Novel Reprogramming Approach Applied to Generation of Cells for Retinal Regeneration
Researchers here report on the application of a novel form of cellular reprogramming that might be more useful than the present standard approach when it comes to generating patient-matched cells and tissues for regenerative medicine. They demonstrate this via the creation of vascular progenitor cells that might be used to treat some forms of retinal degeneration in which blood vessels in that tissue have atrophied.
Scientists began their experiments with a fibroblast - a connective tissue cell - taken from a person with type 1 diabetes. Reprogrammed fibroblasts function as stem cells, with the potential to give rise to all tissues in the body, including blood vessels. The team reprogrammed the fibroblast stem cells to revert to a state that is even more primitive than that of conventional human induced pluripotent stem cells - more like the state of embryonic cells about six days after fertilization. This is when cells are the most "naive," or more capable of developing into any specialized type of cell with a much higher efficiency than conventional human induced pluripotent stem cells.
Researchers used a cocktail mixture of two drugs that other scientists previously used to reprogram stem cells: GSK3β inhibitor CHIR99021, which blocks carbohydrate storage in cells, and MEK inhibitor PD0325901, an experimental anti-cancer drug that can block cancer cell growth. The team had also looked at the potential of a third drug, a PARP inhibitor - a popular anticancer drug used to treat a variety of cancers including those of the ovaries and breast. The team calls the cocktail 3i, named for the three inhibitors.
The research team tracked the reprogrammed stem cells' molecular profile, including measures of proteins such as NANOG, NR5A2, DPPA3 and E-cadherin that guide cell differentiation. That profile appeared similar to that found in so-called naive epiblast cells, the primitive cells that make up an approximately six day-old human embryo. The scientists also found that the stem cells reprogrammed with the 3i cocktail did not have abnormal changes in factors that can alter core DNA, called epigenetics, that typically plague other lab-made versions of naive stem cells. "Interestingly, the 3i cocktail appeared to erase disease-associated epigenetics in the donor cells, and brought them back to a healthy, pristine non-diabetic stem cell state."
Finally, the research team injected cells called vascular progenitors, which were made from the naive stem cells and are capable of making new blood vessels, into the eyes of mice bred to have a form of diabetic retinopathy that results from blood vessels closing off in the retina. They found that the naive vascular progenitors migrated into the retina's innermost tissue layer that encircles the eye, with higher efficiencies than have been reported with vascular cells made from conventional stem cell approaches. The naive vascular cells took root there, and most survived in the retina for the duration of the four-week study.