Dr.Heiko Pult Optometry and Vision Research

Optogenetic gene therapy is a promising mutation-independent treatment that aims to restore visual perception in patients blinded by retinal diseases that cause photoreceptor degeneration. Still, low sensitivity or slow kinetics of currently utilized optogenetic proteins limit the efficacy of such approaches. Here, we evaluated the therapeutic potential of three channelrhodopsin variants: ChRmine, from the algae Rhodomonas lens, ChRmine-T119A, a faster-closing ChRmine variant, and ChroME2s, a second-generation Chronos-based opsin.We expressed these opsins in retinal ganglion cells of rd1 mice, a model of severe retinal degeneration. Single cell electrophysiology demonstrates opsin’s large sensitivity to a range of light intensities as well as opsin-expressing retinal ganglion cells generated action potentials in response to light stimulation. Behavioral tests showed ChRmine-T119A’s efficacy at 360 lux compared to unmodified ChRmine and ChroME2s. ChRmine and ChroME2s did restore light perception at higher light intensities. Additionally, our dose–response study with ChRmine-T119A revealed that lower viral titers were more effective at restoring light sensitivity. Our study demonstrates that these ChRmine- and ChroME-based opsins can enhance vision in late-stage blinding diseases.

Erratum in Invest Ophthalmol Vis Sci. 2014 Sep;55(9):5588. Miller, William S [coorected to Miller, William L].

Mucins adhere to contact lenses (CLs), reflecting the renewal of the preocular fluid and enzymatic activity at the ocular surface. In this study, we aimed to analyze mucin fragmentation on materials new to the ocular surface and investigate whether this correlates with wearing comfort. Lenses were obtained from new CL wearers after 2 weeks each of wearing vifilcon A, followed by senofilcon A, and then by vifilcon A lenses. Symptoms were evaluated using the Ocular Surface Disease Index (OSDI). CLs were extracted in a mixture of guanidinium hydrochloride and radioimmunoprecipitation assay buffer. Mucin mobility was analyzed after electrophoresis, Western blotting, and visualization with antibodies against mucin peptide core. Mobilities, normalized to total reactivity in the lane, were compared between visits for each subject and were expressed as shifts. Mucin (MUC)5AC polymers exceeding 260 kDa were observed in agarose gels; NuPAGE resolved polymers from 260 to 3.5 kDa: when large mucins were detected, the smallest fragments were missing. Fragmentation patterns were significantly different between lens types for MUC1 (analysis of variance, P = 0.006) and MUC4 (P < 0.001) but not for MUC5AC or MUC16 (P > 0.293). Mobility shifts of MUC1 and MUC4 were significantly negatively correlated (Pearson, r = -0.908; P = 0.002). For OSDI scores >15, mucin fragmentation was unchanged, whereas for OSDI scores <15, MUC4 and MUC5AC fragments were longer on vifilcon A than on senofilcon lenses (unpaired t test, P = 0.046), irrespective of the direction of change (analysis of variance, P > 0.366). Changes in MUC1 breakdown were significantly negatively correlated to the overall OSDI score (r = -0.891, P = 0.001). In asymptomatic CL wearers, only changes in mucin fragmentation in response to a new material were consistent and fast, irrespective of CL order. Lack of change seems, therefore, to be connected with discomfort during CL wear.