Photo courtesy of University
of Texas at El Paso
Researchers say they have developed a protocol to prepare human induced pluripotent stem (hiPS) cells using chemically fixed feeder cells.
This method saves time and money by avoiding the need for colony formation of live feeder cells, which is required by current conventional methods.
The new protocol challenges the theory that live feeder cells are required to provide nutrients to growing stem cells.
“We’ve proved an important phenomenon,” said Binata Joddar, PhD, of the University of Texas at El Paso. “And it suggests that these feeder cells, which are difficult to grow, may not be important at all for stem cell growth.”
Dr Joddar and her colleagues described the phenomenon in Journal of Materials Chemistry B.
Using 2.5% glutaraldehyde (GA) or formaldehyde (FA) for 10 minutes, the researchers prepared a niche matrix from autologus human dermal fibroblast (HDF) feeder cells.
They then introduced hiPS cells to the niche matrix, which adhered to and were maintained as colonies on the fixed feeder cells.
The colony doubling times of the cells grown this way were similar to those of hiPS cells grown on mitomycin-C-treated HDF or SNL feeders. (SNL cells are derived from mouse fibroblast STO cells transformed with a neomycin resistance gene.)
But the colony doubling time for the hiPS cells was shorter with the fixed feeder than for cells cultured on laminin-5.
The researchers also discovered that the average number of colonies per passage was signficiantly higher for hiPS cells cultured on fixed feeder cells compared to those cultured without feeders.
They noted hiPS cells cultured on gelatin did not grow beyond the first passage.
The team concluded that the two types of chemically fixed HDF feeder cells (HDF-glutaraldehyde and HDF-formaldehyde) can be used as substitutes for mitomycin-C-treated HDF feeders to culture hiPS cells.
This new method would not extend the doubling time, would save preparation time, and would avoid labor-intensive protocols to prepare.
In addition, after chemical fixation, the feeder cells are non-viable and cannot release active growth factors or chemokines into the cell culture. Therefore, fixed feeder cells can be refrigerated for long-term storage prior to use.
“Because feeder cells don’t need to stay alive in the process, we can store them at room temperature and spend less time cultivating them,” Dr Joddar said.
“This makes me think that we [could] use a nanomanufacturing approach to grow stem cells. We could mimic feeder cells’ nanotopology with 3-D printing techniques and skip using feeder cells altogether in the future.”
