Katrin Hinrichs, DVM, PhD, Dipl. ACT, is a professor and the Patsy Link Chair in Mare Reproductive Studies in the Department of Physiology and Pharmacology at Texas A&M's College of Veterinary Medicine and Biomedical Sciences.
The first equine clones were born in 2003, and now advertisements for cloning can be found in equine magazines, and the AQHA has considered registering cloned foals. There are more than 40 live cloned horses. What have we learned and what concerns do we have?

Foal Health Cloned foals do not seem to suffer the same abnormalities seen in cloned calves and lambs, in which "large offspring syndrome" results in oversized animals and organ system problems.

Our laboratory at Texas A&M has reported on the health of the 14 live foals we have produced. About half of the cloned foals had some issues at birth; the major problems were weakness and maladjustment, contracted tendons in the front legs, and enlarged umbilical remnant, some of which required surgical removal. Two of the foals died, one from pneumonia and one from complications after anesthesia. The remaining foals resolved their problems within a few weeks. Aside from one foal that had bladder stones in its first year, to our knowledge all cloned horses are currently healthy.

Royal Blue Boon clone

Royal Blue Boon (right) and her clone, Royal Blueboon Too, pictured in 2006. Royal Blue Boon died earlier this month.

What is causing problems with cloned foals at birth? Most likely differences in gene expression (how the genes are being used) in the cloned foals. During cloning, the DNA of a skin cell from the donor animal is placed in a host egg (oocyte). The oocyte has to change all the "on/off" instructions attached to this DNA so the DNA can start to express the appropriate genes to make an embryo.

Sometimes these changes are not exactly correct. The problems we have seen at birth seem related to incorrect placental function, which is controlled by a complicated genetic system. There may also be some changes in gene expression with the foal itself; luckily, when the cloned animal makes sperm or eggs, these genetic "on/off" switches are reset, so cloned animals' offspring should be completely normal.

Premature Aging? Dolly the sheep, the first clone of an adult mammal, raised questions as to whether a cloned animal will age prematurely. In Dolly's cells, the chromosomes had shorter caps (telomeres) than expected for her age. Each time the chromosomes replicate during cell division, the telomeres shorten, so old animals have shorter telomeres than young animals. The length of the telomeres is reset in the egg or sperm, so the next generation starts off with long telomeres.

Unfortunately, we did not find out whether Dolly would age prematurely, as at age 6 she contracted a lung virus (along with many of her barnmates) and was euthanized. Studies in cattle and other species showed telomere length was related to the tissue used for cloning: use of mammary gland cells (the cell type used to clone Dolly) and epithelial cells resulted in calves with shorter telomeres, whereas use of fibroblasts (skin connective tissue cells) and other cell types resulted in normal telomere length. Since fibroblasts are used for equine cloning, it is unlikely that telomere length will be a concern.

Mitochondrial DNA Mitochondria are small organelles found in the cytoplasm of every mammalian cell. Mitochondria possess their own tiny genome, coding for about 13 of the estimated 3,000 proteins involved in their function (in comparison, the DNA in the cell nucleus codes for around 50,000 proteins). Mitochondria are inherited along the maternal line: the mitochondria of the oocyte will be the mitochondria of the resulting offspring. When nuclear transfer is performed, the oocyte used for cloning is the major source of mitochondria, although some mitochondria from the donor cell can be present. Therefore the cloned foal will have the nuclear DNA of the genetic donor, but the mitochondrial DNA of the host oocyte, or possibly a mixture of the two.

The impact of this is currently unknown. While diseases associated with abnormal mitochondria have been identified in some species, these are rare and the host oocyte probably has about the same chance of carrying abnormal mitochondria as does the donor animal.

The fact that cloned mares will pass down the host mitochondria is a concern to breed registries. Research is under way to define the amount of mitochondrial variability in different horse breeds.

Originally published October 2009.