Sofia Theodoropoulou

Heat Shock Protein 70 (HSP70) Is Critical for the Photoreceptor Stress Response after Retinal Detachment via Modulating Anti-Apoptotic Akt Kinase

Photoreceptor apoptosis is a major cause of vision loss in many ocular diseases. Significant progress has been made to elucidate the molecular pathways involved in this process, yet little is known about proteins counteracting these apoptotic pathways. It is established that heat shock proteins (HSPs) function as molecular helper proteins (chaperones) by preventing protein aggregation and facilitating refolding of dysfunctional proteins, critical to the survival of all organisms. Here, we investigated the role of HSP70 on photoreceptor survival after experimental retinal detachment (RD) in mice and rats. We found that HSP70 was up-regulated after RD and associated with phosphorylated Akt, thereby preventing its dephosphorylation and further activation of cell death pathways. Administration of quercetin, which inhibits HSP70 and suppresses Akt phosphorylation significantly increased photoreceptor apoptosis. Similarly, RD-induced photoreceptor apoptosis was augmented in mice carrying hypomorphic mutations of the genes encoding HSP70. On the other hand, administration of geranylgeranylacetone, which induces an increase in HSP70 significantly decreased photoreceptor apoptosis after RD through prolonged activation of Akt pathway. Thus, HSP70 may be a favorable potential target to increase photoreceptor cell survival after RD.

Retinoblastoma cells are inhibited by aminoimidazole carboxamide ribonucleotide (AICAR) partially through activation of AMP-dependent kinase

5‐Aminoimidazole‐4‐carboxamide‐1‐β‐4‐ribofuranoside (AICAR), an analog of AMP, is widely used as an activator of AMP‐kinase (AMPK), a protein that regulates the responses of the cell to energy change. We studied the effects of AICAR on the growth of retinoblastoma cell lines (Y79, WERI, and RB143). AICAR inhibited Rb cell growth, induced apoptosis and S‐phase cell cycle arrest, and led to activation of AMPK. These effects were abolished by treatment with dypiridamole, an inhibitor that blocks entrance of AICAR into cells. Treatment with the adenosine kinase inhibitor 5‐iodotubericidin to inhibit the conversion of AICAR to ZMP (the direct activator of AMPK) reversed most of the growth‐inhibiting effects of AICAR, indicating that some of the antiproliferative effects of AICAR are mediated through AMPK activation. In addition, AICAR treatment was associated with inhibition of the mammalian target of rapamycin pathway, decreased phosphorylation of ribosomal protein‐S6 and 4E‐BP1, down‐regulation of cyclins A and E, and decreased expression of p21. Our results indicate that AICAR‐induced activation of AMPK inhibits retinoblastoma cell growth. This is one of the first descriptions of a nonchemotherapeutic drug with low toxicity that may be effective in treating Rb patients.—Theodoropoulou, S., Kolovou, P. E., Morizane, Y., Kayama, M., Nicolaou, F., Miller, J. W., Gragoudas, E., Ksander, B. R., Vavvas, D. G. Retinoblastoma cells are inhibited by aminoimidazole carboxamide ribonucleotide (AICAR) partially through activation of AMP‐dependent kinase. FASEB J. 24, 2620–2630 (2010). www.fasebj.org

Aminoimidazole Carboxamide Ribonucleotide (AICAR) Inhibits the Growth of Retinoblastoma In Vivo by Decreasing Angiogenesis and Inducing Apoptosis

5-Aminoimidazole-4-carboxamide-1-β-4-ribofuranoside (AICAR), an analog of AMP is widely used as an activator of AMP-kinase (AMPK), a protein that regulates the responses of the cell to energy change. Recently, we showed that AICAR-induced AMPK activation inhibits the growth of retinoblastoma cells in vitro by decreasing cyclins and by inducing apoptosis and S-phase arrest. In this study, we investigated the effects of AMPK activator AICAR on the growth of retinoblastoma in vivo. Intraperitoneal injection of AICAR resulted in 48% growth inhibition of Y79 retinoblastoma cell tumors in mice. Tumors isolated from mice treated with AICAR had decreased expression of Ki67 and increased apoptotic cells (TUNEL positive) compared with the control. In addition, AICAR treatment suppressed significantly tumor vessel density and macrophage infiltration. We also showed that AICAR administration resulted in AMPK activation and mTOR pathway inhibition. Paradoxically observed down-regulation of p21, which indicates that p21 may have a novel function of an oncogene in retinoblastoma tumor. Our results indicate that AICAR treatment inhibited the growth of retinoblastoma tumor in vivo via AMPK/mTORC1 pathway and by apoptogenic, anti-proliferative, anti-angiogenesis mechanism. AICAR is a promising novel non-chemotherapeutic drug that may be effective as an adjuvant in treating Retinoblastoma.

