Purpose The development of a technique to induce the transformation of somatic cells to a pluripotent state via the ectopic expression of defined transcription factors was a transformational event in the field of regenerative medicine. The development of this technique also impacted ophthalmology, as patient-specific induced pluripotent stemcells (iPSCs) may be useful resources for some ophthalmological diseases. The lens is a key refractive element in the eye that focuses images of the visual world onto the retina. To establish a new model for drug screening to treat lens diseases and investigating lens aging and development, we examined whether human lens epithelial cells (HLECs) could be induced into iPSCs and if lens-specific differentiation of these cells could be achieved under defined chemical conditions.
Methods hLECs were separated and cultured from a 56-yr-old cataract patient, after cell reprogramming factors (OCT-4,SOX-2,KLF-4) transformation, yielding induced pluripotent stem (iPS) cells with high reprogramming efficiencies. For lens specific differentiation, we applied 3-step induction procedure with defined factors (Noggin, BMP and FGF2), and analysis of 7 lens differentiation markers (PAX6, SOX2, SIX3, CRYAB, CRYAA,, BFSP1, and MIP) in iPS cells and ES cells from d 0 to 30. In addition, we also compared expression of epithelial mesenchymal transition (EMT) markers among HLE-iPS-derived lens cells, human embryonic stem cells (hESCs) and fibroblast-derived iPSCs.
Results HLEC-derived iPS (HLE-iPS) cell colonies were indistinguishable from human ES cells with regards to morphology, gene pattern, pluripotent marker expression, and their ability to generate all embryonic germ layers in vitro and in vivo. Further more, HLE-iPS can be differentiated into large quantities of lens progenitor-like cells with defined factors (Noggin BMP, FGF2), and these cells expressed and accumulated lens-specific markers. Lens specific iPS cells have higher differentiation efficiency than that of ES. In addition, HLE-iPS-derived lens cells exhibited reduced expression of EMT)markers compared with hESCs and fibroblast-derived iPSCs.
Conclusions Our study describes a highly efficient procedure for generating lens progenitor cells from cataract patient HLEC-derived iPSCs. These patient-derived pluripotent cells provide a valuable model for studying the developmental and molecular biological mechanisms that underlie cell determination in lens development and cataract pathophysiology. |
