Polyamine depletion induces G1 and S phase arrest in human retinoblastoma Y79 cells

<p>Abstract</p> <p>Background</p> <p>Polyamines and ornithine decarboxylase (ODC) are essential for cell proliferation. DL-α-difluoromethylornithine (DFMO), a synthetic inhibitor of ODC, induces G₁arrest through dephosphorylation of retinoblastoma protein (pRb). The effect of DFMO on cell growth of pRb deficient cells is not known. We examined the effects of DFMO on pRb deficient human retinoblastoma Y79 cell proliferation and its molecular mechanism.</p> <p>Methods</p> <p>Using cultured Y79 cells, the effects of DFMO were studied by using polyamine analysis, western blot, gel shift, FACS and promoter analysis.</p> <p>Results</p> <p>DFMO suppressed the proliferation of Y79 cells, which accumulated in the G1 and S phase. DFMO induced p27/Kip1 protein expression, p107 dephosphorylation and accumulation of p107/E2F-4 complex in Y79 cells.</p> <p>Conclusion</p> <p>These results indicate that p107 dephosphorylation and accumulation of p107/E2F-4 complex is involved in G₁and S phase arrest of DFMO treated Y79 cells.</p

Glaucomatous vertical vessel density asymmetry of the temporal raphe detected with optical coherence tomography angiography.

Changes in retinal vasculature and ocular circulation may play an important role in the glaucoma development and progression. We evaluated the vertical asymmetry across the temporal raphe of the deep retinal layer vessel density, using swept-source optical coherence tomography angiography (SS-OCTA), and its relationship with the central visual field (VF) loss. Thirty-four eyes of 27 patients with open-angle glaucoma were included. SS-OCTA macular scanning was performed within a 3 × 3 mm (300 × 300 pixels) volume, centred on the fovea. The relationships between the vertical asymmetrical deep retinal vessel density reduction (ADRVD) across the temporal raphe and various ocular parameters were analysed. Twenty-two glaucomatous eyes with ADRVDs had central VF loss. Contrarily, ADRVDs were not found in any of the 12 eyes without central VF loss. Thirteen eyes (59.1%) with central VF loss had ADRVDs topographically corresponding to the central VF loss and macular ganglion cell complex thinning. The glaucomatous eyes with ADRVDs exhibited inferior rather than superior central VF loss (P = 0.032). Thus, ADRVD specifically indicates the glaucomatous central visual loss. Further analysis of ADRVD may improve our understanding on glaucoma pathogenesis, offering new treatment insights

Nasal displacement of retinal vessels on the optic disc in glaucoma associated with a nasally angled passage through lamina cribrosa

To investigate nasal displacement of central retinal vessel (CRV) on the optic nerve head (ONH) in glaucoma in association with its passage through lamina cribrosa (LC). This cross-sectional study included 113 eyes with glaucoma and 60 normal eyes. Horizontal spectral-domain optical coherence tomography (SD-OCT) scans of the ONH were acquired, and point where CRV emerged on the ONH surface was defined as the position of the CRV. Next, radial scans of the ONH were acquired, and angle of the CRV passing through the LC was measured. These parameters were compared between glaucomatous and normal eyes by t-test, and their relationship with possible confounders was assessed by multiple regression analyses. In glaucoma, CRV was significantly more nasally displaced than it was in normal eyes (66.0 +/- 8.6 vs. 54.3 +/- 9.5, P<0.0001), and eyes with more vessel displacement exhibited significantly worse glaucomatous visual field defects (P=0.0004). Greater nasal displacement of the CRV was significantly associated with a more nasally angled path through the LC (rs=0.569, P<0.0001). By using SD-OCT, we confirmed that nasal displacement of the CRV on the ONH was associated with glaucoma and was induced by its nasally angled path through the LC

Cloning and characterization of mr-s, a novel SAM domain protein, predominantly expressed in retinal photoreceptor cells

BACKGROUND: Sterile alpha motif (SAM) domains are ~70 residues long and have been reported as common protein-protein interaction modules. This domain is found in a large number of proteins, including Polycomb group (PcG) proteins and ETS family transcription factors. In this work, we report the cloning and functional characterization of a novel SAM domain-containing protein, which is predominantly expressed in retinal photoreceptors and the pineal gland and is designated mouse mr-s (major retinal SAM domain protein). RESULTS: mr-s is evolutionarily conserved from zebrafish through human, organisms through which the mechanism of photoreceptor development is also highly conserved. Phylogenetic analysis suggests that the SAM domain of mr-s is most closely related to a mouse polyhomeotic (ph) ortholog, Mph1/Rae28, which is known as an epigenetic molecule involved in chromatin modifications. These findings provide the possibility that mr-s may play a critical role by regulating gene expression in photoreceptor development. mr-s is preferentially expressed in the photoreceptors at postnatal day 3–6 (P3-6), when photoreceptors undergo terminal differentiation, and in the adult pineal gland. Transcription of mr-s is directly regulated by the cone-rod homeodomain protein Crx. Immunoprecipitation assay showed that the mr-s protein self-associates mainly through the SAM domain-containing region as well as ph. The mr-s protein localizes mainly in the nucleus, when mr-s is overexpressed in HEK293T cells. Moreover, in the luciferase assays, we found that mr-s protein fused to GAL4 DNA-binding domain functions as a transcriptional repressor. We revealed that the repression activity of mr-s is not due to a homophilic interaction through its SAM domain but to the C-terminal region. CONCLUSION: We identified a novel gene, mr-s, which is predominantly expressed in retinal photoreceptors and pineal gland. Based on its expression pattern and biochemical analysis, we predict that mr-s may function as a transcriptional repressor in photoreceptor cells and in pinealocytes of the pineal gland