Vascular endothelial growth factor inhibitors for treatment of corneal neovascularization: a meta-analysis.

Purpose: To evaluate the therapeutic effect of bevacizumab (Avastin) on corneal neovascularization (NV). Methods: Systematic review and meta-analysis of the literature was performed. Seven eligible clinical human studies and 18 eligible experimental animal studies examining the effectiveness of bevacizumab treatment on corneal NV were included in the meta-analysis. Pertinent publications were identified through a systematic search of PubMed. All references of relevant reviews and eligible articles were also screened, and data were extracted from each eligible study. The random-effects model (of DerSimonian and Laird) was used to combine the results from the selected studies. Heterogeneity was explored using available data. Publication bias was also assessed. Results: A significant reduction of corneal neovascularized area was seen in clinical human studies, with a pooled reduction of 36% [95% confidence interval (CI), 18%–54%] overall, of 32% (95% CI, 10%–54%) for subconjunctival anti–vascular endothelial growth factor injections, and 48% (95% CI, 32%–65%) for topical treatment. Pooled mean change in best-corrected visual acuity showed an improvement in best-corrected visual acuity by 0.04. The summary standardized mean difference in animal studies indicated a statistically significant reduction in the area of corneal NV when treated with bevacizumab compared with the control group by −1.71 (95% CI, −2.12 to −1.30). The subtotal pooled standardized mean differences were −1.83 (95% CI, −2.38 to −1.28) for subconjunctival anti–vascular endothelial growth factor injections and −1.50 (95% CI, −1.88 to −1.12) for topical treatment. Conclusion: Our results suggest that both topical and subconjunctival bevacizumab achieve significant reduction in the area of corneal NV. This meta-analysis provides an evidential basis for the new therapeutic concept of treating corneal NV with antiangiogenic therapy.

Hemodialysis-Induced Alterations in Macular Thickness Measured by Optical Coherence Tomography in Diabetic Patients with End-Stage Renal Disease

Background/Aims: To evaluate changes in macular thickness measured by optical coherence tomography (OCT) during a hemodialysis (HD) session in diabetic patients with end-stage renal disease. Methods: 72 eyes of 36 diabetic patients with and without macular edema were evaluated before and immediately after an HD session. Average and maximum macular thicknesses in the central disk (6 mm in diameter) and total macular volume were measured. Results: In the eyes with diabetic macular edema, maximum macular thickness within the central disk of 6 mm, and mainly in its peripheral parts, was significantly reduced by 31.18 ± 4.18 µm after HD (p < 0.001). Average macular thickness and total macular volume were also significantly reduced (p = 0.003 and 0.015, respectively). In diabetic eyes without edema, maximum macular thickness decreased significantly by 11.21 ± 1.98 µm after HD (p < 0.001), while average macular thickness and total macular volume decreased slightly (p = 0.034, p = 0.043). Best-corrected visual acuity failed to change. We found a significant association of macular thickness changes with osmolality reduction and the presence of macular edema. Conclusion: HD decreases macular thickness in diabetic patients with macular edema, while there exists a less-pronounced effect in diabetic eyes without edema.

