Real-time quantitative monitoring of hiPSC-based model of macular degeneration on Electric Cell-substrate Impedance Sensing microelectrodes

AbstractAge-related macular degeneration (AMD) is the leading cause of blindness in the developed world. Humanized disease models are required to develop new therapies for currently incurable forms of AMD.In this work, a tissue-on-a-chip approach was developed through combining human induced pluripotent stem cells, Electric Cell–substrate Impedance Sensing (ECIS) and reproducible electrical wounding assays to model and quantitatively study AMD. Retinal Pigment Epithelium (RPE) cells generated from a patient with an inherited macular degeneration and from an unaffected sibling were used to test the model platform on which a reproducible electrical wounding assay was conducted to model RPE damage. First, a robust and reproducible real-time quantitative monitoring over a 25-day period demonstrated the establishment and maturation of RPE layers on the microelectrode arrays. A spatially controlled RPE layer damage that mimicked cell loss in AMD disease was then initiated. Post recovery, significant differences (P<0.01) in migration rates were found between case (8.6±0.46μm/h) and control cell lines (10.69±0.21μm/h). Quantitative data analysis suggested this was achieved due to lower cell–substrate adhesion in the control cell line. The ECIS cell–substrate adhesion parameter (α) was found to be 7.8±0.28Ω1/2cm for the case cell line and 6.5±0.15Ω1/2cm for the control. These findings were confirmed using cell adhesion biochemical assays. The developed disease model-on-a-chip is a powerful platform for translational studies with considerable potential to investigate novel therapies by enabling real-time, quantitative and reproducible patient-specific RPE cell repair studies

Enzymatic oxidative dimerization of silymarin flavonolignans

Dimerization of phenolic compounds can potentially enhance their biological (antioxidant) activity. We present here the selective oxidative dimerization of several flavonolignans from Silybum marianum seed extract, namely, silybin A (1a), silybin B (1b), silychristin (3), and silydianin (4) catalyzed by a laccase from Trametes versicolor. Selective benzylation of C-7 OH group of both silybins ensured the priority of the dimerization reaction, avoiding thus polymerization. C C homodimers connected via E-rings of silybin A and B and silydianin dimers were successfully isolated after respective debenzylation. On the contrary, dimerization of 7-O-benzyl silychristin afforded a complex, inseparable mixture ofthe products. All isolated flavonolignan dimers exhibited significantly improved 1,1-diphenyl-2-picrylhydrazyl(DPPH) radical scavenging activity compared to their monomers and, therefore, seem to be promising for further biological studies.This work was supported a bilateral Czech-Italian inter-academic project IT 2012/02 between CNR and AVČR (2013-2015, V.K. + S.R.) and by COST Chemistry CM1303 Action. E.V. acknowledges support from Czech Science Foundation Grant 14-14373P and V.K. acknowledges MSMT Grant LD13041 and research concept of the Institute of MicrobiologyRVO61388971. S.R. acknowledges support from MIUR, Italy, Progetto PRIN 2011 “PROxi”