Conclusion

We end our many-chaptered exploration of the cellular and molecular biology of the immune response by highlighting studies revealing the choreography of immune cells in living tissue. Not only have these images and videos confirmed predictions made by immunologists who did not have the luxury to visualize the cells they were studying, but they have revealed unanticipated features of immune cells: the dramatic swarming behavior of neutrophils, the extensive dendritic processes of germinal center B cells, the dance of B- and T-cell pairs at the follicular boundary, the unusual appearance and activity of resident memory T cells in barrier tissues, to name just a few. We have concluded the chapter by introducing studies that are exploring the behavior of immune cells in pathologic contexts, the featured topic of the next set of chapters, which focus on our growing understanding of the role of immune cells in combating and even causing disease.

REFERENCES

Useful Websites

www.artnscience.us/index.html Immunologist Art Anderson has developed a lovely website that appreciates the importance of integrating excellent immunologic information with images.

https://www.youtube.com/watch?v=FzcTgrxMzZk “The Inner Life of the Cell” is an artistic and informative animation of the intercellular and intracellular molecular events associated with leukocyte extravasation. A shorter version is also available on Youtube (https://www.youtube.com/watch?v=wJyUtbn0O5Y).

www.nature.com/ni/multimedia/index.html This is an archive of images and videos of live immune cells from articles published in Nature and other Nature Publishing Group journals.

www.atlasofscience.org/the-thymus-a-small-organ-with-a-mighty-big-function/ An accessible and visually appealing description of the migratory patterns of T cells.

www.youtube.com/watch?v=XOeRJPIMpSs YouTube hosts many videos of immune cell behavior—in vitro and in vivo. This YouTube video, for instance, shows fluorescently labeled dendritic cells migrating through skin.

Video Links

Videos presented in this chapter can also be found by accessing the associated reference online. Note that some links work better with different browsers.

Video 14-Ov T lymphocytes migrate along the fibroblastic reticular cell network. Supplementary Movie 5 from Bajénoff, M., et al. 2006. Stromal cell networks regulate lymphocyte entry, migration, and territoriality in lymph nodes. Immunity 25:989.

https://dx.doi.org/10.1016/j.immuni.2006.10.011

Video 14-6v Two-photon imaging of live T and B lymphocytes within a mouse lymph node. Movie S3 from Miller, M. J., S. H. Wei, I. Parker, and M. D. Cahalan. 2002. Two-photon imaging of lymphocyte motility and antigen response in intact lymph node. Science 296:1869.

https://doi.org/10.1126/science.1070051

Video 14-7v Dendritic cells (DCs) are present in all lymph-node microenvironments. Supplementary Video 5 from Lindquist, R. L., et al. 2004. Visualizing dendritic cell networks in vivo. Nature Immunology 5:1243.

https://doi.org/10.1038/ni1139

Video 14-8v Lymphocytes exit the lymph node through portals. Supplementary Video 8 from Wei, S. H., et al. 2005. Sphingosine 1-phosphate type 1 receptor agonism inhibits transendothelial migration of medullary T cells to lymphatic sinuses. Nature Immunology 6:1228.

https://doi.org/10.1038/ni1269

Video 14-9v Neutrophils (red) swarming and monocytes (green) following. Video 1 from Lämmermann, T, et al. 2013. Neutrophil swarms require LTB4 and integrins at sites of cell death in vivo. Nature 498:371.

https://doi.org/10.1038/nature12175

Video 14-11v Malarial sporozoites travel to a lymph node and are engulfed by an APC. Movie S2 from Radtke, A. J., et al. 2015. Lymph-node resident CD8+ dendritic cells capture antigens from malaria, sporozoites and induce CD8+ T-cell responses. PLoS Pathogens 11:e1004637.

https://doi.org/10.1371/journal.ppat.1004637

Video 14-12v Dendritic cells crawl toward lymph nodes along lymphatic vessels. Movie S3 from Russo, E., et al. 2016. Intralymphatic CCL21 promotes tissue egress of dendritic cells through afferent lymphatic vessels. Cell Reports 14:1723.

https://doi.org/10.1016/j.celrep.2015.12.067

Video 14-13v B cells capture antigen from macrophages in the subcapsular sinus of the lymph node. Supplementary Movie 5 from Phan, T. G., I. Grigorova, T. Okada, and J. G. Cyster. 2007. Subcapsular encounter and complement-dependent transport of immune complexes by lymph-node B cells. Nature Immunology 8:992.

