.05) vs 0.24 (0.09) mmol/l, respectively, p<0.001). PEDF levels correlated positively with total antioxidant status in the vitreous of patients with PDR (r = 0.37, p<0.05) and in controls (r = 0.41, p<0.05). Further, vitreous levels of PEDF in patients with PDR without vitreous haemorrhage (VH(-)) were significantly (p<0.05) decreased, compared with those in the controls or in patients with PDR with vitreous haemorrhage (VH(+); PDR VH(-), 4.5 (1.1) microg/ml; control, 7.4 (4.1) microg/ml; PDR VH(+) 8.5 (3.6) microg/ml). CONCLUSION: This study demonstrates that PEDF levels are associated with total antioxidant capacity of vitreous fluid in humans, and suggests that PEDF may act as an endogenous antioxidant in the eye and could play a protective role against PDR.