In aging, immune systems falter and the elderly are more susceptible to infection. Age related changes include decreased T-cell activation/proliferation, decreased production of IL-2 and decreased expression of IL-2R?. These are the same key immune changes that are seen in returning astronauts who have experienced altered immune function and increased vulnerability to infection during spaceflights. Loss of immune response in aging and spaceflight has been previously identified in T cells. We have previously identified microgravity induced changes in gene expression, promoter regions, transcription factors and signal transduction pathways in human CD4+ T-cells under normal and altered gravity conditions. Our preliminary data from International Space Station (ISS) described within this proposal show that in early T-cell activation at least one miRNA was upregulated during activation in normal gravity (g), while in microgravity (?g), the induction of the miRNA was muted. Moreover, three protein targets of the miRNA had increased expression in the 1.g environment suggesting a micro RNA mechanism. The changes in immune function in the elderly occur over time and are hard to evaluate. However, the same type of changes in immune response occurs microgravity, both in astronauts and in human T-cells and is easy to measure The central hypothesis is that ?g regulates MiRNA in human T-cells and that weightlessness is a novel model to study the immunosuppression seen in aging. These experiments will contribute to our fundamental understanding of the molecular mechanism of immunosuppression in spaceflight and in aging.
The Specific Aims are the following: (1) Identify gene expression of miRNAs during T-cell activation (using arrays and qRTPCR) under normal gravity. (2) Identify the MiRNA(s) target genes using bioinformatics and verify the changes in expression of those targets messages (qRTPCR). We will test the ability of microgravity to induce/reduce expression of MiRNAs and their targets and verify these results using miRNA overexpression viral constructs or dsRNA. (3) Analyze the protein synthesis of the upregulated/downregulated target genes (SDS PAGE gels and Western Blot proteomics) that are affected by T-cell activation under normal gravity and microgravity conditions. (4) Compare the expression of key MiRNAs candidates and genes after activation in normal gravity and microgravity, vs. that of the lymphocytes from an older population.
Identifying altered microRNAs and their targets in T-cells that have suppressed immune response in spaceflight but normal expression in 1g flight controls will contribute to our understanding of aging and will likely reveal important causes of immunosuppression in the elderly. In recent ISS experiments, one MiRNA is significantly downregulated in ?g flown T cells when compared to onboard 1g controls;with more efficient miRNA collection, several more dysregulated MiRNAs should be identified. Because there are considerable similarities between immunosuppression in astronauts during flight and immunosuppression in the elderly, we expect that our findings will be relevant to the mission of the NIH and be broadly interesting to researchers studying molecular mechanisms of the immune response.
|Chang, Tammy T; Hughes-Fulford, Millie (2014) Molecular mechanisms underlying the enhanced functions of three-dimensional hepatocyte aggregates. Biomaterials 35:2162-71|