Stavros N. Moysidis

Nanoparticle-mediated signaling endosome localization regulates growth cone motility and neurite growth.

Understanding neurite growth regulation remains a seminal problem in neurobiology. During development and regeneration, neurite growth is modulated by neurotrophin-activated signaling endosomes that transmit regulatory signals between soma and growth cones. After injury, delivering neurotrophic therapeutics to injured neurons is limited by our understanding of how signaling endosome localization in the growth cone affects neurite growth. Nanobiotechnology is providing new tools to answer previously inaccessible questions. Here, we show superparamagnetic nanoparticles (MNPs) functionalized with TrkB agonist antibodies are endocytosed into signaling endosomes by primary neurons that activate TrkB-dependent signaling, gene expression and promote neurite growth. These MNP signaling endosomes are trafficked into nascent and existing neurites and transported between somas and growth cones in vitro and in vivo. Manipulating MNP-signaling endosomes by a focal magnetic field alters growth cone motility and halts neurite growth in both peripheral and central nervous system neurons, demonstrating signaling endosome localization in the growth cone regulates motility and neurite growth. These data suggest functionalized MNPs may be used as a platform to study subcellular organelle localization and to deliver nanotherapeutics to treat injury or disease in the central nervous system.

Mitochondrial Dynamics Regulate Growth Cone Motility, Guidance, and Neurite Growth Rate in Perinatal Retinal Ganglion Cells In Vitro

PURPOSE Retinal ganglion cell (RGC) death and failed axonal regeneration after trauma or disease, including glaucomatous and mitochondrial optic neuropathies, are linked increasingly to dysfunctional mitochondrial dynamics. However, how mitochondrial dynamics influence axon growth largely is unstudied. We examined intrinsic mitochondrial organization in embryonic and postnatal RGCs and the roles that mitochondrial dynamics have in regulating neurite growth and guidance. METHODS RGCs were isolated from embryonic day 20 (E20) or postnatal days 5 to 7 (P5-7) Sprague-Dawley rats by anti-Thy1 immunopanning. After JC-1 loading, mitochondria were analyzed in acutely purified RGCs by flow cytometry and in RGC neurites by fluorescence microscopy. Intrinsic axon growth was modulated by overexpressing Krüppel-like family (KLF) transcription factors, or mitochondrial dynamics were altered by inhibiting dynamin related protein-1 (DRP-1) pharmacologically or by overexpressing mitofusin-2 (Mfn-2). Mitochondrial organization, neurite growth, and growth cone motility and guidance were analyzed. RESULTS Mitochondrial dynamics and function are regulated developmentally in acutely purified RGCs and in nascent RGC neurites. Mitochondrial dynamics are modulated differentially by KLFs that promote or suppress growth. Acutely inhibiting mitochondrial fission reversibly suppressed axon growth and lamellar extension. Inhibiting DRP-1 or overexpressing Mfn-2 altered growth cone responses to chondroitin sulfate proteoglycan, netrin-1, and fibronectin. CONCLUSIONS These results support the hypothesis that mitochondria locally modulate signaling in the distal neurite and growth cone to affect the direction and the rate of neurite growth.

Promoting filopodial elongation in neurons by membrane-bound magnetic nanoparticles

Filopodia are 5-10 μm long processes that elongate by actin polymerization, and promote axon growth and guidance by exerting mechanical tension and by molecular signaling. Although axons elongate in response to mechanical tension, the structural and functional effects of tension specifically applied to growth cone filopodia are unknown. Here we developed a strategy to apply tension specifically to retinal ganglion cell (RGC) growth cone filopodia through surface-functionalized, membrane-targeted superparamagnetic iron oxide nanoparticles (SPIONs). When magnetic fields were applied to surface-bound SPIONs, RGC filopodia elongated directionally, contained polymerized actin filaments, and generated retrograde forces, behaving as bona fide filopodia. Data presented here support the premise that mechanical tension induces filopodia growth but counter the hypothesis that filopodial tension directly promotes growth cone advance. Future applications of these approaches may be used to induce sustained forces on multiple filopodia or other subcellular microstructures to study axon growth or cell migration. From the clinical editor: Mechanical tension to the tip of filopodia is known to promote axonal growth. In this article, the authors used superparamagnetic iron oxide nanoparticles (SPIONs) targeted specifically to membrane molecules, then applied external magnetic field to elicit filopodial elongation, which provided a tool to study the role of mechanical forces in filopodia dynamics and function.

