July 9th, 2002
Johns Hopkins Medical Institutions (http://www.hopkinsmedicine.org/)
Johns Hopkins School of Medicine researchers have what is believed to be the first solid evidence that genes in human pluripotent stem cells and their progeny work normally.
The focus of the new study, conducted in the lab of Andrew Feinberg, M.D., was whether Hopkins-developed human embryonic germ cells and their more specialized "daughter" cells would have correct "imprinting," a way cells determine which of the two copies of a gene to use in making proteins. For imprinted genes, which copy is active depends only on which parent it came from.
The scientists showed that imprinting remains normal as the embryonic germ cells, a kind of pluripotent stem cell from fetal tissue, become more specialized, says Feinberg. Earlier reports by researchers studying mouse stem cells had raised concerns that those from humans might not be properly imprinted, casting some doubt on their value for cell-based therapies, he adds.
"Imprinting is normal in human embryonic germ cells," says Feinberg, King Fahd Professor of Medicine in the McKusick-Nathans Institute of Genetic Medicine at Hopkins. "Researchers still have to figure out the logistics and other critical aspects of stem cells to advance to clinical applications, but this work shows that imprinting is not going to be the issue some had anticipated."
Their report, to be published online during the week of July 8 in the Proceedings of the National Academy of Sciences, is likely to cheer researchers hoping that these cells or specialized cells obtained from them will someday be safe and useful for treating diseases.
"It's reassuring to learn that the cells we have derived from primordial germ cells are imprinted normally. This is critical information for their possible safe clinical use in the future," says stem cell pioneer and study co-author John D. Gearhart, Ph.D., the C. Michael Armstrong Professor at Johns Hopkins Medicine and professor of gynecology and obstetrics.
Because only parent-of-origin controls expression of an imprinted gene, when one is passed, say, from a mother to her son to his daughter, the biochemical "mark" that identifies the gene as a maternal or a paternal copy must be removed and a new mark put on. If parental marks were lost in cultured stem cells, as some scientists had proposed was likely, both copies of the imprinted gene would be active.
"The inheritance rules Mendel observed in his pea garden aren't operating here," explains Feinberg. "'Dominant' and 'recessive' don't explain imprinted genes. Instead, for some imprinted genes, only the copy from the mother is used, while for others, only the copy from the father is turned on -- generation after generation."
Believed to be the first to study imprinting in human stem cells, Feinberg and the research team found that the primitive germ cells themselves lacked active imprinting of the four genes studied, but specialized daughter cells had appropriate expression of one copy over another.
"This makes sense," says first author Patrick Onyango, a post doctoral fellow in the McKusick-Nathans Institute, "because some genes are imprinted in the whole organism, and some only in particular tissues. It's probably efficient for a stem cell to have both copies turned on, and then reactivate imprinting to shut off one copy, depending on what type of specialized cell it becomes."
Feinberg and other researchers in the field hypothesize that imprinted genes carry "latent" marks in certain situations -- such as in pluripotent stem cells and in cancers that have both copies of an imprinted gene improperly active.
"In these instances, we've observed that an apparent loss of imprinting can be reversed, either by the cell itself or by giving a drug, in the case of cancer," says Feinberg. "So these imprinted genes, we believe, haven't completely lost the mark that says whether the copy came from mom or dad -- it's still there, it's just not active."
Unfortunately, the idea can't be tested with the human cells, so Feinberg's team developed a mouse model to study the mechanics of imprinting in differentiating cells and developing organisms. By mating males of one kind of mouse with females of another, the researchers created a set of four mouse embryonic germ cell lines in which it's easy to determine whether a gene copy is from the mother or father. With these cells, the researchers can study and manipulate imprinting status, and even see how imprinting changes as the cells differentiate.
Other authors on the paper are Shan Jiang, Hiroshi Uejima, Michael Shamblott and Hengmi Cui, all of The Johns Hopkins University School of Medicine.
Funding for the work was provided by the National Institutes of Health. Funding for the derivation of the human embryonic germ cells used in this study was provided by Geron Inc. (Menlo Park, Calif.). Under a licensing agreement between Geron and The Johns Hopkins University, the University, Shamblott and Gearhart are entitled to a share of sales royalty from Geron. The University, Shamblott and Gearhart own stock in Geron, the sale of which is subject to certain restrictions under University policy. The terms of this arrangement are being managed by the University in accordance with its conflict of interest policies.
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