Creating Healthy, Long-Living Cloned Animals

For this article, James Kling interviewed Eric W. Overström, associate professor biomedical sciences, Tufts University School of Veterinary Medicine, North Grafton, Mass. Data from the Web of Science (ISI, Philadelphia) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age. A. Baguisi, E. Behboodi, D.T. Melican, J.S. Pollock, M.M. Destrempes, C. Cammuso, J.L. Williams, S.D. Nims, C.A. Porter, P. Midura, M.J. Palacios, S.L. Ayres, R.S, Denniston, M.L. Hayes, C.A. Ziomek, H.M. Meade, R.A. Godke, W.G. Gavin, E.W. Overstr?m, Y. Echelard, "Production of goats by somatic cell nuclear transfer," Nature Biotechnology, 17[5]:456-61 May 1999. (Cited in 88 papers) One of the most difficult challenges in producing cloned animals is producing healthy adults. Abnormalities vary with the species. Cattle, for example, often suffer at birth from immature respiratory systems, and are frequently born late, leading to problems with delivery.1,2 The survival rate in cloning attempts also is low: generally only one percent to five percent of embryos typically survive to term, says Eric W. Overström, associate professor, biomedical sciences, Tufts University School of Veterinary Medicine. Most attempts to solve those problems have focused on the nuclear donor cell, which is fused with an egg cell that has had its nucleus removed. "What has come to pass is that whether you take a cell from an adult, from skin, from liver, muscle, or a variety of differentiated tissue types... they can contribute to the formation of a clone, albeit at low efficiency," Overström says. "There was this original thinking that a 'silver bullet' cell type may exist that would be substantially more adept at giving rise to successful cloned offspring. To date, however, there is little evidence to suggest that that is the case." He and his team began to suspect that the state of activation of the egg cell could be a key to improved success. Overström and colleagues bred a transgenic goat with the goal of accessing antithrombin III (ATIII), a protein normally found in human plasma that helps regulate blood clotting, from the animals' milk. "Our thinking was that when you recombine the egg and donor cell and activate it, invariably only a small [percentage of the recombined cells] will begin to move forward," he says. "There appeared to be a bottleneck ... . The donor cells were active, but we were putting them into an inactivated cytoplasm [of the egg]. If we actually activated the egg first--tricked it into thinking that fertilization had taken place--the cytoplasm would (undergo) changes and we thought that may afford greater synchronicity between that cytoplast and donor cell and thereby improve cloning success rates." To produce the cloned embryos, they used fetal cells that were 35 to 40 days old. "We improved the efficiency, and perhaps as important, we produced three live animals that are completely healthy," says Overström. When the animals reached maturity, the team then isolated the ATIII protein from the milk and confirmed its biological activity. "It was the first time, to my knowledge, that a cloned transgenic animal was produced, that the protein was specifically expressed in the milk of an animal, and that it was isolated and demonstrated to be both biologically pure and biologically active," says Overström. Ian Wilmut's group at the Roslin Institute in Scotland had earlier described the production of a cloned, transgenic sheep that expressed a human blood factor, but they did not isolate it, purify it, and characterize its biological activity, he says. According to Overström, this Hot Paper was the first to demonstrate the feasibility "of producing biologically active pharmaceuticals for the treatment of human and animal disease." Overström's lab continues investigating how the state of the egg cell affects cloning success. Overström says only "incremental improvements" have been made since this paper came out. "There haven't been any further developments that jump efficiency from 5 percent to 35 percent, say, and without related health issues. But I'm confident that in the very near future such reports will start to appear, because we know more about how these cellular processes occur." James Kling can be contacted at jkling@nasw.org. References 1.J.P. Renard et al., "Lymphoid hypoplasia and somatic cloning," Lancet, 353:1489-91, 1999. 2.J.R. Hill, et al., "Clinical and pathological featuresof cloned transgenic calves and fetuses (13 case studies)," Theriogenology, 51:1451-65, 1999.

Creating Healthy, Long-Living Cloned Animals

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