RBPMS

Abstract: Ganglion cells, which carry the final neuronal output of the vertebrate retina, collect visual signals from the two preceding layers of nerve cells, bipolar, and amacrine cells, and transmit this information to the brain. The death of retinal ganglion cells (RGCs) and degeneration of their axons in the optic nerve is the cause of vision loss in various optic neuropathies including glaucoma. In experimental rodent models of glaucoma, to evaluate the RGC loss, these cells are commonly retrogradely labeled by injection of tracers such as FluoroGold (FG), DTMR, or DiI into areas of the brain that are targeted by RGCs, primarily superior colliculus (SC), or by exposing the axons in the optic nerve to these dyes. However, both procedures have limitations. Since RGC retrograde labeling with these tracers depends on active axonal transport,1 which has been shown to be affected in animal models of glaucoma,2–6 these labeling techniques do not differentiate between cell loss, axon degeneration and failure of transport. Furthermore, labeling via SC leaves uncounted RGCs projecting to other brain areas. Nevertheless, despite the availability of several antigenic RGC markers, including Thy-1, Brn3, neurofilament and others, retrograde labeling is commonly viewed as the most reliable and accurate way of identifying RGCs. In the present study, we characterized expression of RNA-binding protein with multiple splicing, RBPMS, or hermes in the retina. We present data supporting the use of anti-RBPMS antibodies for quantitative analysis of the number of RGCs, independent of their connectivity to their central target areas. RBPMS was recently identified in a study designed to analyze gene expression profiles in RGCs.7 RBPMS genes (RBPMS and its paralogue RBPMS2) are members of the RRM (RNA recognition motif) family of RNA-binding proteins. Members of the RRM family are involved in the regulation of gene expression at the posttranscriptional level, including pre-mRNA-processing (splicing, capping, and polyadenylation), RNA stability, transport, localization, and translational regulation. The hermes RNA-binding domain is similar to that of the Drosophila couch potato (cpo) and Caenorhabditis elegans Mec-8 genes (see Fig. 1).8 Mutations in cpo lead to several neurologic phenotypes, including bang-sensitive paralysis, seizure susceptibility, and synaptic transmission defects, indicating an important role for cpo in regulating normal function of the nervous system,9 whereas mutations in Mec-8 affect mechanosensory and chemosensory neuronal function.10 Although the exact functions of hermes genes are unknown, it has been reported that RBPMS could be involved in regulation of mRNA translation and localization during Xenopus laevis development.11 RBPMS has also been shown to physically interact with Smad2, -3, and -4,12 which regulate TGF-β signaling13 and with ataxin 1, a protein responsible for spinocerebellar ataxia type 1 due to expansion of a polyglutamine repeat.14 Figure 1. RBPMS amino acid sequences and their homology to Drosophila couch potato (cpo) and RBPMS2 proteins. High consensus, red; low consensus, blue; and neutral, black. The RRM, yellow box; and GGKAEKENTPSEANLQEEEVR, used for antibody production, green box. ...

Abstract: Purpose To determine if bone marrow-derived stem cell (BMSC) small extracellular vesicles (sEV) promote retinal ganglion cell (RGC) neuroprotection in the genetic DBA/2J mouse model of glaucoma for 12 months. Methods BMSC sEV and control fibroblast-derived sEV were intravitreally injected into 3-month-old DBA/2J mice once a month for 9 months. IOP and positive scotopic threshold responses were measured from 3 months: IOP was measured monthly and positive scotopic threshold responses were measured every 3 months. RGC neuroprotection was determined in wholemounts stained with RNA binding protein with multiple splicing (RBPMS), whereas axonal damage was assessed using paraphenylenediamine staining. Results As expected, DBA/2J mice developed chronic ocular hypertension beginning at 6 months. The delivery of BMSC sEV, but not fibroblast sEV, provided significant neuroprotective effects for RBPMS+ RGC while significantly reducing the number of degenerating axons seen in the optic nerve. BMSC sEV significantly preserved RGC function in 6-month-old mice, but provided no benefit at 9 and 12 months. Conclusions BMSC sEV are an effective neuroprotective treatment in a chronic model of ocular hypertension for 1 year, preserving RGC numbers and protecting against axonal degeneration.

Abstract: A photoreceptor cell line, 661W, derived from a mouse retinal tumor that expresses several markers of cone photoreceptor cells has been described earlier. However, these cells can be differentiated into neuronal cells. Here, we report that this cell line expressed certain markers specific to retinal ganglion cells such as Rbpms, Brn3b (Pou4f2), Brn3c (Pou4f3), Thy1 and γ-synuclein (Sncg), and some other markers of neuronal cells (beta-III tubulin, NeuN and MAP2). These cells also expressed Opn1mw, a cone-specific marker and nestin, a marker for neural precursor cells. Two glaucoma-associated mutants of OPTN, E50K and M98K, but not an amyotrophic lateral sclerosis-associated mutant, E478G, induced cell death selectively in 661W cells. However, in a motor neuron cell line, NSC34, E478G mutant of OPTN but not E50K and M98K induced cell death. We conclude that 661W is a retinal ganglion precursor-like cell line, which shows properties of both retinal ganglion and photoreceptor cells. We suggest that these cells could be utilized for exploring the mechanisms of cell death induction and cytoprotection relevant for glaucoma pathogenesis. RGC-5 cell line which probably arose from 661W cells showed expression of essentially the same markers of retinal ganglion cells and neuronal cells as seen in 661W cells.