PURPOSE. To investigate, using in vivo and in vitro models, retinal ganglion cell (RGC) neuroprotective and axon regenerative effects and underlying mechanisms of siRTP801, a translatable small-interfering RNA (siRNA) targeting the mTOR negative regulator RTP801.METHODS. Adult rats underwent optic nerve (ON) crush (ONC) followed by intravitreal siRTP801 or control siRNA (siEGFP) every 8 days, with Brn3a þ RGC survival, GFAP þ reactive gliosis, and GAP43 þ regenerating axons analyzed immunohistochemically 24 days after injury. Retinal cultures, prepared from uninjured animals or 5 days after ONC to activate retinal glia, were treated with siRTP801/controls in the presence/absence of rapamycin and subsequently assessed for RGC survival and neurite outgrowth, RTP801 expression, glial responses, and mTOR activity. Conditioned medium was analyzed for neurotrophin titers by ELISA. RESULTS. Intravitreal siRTP801 enabled 82% RGC survival compared to 45% with siEGFP 24 days after ONC, correlated with greater GAP43þ axon regeneration at 400 to 1200 lm beyond the ONC site, and potentiated the reactive GFAP þ Müller glial response. In culture, siRTP801 had a direct RGC neuroprotective effect, but required GFAP þ activated glia to stimulate neurite elongation. The siRTP801-induced neuroprotection was significantly reduced, but not abolished, by rapamycin. The siRTP801 potentiated the production and release of neurotrophins NGF, NT-3, and BDNF, and prevented downregulation of RGC mTOR activity.CONCLUSIONS. The RTP801 knockdown promoted RGC survival and axon elongation after ONC, without increasing de novo regenerative sprouting. The neuroprotection was predominantly direct, with mTORC1-dependent and -independent components. Enhanced neurite/axon elongation by siRTP801 required the presence of activated retinal glia and was mediated by potentiated secretion of neurotrophic factors.
This is the only murine model of closed globe injury and the only model of retinal trauma with specific photoreceptor cell death. The clinical appearance mirrors that in severe retinal injury after blunt ocular trauma in humans, and the ultrastructural features are consistent with human and animal studies of commotio retinae. After ocular trauma, photoreceptor apoptosis may be prevented and visual outcomes improved by blocking of the cell death pathways.
PURPOSE. To determine if vitreous levels of the pro-fibrotic cytokine transforming growth factor beta2 (TGF-b2) and its opposing regulator decorin predict subsequent proliferative vitreoretinopathy (PVR) development in patients with rhegmatogenous retinal detachment (RRD). METHODS. We examined the effect of TGF-b2 and decorin on epithelial-mesenchymal transition (EMT) and collagen expression in vitro using ARPE-19 cells, and we analyzed extracellular matrix marker expression in PVR membrane and internal limiting membrane patient samples. We performed a prospective noninterventional cohort study, recruiting 125 patients undergoing vitrectomy for RRD and macular hole surgery, measured vitreous levels of TGF-b2 and decorin by ELISA, and followed them up for 6 months. Patients who did not develop PVR were compared to those who did, in order to determine whether vitreous TGF-b2 and decorin levels predicted PVR development. RESULTS. In vitro, TGF-b2 induced EMT and collagen production. Decorin strongly inhibited EMT and collagen production at high levels. PVR membranes expressed high levels of fibrosisassociated proteins, consistent with EMT. Vitreous TGF-b2 levels were unchanged between patients with macular holes and RRD who did or did not subsequently develop PVR. Average decorin levels were higher in the vitreous of RRD patients who subsequently developed PVR compared to those who did not, but at the measured vitreous concentrations (1-2 lg/mL), decorin did not demonstrate an in vitro inhibitory effect on EMT. CONCLUSIONS. In vitro, high concentrations of decorin inhibited EMT and fibrosis. At the levels seen in human vitreous, decorin did not prevent fibrosis or EMT in vitro, and higher initial vitreous decorin levels were associated with the development of postoperative PVR after vitrectomy to treat RRD, but did not reliably predict the outcome.
Current processes for coating titanium implants with ceramics involve very high energy techniques with associated high cost and disadvantages such as heterogeneity of the coatings, phase transformations and inability to coat complex structures. In order to address the above problems, we propose a biomimetic hydroxyapatite coating process with the use of peptides that can bind both on titanium surfaces and hydroxyapatite. The peptides enabled homogeneous coating of a titanium surface with hydroxyapatite. The hydroxyapatite-peptide sandwich coating showed no adverse effects on cell number or collagen deposition. This makes the sandwich coated titanium a good candidate for titanium implants used in orthopaedics and dentistry.
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