Molecular Biotechnology: Principles and Applications - Glick, B. R., & Pasternak, J. J. 2002

Molecular Biotechnology of Microbial Systems
Transgenic Animals
Transgenic Sheep, Goats, and Pigs

Transgenesis experiments in sheep and goats have primarily aimed to turn the Mammary Glands of these animals into biological bioreactors for producing medicinal protein products. Although sheep and goats yield less milk than cows, they still produce hundreds of liters annually. Using a method analogous to that used for generating Transgenic Mice and employing transgenic constructs that contain human genes under the control of mammary gland-specific promoters (Table 19.2), researchers successfully produced transgenic sheep and goats whose milk secreted human Proteins. These proteins were glycosylated and exhibited an activity comparable to that of their human-derived counterparts; however, further studies are required to confirm their complete equivalence. Transgene expression in the mammary gland Cells of sheep and goats caused no adverse effects on either lactating females or their nursing offspring. In contrast, introducing a bovine Growth Hormone transgene under the control of a metallothionein promoter into pigs did result in adverse effects. While hormone levels varied among individuals within the transgenic pig group, the group as a whole showed faster weight gain. Unfortunately, this positive outcome was partially offset by various pathologies, including gastric ulcers, renal failure, lameness, pericarditis, reduced joint mobility, and a predisposition to Pneumonia. The causes of these symptoms remain unknown, though they may be associated with the long-term presence of excess growth hormone in the body, as the transgene was synthesized more or less continuously in these experiments. Transgenic sheep with an increased wool growth rate have also been successfully generated. To achieve this, the cDNA for sheep Insulin-like growth factor 1 was placed under the control of a high-sulfur mouse keratin Gene promoter to ensure cDNA overexpression. Unlike the pigs, these transgenic sheep exhibited no undesirable side effects. Positive results have likewise been achieved in experiments with transgenic pigs. For instance, healthy transgenic pigs were produced carrying the following genetic construct: the regulatory region of the human ß-globin gene, two human a₁-globin genes, and one human ß^A-globin gene. As a result of its expression in porcine Blood Cells, human Hemoglobin was synthesized. Furthermore, substituting the human ß-globin promoter with a porcine promoter resulted in significantly higher yields of human hemoglobin. The human hemoglobin produced by these transgenic pigs shared the same chemical properties as natural human hemoglobin and could be isolated from porcine hemoglobin using standard Chromatography.

Class="center">Table 19.2. Transgenic constructs containing human genes under the control of mammary gland-specific promoters and recipient organisms

These results point to the feasibility of replacing whole blood used in transfusions with human hemoglobin produced via transgenesis. However, isolated hemoglobin does not transport oxygen as efficiently as hemoglobin encapsulated within red blood cells. Moreover, it is rapidly degraded within the recipient animal's body, and its breakdown products are toxic to the Kidneys. Consequently, developing a human blood substitute through transgenesis remains a distant prospect.

Recently, considerable attention has been focused on using animal Organs for xenotransplantation into humans. The primary challenge in interspecies transplantation is hyperacute rejection, which is triggered when host Antibodies bind to carbohydrate antigenic determinants On the surface of the transplanted organ's cells. These bound antibodies provoke a severe inflammatory response (activation of the Complement cascade), leading to massive destruction of the antibody-bearing cells and rapid loss of the graft.

Under natural conditions, the inflammatory response is blocked by specialized proteins on The surface of cells lining the blood vessel walls. These complement-inhibiting proteins are species-specific. It was hypothesized that if a donor animal carried one or more genes for a human complement-inhibiting protein, the transplanted organ would be protected against the initial inflammatory response. To this end, transgenic pigs carrying various human complement inhibitor genes have been generated. Cells from one such animal proved completely resistant to the Components of the complement cascade system. Preliminary experiments involving the transplantation of organs from transgenic pigs into primates demonstrated that graft Tissues sustained less damage and the organs survived slightly longer without rejection. Ultimately, transgenic pigs that carry a human complement inhibitor gene and lack the major porcine Cell surface protein responsible for hyperacute rejection may serve as a viable source of organs for human transplantation.