David Gamm, MD, PhD

Slide of the Week: David Gamm, MD, PhD

Title: Pharmacological modulation of photoreceptor outer segment degradation in Best disease patient-derived hiPSC-RPE

Legend: (a) Representative western blot analysis comparing the amount of the POS protein RHO in untreated and VPA-treated (500 µmol/l) BD hiPSC-RPE at 0 and 24 hours after POS feeding. (b) Quantification of RHO levels 24 hours after POS feeding in untreated versus treated (500 µmol/l VPA, 100 nmol/l RAPA, or 500 µmol/l VPA + 1000 nmol/l RAPA) BD hiPSC-RPE (BD-1 + BD-2). (c) Confocal microscopic images showing accumulation of autofluorescent material (red channel) in untreated (left panels) and VPA-treated (500 µmol/l; right panels) BD-2 hiPSC-RPE in the presence (lower panels) or absence (upper panels) of daily POS feeding for 1.5 months. (d) Quantification of average autofluorescent levels in untreated and VPA-treated BD-2 hiPSC-RPE after daily POS feeding for 1.5 months (note log scale on y-axis). Bar = 50 µM. ACTN = ACTIN control. *p<0.05; **p=0.001. BD, Best disease; hiPSC, human induced pluripotent stem cell; RPE, Retinal pigment epithelium; POS, photoreceptor outer segment; RAPA, Rapamycin; VPA, Valproic acid; RHO, RHODOPSIN.

Citation: Singh R, Kuai D, Guziewicz KE, Meyer J, Wilson M, Lu J, Smith M, Clark E, Verhoeven A, Aguirre G, Gamm DM (2015). Pharmacological modulation of photoreceptor outer segment degradation in a human iPS cell model of inherited macular degeneration. Molecular Therapy. doi:10.1038/mt.2015.141

Abstract: Degradation of photoreceptor outer segments (POS) by retinal pigment epithelium (RPE) is essential for vision, and studies have implicated altered POS processing in the pathogenesis of some retinal degenerative diseases. Consistent with this concept, a recently established hiPSC-RPE model of inherited macular degeneration, Best disease (BD), displayed reduced rates of POS breakdown. Herein we utilized this model to determine (i) if disturbances in protein degradation pathways are associated with delayed POS digestion and (ii) whether such defect(s) can be pharmacologically targeted. We found that BD hiPSC-RPE cultures possessed increased protein oxidation, decreased free-ubiquitin levels, and altered rates of exosome secretion, consistent with altered POS processing. Application of valproic acid (VPA) with or without rapamycin increased rates of POS degradation in our model, whereas application of bafilomycin-A1 decreased such rates. Importantly, the negative effect of bafilomycin-A1 could be fully reversed by VPA. The utility of hiPSC-RPE for VPA testing was further evident following examination of its efficacy and metabolism in a complementary canine disease model. Our findings suggest that disturbances in protein degradation pathways contribute to the POS processing defect observed in BD hiPSC-RPE, which can be manipulated pharmacologically. These results have therapeutic implications for BD and perhaps other maculopathies.

About the investigator: Gamm’s laboratory at the Waisman Center utilizes stem cell technology to investigate the cellular and molecular events that occur during human retinal differentiation and generate cells for use in human retinal disease modeling and cell-based rescue or replacement strategies. To meet these goals, Gamm utilizes a variety of human cell types, including ES and iPS cells, which have the capacity to mimic retinal development and disease, as well as to delineate the genetic “checkpoints” necessary to produce particular retinal cell types. By understanding the behavior of these cell types in vitro and in vivo, Gamm hopes to optimize strategies to delay or reverse the effects of blinding disorders such as retinitis pigmentosa and age–related macular degeneration.

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