The use of induced pluripotent stem cells to reveal pathogenic gene mutations and explore treatments for retinitis pigmentosa.

BACKGROUND: Retinitis pigmentosa (RP) is an inherited human retinal disorder that causes progressive photoreceptor cell loss, leading to severe vision impairment or blindness. However, no effective therapy has been established to date. Although genetic mutations have been identified, the available clinical data are not always sufficient to elucidate the roles of these mutations in disease pathogenesis, a situation that is partially due to differences in genetic backgrounds. RESULTS: We generated induced pluripotent stem cells (iPSCs) from an RP patient carrying a rhodopsin mutation (E181K). Using helper-dependent adenoviral vector (HDAdV) gene transfer, the mutation was corrected in the patient's iPSCs and also introduced into control iPSCs. The cells were then subjected to retinal differentiation; the resulting rod photoreceptor cells were labeled with an Nrl promoter-driven enhanced green fluorescent protein (EGFP)-carrying adenovirus and purified using flow cytometry after 5 weeks of culture. Using this approach, we found a reduced survival rate in the photoreceptor cells with the E181K mutation, which was correlated with the increased expression of endoplasmic reticulum (ER) stress and apoptotic markers. The screening of therapeutic reagents showed that rapamycin, PP242, AICAR, NQDI-1, and salubrinal promoted the survival of the patient's iPSC-derived photoreceptor cells, with a concomitant reduction in markers of ER stress and apoptosis. Additionally, autophagy markers were found to be correlated with ER stress, suggesting that autophagy was reduced by suppressing ER stress-induced apoptotic changes. CONCLUSION: The use of RP patient-derived iPSCs combined with genome editing provided a versatile cellular system with which to define the roles of genetic mutations in isogenic iPSCs with or without mutation and also provided a system that can be used to explore candidate therapeutic approaches.

Epiplakin1 is expressed in the cholangiocyte lineage cells in normal liver and adult progenitor cells in injured liver.

We have previously identified Epiplakin1 (Eppk1) as a gene expressed in pancreatic progenitor cells. Here we studied the expression of Eppk1 in developing and regenerating livers in mice. Eppk1 is initially expressed in the early bipotential hepatoblasts and is later confined to the cholangiocytes. After birth, Eppk1 is expressed in the bile duct. In the livers of mice fed with a choline-deficient ethionine-supplemented (CDE) diet, Eppk1-positive cells dramatically increase in number. The Eppk1-positive cells express A6, thereby indicating that they are hepatic progenitor cells. Other cholangiocyte markers, such as Cytokeratins, E-cadherin, osteopontin and Sox9, are also co-expressed in the hepatic progenitor cells. Some of the Eppk1-positive cells express PCNA, a proliferation marker, thereby suggesting their identities as transient amplifying cells. In conclusion, we have shown that Eppk1 serves as a useful marker for detecting the hepatic progenitor population in the developing and adult liver. The use of Eppk1 as a marker will facilitate studies of mouse hepatic progenitor cells.