Visual and anatomical outcomes of macular epiretinal membrane peeling after previous rhegmatogenous retinal detachment repair.

Purpose: To report outcomes and prognostic factors after epiretinal membrane peeling in patients with previous rhegmatogenous retinal detachment repair. Methods: A consecutive case series. Best-corrected visual acuity and optical coherence tomography characteristics were analyzed before and after epiretinal membrane surgery. Results: Fifty-three eyes were analyzed. Best-corrected visual acuity improved by a mean of 10 letters at 1 month (N = 45; P = 0.001), 15 at 3 months (N = 42; P < 0.001), 11 at 6 months (N = 35; P = 0.001), and 16 at 12 months (N = 33; P < 0.001). The mean optical coherence tomography central foveal thickness decreased by 141 (N = 22; P < 0.001), 185 (N = 24; P < 0.001), 180 (N = 17; P = 0.001), and 151 &mgr;m (N = 9; P = 0.017) at 1, 3, 6, and 12 months, respectively. Better preoperative best-corrected visual acuity correlated with better best-corrected visual acuity at all follow-up visits (P ⩽ 0.001). Intact preoperative inner segment/outer segment junction and external limiting membrane line, but not the change in central foveal thickness or location of fluid, correlated with better postoperative best-corrected visual acuity through 6 months. Conclusion: Epiretinal membrane peeling after previous rhegmatogenous retinal detachment repair resulted in significant improvements in visual acuity and optical coherence tomography thickness, even in eyes with previous macula-involving rhegmatogenous retinal detachment. Better preoperative visual acuity and intact outer retinal layers by optical coherence tomography were the main prognostic factors for visual outcomes.

Acute retinal pigment epithelium detachments after photocoagulation

Purpose: To characterize the morphology of patterned scanning laser (PASCAL) panretinal photocoagulation. Methods: In this prospective cohort study, patients with proliferative diabetic retinopathy or severe nonproliferative diabetic retinopathy with high-risk characteristics, who were treated with PASCAL panretinal photocoagulation as part of their indicated clinical course, were serially imaged with spectral domain optical coherence tomography. Thirty eyes of 25 patients were studied from 1 hour to 21 weeks after laser treatment. Results: Over a quarter (26.1%) of all treatment spots were imaged by spectral domain optical coherence tomography 1 hour after PASCAL panretinal photocoagulation. At 1 hour (±30 minutes) after PASCAL treatment, spectral domain optical coherence tomography demonstrated retinal pigment epithelium detachment in 23 of 27 eyes (85.2%) and in 36.1% of all imaged laser spots. Detachments occurred preferentially at the photocoagulation edges in 48.4% of pigment epithelium detachments (PEDs). Linear regression analysis revealed that average laser power (Pearsons r = 0.671, P < 0.001) and average laser energy (Pearsons r = 0.588, P = 0.001) were significantly associated with PEDs observed 1 hour after treatment. Pigment epithelium detachments occurred at a rate of 9.2% ± 4.9% at an average power of 0 mW to 400 mW, 37.8% ± 9.5% at 401 mW to 800 mW, 42.1% ± 5.6% at 801 mW to 1,200 mW, and 53.6% ± 5.7% at >1,200 mW. By a 1-week follow-up, no PEDs were observed, and the retinal pigment epithelium appeared morphologically similar to its preoperative structure by 3 weeks. Patient characteristics (study eye, sex, race, diagnosis, age, preoperative blood glucose, hemoglobin A1C, duration of diabetes, and body mass index) and other PASCAL parameters (number of laser applications, spot size, pulse duration, and average laser fluence) were not significantly associated with PEDs. Conclusion: Retinal pigment epithelium detachment occurs 1 hour after PASCAL treatment over a wide range of laser settings. Laser power and energy are positively correlated with the occurrence of PEDs, which are no longer observed by 1-week follow-up. Future studies might examine various acute posttreatment time points and directly compare the morphology of PASCAL burns with that of longer pulse–duration laser modalities.