Establishment of an experimental ferret ocular hypertension model for the analysis of central visual pathway damage

Glaucoma optic neuropathy (GON) is a condition where pathogenic intraocular pressure (IOP) results in axonal damage following retinal ganglion cell (RGC) death, and further results in secondary damage of the lateral geniculate nucleus (LGN). Therapeutic targets for glaucoma thus focus on both the LGN and RGC. However, the temporal and spatial patterns of degeneration and the mechanism of LGN damage have not been fully elucidated. Suitable and convenient ocular hypertension (OH) animal models with binocular vision comparable to that of monkeys are strongly needed. The ferret is relatively small mammal with binocular vision like humans - here we report on its suitability for investigating LGN. We developed a new method to elevate IOP by injection of cultured conjunctival cells into the anterior chamber to obstruct aqueous outflow. Histologically, cultured conjunctival cells successfully proliferated to occlude the angle, and IOP was elevated for 13 weeks after injection. Macroscopically, the size of the eye gradually expanded. Subsequent enlargement of optic nerve head cupping and atrophic damage of LGN projected from the OH eye were clearly observed by anterograde staining with cholera toxin B. We believe the ferret may be a promising OH model to investigate secondary degeneration of central nervous system including LGN

Effects of topical latanoprost on optic nerve head circulation in rabbits, monkeys

PURPOSE. To evaluate the effect of topically administrated latanoprost on optic nerve head (ONH) circulation in Dutch rabbits, cynomolgus monkeys, and normal humans. METHODS. The ONH tissue blood velocity (NB ONH ) was determined using the laser speckle method. Latanoprost (0.005%, 30 l) was instilled into one eye, and vehicle into the other eye as a control. In rabbits, NB ONH was measured for 90 minutes after a single instillation and before and after a 7-day once-daily instillation regimen. In monkeys, NB ONH was measured before and after 1, 4, and 7 days of a once-daily instillation regimen. The effect of intravenous indomethacin on the latanoprostinduced NB ONH change was also studied in rabbits and monkeys. In humans, the time-course changes in NB ONH were measured for 4.5 hours before and after a 7-day once-daily instillation regimen. Intraocular pressure (IOP) and systemic parameters were simultaneously studied in each experiment. All measurements were performed by investigators masked to the experimental condition. RESULTS. Latanoprost significantly increased NB ONH 10% to 19% in treated eyes after a single instillation (P ϭ 0.035) or 7-day instillation regimen (P ϭ 0.035) in rabbits, after a 4-day (P ϭ 0.035) or 7-day (P ϭ 0.035) instillation regimen in monkeys, and after a 7-day (P ϭ 0.013) instillation regimen in humans, whereas there were no significant changes in the vehicletreated eyes in any of the experiments (P Ͼ 0.5). Pretreatment with indomethacin (5 mg/kg) abolished the NB ONH increase but not the IOP reduction in latanoprost-treated eyes in rabbits and monkeys. IOP remained unchanged in both eyes in rabbits (P Ͼ 0.4), whereas it significantly decreased only in latanoprost-treated eyes in monkeys (P Ͻ 0.05) and humans (P Ͻ 0.05). CONCLUSIONS. Topical latanoprost significantly increased ONH blood velocity only in treated eyes in rabbits, monkeys, and humans. This effect was independent of the IOP-reducing effect of latanoprost and probably was associated with local penetration of the drug and the production of endogenous prostaglandins. (Invest Ophthalmol Vis Sci. 2001;42:2957-2963 L atanoprost, a recently developed prostaglandin F 2␣ (PGF 2␣ )-related FP receptor agonist compound, 1 is one of the most potent ocular hypotensive eye drops in patients with glaucoma. 2-8 Although intraocular pressure (IOP) is consistently the major risk factor for glaucoma, recent studies indicate that impaired circulation in the optic nerve head (ONH) has a crucial role in glaucomatous optic neuropathy. -11 Previous studies have suggested that topically instilled timolol or betaxolol over a long period can accumulate in the periocular tissues and reach the retrobulbar space in sufficient concentration to have pharmacologic effects on the retrobulbar vessels. 12-14 PGF 2␣ has vasoconstricting or vasodilating effects, depending on its concentration, the nature of the vascular bed, or the animal species. 15-18 Latanoprost also has vasoconstricting effects at higher concentrations 20 Grunwald 21-23 and Gupta et al. In the present study, we examined the effects of not only a single instillation but also a 7-day once-daily instillation regimen of latanoprost on ONH circulation in rabbits, monkeys, and normal humans, by using the noninvasive laser speckle method. MATERIALS AND METHODS Instruments ONH circulation was evaluated using the laser speckle method