This work is an ESA/NASA collaboration between Professor Augusto Cogoli and Professor Millie Hughes-Fulford to study the role of the interleukin-2 receptor in signal transduction of T cells.  The nearly total loss of T cell activation in real microgravity was discovered in a Spacelab in 1983 and was confirmed later by several experiments in space. Thereby, cells appear to undergo apoptosis. Inhibition of IL-2R expression is probably one of the causes of such loss of activation. It is planned to analyze the pathway of mitogenic signal transduction in T cells in true microgravity as well as at variable G levels between 0 and 1xg.

The objectives of this research project are:

§ To investigate selected critical steps of T cell activation

§ To test the hypothesis that a failure in the expression of the IL-2R is   causing the loss of activity in microgravity

§  To identify windows of sensitivity to gravity during the activation process

     The main experimental approach will consist of the activation of cultures of purified peripheral blood cells with T cell mitogens in real and simulated microgravity. The specific IL-2R mRNA will be quantitatively determined with the reverse Transcriptase-polymerase chain reaction (rt-pcr) technology, the insertion of IL-2R in the membrane will be visualized by immunofluorscence and its secretion in the supernatant will be measured by immunoassay.  Signal transduction will be furthered analyzed by determining the early expression of oncogenes.  The Laboratory of Cell Growth (LCG), headed by Dr. Millie Hughes-Fulford is responsible for analysis of gene expression in T cells.

       The STS-107 mission is scheduled for late 2002. In this study, in collaboration with Dr. Augusto Cogoli,  purified human T-cells are flying on the shuttle, the cells are being activated to grow with Conconnavalin A (Con A) and anti-CD-28 while in flight.

Early studies in the Apollo and Skylab eras discovered that lymphocytes from returning astronauts did not activate properly. The logical question was: do lymphocytes change in microgravity, or is this a systemic effect of flight?  Dr. Cogoli, a pioneer in space cell biology, established that isolated lymphocytes do not activate properly in the absence of  gravity. Over the years, he has narrowed down the mechanism to being a defect in the IL-2 signaling. In this study, we are examining the effect of gravity on early gene expression of IL-2 and IL-2R subunits a, b and g. Cells will either have a 1-g force or will be at microgravity.

Progress in 2002 includes identification of kinase signaling pathways in early T-cell activation. Major pathways studied included JNK, p38, PI-3K, AKT and ERK1/2. In addition we are finishing the manuscript describing signaling pathways and the temporal induction of gene expression of IFN-g, IL-2, IL-2Ra, IL2Rg, and control genes.

In addition we have

1.      Continued Hardware testing including quality testing of syringes and Biopack containers.

2.      Prepared EST solution preparation and successfuly analyzed EST samples.

3.      Completed documentation required for Cape and for shipping of samples from Zurich/Cape

4.      Prepared GTIC/ RNA solutions and shipment of equipment to the Cape (solutions will be remade for the Jan 16^th launch).

5.      We have trained the second person for duties at the Cape and for final preparation and documentation of flight operations as well as postflight analysis of the samples.

6.       We have published or have in press 2-3 publications 2002 relating to microgravity and space science studies (see bibliography).

7.      In addition, we have continued to work on ground-based studies of T-cell activation in microgravity.

We have started preparation for gene chip technology analysis of RNA from space activated T-cells in addition to our semi-quantitative analysis of IFN-g, IL-2, IL-2Ra, IL2Rg, and control genes.   The objectives of this research project are: 1) To investigate selected critical steps of T cell activation   2) To test the hypothesis that a failure in the expression of the IL-2R is causing the loss of activity in microgravity  3) To identify windows of sensitivity to gravity during the activation process 4) To establish thresholds of gravi-sensing T cells. 

   The main experimental approach will consist of the activation of cultures of purified peripheral blood cells with T cell mitogens in real and simulated microgravity. The specific IL-2R mRNA will be quantitatively determined with the reverse Transcriptase-polymerase chain reaction (RT-PCR) technology. Signal transduction will be furthered analyzed by determining the early expression of oncogenes. The Laboratory of Cell Growth (LCG), headed by Dr. Millie Hughes-Fulford is responsible for analysis of gene expression in T cells. These studies will add to our knowledge of basic immunology mechanisms of T-cell activation both in space and here on earth.