https://doi.org/10.1038/ni1494

Videos 14-14v1 and 14v2 Simulation of movements of DC and naïve lymphocytes in the absence (Movie 1) and presence (Movie 2) of antigen. From Catron, D. M., A. A. Itano, K. A. Pape, D. L. Mueller, and M. K. Jenkins. 2004. Visualizing the first 50 hr of the primary immune response to a soluble antigen. Immunity 21:341.

http://dx.doi.org/10.1016/j.immuni.2004.08.007

Video 14-14v3 T cells arrest after antigen encounter. Movie S10 from Celli, S., F. Lemaître, and P. Bousso. 2007. Real-time manipulation of T cell-dendritic cell interactions in vivo reveals the importance of prolonged contacts for CD4 T cell activation. Immunity 27:625.

http://dx.doi.org/10.1016/j.immuni.2007.08.018

Video 14-14v4 B cells travel to the border between the follicle and paracortex after being activated by antigen. Video S1 from Okada, T., et al. 2005. Antigen-engaged B cells undergo chemotaxis toward the T zone and form motile conjugates with helper T cells. PLoS Biology 3:e150.

https://doi.org/10.1371/journal.pbio.0030150

Video 14-14v5 Antigen-specific B and T cells interact at the border between the follicle and paracortex. Video S6 from Okada, T., et al. 2005. Antigen-engaged B cells undergo chemotaxis toward the T zone and form motile conjugates with helper T cells. PLoS Biology 3:e150.

https://doi.org/10.1371/journal.pbio.0030150

Video 14-15v1 Germinal center B cells differ in their behavior from naïve B cells. Movie S1 from Allen, C. D., T. Okada, H. L. Tang, and J. G. Cyster. 2007. Imaging of germinal center selection events during affinity maturation. Science 315:528.

https://doi.org/10.1126/science.1136736

Video 14-15v2 T follicular helper cells exiting a germinal center. Movie S1 from Shulman, Z., et al. 2013. T follicular helper cell dynamics in germinal centers. Science 341:673.

https://doi.org/10.1126/science.1241680

Video 14-17v The formation of a tricellular complex in a lymph node during CD8+ T-cell activation. Supplementary Movie 5 from Castellino, F., et al. 2006. Chemokines enhance immunity by guiding naïve CD8+ T cells to sites of CD4+ T cell–dendritic cell interaction. Nature 440:890.

https://doi.org/10.1038/nature04651

Video 14-23v A reticular network is established at the site of infection by Toxoplasma gondii. Movie S11 from Wilson, E. H., et al. 2009. Behavior of parasite-specific effector CD8+ T cells in the brain and visualization of a kinesis-associated system of reticular fibers. Immunity 30:300.

http://dx.doi.org/10.1016/j.immuni.2008.12.013

Video 14-24v Resident memory cells patrolling the dermis of mouse. Movie S1 from Ariotti, S., et al. 2012. Tissue-resident memory CD8+ T cells continuously patrol skin epithelia to quickly recognize local antigen. Proceedings of the National Academy of Sciences USA 109:19739.

