June 8, 1998
Volume 50, No. 33
Emory researchers employ
Using tests described as the new "gold standard" for measuring specific immune system responses to viruses, School of Medicine researchers have found the response of certain cells that provide the front line of protection against invading viruses is considerably larger and much more targeted to specific viral antigens than scientists previously believed.
Although it is well known that T cells multiply in the body in response to viral infection, it has been difficult to measure the number of cells and exactly what they are responding to. The most common test used to measure the response, the limiting dilution analysis (LDA), has led scientists to believe that only 1 to 5 percent of these activated cells, named CD8+, are responding to specific viral antigens, and that most of the T cell response is a general, non-specific one.
Emory Vaccine Center Director Rafi Ahmed, along with Kaja Murali-Krishna, a postdoctoral dellow in Ahmed's laboratory, and John Altman, assistant professor of microbiology and immunology, used three novel tests to examine the CD8+ T cell response to lymphocytic choriomeningitis virus in mice. They found that more than 70 percent of the activated CD8+ T cells were responding to specific antigens of the virus-a number they termed "remarkably high," 20 to 100 times greater than numbers based on LDA testing.
Based on these results, the Emory investigators calculated that virus-specific CD8+ T cells can expand more than 30,000-fold (about 15 cell divisions) in one week with an estimated doubling time of six to eight hours during the peak phase of the response. "The size of the antiviral response is much greater than our previous estimates and radically changes our perception of the T cell response to viruses," said Ahmed.
The research was reported in the journal Immunity.
Nobel laureate Peter Doherty commented in the April 10 issue of Science that the new findings have "triggered a revolution" in our understanding of virus-specific T cell responses by "finally developing accurate methods" to measure these responses. "This research should prove very important for vaccine development because you want a vaccine to induce a good response, and this research allows us to see directly what a good response is," Ahmed noted.
He believes the findings about virus-specific responses will most likely apply to viral infections in general, including infectious mononucleosis, HIV and human T cell leukemia virus type 1, all of which are characterized by massive increases in the number of CD8+ T cells. "It is likely that most of the expanded CD8+ T cells in these infections are specific to those particular viruses also," Ahmed said.
Altman's initial findings have confirmed this prediction for HIV infection, and he continues to actively pursue investigations of the CD8+ T cell response to HIV in chronically infected patients on antiretroviral therapy. One of the tests used in the study-tetramer staining-was developed by him while a researcher at Stanford University. "The beauty of the tetramer technology is that it can be applied to study T cell responses to any pathogen, from influenza to HIV to malaria," Altman said. "And tetramers give you both a more accurate and a much more rapid picture of the immune response," he explains. In his Science article, Doherty said tetramer staining is "set to be the gold standard for quantifying virus-specific CD8+ T cells."
The Emory researchers also tested the CD8+ T cell response using sensitive tests that measure the production of interferon-gamma, a small protein secreted from a cell and produced by virus-specific T cells, that is important in controlling the infection. The interferon-gamma test also demonstrated a strong, virus-specific response by CD8+ T cells.
Studies using the LDA technique have underestimated the virus-specific T cell response, explained Ahmed, partly because it requires culturing the cells for one to two weeks. Only T cells capable of dividing and surviving for that long are counted. Since activated T cells are prone to die when restimulated with viral antigens, many of the antigen-specific T cells either die or do not divide enough to be counted. After the initial CD8+ T cell viral response, about 95 percent of the T cells die and the rest become memory cells poised to respond to future invasions by the same virus. The tests used in the Emory study were also able to detect memory CD8+ T cells specific to the lymphocytic choriomeningitis virus more than one year after infection.
"One of the hardest things to do is to identify T cells responding to a particular pathogen," Ahmed said. "This work greatly revises our thinking on the dynamics of T cell activation and will lead us to re-examine our models of viral-induced T cell proliferation."