Blocking an MS message

Using a mouse system that closely mimics the progressive demyelination found in multiple sclerosis (MS), a condition which afflicts an estimated 300,000 people in the US alone, researchers have developed a promising new form of immunotherapy that may help prevent patient relapses. The scientists, led by Abdolmohamad Rostami and Cris Constantinescu at the University of Pennsylvania (Philadelphia, PA), discovered that the concentration of the cytokine interleukin-12 (IL-12) increased in the mice during experimentally induced relapses. The team reasoned that "if the level of IL-12 was up during the relapses and acute attacks. . .we thought that if we could decrease the level, we might be able to prevent the relapse," explains Rostami. As predicted, mice injected with a monoclonal anti-IL-12 antibody were protected from relapses, and injecting purified IL-12 could induce a relapse. The antibodies did not appear to have any side effects on the mice, and a single injection appeared to provide long-term protection. The researchers, whose work appears in the November 1 issue of the Journal of Immunology (161:5097 –5104, 1998), are now developing similar anti-IL-12 antibodies for testing in humans.

The tet system just got better

The tetracycline (tet) regulatable system for gene induction/repression, while conceptually brilliant, has been fraught with technical difficulties, including high basal levels of expression in the absence of induction. Now Helen Blau and colleagues at the Stanford University School of Medicine have developed a reversible tet-inducible retroviral system (RetroTet-ART) in which activators and repressors are expressed together in cells. Guided by the crystal structure of tetR, a transrepressor was engineered with a dimerization domain distinct from the transactivator. When the transactivator and the transrepressor were expressed in vitro, no complexes containing both proteins were observed in gel retardation and immunoprecipitation studies. This allowed the transrepressor to bind and repress gene expression in the absence of dox, whereas the transactivator binds and induces gene expression only in the presence of dox. The RetroTet-ART system was then tested in cells by reversibly expressing p16, a protein known to arrest growth. In the presence of increasing dox, cell growth decreased. The growth arrest was reversed upon dox withdrawal. Helen Blau says, "The system is remarkably easy to use, as all the components can be introduced into cells in less than two weeks. It will prove a boon when it comes to expressing products toxic to cells, proteins that arrest cell growth, and regulating protein expression in gene therapy and transgenic animals." These findings are reported in Nature Genetics (20: 389–393, 1998).

Endangered egg surrogates

In work that has far-reaching implications for animal husbandry and the preservation of endangered species, researchers report in the October issue of Animal Reproduction Science (53: 265–275, 1998) a technique for transplanting cryopreserved ovarian tissue from elephants into mice. The tissue, collected in South Africa and frozen, was able to undergo normal development in immune-deficient mice, producing mature ovarian follicles and, in at least one case, an oocyte. In the past, female germplasm has been notoriously difficult to preserve, complicating many species conservation efforts. "As far as we can tell, this general procedure should work for any mammalian species except the egg-laying Monotremes. Certainly, we would hope that this procedure would be used for other endangered species," says John Critser, the scientific director of the Cryobiology Research Institute at Indiana University and senior author on the study. The scientists hope to fine-tune the system to reliably produce mature oocytes, which could then be used for in vitro fertilization in breeding programs. While emphasizing that several technical hurdles remain, Critser argues that "genetic material from rare and endangered species that would otherwise be lost can be preserved at this point [by freezing]."

Lilliputian lab

In an effort to make DNA analysis more streamlined and cost effective, a team of engineers and geneticists from the University of Michigan, Ann Arbor have developed a device that can perform enzymatic reactions on DNA and analyze the results in an integrated system the size of a single silicon chip. As reported in Science (Burns et al. 282:484–487, 1998), the device was used to digest and amplify a region of Mycobacterium tuberculosis DNA. The various systems required were built onto the chip using standard photolithographic techniques, in a process conceptually similar to stenciling. "The device is unique due to the fact that it uses complex integrated systems where they all work together simultaneously," says Dr. David Burke, a coauthor of the study. Included on the chip are a nanoliter injector, a sample mixing and positioning system, a temperature-controlled chamber, an electrophoretic separation system, and a fluorescence detector, at a final production cost of approximately six dollars each. Further development of this technology will allow DNA analysis to be performed "on site," whether that means a doctor's office, the rain forest, or the farm.

Taming the sindbis virus

For both cell culture and gene therapy, viruses have long been considered an efficient tool for introducing foreign genetic material into cells. Unfortunately, the viruses most suited to these jobs tend to kill the cells they infect, making them useful only for transient gene expression. To get around this problem, a team of researchers led by microbiologist Charles Rice at the Washington University School of Medicine (St. Louis, MO) has developed a noncytopathic strain of Sindbis, a lytic virus that is already widely used in transient expression systems. The scientists, whose work is described in the October issue of PNAS (95:12989– 12994, 1998), transfected cells with recombinant viral RNA containing a puromycin resistance gene, and then grew the cells in the presence of puromycin. Only cells carrying a noncytopathic mutant viral RNA could survive this double selection, and the resulting mutations were mapped and included in a new gene expression vector. The Washington University team is now focusing on using the system to express genes at physiological levels for experiments in basic cell biology and virology.