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Isolation of the first domestic hESC line in Japan.
Tuesday, 03 June 2003

Researchers in Japan have produced their first domestic human embryonic stem cell (hESC) line. Professor Norio Nakatsuji, of the Institute for Frontier Medical Sciences in Kyoto, told CellNEWS in an exclusive interview that his group had succeeded to isolate hESC’s from one single donated frozen human embryo. Establishing stem cell lines in Japan is important, Prof. Nakatsuji said, so that the cell’s quality and preparation for future clinical application can be assured under Japanese government guidelines.

Frozen and thawed fertilised human egg used for stem cell extraction. Photo by the courtesy of Norio Nakatsuji.

"It is necessary to have human ES cell lines established in Japan from the start, in closely controlled conditions and protocols, for assurance of the cell quality and preparation for future clinical application."

"Also, we distribute the cells free-of-charge and no claim of right for future patent and licensing, so that the user can carry out their research more freely," he said.

"The only regulation is that their research plan to use human ES cells must be approved by government committee’s. These procedures are all prescribed in the Japanese government guidelines for establishment and usage of human ES cells."

The group was very successful in their attempts to grow human stem cells. This, Prof. Nakatsuji explains, could not have been done without their previous experience isolating and growing ESCs from cynomolgus monkey’s (Macaca fascicularis) — known as the crab-eating monkey with a more common name.

"So far, we could use less than ten frozen embryos of middle or lower grades because they were left over from infertility treatment. Of those, only two embryos started cell division after being thawed and cultured," Prof. Nakatsuji said.

The same human embryo at the blastocyst stage after 3 days in culture, used for isolation of hESC’s. Photo by the courtesy of Norio Nakatsuji.

"Of these two, only one reached the blastocyst stage, which fortunately resulted in one ES cell line."

"It had been impossible without our experience and method improvements using cynomolgus monkey blastocysts from which we have produced about 10 ES cell lines. Without it, it should have been impossible to establish one ES cell line from only one blastocyst that we have been able to use so far."

"Naturally, we will continue to produce more lines by using more frozen human embryos, which we can use in the coming weeks. Human ES cells are similar to monkey ones, but there are minor differences, so that we had to modify a little in the sub-culturing and freezing procedures."

Colony of hESCs grown on a feeder layer of mouse cells. Photo by the courtesy of Norio Nakatsuji.

However, the hESC line established this time was grown in the presence of mouse feeder cells, tells Prof. Nakatsuji. This makes them less usable for direct clinical applications.

"We plan to try to do so (grow hESC’s) with human feeder cells or without them in near future," says Prof. Nakatsuji.

When CellNEWS ask Prof. Nakatsuji what is the plan for the nearest future with these hESC lines, he outlines three points:

1. We will distribute ES cells to scientists who have obtained government approval to use human ES cells. It is our obligation from the government guideline and our research plan approved by government in April 2002. Our research has been supported by government funds.

Colony of hESCs at higher magnification. Photo by the courtesy of Norio Nakatsuji.

2. Establishment of cell with human feeder cells or without feeder cells, and search for growth factors for primate ES cells to maintain their pluripotency.
3. Improvement of genetically alteration methods of human ES cells. Gene transfection and gene targeting with homologous recombination to improve the properties of ES cells for research purposes and cell therapy.