Project description:<p>RNA sequencing (RNAseq) of peripheral blood lymphocytes was used to develop a means to assess immune function in a way that can be used in discovery science and applied to patients individually in clinical settings. The premise is that profiles of RNA present in immune cells is reflective of the combined influence of genetic and environmental variation on immune potential of individuals and that this potential can be tapped to understand human immunity in a variety of biological contexts. CD4+ cells were isolated from fresh whole blood via positive magnetic bead selection and cell lysates were prepared using Qiazol (QIAGEN) and stored at -80ºC for 3 to 8 months. RNA was extracted in batches for cDNA library preparation and RNA-Seq. For this study, we developed standard operating procedures for handling human blood samples and determined: a) the best way to enrich for CD4+ T cells from whole blood and yield high quality RNA, b) the sensitivity of this RNA profiling strategy, and c) the reproducibility of generated immune profiles from healthy subjects. We then developed bioinformatics processes to establish immune response signatures and immune response phenotypes within cohorts of individuals.</p>
Project description:Natural killer (NK) cells are leukocytes of the innate immune system and play a central role in the control of cancer metastases. NK cells and other innate immune cells often do not function well in patients with cancer and are also profoundly suppressed after cancer surgery. Dr. Auer’s Lab and others have shown that NK cells are critically important in the clearance of tumor metastases and that their impairment can be recovered with immune therapy augmenting the innate immune system. Several studies suggest that cancer patients have depressed NK cell cytotoxicity as compared to healthy controls but that following resection of the cancer, NK cell cytotoxicity returns to normal levels. In this observational study, the investigators will measure NK cell cytotoxicity by the gold standard method (51Cr, a chromium51 release assay) and by a new interferon-ɣ (IFN-ɣ) based assay (NK-Vue) in healthy humans and colorectal cancer (CRC) surgery patients seen a The Ottawa Hospital. The results of this study will determine if the NK-Vue is able to discriminate between healthy human volunteers and newly diagnosed cancer patients and is sufficiently sensitive to detect transient NK cell suppression immediately following surgery.
Project description:Immune cell-specific expression is one indication of the importance of a gene's role in the immune response. In order to identify such patterns, we set out to broadly profile gene expression in a variety of immune cells. We isolated twelve different types of human leukocytes from peripheral blood and bone marrow, treated them to induce activation and/or differentiation, and profiled their gene expression before and after treatment. The twelve cell types are: B cells, CD14+ cells, CD4+ CD45RO+ CD45RA- T cells, CD4+ T cells, CD8+ T cells, IgG/IgA memory B cells, IgM memory B cells, Monocytes, NK cells, Neutrophils, Plasma cells from bone marrow, and Plasma cells from PBMC.
