BackgroundStudy of age related macular degeneration (AMD) has been hampered by lack of human models that represent the complexity of the disease. Here we have developed a human in vitro disease model of AMD to investigate the underlying AMD disease mechanisms.MethodsGeneration of iPSCs from retinal pigment epithelium (RPE) of AMD donors, age-matched normal donors, skin fibroblasts of a dry AMD patient, and differentiation of iPSCs into RPE (AMD RPE-iPSC-RPE, normal RPE-iPSC-RPE and AMD Skin-iPSC-RPE, respectively). Immunostaining, cell viability assay and reactive oxygen species (ROS) production under oxidative stress conditions, electron microscopy (EM) imaging, ATP production and glycogen concentration assays, quantitative real time PCR, western blot, karyotyping.ResultsThe AMD RPE-iPSC-RPE and AMD Skin-iPSC-RPE present functional impairment and exhibit distinct disease phenotypes compared to RPE-iPSC-RPE generated from normal donors (Normal RPE-iPSC-RPE). The AMD RPE-iPSC-RPE and AMD Skin-iPSC-RPE show increased susceptibility to oxidative stress and produced higher levels of reactive oxygen species (ROS) under stress in accordance with recent reports. The susceptibility to oxidative stress-induced cell death in AMD RPE-iPSC-RPE and Skin-iPSC-RPE was consistent with inability of the AMD RPE-iPSC-RPE and Skin-iPSC-RPE to increase SOD2 expression under oxidative stress. Phenotypic analysis revealed disintegrated mitochondria, accumulation of autophagosomes and lipid droplets in AMD RPE-iPSC-RPE and AMD Skin-iPSC-RPE. Mitochondrial activity was significantly lower in AMD RPE-iPSC-RPE and AMD Skin-iPSC-RPE compared to normal cells and glycogen concentration was significantly increased in the diseased cells. Furthermore, Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), a regulator of mitochondrial biogenesis and function was repressed, and lower expression levels of NAD-dependent deacetylase sirtuin1 (SIRT1) were found in AMD RPE-iPSC-RPE and AMD Skin-iPSC-RPE as compared to normal RPE-iPSC-RPE.ConclusionsOur studies suggest SIRT1/PGC-1α as underlying pathways contributing to AMD pathophysiology, and open new avenues for development of targeted drugs for treatment of this devastating neurodegenerative disease of the visual system.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-016-1101-8) contains supplementary material, which is available to authorized users.

The development and progression of diabetic nephropathy is dependent on glucose homeostasis and many other contributing factors. In the present study, we examined the effect of nitecapone, an inhibitor of the dopaminemetabolizing enzyme catechol-O-methyl transferase (COMT) and a potent antioxidant, on functional and cellular determinants of renal function in rats with streptozotocin-induced diabetes. Administration of nitecapone to diabetic rats normalized urinary sodium excretion in a manner consistent with the dopaminedependent inhibition of proximal tubule Na,K-ATPase activity. Hyperfiltration, focal glomerulosclerosis, and albuminuria were also reversed by nitecapone, but in a manner that is more readily attributed to the antioxidant potential of the agent. A pattern of elevated oxidative stress, measured as CuZn superoxide dismutase gene expression and thiobarbituric acid-reactive substance content, was noted in diabetic rats, and both parameters were normalized by nitecapone treatment. In diabetic rats, activation of glomerular protein kinase C (PKC) was confirmed by isoform-specific translocation and Ser23 phosphorylation of the PKC substrate Na,KATPase. PKC-dependent changes in Na,K-ATPase phosphorylation were associated with decreased glomerular Na,K-ATPase activity. Nitecapone-treated diabetic rats were protected from these intracellular modifications. The combined results suggest that the COMTinhibitory and antioxidant properties of nitecapone provide a protective therapy against the development of diabetic nephropathy. Diabetes 49:1381-1389, 2000

Na¤,K¤-ATPase (NKA) is an ubiquitous membrane bound protein that transports Na¤ and K¤ across the plasma membrane against their electrochemical gradients (Skou & Esmann, 1992). It plays a pivotal role for many basic cellular functions such as active transport of certain solutes, regulation of cell volume, and restoration of the membrane potential. An altered enzyme activity occurs in a number of clinical disorders, such as hypertension and diabetes (for a review see Laski & Kurtzman, 1996). Therefore, elucidating the regulatory mechanisms that underlie the control of NKA activity has become an important issue.There is overwhelming evidence that the activity of NKA can be modulated by first messengers as well as by signalling pathways that involve activation of cAMPdependent protein kinase (PKA) andÏor protein kinase C (PKC) (for a review see Ewart & Klip, 1995). It is well established that NKA from several species can be phosphorylated by PKA (