https://doi.org/10.1073/pnas.1208927109

STUDY QUESTIONS

  1. You want to track the behavior of T cells specific for the influenza virus in a mouse lymph node. You have a mouse whose cells all express yellow fluorescent protein (YFP). You decide to isolate T cells from this mouse and introduce them into a mouse that has been immunized with influenza, as well as into a control mouse that was given no antigen. You look at the lymph nodes of both mice, expecting to see a difference in the behavior of the cells. However, you do not see much of a difference. At first you wonder if all you read is true—perhaps T cells do not arrest when they encounter antigen! But then you realize that your experimental design was flawed. What was the problem?
  2. Indicate whether each of the following statements is true or false. If you think a statement is false, explain why.
    1. Chemokines are chemoattractants for lymphocytes but not other leukocytes.
    2. T cells, but not B cells, express the chemokine receptor CCR7.
    3. Antigen can come into the lymph node only if it is associated with an antigen-presenting cell.
    4. Lymphocytes increase their motility after they engage dendritic cells (DCs) expressing an antigen to which they bind.
    5. T cells crawl along the follicular DC network as they scan DCs for antigen in the lymph node.
    6. Lymphocytes make use of reticular networks only in secondary lymphoid organs.
    7. Leukocyte extravasation follows this sequence: adhesion, chemokine activation, rolling, transmigration.
    8. All secondary lymphoid organs contain high-endothelial venules (HEVs).
  3. Provide an example of lymphocyte chemotaxis during an immune response.
  4. Describe where CD8+ T cells and B cells receive T-cell help within a secondary lymphoid organ.
  5. How might the behavior of an antigen-specific CD8+ T cell differ in the lymph node of a CCL3-deficient animal versus a wild-type animal?
  6. Extravasation of neutrophils and of lymphocytes occurs by generally similar mechanisms, although some differences distinguish the two processes.
    1. List in order the basic steps in leukocyte extravasation.
    2. Which step requires chemokine activation and why?
    3. Naïve lymphocytes generally do not enter tissues other than the secondary lymphoid organs. What confines them to this system?
  7. Naïve T and naïve B-cell subpopulations migrate preferentially into different parts of the lymph node. What is the basis for this compartmentalization? Identify both structural and molecular influences.
  8. True or false? Germinal center B cells differ in morphology and motility from other B cells in the follicle.
  9. Place a check mark next to the molecules that interact with each other.
    1. Chemokine and L-selectin
    2. E-selectin and L-selectin
    3. CCL19 and CCR7
    4. ICAM and chemokine
    5. Chemokine and G protein–coupled receptor
    6. BCR and MHC
    7. TCR and MHC
  10. Predict how a deficiency in each of the following would affect T-cell and B-cell trafficking in a lymph node during a response to antigen. How might they affect an animal’s health?
    1. CD62L
    2. CCR7
    3. CCR5
    4. S1P1
  11. Match the following type of cells with their location in the body. Note that (1) more than one cell type can be associated with a location and (2) a cell type might be found in more than one location.

    Cell types: Resident memory T cell, central memory T cell, effector T cell, plasma cell, naïve lymphocyte, dendritic cell, CD169+ macrophage, TFH cell

    Locations: Secondary lymphoid organs, barrier organs, T-cell zones of secondary lymphoid organs, sinuses of secondary lymphoid organs, bone marrow, follicle, blood, lymphatic vessels, brain

ANALYZE THE DATA

A paper (Gebhardt, T., et al. 2011. Different patterns of peripheral migration by memory CD4+ and CD8+ T cells. Nature 477:216) presented some of the first dynamic imaging data directly comparing the behavior of memory CD4+ and CD8+ T cells in response to infection. Their results were surprising.

Gebhardt et al.’s experimental strategy: The investigators infected the skin of mice with herpes simplex virus and adoptively transferred CD4+ T cells specific for the virus as well as CD8+ T cells, both of which expressed T-cell receptors specific for the virus. They observed the movements of these cells in the infected skin during the effector phase of the immune response (8 days after infection) as well as 5 or more weeks later when the only cells in the tissue would be memory cells. Remember that skin has two layers: an outer layer (epidermis) and an inner layer (dermis).

Their results: During the effector phase, both CD4+ and CD8+ T cells initially moved similarly through the dermis. However, gradually, these two cell populations distributed themselves differently: CD4+ T cells stayed in the dermis, CD8+ T cells moved to the epidermis (!).

The investigators then looked at the memory cell populations in infected skin weeks later. What they saw is depicted in the supplementary movie 4 (memory CD8+ T cells [red], memory CD4+ T cells [green]): www.nature.com/nature/journal/v477/n7363/full/nature10339.html#supplementary-information

Your assignment: Take a look at this time-lapse video and read its legend provided in the original paper.

  1. Describe what you see, identifying at least two specific differences between CD4+ and CD8+ T-cell behavior.
  2. Propose a molecular difference that could explain one of these distinctions.
  3. Advance one hypothesis about the adaptive value of the difference(s) you observed. That is, what advantage (if any) may such a difference in CD4+ and CD8+ T-cell behavior provide an animal responding to a skin infection?

EXPERIMENTAL DESIGN QUESTION

You want to directly test claims that CCR5 is important for the localization of naïve B cells to the follicles of a lymph node. You have all the reagents that you need to perform dynamic imaging in live tissue, including (1) an anti-CCR5 antibody that you know will block the interactions between CCR5 and its ligand (and can be injected), and (2) a CCR5-deficient (CCR5−/−) mouse. Design an experiment that will definitively test these claims. Define what you will measure, and sketch one figure predicting your results.

CLINICAL FOCUS QUESTION

Leukocyte adhesion deficiency I (LAD I) is a rare genetic disease that results from a defect or deficiency in CD18. Patients with this condition usually do not live past childhood because they cannot fight off bacterial infections. Why? Given what you have learned in this chapter, advance a proposal explaining how this disease impairs the immune system’s ability to fight bacterial infections. Be specific and concise. What approaches might be taken to treat this disease?