Transcriptional, proteomic, and functional analysis of cytokine secreting plasmacytoid dendritic cells
Abstract/Contents
- Abstract
- Plasmacytoid dendritic cells (pDCs) are a rare subset of immune cells characterized by their ability to secrete large amounts of interferon alpha (IFN[alpha]) in response to Toll Like Receptor (TLR) activation by viral and endogenous nucleic acids. In these studies, we developed novel techniques that fundamentally change how cellular biologists can detect and assess protein and gene expression in cells based on function instead of phenotype. These techniques, which allow for transcript profiling of cells sorted by intracellular cytokine staining, and high throughput single cell detection of RNA transcript, were used to examine the role of pDC subsets that differ by IFN[alpha] and TNF secretion. Transcript profiling of these subsets revealed that after activation, there are two subsets of pDCs; cytokine secreting pDCs which express inflammatory and tolerogenic mediators, and quiescent noncytokine secreting pDCs, which maintain mRNA profile of unstimulated pDCs. Single cell RNAflow demonstrated that gene expression in subsets of pDCs from FLT3L DC (FLDC) cultures, bone marrow, and spleen correlates with cytokine staining. From our transcript profile data, we generated a list of candidate surface markers for cytokine secreting pDCs, which we screened by CyTOF and present here. Finally, we performed functional analysis of CD4- and CD4+ pDC subsets, and demonstrated that CD4- pDCs contain more activateable pDCs which take up antigen and induce antigen specific T cell proliferation. Taken together, these studies suggest that upon TLR activation, concomitant expression of rate limiting signaling genes leads to stochastic pDC expression of IFN[alpha] and TNF, leaving a larger pool of non cytokine secreting pDCs that maintain their pDC phenotype and expression pattern, and which may be activated at a later time. These studies also demonstrate the utility of screening based on function (cytokine secretion) instead of phenotype, which in this case, allowed for more complete assessment of variation between cells of different function.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2012 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Cheung, Regina Kar Wuen |
|---|---|
| Associated with | Stanford University, Department of Immunology. |
| Primary advisor | Utz, Paul |
| Thesis advisor | Utz, Paul |
| Thesis advisor | Engleman, Edgar G |
| Thesis advisor | Nolan, Garry P |
| Thesis advisor | Robinson, William (William Hewitt) |
| Thesis advisor | Steinman, Lawrence |
| Advisor | Engleman, Edgar G |
| Advisor | Nolan, Garry P |
| Advisor | Robinson, William (William Hewitt) |
| Advisor | Steinman, Lawrence |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Regina Kar Wuen Cheung. |
|---|---|
| Note | Submitted to the Department of Immunology. |
| Thesis | Ph.D. Stanford University 2012 |
| Location | electronic resource |
Access conditions
- Copyright
- © 2012 by Regina Kar Wuen Cheung
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
Also listed in
Loading usage metrics...