Effects of metformin on retinoblastoma growth in vitro and in vivo

Recent studies suggest that the anti-diabetic drug metformin may reduce the risk of cancer and have anti-proliferative effects for some but not all cancers. In this study, we examined the effects of metformin on human retinoblastoma cell proliferation in vitro and in vivo. Two different human retinoblastoma cell lines (Y79, WERI) were treated with metformin in vitro and xenografts of Y79 cells were established in nu/nu immune-deficient mice and used to assess the effects of pharmacological levels of metformin in vivo. Metformin inhibited proliferation of the retinoblastoma cells in vitro. Similar to other studies, high concentrations of metformin (mM) blocked the cell cycle in G0–G1, indicated by a strong decrease of G1 cyclins, especially cyclin D, cyclin-dependent kinases (4 and 6), and flow cytometry assessment of the cell cycle. This was associated with activation of AMPK, inhibition of the mTOR pathways and autophagy marker LC3B. However, metformin failed to suppress growth of xenografted tumors of Y79 human retinoblastoma cells in nu/nu mice, even when treated with a maximally tolerated dose level achieved in human patients. In conclusion, suprapharmacological levels (mM) of metformin, well above those tolerated in vivo, inhibited the proliferation of retinoblastoma cells in vitro. However, physiological levels of metformin, such as seen in the clinical setting, did not affect the growth of retinoblastoma cells in vitro or in vivo. This suggests that the potential beneficial effects of metformin seen in epidemiological studies may be limited to specific tumor types or be related to indirect effects/mechanisms not observed under acute laboratory conditions.

The Effect of Alpha Antagonists on Pupil Dynamics: Implications for the Diagnosis of Intraoperative Floppy Iris Syndrome

PURPOSE To assess pupil dynamics quantitatively in relation to the use of α1-adrenoceptor antagonists, which contribute to the features of intraoperative floppy iris syndrome, using a new, hand-held, digital pupillometer. DESIGN Prospective case-control study. METHODS We studied 15 and 25 patients administered tamsulosin and alfuzosin, respectively, as well as 25 control patients. Resting pupil diameter and subsequent contraction, latency, constriction velocity, and dilation velocity were recorded using an electronic pupillometer. All pupil measurements were performed before and after pharmacologic dilation. RESULTS In predilation pupillary measurements, we detected a significant decrease in maximum pupillary diameter by 0.50±0.19 mm (P=.011) and in the mean percentage of diameter reduction after stimulation (5.23±2.42%, P=.035) in the tamsulosin group. Alfuzosin also induced a significant decrease in maximum pupillary diameter (0.49±0.17 mm, P=.005). Constriction velocity was significantly reduced by 0.70±0.20 m/s (P=.001) in the tamsulosin group and by 0.54±0.18 m/s (P=.004) in the alfuzosin group. In terms of postdilation measurements, maximum and minimum pupil diameters were reduced significantly only in the tamsulosin group (by 1.09±0.31 mm [P=.001] and by 0.89±0.36 mm [P=.016], respectively). CONCLUSIONS We describe a reliable, accurate, and rapid method to acquire quantitative pupil measurements and identify the tendency for intraoperative floppy iris syndrome before cataract surgery after the use of alfuzosin and tamsulosin. This investigation also analyzed the similarities and differences induced by the 2 drugs in predilation and postdilation pupil dynamics, demonstrating that tamsulosin is more potent than alfuzosin in inducing intraoperative floppy iris syndrome.

Triamcinolone acetonide as an adjuvant to membrane peeling surgery: a pilot study.

BACKGROUND AND OBJECTIVE To evaluate the clinical outcome and complications of intravitreal injections of triamcinolone acetonide as adjuvant to reduce postoperative macular edema in patients undergoing pars plana vitrectomy for epiretinal membranes. PATIENTS AND METHODS This retrospective comparative study included 22 patients (22 eyes) who underwent pars plana vitrectomy with membrane peeling for the treatment of idiopathic epiretinal membrane. Fifteen eyes (15 patients) received an intravitreal injection of 4 mg (0.1 cc) of triamcinolone acetonide at the end of surgery, and no injection was performed for 7 eyes (7 patients). Main outcome measures were visual acuity and intraocular pressure. Minimum follow-up was 3 months. RESULTS Twenty-two eyes of 22 patients were included in the study. The follow-up ranged from 3 to 12 months. Visual acuity improved in both groups at 3 months: logarithm of the minimum angle of resolution -0.26 ± 0.19 in the triamcinolone acetonide group (P = .001) and -0.26 ± 0.13 in the control group (P = .002). However, there was no statistically significant difference in visual acuity improvement 1, 3, and 12 months postoperatively in the triamcinolone acetonide group compared with the control group (P = .79, = .94, and = .21, respectively). There was no significant difference in intraocular pressure change between the two groups during the follow-up period (P > .05). CONCLUSION The current pilot study suggests that postoperative intravitreal injection of triamcinolone acetonide does not lead to better visual outcomes in patients undergoing pars plana vitrectomy for the treatment of idiopathic epiretinal membranes.